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Accurate Identification and Treatment of Emergent Cardiac Events

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Accurate Identification and Treatment of Emergent Cardiac Events

Author: Larry H Bernstein, MD, FCAP
In the immediately preceding article, I discussed the difficulties in predicting long-term safety for developing drugs, and the cost of failure in early identification.

It is not the same scale of issue as for the patient emergently presenting to the ED. Despite enormous efforts to reduce the development of and the complications of acute ischemia related cardiac events, the accurate diagnosis of the patient presenting to the emergency room is still, as always, reliant on clinical history, physical examination, effective use of the laboratory, and increasingly helpful imaging technology. The main issue that we have a consensus agreement that PLAQUE RUPTURE is not the only basis for a cardiac ischemic event. The introduction of high sensitivity troponin tests has made it no less difficult after throwing out the receiver-operator characteristic curve (ROC) and assuming that any amount of cardiac troponin released from the heart is pathognomonic of an acute ischemic event. This has resulted in a consensus agreement that

ctn measurement at a coefficient of variant (CV) measurement in excess of 2 Std dev of the upper limit of normal is a “red flag”
signaling AMI? or other cardiomyopathic disorder

This is the catch. The ROC curve established AMI in ctn(s) that were accurate for NSTEMI – (and probably not needed with STEMI or new Q-wave, not previously seen) –

ST-depression
T-wave inversion
- in the presence of other findings
- suspicious for AMI

Wouldn’t it be nice if it was like seeing a robin on your lawn after a harsh winter? Life isn’t like that. When acute illness hits the patient may well present with ambiguous findings. We are accustomed to relying on

clinical history
family history
co-morbidities, eg., diabetes, obesity, limited activity?, diet?
1. stroke and/or peripheral vascular disease
2. hypertension and/or renal vascular disease
3. aortic atherosclerosis or valvular heart disease
  - these are evidence, and they make up syndromic classes
Electrocardiogram – 12 lead EKG (as above)
Laboratory tests
1. isoenzyme MB of creatine kinase (CK)… which declines after 12-18 hours
2. isoenzyme-1 of LD if the time of appearance is > day-1 after initial symptoms (no longer used)
3. cardiac troponin cTnI or cTnT
  - genome testing
  - advanced analysis of EKG

This may result in more consults for cardiologists, but it lays the ground for better evaluation of the patient, in the long run. When you look at the amount of information that has to be presented to the physician, there is serious need for improvement in the electronic medical record to benefit the patient and the caregivers. Recently, we have a publication on a new test that has been evaluated, closely related to the C-reactive protein (CRP), a test that has generated much discussion over the effect of treatment for patients who have elevated CRP in the absence of increased LDL cholesterol, diabetes, or obvious atherosclerotic comorbidities. The serum pentraxin 3 test is related to cell mediated immunity, and an evaluation has been published in the Journal of Investigative Medicine.

Journal of Investigative Medicine Feb 2013; 61 (2): 278–285.
http://dx.doi.org/10.231/JIM.0b013e31827c2971

Serum Pentraxin 3 Levels Are Associated With the Complexity and Severity of Coronary Artery Disease in Patients With Stable Angina Pectoris
Karakas, Mehmet Fatih MD*; Buyukkaya, Eyup MD*; Kurt, Mustafa MD*; et al.
From the Departments of Cardiology and,Clinical Biochemistry, Mustafa Kemal University, Tayfur Ata Sokmen Medical School, Hatay, Turkey.
Reprints: Mehmet Fatih Karakas, MD, Antakya 31005, Turkey. E-mail: mfkarakas@hotmail.com.

Abstract
Background: Atherosclerosis is a complex inflammatory process. Although pentraxin 3 (PTX-3), a newly identified inflammatory marker, was associated with adverse outcomes in stable angina pectoris,

an association between PTX-3 and the complexity of coronary artery disease (CAD) has not been reported.

The aim of the present study is to assess

the association between the level of PTX-3 and
the complexity and severity of CAD assessed with
SYNTAX and Gensini scores in patients with stable angina pectoris.

Methods: The study population is 2 groups:

161 patients with anginal symptoms and evidence of ischemia
- who underwent coronary angiography and
50 age- and sex- matched control subjects without evidence of ischemia .

Patients were grouped into 3 groups according to the complexity and severity of coronary lesions

assessed by the SYNTAX score (30 patients with a SYNTAX score of 0 were excluded).

Serum PTX-3 and high-sensitivity C-reactive protein (hs-CRP) levels were measured in both groups.

Results: The PTX-3 levels demonstrated

an increase from low to high SYNTAX groups (r = 0.72, P < 0.001).

Whereas the low SYNTAX group had statistically significantly higher PTX-3 levels when compared with the control group (0.50 ± 0.01 vs 0.24 ± 0.01 ng/mL, P < 0.001),

the hs-CRP levels were not different (0.81 ± 0.42 vs 0.86 ± 0.53 mg/dL, P = 0.96).
but the intermediate SYNTAX group had higher hs-CRP levels compared with the low SYNTAX group (1.3 ± 0.66 vs 0.86 ± 0.53 mg/dL, P = 0.002).

Serum PTX-3 levels and hs-CRP levels were both correlated with the SYNTAX scores and Gensini scores (for SYNTAX: r = 0.87 [P < 0.001] and r = 0.36 [P = 0.01]; for Gensini: r = 0.75 [P < 0.001] and r = 0.27 [P = 0.002], respectively), and

according to the results of univariate and multivariate analyses, for “intermediate and high” SYNTAX scores, age, diabetes mellitus, low-density lipoprotein cholesterol, hs-CRP, and PTX-3
were found to be independent predictors, whereas
for the presence of “high” SYNTAX score only PTX-3 was found to be an independent predictor.
The receiver operating characteristic curve analysis further revealed that the PTX-3 level was
- a strong indicator of high SYNTAX score with an area under the curve of 0.91 (95% confidence interval, 0.86–0.96).

Conclusions: Pentraxin 3, a novel inflammatory marker, was more tightly associated with the complexity and severity of CAD than hs-CRP and

it was found to be an independent predictor for high SYNTAX score.

The association between atherosclerosis and inflammation has been more understood during recent years. Currently, atherosclerosis is considered as a complex inflammatory process in which

leukocytes and inflammatory markers are involved.1

Several inflammatory markers

high-sensitivity C-reactive protein (hs-CRP),
fibrinogen, and
complement C3…. are associated with cardiovascular events.1–5

Pentraxin 3 (PTX-3), that resembles CRP both in structure and function,1 is produced both by

hematopoietic cells such as macrophages, dendritic cells, neutrophils, and by
nonhematopoietic cells such as fibroblasts and vascular endothelial cells.2

Plasma PTX-3 levels may be elevated in patients with

vasculitis,6
acute myocardial infarction,7,8 and
systemic inflammation or sepsis,9
psoriasis,
unstable angina pectoris, and
heart failure.10–13

Dubin et al14 reported that PTX-3 levels are associated with with adverse outcomes in stable angina pectoris (SAP). Despite reports about the association of PTX-3 and coronary artery disease (CAD),

an association between the level of PTX-3 and the complexity and severity of CAD is not established.15,16 Thus, the aim of this study was

to assess the association between the level of PTX-3 and the complexity and severity of CAD assessed with SYNTAX and Gensini scores in SAP patients.

MATERIALS AND METHODS

Of 211 patients were prospectively recruited, 161 SAP patients with evidence of ischemia (positive treadmill or myocardial perfusion scan) underwent coronary angiography for suspected CAD, and 50 age- and sex- matched outpatient subjects with a negative treadmill or myocardial perfusion scan test were taken as the control group. Patients were excluded if they had

acute coronary syndrome
history of previous myocardial infarction;
coronary artery bypass grafting or percutaneous coronary intervention;
secondary hypertension (HT);
renal failure;
hepatic failure;
chronic obstructive lung disease and/or
manifest heart disease, such as
- cardiac failure (left ventricular ejection fraction <50%),
- atrial fibrillation, and
- moderate to severe cardiac valve disease; and
- SYNTAX score of zero

Similarly, patients were excluded with

infection,
acute stress, or chronic systemic inflammatory disease and
those who had been receiving medications affecting the number of leukocytes .

Thirty patients were excluded from the study because the coronary angiograms revealed normal coronary arteries (SYNTAX score of 0). All the participants included in the study were informed about the study, and they voluntarily consented to participate. The Serum PTX-3 level was measured on blood samples collected after 12-hour fast just prior to coronary angiography and kept at −80°C until the assays were performed. PTX3 was measured by enzyme immunoassay (EIA) using quantitative kit (human PTX-3/TSG-14 immunoassay, DPTX30; R&D Systems, Inc, Minneapolis, MN). The intra-assay and interassay coefficients of variation ranged from 3.8% to 4.4% and 4.1% to 6.1%, respectively (minimum detectable concentration, 0.025 ng/mL). High-sensitivity CRP was measured in serum by EIA (Immage hs-CRP EIA Kit; Beckman Coulter Inc, Brea, CA). Transthoracic echocardiography was performed, and biplane Simpson’s ejection fraction (%) was calculated before coronary angiography. Hypertension was defined as having at least 2 blood pressure measurements greater than 140/90 mm Hg or using antihypertensive drugs, whereas diabetes mellitus (DM) was defined as having at least 2 fasting blood sugar measurements greater than 126 mg/dL or using antidiabetic drugs. Smoking was categorized into current smokers and nonsmokers. Nonsmokers included ex-smokers who had quit smoking for at least 6 months before the study. Body mass index (BMI) values were calculated based on the height and weight of each patient. Medications used before the coronary angiography were noted. The study was approved by the local ethics committee.
SYNTAX and Gensini Scores
To grade the complexity of CAD, the SYNTAX score was used. Each coronary lesion with a stenosis diameter of 50% or greater in vessels of 1.5 mm or greater was scored. Parameters used in the SYNTAX scoring are shown in Table 1. The latest online updated version (2.11) was used in the calculation of the SYNTAX scores (www.syntaxscore.com).17 The SYNTAX score was classified as follows:

low SYNTAX score (≤22),
intermediate SYNTAX score (23–32)
high SYNTAX score (≥33).

Table 1 http://images.journals.lww.com/jinvestigativemed/LargeThumb.00042871-201302000-00007.TT1.jpeg

The severity of CAD was determined by the Gensini score, which

measures the extent of coronary stenosis according to degree and location.18

In the Gensini scoring system,

larger segments are more heavily weighted ranging from 0.5 to 5.0
- left main coronary artery × 5;
- proximal segment of the left anterior descending coronary artery [LAD] × 2.5;
- proximal segment of the circumflex artery × 2.5;
- midsegment of the LAD × 1.5;
- right coronary artery distal segment of the LAD,
- posterolateral artery, and obtuse marginal artery × 1;
- and others × 0.5.

The narrowing of the coronary artery lumen is rated

2 for 0% to 25% stenosis,
4 for 26% to 50%,
8 for 51% to 75%,
16 for 76% to 90%,
32 for 91% to 99%,
64 for 100%.

The Gensini index is the sum of the total weights for each segment. All angiographic variables of the SYNTAX and Gensini score were computed by

2 experienced cardiologists who were blinded to the procedural data and clinical outcomes.

The final decision was reached by consensus when a conflict occurred.The number of diseased vessels with

greater than 50% luminal stenosis was scored from 1 to 3 (namely, 1-, 2-, or 3-vessel disease), and
a lesion greater than 50% in the left main coronary artery was regarded as a 2-vessel disease.

Statistical Analyses

Statistical analyses were conducted with SPSS 17 (SPSS Inc, Chicago, IL) software package program.
Continuous variables were expressed as mean ± SD or median ± interquartile range values, whereas categorical variables were presented as percentages.
The differences between normally distributed numeric variables were evaluated by Student t test or 1-way analysis of variance, whereas

non–normally distributed variables were analyzed by Mann-Whitney U test or Kruskal-Wallis variance analysis as appropriate.

χ2 Test was used for the comparison of categorical variables. Pearson test was used for correlation analysis.
To determine the independent predictors of “intermediate and high” SYNTAX scores and only “high” SYNTAX scores,

2 different sets of univariate and multivariate analyses were performed
- (in the first model SYNTAX cutoff was 22, whereas
- in the second model SYNTAX cutoff was 33).

The standardized parameters that were found to have a significance (P < 0.10) in the univariate analysis were evaluated by stepwise logistic regression analysis.
Ninety-five percent confidence interval (CI) and odds ratio (OR) per SD increase were presented together. Interobserver and intraobserver variability for SYNTAX scores

was done by Bland-Altman analysis.

An exploratory evaluation of additional cut points was performed using the receiver operating characteristic (ROC) curve analysis.
All the P values were 2-sided, and a P < 0.05 was considered as statistically significant.
RESULTS
Baseline Characteristics
In total, 181 patients (50.2 ± 6.5 years, 52.5% were composed of males) were included in the study. Baseline clinical, angiographic, and laboratory characteristics of the patients
relative to SYNTAX score groups are shown in Table 2. Age, sex, HT, DM, BMI, and medication were not different between the groups. Baseline clinical and laboratory characteristics
of patients according to PTX-3 quartiles are shown in Table 3. The Bland-Altman analysis revealed that the degrees of intraobserver and interobserver variability for SYNTAX score
and Gensini score readings were 5% and 6% for SYNTAX and 8% and 9% for Gensini, respectively.
Table 2 http://images.journals.lww.com/jinvestigativemed/Original.00042871-201302000-00007.TT2.jpeg
Table 3 http://images.journals.lww.com/jinvestigativemed/Original.00042871-201302000-00007.TT3.jpeg

The PTX-3 levels demonstrated an increase from the low SYNTAX group to the high SYNTAX group (r = 0.87, P < 0.001).
The low SYNTAX group had statistically significantly higher PTX-3 levels when compared with the control group (0.50 ± 0.01 vs 0.24 ± 0.01 ng/mL, P < 0.001); similarly,
the PTX-3 levels were higher in the high SYNTAX group than in both

the intermediate SYNTAX group (0.84 ± 0.08 vs 0.55 ± 0.01 ng/mL, P < 0.001) and
the low SYNTAX group (0.84 ± 0.08 vs 0.50 ± 0.01 ng/mL, P < 0.001).
there was no difference in levels of PTX-3 between the low and the intermediate SYNTAX group (0.50 ± 0.01 vs 0.55 ± 0.01 ng/mL, P = 0.09).

On the other hand, there was no difference in levels of hs-CRP between the control and the low SYNTAX group (0.81 ± 0.42 vs 0.86 ± 0.53 mg/dL, P = 0.96).
The intermediate SYNTAX group had statistically significantly higher hs-CRP levels

compared with the low SYNTAX group (1.3 ± 0.66 vs 0.86 ± 0.53 mg/dL, P = 0.002);
the hs-CRP levels were not different between the high SYNTAX group
- and the intermediate SYNTAX group. (1.3 ± 0.66 vs 1.3 ± 0.43 mg/dL, P = 0.99).

Univariate correlation analysis revealed a positive correlation between serum PTX-3 levels and hs-CRP levels with

the SYNTAX and Gensini scores
- for SYNTAX: r = 0.87 [P < 0.001] and r = 0.36 [P = 0.01];
- for Gensini: r = 0.75 [P < 0.001] and r = 0.27 [P = 0.002], (Fig. 1).

In addition to that, the Gensini and SYNTAX scores are found to be well correlated with each other (r = 0.80, P < 0.001).
When the SYNTAX score was taken as continuous variable, multivariate linear regression analysis revealed that

the SYNTAX score was correlated with PTX-3 and hs-CRP (for PTX-3: β = 0.84 [P < 0.001]; hs-CRP: β =0.08 [P = 0.032]).

Figure 1 http://images.journals.lww.com/jinvestigativemed/Original.00042871-201302000-00007.FF1.jpeg

For determining the predictors of intermediate and high SYNTAX scores and only-high SYNTAX scores,

2 different sets of univariate and multivariate analyses were performed among the patients who underwent coronary angiography.

For predicting the intermediate and high SYNTAX scores, the SYNTAX score was dichotomized into

high (score ≥22) and
low (<22) groups,

whereas for predicting the only-high SYNTAX scores, the SYNTAX score was dichotomized into

2 groups with a score of 33 or greater and a score of less than 33.

In the first multivariate analysis (where SYNTAX cutoff was 22), the parameters showing significance in the univariate analysis

age,
sex,
HT,
DM,
low-density lipoprotein cholesterol [LDL-C],
hs-CRP,
PTX-3

were evaluated by multivariate analysis to determine the

independent predictors of intermediate and high SYNTAX scores.

In the univariate analysis, higher values of

age (OR, 1.5 [95% CI, 1.1–2.0]; P = 0.01),
LDL-C (OR, 1.3 [95% CI, 0.98–1.8]; P = 0.068),
hs-CRP (OR, 2.6 [95% CI, 1.8–3.8]; P < 0.001), and
PTX-3 (OR, 13.6 [95% CI, 6.4–28.9]; P < 0.001)
- were associated with higher SYNTAX scores,
HT (OR, 0.44 [95% CI, 0.24–0.80]; P = 0.008) and
DM (OR, 0.48 [95% CI, 0.25–0.91]; P = 0.02)
- were associated with lower SYNTAX scores.

In the multivariate analysis – age, DM, LDL-C, hs-CRP, and PTX-3 – were found to be

independent predictors of “intermediate to high” SYNTAX score (Table 4).

Increased

age (OR, 2.5 [95% CI, 1.3–4.8]; P = 0.007),
LDL-C (OR, 2.8 [95% CI, 1.5–5.2]; P = 0.001),
hs-CRP (OR, 3.3 [95% CI, 1.8–6.1]; P < 0.001), and
PTX-3 (OR, 35.4 [95% CI, 10.1–123.6]; P < 0.001)
- were associated with increased SYNTAX scores,

whereas DM (OR, 0.08 [95% CI, 0.02–0.33]; P < 0.001) was associated with lower SYNTAX score (Table 4).

In the second univariate and multivariate analyses (where SYNTAX cutoff was 33),

the parameters that showed significance in the univariate analysis were age, LDL-C, glucose, hs-CRP, and PTX-3.
In the univariate analysis, increased
- age (OR, 1.5 [95% CI, 1.0–2.3]; P = 0.05),
- LDL-C (OR, 1.5 [95% CI, 0.97–2.2]; P = 0.07),
- hs-CRP (OR, 1.4 [95% CI, 0.97–2.1]; P = 0.072), and
- PTX-3 (OR, 18.5 [95% CI, 6.6–51.8]; P < 0.001)
  - were found to be associated with increased SYNTAX scores.

When these parameters were evaluated with multivariate analysis, only PTX-3 (OR, 18.4 [95% CI, 6.2–54.2]; P < 0.001)

was found to be an independent predictor for high SYNTAX score (Table 4).

Table 4 http://images.journals.lww.com/jinvestigativemed/Original.00042871-201302000-00007.TT4.jpeg

The ROC curve analysis further revealed that the PTX-3 level was a strong indicator of high SYNTAX score with

an area under the curve (AUC) of 0.91 (95% CI, 0.86–0.96) (Fig. 2).

The optimal cutoff of PTX-3 for the high SYNTAX score was 0.75 ng/mL.
Sensitivity, specificity, positive predictive value, and negative predictive value to identify high SYNTAX score for the PTX-3 level

were 90%, 84%, 97%, and 60%, respectively.
the ROC curve analysis of PTX-3 for intermediate-high SYNTAX score revealed that the AUC value was 0.82 (95% CI, 0.75–0.89).

The optimal threshold of PTX-3 level that

maximized the combined specificity and sensitivity to predict
- intermediate to high SYNTAX score was 0.73 ng/mL.

For the cutoff value of 0.73 ng/mL, sensitivity, specificity, positive predictive value, and negative predictive value

to identify intermediate-high SYNTAX score were 56%, 98%, 97%, and 56%, respectively.

Figure 2 http://images.journals.lww.com/jinvestigativemed/Original.00042871-201302000-00007.FF2.jpeg

In the ROC analysis of hs-CRP for high SYNTAX scores, the AUC value was found to be 0.68 (95% CI, 0.59–0.77; P < 0.001).
The optimal threshold of hs-CRP that maximized the combined specificity and sensitivity to predict for high SYNTAX scores was 0.89 mg/dL.
Similarly, the ROC analysis of hs-CRP for the intermediate-high SYNTAX scores revealed an AUC of 0.74 (95% CI, 0.65–0.83; P = 0.001).
The cutoff value of hs-CRP to predict the intermediate-high SYNTAX scores with a maximized sensitivity and specificity was 0.66 mg/dL.
DISCUSSION
In this particular study, we investigated the relationship between the serum PTX-3 level and the severity of CAD

assessed by SYNTAX and Gensini scores in patients with SAP.

The PTX-3, was significantly higher than control group in the patients with CAD, and the serum PTX-3 levels

were associated with the SYNTAX and Gensini scores.

When compared with the hs-CRP, the PTX-3 was found to be more tightly associated with the complexity and severity of CAD in the patients with SAP.
Pentraxin 3, an acute-phase reactant that is functionally and structurally similar to CRP,1 is produced both by different kinds of cells such as

macrophages, dendritic cells, neutrophils, fibroblasts, and vascular endothelial cells.2
Pentraxin 3 is released following the inflammatory stimuli19; therefore, it may reflect the local inflammatory status in tissues.20

Serum PTX-3 levels were shown to be elevated in patients with

vasculitis,6 acute myocardial infarction,7,8 and systemic inflammation or sepsis,9 psoriasis, unstable angina pectoris, and heart failure.10–13

Higher PTX3 levels were reported to be associated with worse cardiovascular outcomes

after acute coronary syndromes,8,21
in the elderly people without known cardiovascular disease22 and
associated with overall mortality in patients with stable coronary disease,
independent of systemic inflammation.14

There are 2 reports investigating the association of PTX-3 level and the atherosclerotic burden.15,16 In one of these reports,

Knoflach et al.15 took B-mode ultrasonography as the atherosclerosis index.

They did not provide any information about coronary anatomy, and in the other report, Soeki et al.16 evaluated 40 patients who

underwent coronary angiography and measured their Gensini scores.

However, in none of the studies were the SYNTAX score and Gensini score used together to assess the degree of coronary atherosclerotic burden.
To our knowledge, this is the first report that showed the association of PTX-3 levels with the complexity and severity of CAD assessed by

SYNTAX and Gensini scores in patients with stable coronary disease.

Chronic low-grade inflammation has been thought to play a major role in the pathogenesis of atherosclerosis.23,24 Previous studies have reported that

levels of inflammatory markers such as hs-CRP, interleukin 6, and so on were increased in atherosclerosis.25

In the present study, both the SYNTAX and the Gensini scores were found to be correlated with serum PTX-3 and hs-CRP levels,

which in turn might reflect the degree of inflammation.

The SYNTAX score is an important tool in the classification of complex CAD26 and can give predictive information about short- and long-term outcomes

in patients with stable CAD who undergo percutaneous coronary intervention.27–30

Although the SYNTAX score is currently used for assessing the angiographic complexity of CAD rather than the severity of coronary atherosclerotic burden,

because more complex lesions tend to have more atherosclerotic burden,
the SYNTAX scores may also reflect the severity of coronary atherosclerotic burden.

The Gensini score, a well-known and widely used scoring system to evaluate the severity of CAD,18 was measured and

found to be well correlated with the SYNTAX score,
- which supports the idea that angiographically more complex lesions tend to have more atherosclerotic burden.

When compared with the hs-CRP,

the PTX-3 seems to be more tightly associated with coronary disease burden (r = 0.36 vs r = 0.87).

We found out that the serum PTX-3 levels were higher than those in the control group, even in the low SYNTAX group.
On the other side, the serum hs-CRP levels were not different in the control and the low SYNTAX groups.
It was reported that the leukocytes mainly found in the coronary artery lumen are the neutrophils.31
It is also known that PTX-3 is stored in specific granules of neutrophils and released in response to inflammatory signals.32
The reason why serum PTX-3 levels seem more tightly associated with the coronary disease burden

when compared with serum hs-CRP levels may be the association of the
on-site presence of neutrophils and local inflammatory signal–triggered release of PTX-3.

On the other hand, some human studies revealed that PTX-3 was produced more in areas of atherosclerosis and may contribute to its pathogenesis.31
Some other studies suggested that PTX-3 may be part of a protective mechanism in

vascular repair via inhibiting fibroblast growth factor 2 or some other growth factors responsible for smooth muscle proliferation.33,34

But still, the exact role of PTX-3 in the pathophysiology of atherosclerosis seems to be obscure for the time being. It is well established that atherosclerosis
has an inflammatory background in most of the cases. In addition to that, high blood CRP level is known as an indicator of future cardiovascular disease risk
even in healthy individuals.35 According to the results of univariate and multivariate analyses, for intermediate and high SYNTAX scores,

age, DM, LDL-C, hs-CRP, and PTX-3 were found to be independent predictors, whereas for the presence of
high SYNTAX score, only PTX-3 was found to be an independent predictor.

Because of the tighter association with atherosclerotic burden and the on-site vascular presence,

PTX-3 may be a promising candidate marker for vascular inflammation and future cardiovascular events.

LIMITATIONS
The major limitation of the current study is the number of patients included. It would be better to include more patients to increase the statistical power.

Besides, the SYNTAX and Gensini scores give us an idea about the complexity and severity of coronary atherosclerosis; however,
with coronary angiography alone, it is not possible to understand the extent of coronary plaque. In addition to that, the coronary anatomy of the
control group was not known, which was another limitation. Our selected population was free of other confounders of systemic inflammation, and
we did not have data about inflammatory markers other than hs-CRP, such as interleukin 6, tumor necrosis factor α, and so on, which may be accepted
as a limitation. Another limitation of the current study is that because there was no long-term follow-up of the patients, it did not provide any prognostic
data in terms of future cardiovascular events.
CONCLUSIONS
Pentraxin 3, a novel inflammatory marker, is associated with the complexity and severity of the CAD assessed by the SYNTAX and the Gensini scores in patients with SAP and seems to be more tightly associated with coronary atherosclerotic burden than hs-CRP.

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14. Dubin R, Li Y, Ix JH, et al.. Associations of pentraxin-3 with cardiovascular events, incident heart failure, and mortality among persons with coronary heart disease: data from the Heart and Soul Study. Am Heart J. 2012; 163: 274–279.
16. Soeki T, Niki T, Kusunose K, et al.. Elevated concentrations of pentraxin 3 are associated with coronary plaque vulnerability. J Cardiol. 2011; 58: 151–157.
17. SYNTAX working group. SYNTAX score calculator. Available at http://www.syntaxscore.com. Accessed May 20, 2012.
18. Gensini GG. A more meaningful scoring system for determining the severity of coronary heart disease. Am J Cardiol. 1983; 51: 606.
20. Mantovani A, Garlanda C, Bottazzi B, et al.. The long pentraxin PTX3 in vascular pathology. Vascul Pharmacol. 2006; 45: 326–330.
21. Matsui S, Ishii J, Kitagawa F, et al.. Pentraxin 3 in unstable angina and non-ST-segment elevation myocardial infarction. Atherosclerosis. 2010; 210: 220–225.
22. Jenny NS, Arnold AM, Kuller LH, et al.. Associations of pentraxin 3 with cardiovascular disease and all-cause death: the Cardiovascular Health Study. Arterioscler Thromb Vasc Biol. 2009; 29: 594–599.
26. Serruys PW, Morice MC, Kappetein AP, et al.. Percutaneous coronary intervention versus coronary-artery bypass grafting for severe coronary artery disease. N Engl J Med. 2009; 360: 961–972.
27. van Gaal WJ, Ponnuthurai FA, Selvanayagam J, et al.. The SYNTAX score predicts peri-procedural myocardial necrosis during percutaneous coronary intervention. Int J Cardiol. 2009; 135: 60–65.
28. Lemesle G, Bonello L, de Labriolle A, et al.. Prognostic value of the SYNTAX score in patients undergoing coronary artery bypass grafting for three-vessel coronary artery disease. Catheter Cardiovasc Interv. 2009; 73: 612–617.
29. Capodanno D, Di Salvo ME, Cincotta G, et al.. Usefulness of the SYNTAX score for predicting clinical outcome after percutaneous coronary intervention of unprotected left main coronary artery disease. Circ Cardiovasc Interv. 2009; 2: 302–308.
30. Kim YH, Park DW, Kim WJ, et al.. Validation of SYNTAX (Synergy between PCI with Taxus and Cardiac Surgery) score for prediction of outcomes after unprotected left main coronary revascularization. JACC Cardiovasc Interv. 2010; 3: 612–623.
32. Jaillon S, Peri G, Delneste Y, et al.. The humoral pattern recognition receptor PTX3 is stored in neutrophil granules and localizes in extracellular traps. J Exp Med. 2007; 204: 793–804.
33. Inforzato A, Baldock C, Jowitt TA, et al.. The angiogenic inhibitor long pentraxin PTX3 forms an asymmetric octamer with two binding sites for FGF2. J Biol Chem. 2010; 285: 17681–17692.
34. Camozzi M, Zacchigna S, Rusnati M, et al.. Pentraxin 3 inhibits fibroblast growth factor 2–dependent activation of smooth muscle cells in vitro and neointima formation in vivo. Arterioscler Thromb Vasc Biol. 2005; 25: 1837–1842.
35. Koenig W, Sund M, Frohlich M, et al.. C-Reactive protein, a sensitive marker of inflammation, predicts future risk of coronary heart disease in initially healthy middle-aged men: results from the MONICA (Monitoring Trends and Determinants in Cardiovascular Disease) Augsburg Cohort Study, 1984 to 1992. Circulation. 1999; 99: 237–242.
Keywords: pentraxin 3; coronary artery disease; SYNTAX score; hs-CRP; inflammation

This is not the only recent finding that adds to the ability to evaluate these patients. An as yet unpublished paper, expected to be published soon reports on

QRS fragmentation as a Prognostic test in Acute Coronary Syndrome, and this reviewer expects the work to have a high impact. The authors state that
QRS complex fragmentation is a promising bed-side test for assessment of prognosis in those patients. Presence of fragmented QRS in surface ECG during ACS

represents myocardial scar or fibrosis and reflect severity of coronary lesions and a correlation between fQRS and depression of Lv function is established.

There are still other indicators that need to be considered, such as the mean arterial blood pressure.

There has been review and revisions of the guidelines for treatment of UA/NSTEMI within the last year, with differences being resolved among the Europeans and US.

Guidelines Updated for Unstable Angina/Non-ST Elevation Myocardial Infarction
According to the current study by Jneid and colleagues, new evidence is available on the management of unstable angina. This report replaces the 2007 American College of Cardiology Foundation/American Heart Association (ACC/AHA) Guidelines for the Management of Patients With Unstable Angina/Non–ST-Elevation Myocardial Infarction (UA/NSTEMI) that were updated by the 2011 guidelines.

This guideline was reviewed by

2 official reviewers each nominated by the ACCF and the AHA, as well as
1 or 2 reviewers each from the American College of Emergency Physicians; the Society for Cardiovascular Angiography and Interventions; and the Society of Thoracic Surgeons; and
29 individual content reviewers, including members of the ACCF Interventional Scientific Council.

The recommendations in this focused update are considered current

until they are superseded in another focused update or the full-text guideline is revised, and are official policy of both the ACCF and the AHA.

STUDY SYNOPSIS AND PERSPECTIVE
American cardiology societies have caught up with the European Society of Cardiology by

issuing their second update to the UA/NSTEMI guidelines in 18 months,
with the 2012 focused update replacing the 2011 guidelines [1].

The new recommendations include ticagrelor (Brilinta) as one of the options for antiplatelet therapy alongside prasugrel (Effient) and clopidogrel, bringing them in line with European.
The European guidance, however, gave precedence to the new antiplatelets over clopidogrel, whereas the American update “places ticagrelor on an equal footing with the other two antiplatelets available–
this is the main reason for the update,” lead author Dr Hani Jneid (Baylor College of Medicine, Houston, TX), told heartwire . “Doctors now have a choice for second-line therapy after aspirin, depending on

the patient’s clinical scenario,
physician preference, and cost,”
- now that clopidogrel is available generically.

The US decision to recommend

first prasugrel–in its 2011 update to the UA/NSTEMI guidelines–and
now ticagrelor as equivalent antiplatelet therapy choices to clopidogrel after aspirin
- puts it somewhat at odds with the Europeans,
- who reserve clopidogrel use for those who cannot take the newer agents.

The reason for the Americans differing stance is that because while they are faster acting and more potent–

the cost-effectiveness of the new agents is not known.
it isn’t clear how the efficacy observed in pivotal clinical trials of these agents is going to translate into real-world benefit,
and issues such as bleeding with prasugrel and compliance with a twice-daily drug such as ticagrelor remain concerns.

Bulk of 2012 Update on How to Use Ticagrelor
The 2012 ACCF/AHA focused update for the management of UA/NSTEMI stresses that

all patients at medium/high risk should receive dual antiplatelet therapy on admission,
with aspirin being first-line, indefinite therapy.

The bulk of the update centers on how to use ticagrelor which–

like prasugrel or clopidogrel–
can be added to aspirin for up to 12 months (or longer, at the discretion of the treating clinician).

Jneid notes it’s important to remember that prasugrel can only be used in the cath lab

in patients undergoing percutaneous coronary intervention (PCI),
whereas ticagrelor, like clopidogrel, can be used in medically managed or PCI patients.

And he emphasizes that, in line with the FDA’s black-box warning on ticagrelor,

this new antiplatelet agent must only be administered with a “baby” dose of aspirin (81 mg in the US).

The 81-mg aspirin dose is also considered a reasonable option in preference to a higher maintenance dose of 325 mg in

any acute coronary syndrome (ACS) patient following PCI, he adds, as
this strategy is believed to result in equal efficacy and lower bleeding risk.

With regard to how long antiplatelet therapy should be stopped before planned cardiac surgery, the recommendation is

five days for ticagrelor–the same as that advised for clopidogrel.
and seven days prior to surgery for prasugrel.

Jneid also highlights other important recommendations from the 2011 focused update carried over to 2012:

It is “reasonable” to proceed with cardiac catheterization and revascularization within

12–24 hours of admission in initially stable, very high-risk patients with ACS.

An invasive strategy is “reasonable” in patients with

mild and moderate chronic kidney disease.

In those with diabetes hospitalized with ACS, insulin use should target glucose levels <180 mg/dL,

a less-intensive reduction than previously recommended.

Platelet function or genotype testing for clopidogrel resistance are both considered “reasonable”

if clinicians think the results will alter management,
but Jneid acknowledged that “there is not much evidence to support these assays” .

Committee Encourages Participation in Registries
Jneid observes that unstable angina and NSTEMI are “very common” conditions that carry a high risk of death and recurrent heart attacks,

which is why “the AHA and ACCF constantly update their guidelines so that physicians can provide patients with
the most appropriate, aggressive therapy with the goal of improving health and survival.”

To this end, he notes that the writing panel encourages

clinicians and hospitals to participate in quality-of-care registries designed
to track and measure outcomes, complications, and
adherence to evidence-based medicines.

Conflicts of interest for the writing committee are listed in the paper.

References

Jneid H, Anderson JL, Wright SR, et al. 2012 ACCF/AHA focused update on the guideline for the management of patients with unstable angina/non-ST elevation myocardial infarction (Updating the 2007 guideline and replacing the 2011 focused update): A report of the ACCF/AHA.
Circulation 2012; Available at: http://circ.ahajournals.org/ http://dx.doi.org/10.1161/CIR0b013e3182566fleo
source http://www.medscape.org

Implantable Synchronized Cardiac Assist Device Designed for Heart Remodeling: Abiomed’s Symphony
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Obstructive Coronary Artery Disease diagnosed by RNA levels of 23 genes – CardioDx, a Pioneer in the Field of Cardiovascular Genomic Diagnostics

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FDA Pending 510(k) for The Latest Cardiovascular Imaging Technology

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The Electronic Health Record: How far we have travelled, and where is journey’s end?
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New Definition of MI Unveiled, Fractional Flow Reserve (FFR)CT for Tagging Ischemia

http://pharmaceuticalintelligence.com/2012/08/27/new-definition-of-mi-unveiled-fractional-flow-reserve-ffrct-for-tagging-ischemia/

Assessing Cardiovascular Disease with Biomarkers

http://pharmaceuticalintelligence.com/2012/12/25/assessing-cardiovascular-disease-with-biomarkers/

Coronary CT Angiography versus Standard Evaluation in Acute Chest Pain

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Dilated Cardiomyopathy: Decisions on implantable cardioverter-defibrillators (ICDs) using left ventricular ejection fraction (LVEF) and Midwall Fibrosis: Decisions on Replacement using late gadolinium enhancement cardiovascular MR (LGE-CMR)

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Acute Chest Pain/ER Admission: Three Emerging Alternatives to Angiography and PCI
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The potential contribution of informatics to healthcare is more than currently estimated

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PCI Outcomes, Increased Ischemic Risk associated with Elevated Plasma Fibrinogen not Platelet Reactivity

http://pharmaceuticalintelligence.com/2013/01/10/pci-outcomes-increased-ischemic-risk-associated-with-elevated-plasma-fibrinogen-not-platelet-reactivity/

Amplifying Information Using S-Clustering and Relationship to Kullback-Liebler Distance: An Application to Myocardial Infarction

http://pharmaceuticalintelligence.com/2012/09/22/amplifying-information-using-s-clustering/

Percutaneous Endocardial Ablation of Scar-Related Ventricular Tachycardia

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Vascular Medicine and Biology: CLASSIFICATION OF FAST ACTING THERAPY FOR PATIENTS AT HIGH RISK FOR MACROVASCULAR EVENTS Macrovascular Disease – Therapeutic Potential of cEPCs

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Ablation Devices Market to 2016 – Global Market Forecast and Trends Analysis by Technology, Devices & Applications

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Endothelial Dysfunction, Diminished Availability of cEPCs, Increasing CVD Risk for Macrovascular Disease – Therapeutic Potential of cEPCs

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Positioning a Therapeutic Concept for Endogenous Augmentation of cEPCs — Therapeutic Indications for Macrovascular Disease: Coronary, Cerebrovascular and Peripheral

http://pharmaceuticalintelligence.com/2012/08/29/positioning-a-therapeutic-concept-for-endogenous-augmentation-of-cepcs-therapeutic-indications-for-macrovascular-disease-coronary-cerebrovascular-and-peripheral/

Cardiovascular Outcomes: Function of circulating Endothelial Progenitor Cells (cEPCs): Exploring Pharmaco-therapy targeted at Endogenous Augmentation of cEPCs

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Competition in the Ecosystem of Medical Devices in Cardiac and Vascular Repair: Heart Valves, Stents, Catheterization Tools and Kits for Open Heart and Minimally Invasive Surgery (MIS)

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Effects of Intensive Blood Pressure Lowering on Cardiovascular and Renal Outcomes: A Systematic Review and Meta-Analysis. (zedie.wordpress.com)
Measure of coronary artery calcium linked with improved prediction of cardiovascular disease risk (eurekalert.org)
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Age-standardised disability-adjusted life year (DALY) rates from Cardiovascular diseases by country (per 100,000 inhabitants). (Photo credit: Wikipedia)

English: Cardiovascular disease: PAD therapy with stenting Deutsch: PAVK Therapie: Kathetertherapie mit stenting (Photo credit: Wikipedia)

Micrograph of an artery that supplies the heart with significant atherosclerosis and marked luminal narrowing. Tissue has been stained using Masson’s trichrome. (Photo credit: Wikipedia)

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Predicting Drug Toxicity for Acute Cardiac Events

Posted in Advanced Drug Manufacturing Technology, Bio Instrumentation in Experimental Life Sciences Research, Computational Biology/Systems and Bioinformatics, Disease Biology, Small Molecules in Development of Therapeutic Drugs, FDA Regulatory Affairs, Genomic Testing: Methodology for Diagnosis, Health Law & Patient Safety, International Global Work in Pharmaceutical, Molecular Genetics & Pharmaceutical, Pharmaceutical Analytics, Pharmaceutical Industry Competitive Intelligence, Pharmaceutical R&D Investment, Pharmacogenomics, Pharmacotherapy of Cardiovascular Disease, Population Health Management, Genetics & Pharmaceutical, Regulated Clinical Trials: Design, Methods, Components and IRB related issues, Statistical Methods for Research Evaluation, Technology Transfer: Biotech and Pharmaceutical, tagged Cardiotoxicity, Drug development, ECG, Electrocardiography, Investigational New Drug, NCE, New chemical entity, QT interval on March 15, 2013| 5 Comments »

Predicting Drug Toxicity for Acute Cardiac Events

Reporter: Larry H Bernstein, MD, FACP

http://pharmaceuticalintelligence.com/?p=10679/Predicting Drug Toxicity for Acute Cardiac Events

Pharmaceuticals Dilemma

The pharmaceutical industry has, as the clinical diagnostics industry, consolidated, and seen new entries that are at some time merged into an established giant, needing resources to grow. In the past, it was considered essential for a scientific commercial entity to invest at least 8 percent of budget to R&D. However, the cost of manufacturing has gone down, but a large part of the budget outside of manufacture has to be taken up with, maybe a few exceptions, development, validation in clinical trials, and marketing. This leaves the situation precarious without a basic research base, and has lead to consortia between academic centers, the federal governmant, and the industries. I can’t venture into the role of Wall Street Investment and Venture Capital in the process of innovation, proprietary rights to discoveries, and viability. A large problem they encounter really comes down to complexity of the biomedical reality, that keeps peeling off layers like an onion, exposing new problems to deal with. As a result, we have seen repeated recalls of drugs that were blockbusters, over the last 2 decades. To date, every “miracle” drug to manage sepsis and the several cardiac related drugs have resulted in unexpected toxicities.
One of the leading causes of drug attrition during development is cardiac toxicity, which has a serious impact on cost and can impact getting new drugs to patients. Detecting cardiovascular safety issues earlier in the drug development program

would produce significant benefits for pharmaceutical companies and, ultimately, public health, but
- the reduction of therapeutic toxicities will not be easy and depends on the
- emergence of genomic-based personalized medicine.

Comprehensive cardiovascular and electrophysiology assessments are routinely conducted in vivo and in vitro early in the preclinical or lead optimization phases of drug development. For example,

the isolated perfused guinea pig heart preparation (classically called the Langendorff preparation)
can be used to screen a series of related new chemical entities (NCE)

in the lead optimization phase for preliminary information on the relative effects on contractility and rhythm.
Additionally, intact animal non-GLP studies—generally conducted in anesthetized, non-recovery models—are designed to assess

effects of NCEs on a range of acute hemodynamic and cardiac parameters such as
- heart rate,
- blood pressure,
- electrocardiogram (ECG),
- ventricular contractility,
- vascular resistance,
- cardiac output, etc.

These studies employ small numbers of animals, but may allow termination of research into NCEs with obvious cardiovascular side effects. These preparations also provide information on the involvement of the

autonomic nervous system in the cardiovascular responses of the NCE.

Such effects can be important determinants in the total cardiovascular response to an NCE, and this information cannot be obtained with any known in vitro method.
But what if there are dangers that are not predictable in the short term because of the time span under which the effects can be viewed? The effects themselves are a result of interactions between

the drug,
endothelial cell receptors,
and/or imbalance in oxidative stress promoters and suppressors,
and involve signaling pathways.

That is a difficult challenge that may only be realized

by rapidly advancing knowledge at the molecular cell level.

The ICH S7A and ICH S7B guidelines provide

guidance on important physiological systems and
assessment of pharmaceuticals on
- ventricular repolarization and
- proarrhythmic risk.

The guidelines were designed to protect patients from potential adverse effects of pharmaceuticals. Since these guidelines were issued in 2000 and 2005, respectively,

cardiac safety study designs have been realigned
to identify potential concerns prior to administering the first dose to humans.

It is now routine for all NCEs to be evaluated using an

in vitro Ikr assay such as the hERG voltage patch clamp assay to assess for
- the potential for QT interval prolongation.

Systems have evolved to screen large numbers of compounds

using automated high-throughput patch clamp systems early in the lead
optimization/drug discovery phase.

This is a cost effective method for determining an initial go/no-go gate. Once a compound has progressed to

the development phase, it can once again be assessed with the hERG assay
utilizing the gold standard manual patch clamp assay.

If the NCE under investigation is a cardiovascular therapy, then

pharmacological characterization should occur
early in the lead development process.

In addition to the techniques just discussed,

a variety of “disease models” are available to help determine
- whether the NCE will be efficacious in a clinical setting.

However sound the in vitro data used in screening and selection process (e.g., receptor-binding studies),

NCEs that have been shown to be active in at least one in vivo model (e.g,. salt-sensitive Dahl rat model)
have a higher likelihood of clinical success.

Once a lead is identified, it should still go through the generalized safety characterization discussed earlier.
The in vivo study designs for NCEs reaching the development phase to support the Investigational New Drug (IND) application (just prior to the first human dose) require acquisition of

heart rate,
blood pressure, and
ECG data

using an appropriate species
at and above clinically relevant doses.

The trend in the industry for these regulatory-driven studies has been to

utilize animals surgically instrumented with telemetry devices that
can acquire the required parameters.

The advantage of using instrumented animals over anesthetized animals is that

data can be acquired from freely moving animals over greater periods of time
without anesthetic in the test system,
- which has the potential to confound and perturb results interpretation.

Appropriate dose selection relative to those used in the clinic provides valuable information about

potential acute cardiac events and
how they may impact trial participants.

The obvious limitation here is that the method of observation is essentially

the same or less than that which is used in clinical practice,
relying mainly on classical physiology to detect
- inherently deep seated processes.

But it is not the same scale of issue as for the patient emergently presenting to the ED. Despite enormous efforts to reduce the development of and the complications of acute ischemia related cardiac events,

the accurate diagnosis of the patient presenting to the emergency room is still, as always, reliant on

clinical history,
physical examination,
effective use of the laboratory, and
increasingly helpful imaging technology.

and age, sex, diet, and ability to carry out the activities of daily living before treatment and 6 months to a year after discharge are relevant.

The main issue that we have a consensus agreement that PLAQUE RUPTURE is not the only basis for a cardiac ischemic event. There will be more to say about this.
Animal studies
Telemetry-instrumented animals can be used as screening tools earlier in the drug selection phase. Colonies of animals that can be reused, following a suitable wash-out period,
provide an excellent resource for screening compounds to detect unwanted side effects. The use of these animals

coupled with
recent advances in software-analysis systems allow for rapid data turnaround,
- enables scientists to quickly determine if there are any potentially unwanted signals.

If any effects are detected on, for example, blood pressure or QT interval, then the decision to

either shelve the drug or
conduct additional studies

can be made before advancing any further in the developmental phase. While this is very good for observing large effects, is it really sufficient for avoidance of late phase failure?

Interestingly, the experience that has been acquired since the approval of the ICH guidelines

has allowed pharmaceutical companies to temper their response to finding a potentially unwanted signal.
Rather than permanently shelve libraries of compounds that, for example, were
found to be positive in the hERG assay—common practice when the 2005 guidelines came into being—
- companies can now determine a risk potential based on knowledge gained with the intact animal studies.

Similarly, if changes in hemodynamic parameters are detected, there are follow-up experiments employing anesthetized or telemetry models that include additional measurements like

left ventricular pressure.
These experiments can be utilized to further assess their potential clinical impact
by examining effects on
myocardial contractility,
relaxation, and
conduction velocity.
These techniques primarily address acute effects: those following a single exposure.
Chronic effects—those seen with long-term administration of the NCE to an intact organism—are difficult to obtain in early development, but are routinely monitored during safety studies,
are conducted non-clinically during Phase 1 and 2 of the development process.

ECGs typically are collected to evaluate the chronic cardiac effects in non-rodent species during these studies. It is recommended that
- JET (jacketed external telemetry) techniques, which permit the recording of ECG’s—
- but not blood pressure—

be applied in freely moving animals. If chronic effects are discovered,

follow-up experiments can be conducted with any of the techniques mentioned in this article.

As the focus on cardiac safety has matured over the last 10 years, the Safety Pharmacology Society has led efforts to establish an approach

to determine best practices for conducting key preclinical cardiovascular assessments in drug development.
to provide sensitive preclinical assays that can detect high-probability safety concerns.

Parallel efforts have been made to more accurately assess the translation of preclinical cardiovascular data into

clinical outcomes and
to encourage collaborations
- between preclinical and clinical scientists involved in cardiac safety assessment.

This has been conducted under the umbrella of the International Life Science Institute–Health and Environmental Services Institute (ILSI-HESI) consortium, which has bought together

industrial,
academic, and
government scientists
- to discuss and determine what steps are necessary
- to establish an integrated cardiovascular safety assessment program.

The goal is to provide better ways of predicting potential adverse events, allowing for earlier detection of cardiovascular safety issues and reducing the number of clinical trial failures.
http://www.dddmag.com/articles/2012/08/predicting-potential-cardiac-events?et_cid=2816494&et_rid=45527476&linkid=http%3a%2f%2fwww.dddmag.com%2farticles%2f2012%2f08%2fpredicting-potential-cardiac-events.

A recent poster presentation I think makes a good statement of advances that should move us forward:

http://www.biotechniques.com/multimedia/archive/00178/BTN_0311-March_Post_178205a.pdf

Another possibility is genetic testing to determine the likelihood of stroke, for example Corus CAD is

a shoebox-size kit that uses a simple blood draw to measure the RNA levels of 23 genes.
it creates an algorrhytm-based score that determines the likelihood that a patient has obstructive coronary artery disease.

http://pharmaceuticalintelligence.com/2012/08/14/obstructive-coronary-artery-disease-diagnosed-by-rna-levels-of-23-genes-cardiodx-heart-disease-test-wins-medicare-coverage/
“By providing Medicare beneficiaries access to Corus CAD, this coverage decision enables patients to avoid unnecessary procedures and risks associated with cardiac imaging and elective invasive angiography, while helping payers address an area of significant healthcare spending,” CardioDx President and CEO David Levison said in a press release.
This discussion will be followed with a discussion of the evaluation of the patient acutely presenting with symptoms and signs that are suggestive of either acute pulmonary or cardiac disease, or both, that may be suggestive of a non ST elevation AMI. It becomes more difficult if ST depression or T-wave inversion is not detected.
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English: QT interval corrected by heart rate. (Photo credit: Wikipedia)

Schematic diagram of normal sinus rhythm for a human heart as seen on ECG (with English labels). (Photo credit: Wikipedia)

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Introduction to Nanotechnology and Alzheimer disease

Posted in Alzheimer's Disease, Biomarkers & Medical Diagnostics, BioSimilars, Cerebrovascular and Neurodegenerative Diseases, Disease Biology, Small Molecules in Development of Therapeutic Drugs, Nanotechnology for Drug Delivery, Pharmacotherapy and Cell Activity, Technology Transfer: Biotech and Pharmaceutical, tagged AD, AD biomarkers, ADDL, Aβ production, Alzheimers Disease, Anti-amyloid protections, Bio-Barcode, CSF biomarkers, nanotechnology, Neuroprotection, neurotoxicity, oxidative stress, plaues on March 14, 2013| 4 Comments »

Author: Tilda Barliya PhD

Alzheimer disease (AD) is among the most common brain disorders affecting the elderly population the world over, and is projected to become a major health problem with grave socio-economic implications in the coming decade (1a, 1b). Alzheimer’s disease arises in large part from the body’s inability to clear these naturally occurring proteins. As amyloid beta levels increase, they tend to aggregate and contribute to the brain “plaques” found in Alzheimer’s disease. There are still no effective treatments to prevent, halt, or reverse Alzheimer’s disease, but research advances over the past three decades could change this gloomy picture. Genetic studies demonstrate that the disease has multiple causes (2). Interdisciplinary approaches have been used to reveal the molecular mechanism of the disease including; biochemistry, molecular and cell biology and transgenic mice models. Progress in chemistry, radiology, and systems biology is beginning to provide useful biomarkers, and the emergence of personalized medicine is poised to transform pharmaceutical development and clinical trials. However, investigative and drug development efforts should be diversified to fully address the multifactoriality of the disease (2). A nice research review shows for example, the effects of cancer drugs on AD treatment (3).

Nanotechnology Solutions for Alzheimer

Dr. Amir Nazem and Dr. G. Ali Mansoori described in their paper “Nanotechnology Solutions for Alzheimer’s Disease: Advances in Research Tools, Diagnostic Methods and Therapeutic Agents”
that he development of nanotechnology approaches for early-stage diagnosis of AD is quite promising but acknowledge that scientists are still at the very beginning of the ambitious project of designing effective drugs and methods for the regeneration of the central nervous system (4). Figure 1- Nanotechnology solutions of AD.

Applications of nanotechnology in AD therapy including:

Nanodiagnostics including imaging
Targeted drug delivery and controlled release
Regenerative medicine

These inclued: neuroprotections against oxidative stress anti-amyloid therapeutics, neuroregeneration and drug delivery beyond the blood brain barrier (BBB) are discussed and analyzed.

All of these applications could improve the treatment approach of AD and other neurodegenerative diseases.

Nanotechnology and Diagnostics:

The diagnosis of AD during life remains difficult and a definite diagnosis of AD relies on histopathological confirmation at post-mortem or by cerebral biopsy. An early clinical diagnosis can be made if patients are tested by trained neuropsychologists. The great problem is not that mild cognitive impairment (MCI) cannot be diagnosed, but that the patients do not see doctor until severely affected (5).

During the last decade, research efforts have focused on developing cerebrospinal fluid (CSF) biomarkers for AD. The diagnostic performance of the CSF biomarkers: Tau protein, the 42-amino acid form of beta amyloid (Aβ42) and Amyloid Precursor Protein are of great importance. One possible biomarker for Alzheimer’s is amyloid beta-derived diffusible ligands (ADDL). The correlation of CSF ADDL levels with disease state offers promise for improved AD diagnosis and early treatment. Singh et al have developed ADDL-specific monoclonal antibodies with an ultrasensitive, nanoparticle-based protein detection strategy termed biobarcode amplification (BCA) (5).

The BCA strategy used by Klein, Mirkin and coworkers makes clever use of nanoparticles as DNA carriers to enable millionfold improvements over ELISA sensitivity. CSF is first exposed to monoclonal anti-ADDL antibodies bound to magnetic microparticles. After ADDL binding, the microparticles are separated with a magnetic field and washed before addition of secondary antibodies bound to DNA:Au nanoparticle conjugates. These conjugates conatin covalently bound DNA as well as complementary “barcode” DNA that is attached via hybridization. Unreacted antibody:DNA:Au nanoparticle conjugates are removed during second magnetic separation, after which elevated temperature and low-salt conditions release the barcode DNA for analysis.

“Such a sensor must be able to transmit any biomarker detection event to an external device that records the transmitted signals and reports an estimated amount for the concentration of AD biomarkers in the CSF. Of course, in order to send such biosensor to a place exposing with CSF, it is necessary to design noninvasive approaches.” (4)

Nanotechnology and treatment:

Presently there exist no therapeutic methods available for curing AD [84]. The cure for AD would require therapeutics that will cease the disease progress and will reverse its resultant damages. Today, common medications for AD are symptomatic and aim at the disrupted neurotransmission between the degenerated neurons. Examples of such medications are acetylcholine esterase inhibitors, including tacrine, donepezil, rivastigmine and galantamine (4).

Design of each mechanistic therapeutic is for targeting a different stage of the AD pathogenetic process and therefore help to cease the progress of the disease. Currently there are 5 mechanistic therapeutic molecular approaches:

Inhibition of Aβ production;
Inhibition of Aβ oligomerization,
Anti-inﬂammation,
Cholesterol homeostasis modulating;
Metal chelation

The nanotechnology approaches are:

Drug discovery and monitoring
Controlled release
Targeted drug delivery

For example: Neuroprotection

Oxidative stress and amyloid induced toxicity are two basic toxicity processes in AD pathogenesis.

Oxidative stress protection:

Fullerene is a nanotechnology building block and can be used to design neuroprotective compounds. It’s chemical structure is known for it’s anti-oxidative and free-scavenger potentials. Applications of functionalized fullerene derivatives including carboxyfullerene and hydroxyfullerene (fullerenols), are promising in discovery of new drugs for AD; however further research on their pharmacodynamic and pharmacokinetic properties is necessary.

Anti-amyloid protections:

Nanotechnology has recently offered some antiamyloid neuroprotective approaches against the cellular and synaptic toxicity of oligomeric and ﬁbrillar (polymeric) Aβ species. The current ongoing nanotechnology research categories on anti-amyloid neuroprotective approaches are the following three:

Prevention from assembly of Aβ monomers
Breaking and resolubilization of the oligomeric or ﬁbrillar (polymeric) Aβ species
Prevention from toxic effects of Aβ

Summary:

AD is a very common disease worldwide, Solving the major problems of early diagnosis and effective cure for AD requires interdisciplinary research efforts. Research on the basic pathogenetic mechanisms of the disease has provided new insight for designing diagnostic and therapeutic methods. Nanotechnology has great potential in aiding and providing tools for diagnosing and treating AD. However, these research combining nanotechnology is still at very early stages and continuous understanding of the disease, neuronal protection and regeneration are needed in order to alleviate the symptoms of the disease.

Ref.
1a. D. G. Georganopoulou et al., “Nanoparticle-based Detection in Cerebral Spinal Fluid of a Soluble Pathogenic Biomarker for Alzheimer’s Disease”, Proc. Natl Acad Sci., 102 (2005) 2273-2276

1b D.A. Davis, W. Klein and L. Chang, “Nanotechnology-based Approaches to Alzheimer’s Clinical Diagnostics”, Nanoscape, 3 (2006) 13-17.
Read more: http://www.nanowerk.com/spotlight/spotid=23726.php#ixzz2NWlx6jYa

2. Huang Y and Mucke L. Alzheimer mechanisms and therapeutic strategies. Cell. 2012 Mar 16;148(6):1204-22.

http://www.cell.com/abstract/S0092-8674(12)00278-4

http://www.ncbi.nlm.nih.gov/pubmed/22424230

3. Cancer Drug Shows Promise in Alzheimer’s Treatment: Helps clear plaque and improve brain function in mice. Alzheimer’s Disease Research is a program of the American Health Assistance Foundation. http://www.nanowerk.com/spotlight/spotid=5262.php

4. Amir Nazem¹, G. Ali Mansoori. Nanotechnology solutions for Alzheimer’s disease: advances in research tools, diagnostic methods and therapeutic agents. J Alzheimers Dis. 2008 Mar;13(2):199-223. http://www.ncbi.nlm.nih.gov/pubmed/18376062?dopt=Abstract.

Full text: http://www.uic.edu/labs/trl/1.OnlineMaterials/08-Nanotechnology_Solutions_for_Alzheimer’s_Disease.pdf

5. Shinjini Singh, Mritunjai Singh, I. S. Gambhir*. Nanotechnology for Alzheimer’s Disease Detection. Digest Journal of Nanomaterials and Biostructures Vol. 3, No.2, June 2008, p. 75 – 79 .

http://www.chalcogen.infim.ro/Singh-Gambhir.pdf

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AMPK Is a Negative Regulator of the Warburg Effect and Suppresses Tumor Growth In Vivo

Posted in Biological Networks, Gene Regulation and Evolution, BioSimilars, CANCER BIOLOGY & Innovations in Cancer Therapy, Cell Biology, Signaling & Cell Circuits, Chemical Biology and its relations to Metabolic Disease, Chemical Genetics, Disease Biology, Small Molecules in Development of Therapeutic Drugs, Genome Biology, Health Economics and Outcomes Research, Metabolomics, Molecular Genetics & Pharmaceutical, Nutrigenomics, Personalized and Precision Medicine & Genomic Research, tagged Allosteric regulation, AMP-activated protein kinase, AMPK, Aviva Lev-Ari, Biology, Cancer - General, Cell Biology, cell proliferation, cellular metabolism, cellular stress, Chemotherapy, DNA, gene expression, genetics, health, medicine, Metabolism, mutation, Myc, redox, research, RNA, Stem cell, tumor response, tumor suppressor, Warburg, Warburg Effect on March 12, 2013| 5 Comments »

AMPK Is a Negative Regulator of the Warburg Effect and Suppresses Tumor Growth In Vivo

Reporter-Curator: Stephen J. Williams, Ph.D.

Word Cloud by Daniel Menzin

There has been a causal link between alterations in cellular metabolism and the cancer phenotype. Reorganization of cellular metabolism, marked by a shift from oxidative phosphorylation to aerobic glycolysis for cellular energy requirements (Warburg effect), is considered a hallmark of the transformed cell. In addition, if tumors are to survive and grow, cancer cells need to adapt to environments high in metabolic stress and to avoid programmed cell death (apoptosis). Recently, a link between cancer growth and metabolism has been supported by the discovery that the LKB1/AMPK signaling pathway as a tumor suppressor axis[1].

LKB1/AMPK/mTOR Signaling Pathway

The Liver Kinase B1 (LKB1)/AMPK AMP-activated protein kinase/mammalian Target of Rapamycin Complex 1 (mTORC1) signaling pathway links cellular metabolism and energy status to pathways involved in cell growth, proliferation, adaption to energy stress, and autophagy. LKB1 is a master control for 14 other kinases including AMPK, a serine-threonine kinase which senses cellular AMP/ATP ratios. In response to cellular starvation, AMPK is allosterically activated by AMP, leading to activation of ATP-generating pathways like fatty acid oxidation and blocking anabolic pathways, like lipid and cholesterol synthesis (which consume ATP). In addition, AMPK regulates cell growth, proliferation, and autophagy by regulating the mTOR pathway. AMPK activates the tuberous sclerosis complex 1/2, which ultimately inhibits mTORC1 activity and inhibits protein translation. This mTOR activity is dis-regulated in many cancers.

LKB1/AMPK in Cancer

Somatic mutations of the STK11 gene encoding LKB1 are detected in lung and cervical cancers
Therefore LKB1 may be a strong tumor suppressor
Pharmacologic activation of LKB1/AMPK with metformin can suppress cancer cell growth

In a recent Cell Metabolism paper[2], Brandon Faubert and colleagues describe how AMPK activity reduces aerobic glycolysis and tumor proliferation while loss of AMPK activity promotes tumor proliferation by shifting cells to aerobic glycolysis and increasing anabolic pathways in a HIF1-dependent manner.

The paper’s major findings were as follows:

Loss of AMPKα1 cooperates with the Myc oncogene to accelerate lymphomagenesis
AMPKα dysfunction enhances aerobic glycolysis (Warburg effect)
Inhibiting HIF-1α reverses the metabolic effects of AMPKα loss
HIF-1α mediates the growth advantage of tumors with reduced AMPK signaling

Summary

AMPK is a metabolic sensor that helps maintain cellular energy homeostasis. Despite evidence linking AMPK with tumor suppressor functions, the role of AMPK in tumorigenesis and tumor metabolism is unknown. Here we show that AMPK negatively regulates aerobic glycolysis (the Warburg effect) in cancer cells and suppresses tumor growth in vivo. Genetic ablation of the α1 catalytic subunit of AMPK accelerates Myc-induced lymphomagenesis. Inactivation of AMPKα in both transformed and nontransformed cells promotes a metabolic shift to aerobic glycolysis, increased allocation of glucose carbon into lipids, and biomass accumulation. These metabolic effects require normoxic stabilization of the hypoxia-inducible factor-1α (HIF-1α), as silencing HIF-1α reverses the shift to aerobic glycolysis and the biosynthetic and proliferative advantages conferred by reduced AMPKα signaling. Together our findings suggest that AMPK activity opposes tumor development and that its loss fosters tumor progression in part by regulating cellular metabolic pathways that support cell growth and proliferation.

Below is the graphical abstract of this paper.

(Photo credit reference(2; Faubert et. al) permission from Elsevier)

However, this regulation of tumor promotion by AMPK may be more complicated and dependent on the cellular environment.

Nissam Hay from the University of Illinois College of Medicine, Chicago, Illinois, USA and his co-workers Sang-Min Jeon and Navdeep Chandel were investigating the mechanism through which LKB1/AMPK regulate the balance between cancer cell growth and apoptosis under energy stress[3]. In their system, the loss of function of either of these proteins makes cells more sensitive to apoptosis in low glucose environments, and cells deficient in either AMPK or LKB1 were shown to be resistant to oncogenic transformation. Whereas previous studies showed (as above) AMPK opposes tumor proliferation in a HIF1-dependent manner, their results showed AMPK could promote tumor cell survival during periods of low glucose or altered redox status.

The researchers incubated LKB1-deficient cancer cells in the presence of either glucose or one of the non-metabolizable glucose analogues 2-deoxyglucose (2DG) and 5-thioglucose (5TG), and found that 2DG, but not 5TG, induced the activation of AMPK and protected the cells from apoptosis, even in cells that were deficient in LKB1.

The authors demonstrated that glucose deprivation depleted NADPH levels, increased H₂O₂ levels and increased cell death, and that this was accelerated in cells deficient in the enzyme glucose-6-phosphate dehydrogenase. Anti-oxidants were also found to inhibit cell death in cells deficient in either AMPK or LKB1.

Knockdown or knockout of either LKB1 or AMPK in cancer cells significantly increased levels of H₂O₂but not of peroxide (O₂^–) during glucose depletion. The glucose analogue 2DG was able to activate AMPK and maintain high levels of NADPH and low levels of H₂O₂ in these cells.

The nucleotide coenzyme NADPH is generated in the pentose phosphate pathway and mitochondrial metabolism, and consumed in H₂O₂ elimination and fatty acid synthesis. If glucose is limited mitochondrial metabolism becomes the major source of NADPH, supported by fatty acid oxidation. AMPK is known to be a regulator of fatty acid metabolism through inhibition of two acetyl-CoA carboxylases, ACC1 and ACC2.

Short interfering RNAs (siRNAs) to knock down levels of both ACC1 and ACC2 in A549 cancer cells and found that only ACC2 knockdown significantly increased peroxide accumulation and apoptosis, while over-expression of mutant ACC1 and ACC2 in LKB1-proficient cells increased H₂O₂ and apoptosis.

Therefore, it was concluded AMPK acts to promote early tumor growth and prevent apoptosis in conditions of energy stress through inhibiting acetyl-CoA carboxylase activity, thus maintaining NADPH levels and preventing the build-up of peroxide in glucose-deficient conditions.

This may appear to be conflicting with the previous report in this post however, it is possible that these reports reflect differences in the way cells respond to various cellular stresses, be it hypoxia, glucose deprivation, or changes in redox status. Therefore a complex situation may arise:

AMPK promotes tumor progression under glucose starvation
AMPK can oppose tumor proliferation under a normoxic, HIF1-dependent manner
Could AMPK regulation be different in cancer stem cells vs. non-stem cell?

References:

1. Green AS, Chapuis N, Lacombe C, Mayeux P, Bouscary D, Tamburini J: LKB1/AMPK/mTOR signaling pathway in hematological malignancies: from metabolism to cancer cell biology. Cell Cycle 2011, 10(13):2115-2120.

2. Faubert B, Boily G, Izreig S, Griss T, Samborska B, Dong Z, Dupuy F, Chambers C, Fuerth BJ, Viollet B et al: AMPK is a negative regulator of the Warburg effect and suppresses tumor growth in vivo. Cell metabolism 2013, 17(1):113-124.

3. Jeon SM, Chandel NS, Hay N: AMPK regulates NADPH homeostasis to promote tumour cell survival during energy stress. Nature 2012, 485(7400):661-665.

Other posts on this site related to Warburg Effect and Cancer include:

Otto Warburg, A Giant of Modern Cellular Biology

Is the Warburg Effect the cause or the effect of cancer: A 21st Century View?

Breast Cancer and Mitochondrial Mutations

AKT signaling variable effects

Bibliographies on Genomics by Subject Matter

Volume One: Genomics Orientations for Individualized Medicine

The Initiation and Growth of Molecular Biology and Genomics – Part I

Ubiquitin-Proteosome pathway, Autophagy, the Mitochondrion, Proteolysis and Cell Apoptosis: Part III

Novel Cancer Hypothesis Suggests Antioxidants Are Harmful

Mitochondrial Damage and Repair under Oxidative Stress

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Liver Endoplasmic Reticulum Stress and Hepatosteatosis

Posted in Biological Networks, Gene Regulation and Evolution, Biomarkers & Medical Diagnostics, Cell Biology, Signaling & Cell Circuits, Chemical Biology and its relations to Metabolic Disease, Computational Biology/Systems and Bioinformatics, Disease Biology, Small Molecules in Development of Therapeutic Drugs, Genome Biology, International Global Work in Pharmaceutical, Liver & Digestive Diseases Research, Metabolomics, Molecular Genetics & Pharmaceutical, Nutrigenomics, Nutrition, Nutritional Supplements: Atherogenesis, lipid metabolism, Personalized and Precision Medicine & Genomic Research, Pharmaceutical Industry Competitive Intelligence, Population Health Management, Genetics & Pharmaceutical, Population Health Management, Nutrition and Phytochemistry, Scientist: Career considerations, Systemic Inflammatory Response Related Disorders, Technology Transfer: Biotech and Pharmaceutical, tagged alcohol consumption, Biochemistry and Molecular Biology, Cholesterol, ChREBP, dyslipidemia, Endoplasmic reticulum, ER stress, ERS, Foufelle F Inserm, gene expression, Glycolysis, hepatic steatosis, IL-6, inflammatory cytokin, Insulin, lipid biosynthesis, metabolic syndrome, NAFLD, NASH, NFkB, Non-alcoholic fatty liver disease, Phosphorylation, signaling pathways, signaling protein, SREPB, TNF-a, Transcription factor, type 2 diabetes, Unfolded protein response on March 10, 2013| 2 Comments »

Liver Endoplasmic Reticulum Stress and Hepatosteatosis

Larry H Bernstein, MD, FCAP

1. Absence of adipose triglyceride lipase protects from hepatic endoplasmic reticulum stress in mice.

Fuchs CD, Claudel T, Kumari P, Haemmerle G, et al.
LabExpMol Hepatology, Medical Univ of Graz, Austria.
Hepatology. 2012 Jul;56(1):270-80. http://dx.doi.org/10.1002/hep.25601. Epub 2012 May 29.

Nonalcoholic fatty liver disease (NAFLD) is characterized by

triglyceride (TG) accumulation and
endoplasmic reticulum (ER) stress.

Fatty acids (FAs) may trigger ER stress, therefore,

the absence of adipose triglyceride lipase (ATGL/PNPLA2)-
- the main enzyme for intracellular lipolysis,
releasing FAs, and
closest homolog to adiponutrin (PNPLA3)

recently implicated in the pathogenesis of NAFLD-

could protect against hepatic ER stress.

Wild-type (WT) and ATGL knockout (KO) mice

were challenged with tunicamycin (TM) to induce ER stress.

Markers of hepatic

lipid metabolism,
ER stress, and
inflammation were explored
- for gene expression by
- serum biochemistry,
- hepatic TG and FA profiles,
- liver histology,
- cell-culture experiments were performed in Hepa1.6 cells
after the knockdown of ATGL before FA and TM treatment.

TM increased hepatic TG accumulation in ATGL KO, but not in WT mice. Lipogenesis and β-oxidation
were repressed at the gene-expression level (sterol regulatory element-binding transcription factor 1c,
fatty acid synthase, acetyl coenzyme A carboxylase 2, and carnitine palmitoyltransferase 1 alpha) in
both WT and ATGL KO mice. Genes for very-low-density lipoprotein (VLDL) synthesis (microsomal
triglyceride transfer protein and apolipoprotein B)

were down-regulated by TM in WT
and even more in ATGL KO mice,
which displayed strongly reduced serum VLDL cholesterol levels.

ER stress markers were induced exclusively in TM-treated WT, but not ATGL KO, mice:

glucose-regulated protein,
C/EBP homolog protein,
spliced X-box-binding protein,
endoplasmic-reticulum-localized DnaJ homolog 4, and
inflammatory markers Tnfα and iNos.

Total hepatic FA profiling revealed a higher palmitic acid/oleic acid (PA/OA) ratio in WT mice.
Phosphoinositide-3-kinase inhibitor-

known to be involved in FA-derived ER stress and
blocked by OA-
was increased in TM-treated WT mice only.

In line with this, in vitro OA protected hepatocytes from TM-induced ER stress. Lack of ATGL may protect from
hepatic ER stress through alterations in FA composition. ATGL could constitute a new therapeutic strategy
to target ER stress in NAFLD.
PMID: 22271167 Diabetes Obes Metab. 2010 Oct;12 Suppl 2:83-92.
http://dx.doi.org/10.1111/j.1463-1326.2010.01275.x.

2. Hepatic steatosis: a role for de novo lipogenesis and the transcription factor SREBP-1c.
Ferré P, Foufelle F. INSERM, and Université Pierre et Marie Curie-Paris, Paris, France. PMID: 21029304

Excessive availability of plasma fatty acids and lipid synthesis from glucose (lipogenesis) are important determinants of steatosis.
Lipogenesis is an insulin- and glucose-dependent process that is under the control of specific transcription factors,

sterol regulatory element binding protein 1c (SREBP-1c), activated by
insulin and carbohydrate response element binding protein (ChREBP) activated by glucose.

Insulin induces the maturation of SREBP-1c in the endoplasmic reticulum (ER).

SREBP-1c in turn activates glycolytic gene expression,
- allowing glucose metabolism, and
- lipogenic genes in conjunction with ChREBP.

Lipogenesis activation in the liver of obese markedly insulin-resistant steatotic rodents is then paradoxical.
It appears the activation of SREBP-1c and thus of lipogenesis is

secondary in the steatotic liver to an ER stress.

The ER stress activates the

cleavage of SREBP-1c independent of insulin,
explaining the paradoxical stimulation of lipogenesis
in an insulin-resistant liver.

Inhibition of the ER stress in obese rodents

decreases SREBP-1c activation and lipogenesis and
improves markedly hepatic steatosis and insulin sensitivity.
ER is thus worth considering as a potential therapeutic target for steatosis and metabolic syndrome.

3. SREBP-1c transcription factor and lipid homeostasis: clinical perspective
Ferré P, Foufelle F
Inserm, Centre de Recherches Biomédicales des Cordeliers, Paris, France.
Horm Res. 2007;68(2):72-82. Epub 2007 Mar 5. PMID:17344645

Insulin has long-term effects on glucose and lipid metabolism through its control on the expression of specific genes.
In insulin sensitive tissues and particularly in the liver,

the transcription factor sterol regulatory element binding protein-1c (SREBP-1c) transduces the insulin signal, which is
synthetized as a precursor in the membranes of the endoplasmic reticulum
which requires post-translational modification to yield its transcriptionally active nuclear form.

Insulin activates the transcription and the proteolytic maturation of SREBP-1c, which induces the

expression of a family of genes
involved in glucose utilization and fatty acid synthesis and
can be considered as a thrifty gene.

Since a high lipid availability is

deleterious for insulin sensitivity and secretion,
a role for SREBP-1c in dyslipidaemia and type 2 diabetes
has been considered in genetic studies.

SREBP-1c could also participate in

hepatic steatosis observed in humans
related to alcohol consumption and
hyperhomocysteinemia
concomitant with a ER-stress and
insulin-independent SREBP-1c activation.

4. Hepatic steatosis: a role for de novo lipogenesis and the transcription factor SREBP-1c
Ferré P, Foufelle F
INSERM, Centre de Recherches des Cordeliers and Université Pierre et Marie Curie-Paris, Paris, France.
Diabetes Obes Metab. 2010 Oct;12 Suppl 2:83-92. PMID: 21029304
http://dx.doiorg/10.1111/j.1463-1326.2010.01275.x.

Lipogenesis in liver steatosis is

an insulin- and glucose-dependent process
under the control of specific transcription factors,
sterol regulatory element binding protein 1c (SREBP-1c),
activated by insulin and carbohydrate response element binding protein (ChREBP)

Insulin induces the maturation of SREBP-1c in the endoplasmic reticulum (ER).
SREBP-1c in turn activates glycolytic gene expression, allowing –

glucose metabolism in conjunction with ChREBP.

activation of SREBP-1c and lipogenesis is secondary in the steatotic liver to ER stress, which

activates the cleavage of SREBP-1c independent of insulin,
explaining the stimulation of lipogenesis in an insulin-resistant liver.
Inhibition of the ER stress in obese rodents decreases SREBP-1c activation and improves
hepatic steatosis and insulin sensitivity.

ER is thus a new partner in steatosis and metabolic syndrome

5. Pharmacologic ER stress induces non-alcoholic steatohepatitis in an animal model
Jin-Sook Leea, Ze Zhenga, R Mendeza, Seung-Wook Hac, et al.
Wayne State University SOM, Detroit, MI
Toxicology Letters 20 May 2012; 211(1):29–38 http://dx.doi.org/10.1016/j.toxlet.2012.02.017

Endoplasmic reticulum (ER) stress refers to a condition of

accumulation of unfolded or misfolded proteins in the ER lumen, which is known to
activate an intracellular stress signaling termed
Unfolded Protein Response (UPR).

A number of pharmacologic reagents or pathophysiologic stimuli

can induce ER stress and activation of the UPR signaling,
leading to alteration of cell physiology that is
associated with the initiation and progression of a variety of diseases.

Non-alcoholic steatohepatitis (NASH), characterized by hepatic steatosis and inflammation, has been considered the
precursor or the hepatic manifestation of metabolic disease. In this study, we delineated the

toxic effect and molecular basis
by which pharmacologic ER stress,
induced by a bacterial nucleoside antibiotic tunicamycin (TM),
promotes NASH in an animal model.

Mice of C57BL/6J strain background were challenged with pharmacologic ER stress by intraperitoneal injection of TM. Upon TM injection,

mice exhibited a quick NASH state characterized by
hepatic steatosis and inflammation.

TM-treated mice exhibited an increase in –

hepatic triglycerides (TG) and a –
decrease in plasma lipids, including
plasma TG,
plasma cholesterol,
high-density lipoprotein (HDL), and
low-density lipoprotein (LDL),

In response to TM challenge,

cleavage of sterol responsive binding protein (SREBP)-1a and SREBP-1c,
the key trans-activators for lipid and sterol biosynthesis,
was dramatically increased in the liver.

Consistent with the hepatic steatosis phenotype, expression of

some key regulators and enzymes in de novo lipogenesis and lipid droplet formation was up-regulated,
while expression of those involved in lipolysis and fatty acid oxidation was down-regulated
in the liver of mice challenged with TM.

TM treatment also increased phosphorylation of NF-κB inhibitors (IκB),

leading to the activation of NF-κB-mediated inflammatory pathway in the liver.

Our study not only confirmed that pharmacologic ER stress is a strong “hit” that triggers NASH, but also demonstrated

crucial molecular links between ER stress,
lipid metabolism, and
inflammation in the liver in vivo.

Highlights
► Pharmacologic ER stress induced by tunicamycin (TM) induces a quick NASH state in vivo.
► TM leads to dramatic increase in cleavage of sterol regulatory element-binding protein in the liver.
► TM up-regulates lipogenic genes, but down-regulates the genes in lipolysis and FA oxidation.
► TM activates NF-κB and expression of genes encoding pro-inflammatory cytokines in the liver.
Abbreviations
ER, endoplasmic reticulum; TM, tunicamycin; NASH, non-alcoholic steatohepatitis; NAFLD,
non-alcoholic fatty liver disease; TG, triglycerides; SREBP, sterol responsive binding protein;
NF-κB, activation of nuclear factor-kappa B; IκB, NF-κB inhibitor
Keywords: ER stress; Non-alcoholic steatohepatitis; Tunicamycin; Lipid metabolism; Hepatic inflammation
Figures and tables from this article:

Fig. 1. TM challenge alters lipid profiles and causes hepatic steatosis in mice. (A) Quantitative real-time RT-PCR analysis of liver mRNA isolated from mice challenged with TM or vehicle control. Total liver mRNA was isolated at 8 h or 30 h after injection with vehicle or TM (2 μg/g body weight) for real-time RT-PCR analysis. Expression values were normalized to β-actin mRNA levels. Fold changes of mRNA are shown by comparing to one of the control mice. Each bar denotes the mean ± SEM (n = 4 mice per group); **P < 0.01. Edem1, ER degradation enhancing, mannosidase alpha-like 1. (B) Oil-red O staining of lipid droplets in the livers of the mice challenged with TM or vehicle control (magnification: 200×). (C) Levels of TG in the liver tissues of the mice challenged with TM or vehicle control. (D) Levels of plasma lipids in the mice challenged with TM or vehicle control. TG, triglycerides; TC, total plasma cholesterol; HDL, high-density lipoproteins; VLDL/LDL, very low and low density lipoproteins. For C and D, each bar denotes mean ± SEM (n = 4 mice per group); *P < 0.05; **P < 0.01.

Fhttp://ars.els-cdn.com/content/image/1-s2.0-S0378427412000732-gr1.jpgigure options

Fig. 2. TM challenge leads to a quick NASH state in mice. (A) Histological examination of liver tissue sections of the mice challenged with TM (2 μg/g body weight) or vehicle control. Upper panel, hematoxylin–eosin (H&E) staining of liver tissue sections; the lower panel, Sirius staining of collagen deposition of liver tissue sections (magnification: 200×). (B) Histological scoring for NASH activities in the livers of the mice treated with TM or vehicle control. The grade scores were calculated based on the scores of steatosis, hepatocyte ballooning, lobular and portal inflammation, and Mallory bodies. The stage scores were based on the liver fibrosis. Number of mice examined is given in parentheses. Mean ± SEM values are shown. P-values were calculated by Mann–Whitney U-test.

http://ars.els-cdn.com/content/image/1-s2.0-S0378427412000732-gr2.jpg

Fig. 3. TM challenge significantly increases levels of cleaved/activated forms of SREBP1a and SREBP1c in the liver. Western blot analysis of protein levels of SREBP1a (A) and SREBP1c (B) in the liver tissues from the mice challenged with TM (2 μg/g body weight) or vehicle control. Levels of GAPDH were included as internal controls. For A and B, the values below the gels represent the ratios of mature/cleaved SREBP signal intensities to that of SREBP precursors. The graph beside the images showed the ratios of mature/cleaved SREBP to precursor SREBP in the liver of mice challenged with TM or vehicle. The protein signal intensities shown by Western blot analysis were quantified by NIH imageJ software. Each bar represents the mean ± SEM (n = 3 mice per group); **P < 0.01. SREBP-p, SREBP precursor; SREBP-m, mature/cleaved SREBP.

http://ars.els-cdn.com/content/image/1-s2.0-S0378427412000732-gr3.jpg

Fig. 4. TM challenge up-regulates expression of genes involved in lipogenesis but down-regulates expression of genes involved in lipolysis and FA oxidation. Quantitative real-time RT-PCR analysis of liver mRNAs isolated from the mice challenged with TM (2 μg/g body weight) or vehicle control, which encode regulators or enzymes in: (A) de novo lipogenesis: PGC1α, PGC1β, DGAT1 and DGAT2; (B) lipid droplet production: ADRP, FIT2, and FSP27; (C) lipolysis: ApoC2, Acox1, and LSR; and (D) FA oxidation: PPARα. Expression values were normalized to β-actin mRNA levels. Fold changes of mRNA are shown by comparing to one of the control mice. Each bar denotes the mean ± SEM (n = 4 mice per group); **P < 0.01. (E and F) Isotope tracing analysis of hepatic de novo lipogenesis. Huh7 cells were incubated with [1-14C] acetic acid for 6 h (E) or 12 h (F) in the presence or absence of TM (20 μg/ml). The rates of de novo lipogenesis were quantified by determining the amounts of [1-14C]-labeled acetic acid incorporated into total cellular lipids after normalization to cell numbers.

http://ars.els-cdn.com/content/image/1-s2.0-S0378427412000732-gr4.jpg

Fig. 5. TM activates the inflammatory pathway through NF-κB, but not JNK, in the liver. Western blot analysis of phosphorylated Iκ-B, total Iκ-B, phosphorylated JNK, and total JNK in the liver tissues from the mice challenged with TM (2 μg/g body weight) or vehicle control. Levels of GAPDH were included as internal controls. The values below the gels represent the ratios of phosphorylated protein signal intensities to that of total proteins.

http://ars.els-cdn.com/content/image/1-s2.0-S0378427412000732-gr5.jpg

Fig. 6. TM induces expression of pro-inflammatory cytokines and acute-phase responsive proteins in the liver. Quantitative real-time RT-PCR analyses of liver mRNAs isolated from the mice challenged with TM (2 μg/g body weight) or vehicle control, which encode: (A) pro-inflammatory cytokine TNFα and IL6; and (B) acute-phase protein SAP and SAA3. Expression values were normalized to β-actin mRNA levels. Fold changes of mRNA are shown by comparing to one of the control mice. (C–E) ELISA analyses of serum levels of TNFα, IL6, and SAP in the mice challenged with TM or vehicle control for 8 h ELISA. Each bar denotes the mean ± SEM (n = 4 mice per group); *P < 0.05, **P < 0.01.

http://ars.els-cdn.com/content/image/1-s2.0-S0378427412000732-gr6.jpg

Corresponding author at: Center for Molecular Medicine and Genetics, Wayne State University School of Medicine, 540 E. Canfield Avenue, Detroit, MI 48201, USA. Tel.: +1 313 577 2669; fax: +1 313 577 5218.

The SREBP regulatory pathway. Brown MS, Goldstein JL (1997). “The SREBP pathway: regulation of cholesterol metabolism by proteolysis of a membrane-bound transcription factor”. Cell 89 (3) : 331–340. doi:10.1016/S0092-8674(00)80213-5. PMID 9150132. (Photo credit: Wikipedia)

English: Structure of the SREBF1 protein. Based on PyMOL rendering of PDB 1am9. (Photo credit: Wikipedia)

The SREBP regulatory pathway (Photo credit: Wikipedia)

English: Diagram of rough endoplasmic reticulum by Ruth Lawson, Otago Polytechnic. (Photo credit: Wikipedia)

Micrograph demonstrating marked (macrovesicular) steatosis in non-alcoholic fatty liver disease. Masson’s trichrome stain. (Photo credit: Wikipedia)

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Imaging-biomarkers is Imaging-based tissue characterization

Posted in Advanced Drug Manufacturing Technology, Bio Instrumentation in Experimental Life Sciences Research, Biomarkers & Medical Diagnostics, CANCER BIOLOGY & Innovations in Cancer Therapy, Cancer Prevention: Research & Programs, Computational Biology/Systems and Bioinformatics, Disease Biology, Small Molecules in Development of Therapeutic Drugs, Drug Delivery Platform Technology, Ecosystems & Industrial Concentration in the Medical Device Sector, FDA Regulatory Affairs, FDA, CE Mark & Global Regulatory Affairs: process management and strategic planning - GCP, GLP, ISO 14155, Genomic Testing: Methodology for Diagnosis, Health Economics and Outcomes Research, HealthCare IT, Imaging-based Cancer Patient Management, International Global Work in Pharmaceutical, Medical Devices R&D and Inventions, Medical Devices R&D Investment, Medical Imaging Technology, Image Processing/Computing, MRI, CT, Nuclear Medicine, Ultra Sound, Personalized and Precision Medicine & Genomic Research, Pharmaceutical R&D Investment, Pharmacotherapy and Cell Activity, Population Health Management, Genetics & Pharmaceutical, Regulated Clinical Trials: Design, Methods, Components and IRB related issues, Technology Transfer: Biotech and Pharmaceutical, tagged bio-markers, biomarkers, Cancer - General, cancer management, imaging-bio-markers, MRI, MRI sequence, research, tissue characterisation on March 10, 2013| 3 Comments »

Imaging-biomarkers is Imaging-based tissue characterization

Author – Writer: Dror Nir, PhD

Article 11.1 Introduction by Dror Nir PhD

For everyone who is skeptical about the future role of imaging-based tissue chracterisation in the management of cancer, the following “Statement paper” ESR statement on the stepwise development of imaging biomarkers published online: 9 February 2013, by the European Society of Radiology (ESR), should provide substantial reassurance that this kind of technology will become a must! In support of this claim I quote the following information:

“The European Society of Radiology and its related European Institute for Biomedical Imaging Research (EIBIR) should have a relevant role in coordinating future developments of biomarkers and in the assessment and validation of imaging biomarkers as surrogate end points.

Acknowledgements

This paper was kindly prepared by the ESR Subcommittee on Imaging Biomarkers (Chairperson: Bernard Van Beers. Research Committee Chairperson: Luis Martí-Bonmatí. Members: Marco Essig, Thomas Helbich, Celso Matos, Wiro Niessen, Anwar Padhani, Harriet C. Thoeny, Siegfried Trattnig, Jean-Paul Vallée. Co-opted members: Peter Brader, Nicolas Grenier) on behalf of the European Society of Radiology (ESR) and with the help of Sabrina Doblas, INSERM U773, Paris, France.

It was approved by the ESR Executive Council in December 2012..”

According to ESR: “There is increasing interest in developing the quantitative imaging of biomarkers in personalised medicine”. In this perspective, “Biomarkers” are tissue properties that can be quantitatively and reproducibly measured by imaging devices. One example for a major unmet need, which I found to be most interesting is the imaging-based detection of tumor invasiveness.

Quoting from the paper: ” Biomarkers are defined as “characteristics that are objectively measured and evaluated as indicators of normal biological processes, pathological processes, or pharmaceutical responses to a therapeutic intervention” [1]. Broadly, biomarkers fall into two categories: bio-specimen biomarkers, including molecular biomarkers and genetic biomarkers, and bio-signal biomarkers or imaging biomarkers. Bio-specimen biomarkers are obtained by removing a sample from a patient. Examples of these molecular biomarkers are genes and proteins detected from fluids or tissue samples. Bio-signal biomarkers remove no material from the patient, but rather detect and analyse an electromagnetic, photonic or acoustic signal emitted by the patient [2]. These imaging biomarkers have the advantage of being non-invasive, spatially resolved and repeatable [3]. They are of particular interest if they can overcome the limitations of the established histological “gold standards”. Indeed, invasive reference examinations, such as biopsy, can be inconclusive, are non-representative of the whole tissue (which is a tremendous limitation when assessing malignant tumours, which are known to be heterogeneous) and possess non-negligible levels of mortality and morbidity.

Genetic biomarkers indicate whether a disease may occur, but they are usually inefficient to assess the presence and stage of a disease. Similar to molecular biomarkers, imaging biomarkers can be used for early detection of diseases, staging and grading, and predicting or assessing the response to treatment [3]. Accordingly, because of their relative lower cost compared with imaging, molecular biomarkers may be more appropriate for disease screening and early detection than imaging biomarkers. With their high sensitivity, molecular biomarkers could also detect subclinical stages of disease before any morphological or functional change is detectable on imaging. In contrast, imaging biomarkers are often more useful than molecular biomarkers for disease staging, and also grading and for assessing tumour response, because localised information is crucial.”

The main messages ESR wishes to deliver in this paper are that:

• Using imaging-biomarkers to streamline drug discovery and disease progression will drive a huge advancement in healthcare.

• The clinical qualification and validation of imaging biomarkers technology pose challenges, mainly in establishing the accuracy and reproducibility of such techniques. In that respect, agreements on standards and evaluation methods (e.g. clinical studies design) is imperative.

• There should be high motivation to pursue the development of imaging-biomarkers as the “clinical value of new biomarkers is of the highest priority in terms of patient management, assessing risk factors and disease prognosis.”

The paper deals to a great extent with the requirements on accuracy, reproducibility, standardization and quality control from the process of developing imaging-biomarkers:

“Accuracy: Before being routinely used in the clinic, imaging biomarkers must be validated. Determining the accuracy implies calculating the sensitivity and specificity of the biomarker when compared with a biological process, such as tumour necrosis, which can be assessed at histopathological examination… [6–9] [10, 11]

Reproducibility: Repeatability (measurements at short intervals on the same subjects using the same equipment in the same centres) and reproducibility (measurements at short intervals on the same subjects using different facilities in the same and different centres) studies must be conducted for image acquisition and image analysis…. Reproducibility studies are now very often included in scientific papers, as advised by the “standards for reporting of diagnostic accuracy” (STARD) criteria and should ideally include Bland-Altman plots and results of coefficients of repeatability [16, 17].

Standardisation: Standardisation relates to the establishment of norms or requirements about technical aspects. In the development of imaging biomarkers, two main aspects should be considered: Standardisation of image acquisition and Standardisation of image analysis… [18] [19–21] [22] [27, 28] [31–33]

Quality control: Adequate phantoms could be used to validate, on a day-to-day basis, that the biomarker stays robust and to avoid any drift in the machine, acquisition or processing protocol…. [34] [30, 35] [36] [37] [23].”

The proposed development workflow:

“Similar to new drugs, the development of biomarkers has to pass along a pipeline going from discovery, through verification in different laboratories, validation and qualification before they can be used in clinical routine. Validation includes the determination of the accuracy and the precision (reproducibility) of the biomarker and standardisation concerns both acquisition and analysis. Qualification, defined as a “graded, fit-for-purpose evidentiary process linking a biomarker with biological processes and clinical end-points”, is a validation process in large cohorts of patients involving multiple centres, similar to phase III clinical trials, to obtain regulatory approval as surrogate endpoints [4]. A more extensive path to biomarker development has been reported [5]. The first step is the proof of concept, which defines any specific change relevant to the disease that can be studied using the available imaging and computational techniques. The relationship between this change and the presence, grading and response to treatment of the disease constitutes the proof of mechanism. The images needed to extract the biomarker must be appropriate (in terms of resolution, signal and contrast behaviour). Preparation of images relates to improving the data before the analysis (such as segmentation, filtering, interpolation or registration). The analysis and modelling of the signal by computational numerical adjustment of a mathematical model allow extracting the needed information (such as structural, physical, chemical, biological and functional properties). After this voxel-by-voxel computation, the spatial distribution of the biomarker can be depicted by parametric images, defined as derived secondary images which pixels represent the distribution values of a given parameter. Multivariate parametric images obtained by statistical modelling of the relevant parameters allow the reduction of data and a clear definition of the defined disease target. The abnormal values should be defined and measured through histogram analysis. A pilot test on a small sample of subjects, with and without the disease, has to be performed to validate the process—also called proof of principle—and to evaluate the influence of potential variations related to age, sex or any other source of biases. Finally, proofs of efficacy and effectiveness on larger and well-defined series of patients will show the ability of a biomarker to measure the clinical endpoint (Fig. 1).”

Steps for the development of imaging biomarkers (adapted from [5])

The authors admit that the requirement posed on development of imaging-biomarkers represents a huge challenge and they try to offer ideas, mainly taken from the “MRI experience” to overcome certain hurdles. There is one important point on which they do not discuss: the definition of appropriate reference test. It is my own experience, based on many study protocols I developed in the past decade, that without reaching an agreement on that point, the development of imaging-biomarkers will just move in circles. Note, that today’s most “acceptable” reference test is histopathology, which everyone admits (as well mentioned in this paper); suffers many limitations. When it comes to validating imaging-biomarkers, the need to accurately match imaging products with histopathology is an additional major hurdle.

This is why, I see as a necessary step, to develop “real-time” imaging based tissue characterization combined with in-situ imaging-based histology.

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Alzheimer’s Disease Conundrum – Are We Near the End of the Puzzle?

Posted in Advanced Drug Manufacturing Technology, Biological Networks, Gene Regulation and Evolution, Biomarkers & Medical Diagnostics, Cell Biology, Signaling & Cell Circuits, Cerebrovascular and Neurodegenerative Diseases, Chemical Genetics, Disease Biology, Small Molecules in Development of Therapeutic Drugs, Genome Biology, Genomic Testing: Methodology for Diagnosis, Human Immune System in Health and in Disease, Innovations in Neurophysiology & Neuropsychology, International Global Work in Pharmaceutical, Medical Imaging Technology, Image Processing/Computing, MRI, CT, Nuclear Medicine, Ultra Sound, Molecular Genetics & Pharmaceutical, Pharmaceutical Industry Competitive Intelligence, Proteomics, Regulated Clinical Trials: Design, Methods, Components and IRB related issues, Technology Transfer: Biotech and Pharmaceutical, tagged Alois Alzheimer, Alzheimer, Alzheimer Disease, Alzheimers Disease, ApoE, BCHE brain imaging, Beta amyloid, biomarkers, cellular inflammatiory response, CLU, conitive loss, Decode Genetics, dementia, Icelanders, innovative applications, Kári Stefánsson, mapping, Massachusetts General Hospital, Picalm, plaque regression, Tau protein on March 9, 2013| 1 Comment »

Alzheimer’s Genomic Diagnosis and Treatment

Curator: Larry H Bernstein, MD, FCAP

Gene Mutation Protects Against Alzheimer’s

by Greg Miller on 11 July 2012
Brain preserver. A newly discovered gene mutation appears to protect against Alzheimer’s disease. Credit: Alzheimer’s Disease Education and Referral Center/NIA/NIH
http://news.sciencemag.org/sciencenow/2012/07/gene-mutation-protects-against-a.html

A rare mutation that alters a single letter of the genetic code protects people from the

memory-robbing dementia of Alzheimer’s disease.

The DNA change may inhibit the buildup of β amyloid, the

protein fragment that forms the hallmark plaques in the brains of Alzheimer’s patients.
The mutation affects a gene called APP,
which encodes a protein that gets broken down into pieces,
including β amyloid.

Researchers previously identified more than 30 mutations to APP, none of them good. Several of these changes increase β amyloid formation and cause

• a devastating inherited form of Alzheimer’s that afflicts people in their 30s and 40s—

• much earlier than the far more common “late-onset” form of Alzheimer’s

that typically strikes people their 70s and 80s.

The new mutation, discovered from whole-genome data from 1795 Icelanders for variations in APP that protect against Alzheimer’s, appears to do the opposite. The mutation interferes with one of the enzymes that breaks down the APP protein and causes a 40% reduction in β amyloid formation

New pharmacological strategies for treatment of Alzheimer’s disease: focus on disease modifying drugs.
Salomone S, Caraci F, Leggio GM, Fedotova J, Drago F.
University of Catania, Viale Andrea Doria 6, Catania, Italy.
Br J Clin Pharmacol. 2012 Apr;73(4):504-17. doi: 10.1111/j.1365-2125.2011.04134.x.

Current approved drug treatments for Alzheimer disease (AD) include

cholinesterase inhibitors (donepezil, rivastigmine, galantamine) and the
NMDA receptor antagonist memantine.

These drugs provide symptomatic relief but poorly affect the progression of the disease. Drug discovery has been directed, in the last 10 years, to develop ‘disease modifying drugs’ hopefully able to counteract the progression of AD. Because in a chronic, slow progressing pathological process, such as AD, an early start of treatment enhances the chance of success,

it is crucial to have biomarkers for early detection of AD-related brain dysfunction,
- usable before clinical onset.

Reliable early biomarkers need therefore to be prospectively tested for predictive accuracy,

with specific cut off values validated in clinical practice.

Disease modifying drugs developed so far include drugs to

reduce β amyloid (Aβ) production,
drugs to prevent Aβ aggregation,
drugs to promote Aβ clearance,
drugs targeting tau phosphorylation and assembly

None of these drugs has demonstrated efficacy in phase 3 studies. The failure of clinical trials with disease modifying drugs raises a number of questions, spanning from

methodological flaws to
fundamental understanding of AD pathophysiology and biology.

Diagnostic criteria applicable to presymptomatic stages of AD have now been published.

These new criteria may impact on drug development, such that future trials on disease modifying drugs will include populations susceptible to AD, before clinical onset. http://www.ncbi.nlm.nih.gov/pubmed/22035455

Gene mutation defends against Alzheimer’s disease
Rare genetic variant suggests a cause and treatment for cognitive decline.
Ewen Callaway 11 July 2012
http://www.nature.com/news/gene-mutation-defends-against-alzheimer-s-disease-1.10984

J. NIETH/CORBIS
Almost 30 million people live with Alzheimer’s disease worldwide, a staggering health-care burden that is expected to quadruple by 2050. Yet doctors can offer no effective treatment, and scientists have been unable to pin down the underlying mechanism of the disease.
Research published this week offers some hope on both counts – few people carry a genetic mutation that naturally prevents them from developing the condition – 0.5% of Icelanders have a protective gene, as are 0.2–0.5% of Finns, Swedes and Norwegians. Icelanders who carry it have a 50% better chance of reaching age 85, are more than five times more likely to reach it 85 without Alzheimer’s. The mutation seems to put a brake on the milder mental deterioration that most elderly people experience. Carriers are about 7.5 times more likely than non-carriers to reach the age of 85 without major cognitive decline, and perform better on the cognitive tests that are administered thrice yearly to Icelanders who live in nursing homes.
The discovery not only confirms the principal suspect that is responsible for Alzheimer’s, it also suggests that the disease could be

an extreme form of the cognitive decline seen in many older people.

The mutation — the first ever found to protect against the disease — lies in a gene that produces

amyloid-β precursor protein (APP),
which has an unknown role in the brain

APP was discovered 25 years ago in patients with rare,

inherited forms of Alzheimer’s that strike in middle age.
In the brain, APP is broken down into a smaller molecule called amyloid-β.

Visible clumps, or plaques, of amyloid-β found in the autopsied brains of patients are a hallmark of Alzheimer’s.
Scientists have long debated whether the plaques are a cause of the neurodegenerative condition

or a consequence of other biochemical changes associated with the disease.

The latest finding supports other genetics studies blaming amyloid-β, according to Rudolph Tanzi, a neurologist at the Massachusetts General Hospital in Boston and a member of one of the four teams that discovered APP’s role in the 1980s.
If amyloid-β plaques were confirmed as the cause of Alzheimer’s, it would bolster efforts to develop drugs that block their formation, says Kári Stefánsson, chief executive of deCODE Genetics in Reykjavik, Iceland, who led the latest research. He and his team first discovered the mutation by comparing the complete genome sequences of 1,795 Icelanders with their medical histories. The researchers then studied the variant in nearly 400,000 more Scandinavians.
This suggests that Alzheimer’s disease and cognitive decline are two sides of the same coin, with a common cause — the build-up of amyloid-β plaques in the brain, something seen to a lesser degree in elderly people who do not develop full-blown Alzheimer’s. A drug that mimics the effects of the mutation, might slow cognitive decline as well as prevent Alzheimer’s.
Stefánsson and his team discovered that the mutation introduces a single amino-acid alteration to APP. This amino acid is close to the site where an enzyme called

β-secretase 1 (BACE1) ordinarily snips APP into smaller amyloid-β chunks —
and the alteration is enough to reduce the enzyme’s efficiency.

Stefánsson’s study suggests that blocking β-secretase from cleaving APP has the potential to prevent Alzheimer’s, but Philippe Amouyel, an epidemiologist at the Pasteur Institute in Lille, France, says “it is very difficult to identify the

precise time when this amyloid toxic effect could still be modified”.

“If this effect needs to be blocked as early as possible in life to protect against Alzheimer’s disease, we will need to propose a new design for clinical trials” to identify an effective treatment.

The results demonstrate that whole-genome sequencing can uncover very rare mutations that might offer insight into common diseases.

disease risk, may be determined by genetic variants that slightly tilt the odds of developing disease
In this case a rare mutant may provide very key mechanistic insights into Alzheimer’s

Jonsson, T. et al. Nature http://dx.doi.org/10.1038/nature11283 (2012).
Kang, J. et al. Nature 325, 733–736 (1987).
Goldgaber, D., Lerman, M. I., McBride, O. W., Saffiotti, U. & Gajdusek, D. C. Science 235, 877–880 (1987).

BHCE genetic data combined with brain imaging using agent florbetapir connects the BHCE gene to AD plaque buildup. BHCE is an enzyme that breaks down acetylcholine in the brain, which is depleted early in the disease and results in memory loss. http://www.genengnews.com/

New Alzheimer’s Genes Found
Gigantic Scientific Effort Discovers Clues to Treatment, Diagnosis of Alzheimer’s Disease
By Daniel J. DeNoon
WebMD Health News Reviewed by Laura J. Martin, MD
http://www.webmd.com/alzheimers/news/20110403/new-alzheimers-genes-found

A massive scientific effort has found five new gene variants linked to Alzheimer’s disease. The undertaking involved analyzing the genomes of nearly 40,000 people with and without Alzheimer’s. This study was undertaken by two separate research consortiums in the U.S. and in Europe, which collaborated to confirm each other’s results.
Four genes had previously been linked to Alzheimer’s. Three of them affect only the risk of relatively rare forms of Alzheimer’s. The fourth is APOE, until now the only gene known to affect risk of the common, late-onset form of Alzheimer’s. Roughly 27% of Alzheimer’s disease can be attributed to the five new gene variants. Even though Alzheimer’s is a very complex disease, the new findings represent a large chunk of Alzheimer’s risk, according to Margaret A. Pericak-Vance, PhD, of the U.S. consortium –

20% of the causal risk of Alzheimer’s disease and
32% of the genetic risk.

Alzheimer’s Tied to Mutation Harming Immune Response
By GINA KOLATA Published: November 14, 2012 in NY Times
http://www.nytimes.com/2012/11/15/health/gene-mutation-that-hobbles-immune-response-is-linked-to-alzheimers.html?_r=0
Alzheimer’s researchers and drug companies have for years concentrated on one hallmark of Alzheimer’s disease: the production of toxic shards of a protein that accumulate in plaques on the brain.
Two groups of researchers working from entirely different starting points have converged on a mutated gene involved in another aspect of Alzheimer’s disease:

the immune system’s role in protecting against the disease.

The mutation is suspected of interfering with

the brain’s ability to prevent the buildup of plaque.

When the gene is not mutated, white blood cells in the brain spring into action,

gobbling up and eliminating the plaque-forming toxic protein, beta amyloid.

As a result, Alzheimer’s can be staved off or averted. People with the mutated gene have a threefold to fivefold increase in the likelihood of developing Alzheimer’s disease in old age.

Comparing Differences

Dr. Julie Williams’s, Cardiff, Wales (European team leader) report identified CLU and Picalm. A second study published in Nature Genetics, by Philippe Amouyel from Institut Pasteur de Lille in France, pinpointed CLU and CR1. The greatest inherited risk comes from the APOE gene, discovered in 1993 by a team led by Allen Roses, now director of the Deane Drug Discovery Institute at Duke UMC, in Durham, North Carolina.
The findings “are beginning to give us insight into the biology, but I don’t think you can expect treatments overnight,” Dr. Michael Owen (Cardiff, Wales) said. Instead, the genes will show a mosaic of risk, and “the key issue is what hand of cards you’re dealt,” he said.

Promise for Early Diagnosis
BHCE genetic data combined with brain imaging using agent florbetapir connects the BHCE gene to AD plaque buildup. BHCE is an enzyme that breaks down acetylcholine in the brain, which is depleted early in the disease and results in memory loss.

Dr. Bernstein’s comments:

There has been a long history of failure of drugs to slow down the progression of Alzheimer’s. Regression of the plaques has not corresponded with retention of cognitive ability, which has been behind the arguments over beta amyloid or tau.
We now have two particularly interesting mutations –
1. ApoE gene mutation that increases risk
2. APP mutation that quite dramatically affects retention of cognition

Alzheimer’s Disease: Green Tea Extract Found To Interfere With The Formation Of Amyloid Plaques (medicalnewstoday.com)
Aging Brain – The Specific Gene That Causes It (belmarrahealth.com)
The Pathogenic Aβ43 Is Enriched in Familial and Sporadic Alzheimer Disease (plosone.org)
Damaged blood vessels loaded with amyloid worsen cognitive impairment in Alzheimer’s disease (eurekalert.org)
Major step toward an Alzheimer’s vaccine (nextbigfuture.com)
Damaged blood vessels loaded with amyloid worsen cognitive impairment in Alzheimer’s disease (medicalxpress.com)
Top Signs You are at Risk for Alzheimer’s (belmarrahealth.com)
A Reliable Way to Detect Endogenous Murine β-Amyloid (plosone.org)

β-amyloid fibrils. (Photo credit: Wikipedia)

English: PET scan of a human brain with Alzheimer’s disease (Photo credit: Wikipedia)

Depiction of amyloid precursor protein processing, created by I. Peltan Ipeltan (Photo credit: Wikipedia)

English: Diagram of how microtubules desintegrate with Alzheimer’s disease Français : La protéine Tau dans un neurone sain et dans un neurone malade Español: Esquema que muestra cómo se desintegran los microtúbulos en la enfermedad de Alzheimer (Photo credit: Wikipedia)

English: Histopathogic image of senile plaques seen in the cerebral cortex in a patient with presenile onset of Alzheimer disease. Bowdian stain. The same case as shown in a file “Alzheimer_dementia_(1)_presenile_onset.jpg”. (Photo credit: Wikipedia)

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Biological Therapeutics for Asthma

Posted in Advanced Drug Manufacturing Technology, Biological Networks, Gene Regulation and Evolution, Disease Biology, Small Molecules in Development of Therapeutic Drugs, Genome Biology, Human Immune System in Health and in Disease, International Global Work in Pharmaceutical, Pharmaceutical Industry Competitive Intelligence, tagged Allergy, Antibodies, Asthma, Biology, Food and Drug Administration, Immunoglobulin E, Immunology, Johns Hopkins School of Medicine, Monoclonal antibodies, Therapy on March 8, 2013| 1 Comment »

Biological Therapeutics for Asthma

Curator: Larry H Bernstein, MD, FCAP

Update on Biological Therapeutics for Asthma

Marisha L. Cook, MD, and Bruce S. Bochner, MD
Department of Medicine, Division of Allergy and Clinical Immunology
Johns Hopkins University School of Medicine, Baltimore, MD

BASIC AND CLINICAL TRANSLATIONAL SCIENCE IN ALLERGY, ASTHMA AND IMMUNOLOGY
WAO Journal 2010; 3:188–194
Difficulty in managing severe asthma has encouraged research about its pathobiology and treatment options. Novel biologic therapeutics are being developed for the treatment of asthma and are of potential use for severe refractory asthma, especially where the increased cost of such agents is more likely justified. This review summarizes currently approved (omalizumab) and investigational biologic agents for asthma, such as

antibodies,
soluble receptors,
other protein-based antagonists,

and highlight recent published data on efficacy and safety of these therapies in humans. As these newer agents with highly targeted pharmacology are tested in asthma,

we are also poised to learn more about the role of cytokines and other molecules in the pathophysiology of asthma.

Key Words: asthma, biologic therapies, cytokines, monoclonal antibodies

Despite the well-known and fairly consistent efficacy of

drugs such as inhaled corticosteroids, leukotriene modifiers

and 2 agonists for the majority of asthmatics, as many as

10% suffer from severe disease inadequately controlled by

conventional therapy. Severe and sustained symptoms lead to

poor quality of life, disproportionate use of health care

resources, and significant adverse effects. Novel biologic therapeutics are being developed for the treatment of asthma and are of potential use for severe refractory asthma, especially where the increased cost of such agents is more likely justified.
This review will briefly summarize what is meant by “biologic therapies” and then highlight recent published data on efficacy and safety of these therapies for asthma.

WHAT ARE BIOLOGIC THERAPIES?

Biologic therapies have revolutionized the treatment of many diseases including asthma. By definition, the term “biologics” or “biologicals” include a variety of protein based therapeutics, such as antibodies, soluble receptors (eg,etanercept), recombinant protein-based receptor antagonists (eg, pitrakinra) and other related structures. Their main advantages include the duration of action and highly specific and strong binding to the target of interest; their main disadvantages are the cost and need for parenteral administration. Most biologicals in clinical use are antibodies, and their generic names contain standard nomenclature as a suffix to

indicate their origins (Fig. 1). Initially, pure murine antibodies were created with hybridoma technology, generating therapies that were 100% mouse with generic names given the suffix “momab” (eg, ibritumomab); however, immunogenicity of mouse antibodies in human subjects caused reduced efficacy and increased risk of infusion reactions including anaphylaxis and death. To reduce immunogenicity, chimeric antibodies

(“ximabs” like rituximab) were engineered. These antibodies are a marriage of murine variable regions combined with human constant regions, creating antibodies that are 80% human. These were a step forward but still had the potential for being immunogenic. Humanized monoclonal antibodies (“zumabs” such as omalizumab) go one step further, where now only the hypervariable regions of the mouse antibody are retained,

while the remaining 95% of the antibody is molecularly replaced by human sequences.

In the latest approach, fully human antibodies (“umabs” such as adalimumab) can be created by using phage display technology and molecular biology or more directly by immunizing mice that have had their immunoglobulin genes replaced with human versions. Newer artificial antibody structures such as bispecific antibodies, mix 2 separate arms with 2 different binding specificities to target 2 different types of antigens [eg, a single antibody where one arm binds interleukin (IL)-4 and the other arm binds IL-13]. Standard nomenclature for mAbs identifies their source with the last 4 or 5 letters: -omab, murine: –ximab, chimeric: -zumab, humanized: and –umab, human. The middle part of the name reflects the disease indication for which the mAb was initially intended: -lim for immune and inflammatory diseases, -cir for cardiovascular disorders, and -tu for tumors or neoplastic conditions. The first 3 or 4 letters may be chosen by the sponsor. Modified (by adding the structure of a bispecific antibody) . In general, FDA-approved mAbs have emerged between 10 and 12 years after the date that the new technologies on which they were based were reported in the scientific literature. None of these newer antibody structures have been tried in asthma, so the remainder of this review will focus on available data with standard biologicals.
Here is a listing of the key focus on biomolecules for therapeutics:
IL-4

It induces the IgE isotype switch and up-regulates expression of vascular cell adhesion molecule-1 on endothelium and a variety of TH2 chemokines, thus promoting recruitment of T lymphocytes, monocytes, basophils, and eosinophils to sites of allergic inflammation. A clinical trial studied the soluble recombinant human IL-4 receptor (IL-4R), Nuvance in asthma. Nuvance inhibited a decline in FEV1 during inhaled corticosteroid withdrawal and was overall well tolerated.2,3 However, in subsequent clinical trials in patients taking only beta agonist, soluble IL-4R failed to demonstrate significant clinical efficacy. A phase I randomized double blind placebo controlled study evaluated the effects of pascolizumab, a humanized anti-IL-4 antibody, in 24 patients with mild to moderate asthma. Pascolizumab was well tolerated and no serious adverse events occurred.5 However, a phase IIa clinical trial in steroid-naive, mild to moderate asthmatics, did not demonstrate clinical efficacy. Because the IL-4 targeting studies have failed to demonstrate clinical efficacy, one can justify concluding that either IL-4R is not an effective therapeutic target in asthma.

TNFa

Tumor necrosis factor (TNF) is a multifunctional proinflammatory cytokine produced by inflammatory cells including monocytes, macrophages, mast cells, smooth muscle cells, and epithelial cells. TNF may initiate airway inflammation by up-regulating adhesion molecules, mucin hypersecretion, and airway remodeling, and by synergizing with TH2 cytokines. Berry et al demonstrated that severe refractory asthmatics have evidence of up-regulation of TNF as compared with healthy controls and mild asthmatics. Entanercept was evaluated in a small, randomized, double-blind placebo-controlled crossover study in 10 patients with severe refractory asthma and elevated TNF levels, 10 patients with mild to moderate asthma, and 10 control patients. Entanercept treatment was associated with improved FEV1, asthma related quality of life, and the concentration of methacholine needed to provoke a 20% decrease in FEV1. No serious adverse reactions were noted. In another double-blind, placebo-controlled, parallel group study, 38 patients with moderate asthma on inhaled corticosteroids were treated with infliximab. Although infliximab treatment did not improve the primary end point of morning peak expiratory flow, it decreased diurnal variation of the peak expiratory flow rate and asthma exacerbations. No serious adverse events were noted. Golimumab was recently evaluated in the largest randomized, double-blind, placebo-controlled study in 309 patients with severe, uncontrolled asthma. No significant differences were observed for the change in FEV1 or exacerbations. However, several serious adverse events occurred. There is no clear role for TNF in perpetuating asthma or asthma exacerbations.

CD4

CD4 T cells are likely to be involved as a source of proinflammatory cytokines in asthma. Keliximab is a monoclonal antibody that causes a transient reduction in the number of CD4 T cells. A double blind, randomized, placebo controlled study with 22 severe oral corticosteroid dependent asthmatics patients was completed. A subset of patients received the highest dose of keliximab (3.0 mg/kg). There was significant improvement of peak expiratory flow rates in the high dose treatment arm. However, CD4 T cells remained transiently reduced 14 days postinfusion, raising safety concerns.

CD23

CD23 is a low-affinity immunoglobulin E receptor (FcRII) and is important in regulating IgE production. IDEC-152 is a chimeric monoclonal antibody directed against CD23. CD23 is expressed on

T and B cells,
neutrophils,
monocytes, and
macrophages.

CD23 is overexpressed in allergic disease and may be involved in IgE overproduction,

which can lead to mast cell degranulation.

A phase I dose escalating placebo-controlled study in 30 asthmatics demonstrated that

IDEC-152 caused a dose-dependent reduction in serum IgE concentrations.
- No significant adverse events were reported

CD25

Airway inflammation is associated with activated CD25 T cells, IL-2, and soluble IL-2 receptors. Daclizumab is a humanized monoclonal antibody directed against the alpha subunit of the high affinity IL-2 receptor (CD25). This inhibits IL-2 binding and release of inflammatory cytokines. A randomized, double-blind, placebo-controlled, parallel group study was performed (115 patients, 88 to the treatment arm, 27 to placebo)to evaluate the efficacy of daclizumab in patients with moderate to severe asthma poorly controlled on inhaled corticosteroids. Treatment with daclizumab led to improvements in FEV1, daytime asthma symptoms, and rescue 2 agonist use,but the effects were modest.

IgE

Omalizumab is a humanized monoclonal anti-IgE antibody that binds free circulating IgE and prevents the interaction between IgE and high affinity (FcRI) and low affinity (FcRII) IgE receptors on inflammatory cells. Omalizumab also down-regulates the surface expression of FcRI on basophils, mast cells, and dendritic cells. Omalizumab decreases free IgE levels and reduces FcRI receptor expression on mast cells and basophils. This results in decreased mast cell activation and sensitivity, leading to a reduction in eosinophil influx and activation. Anti-IgE treatment with omalizumab might result in decreased mast cell survival. Omalizumab also reduces dendritic cell FcRI receptor expression. The primary end point in a phase III randomized prospective trial was the number of exacerbation episodes during the steroid reduction period and the stable steroid period. During the stable steroid phase, fewer omalizumab subjects than placebo subjects experienced one or more exacerbations (14.6 vs. 23.3%; P 0.009). During the steroid reduction phase, the omalizumab group had fewer subjects with exacerbations (21.3 vs. 32.3%; P 0.04). The median reduction in inhaled corticosteroid dose was significantly greater in the omalizumab group than in the placebo group (75 vs. 50%; P 0.001). The efficacy of omalizumab was demonstrated in other clinical trials including INNOVATE. INNOVATE was a double-blind, parallel-group study in which 419 subjects were randomized to receive omalizumab or placebo for 28 weeks. The omalizumab group had a 26% reduction in the rate of clinically significant exacerbations compared with placebo (.68 vs. .91, P 0.042). A recent omalizumab observational study of 280 subjects demonstrates similar findings. After 6 months, they found a reduction in daily symptoms by 80%, nocturnal symptoms by 86%, asthma exacerbations by 82%, hospitalizations by 76%, unscheduled health care visits by 81%, and improvement in quality of life (Mini Asthma Quality of Life Questionnaire increased from 2.9 to 4.5 after 6 months of treatment).

Examining the effects of biologic agents provides unique and valuable insight into the pathobiology of asthma. Furthermore, it is an ideal opportunity to identify mechanisms inherent to severe refractory asthma. The development of biologic agents has been a slow and arduous process; however, a substantial amount of progress has been achieved. Although omalizumab is an expensive medical treatment, therapy may be cost effective in patients with uncontrolled severe persistent allergic asthma because the majority of the economic burden is in this population. Hopefully ongoing efforts with biologicals will lead to improved management options for our most severe asthma patients.

More information is available from the article: World Allergy Organ J. 2010;3(6):188–194. http://dx.doi.org/10.1097/WOX.0b013e3181e5ec5a
PMCID: PMC2922052 NIHMSID: NIHMS221446
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2922052/figure/F2/ http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2922052/bin/waoj-3-188-g002.gif http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2922052/figure/F3/ http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2922052/bin/waoj-3-188-g003.gif
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2922052/

#Shopping# : Therapeutic antibody engineering: Current and future advances driving the strongest growth area in the pharmaceutical industry (Woodhead Publishing Series in Biomedicine) (lacking29inv.wordpress.com)
Novartis has been confirmed as key speaker for SMi Group’s 9th Asthma & COPD event (prweb.com)
Research and Markets: Monoclonal Antibodies Market in Rheumatoid Arthritis to 2018 – Market … (biomedreports.com)
BioLineRx’s EDP-14, for Treatment of Severe and Persistent Asthma, to Enter the Company’s Main Thera (dailyfinance.com)

English: Overview of hybridoma technology and monoclonal antibody creation (Photo credit: Wikipedia)

Mast cells are involved in allergy. Allergies such as pollen allergy are related to the antibody known as IgE. Like other antibodies, each IgE antibody is specific; one acts against oak pollen, another against ragweed. (Photo credit: Wikipedia)

Emil von Behring (Photo credit: Wikipedia)

Diagram showing the production of monoclonal antibodies via hybridoma technology (Photo credit: Wikipedia)

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In focus: Melanoma therapeutics

Posted in CANCER BIOLOGY & Innovations in Cancer Therapy, Cell Biology, Signaling & Cell Circuits, Chemical Biology and its relations to Metabolic Disease, Disease Biology, Small Molecules in Development of Therapeutic Drugs, FDA Regulatory Affairs, Human Immune System in Health and in Disease, Molecular Genetics & Pharmaceutical, Pharmaceutical Analytics, Regulated Clinical Trials: Design, Methods, Components and IRB related issues, tagged Aldesleukin, American Cancer Society, angiogenesis, biochemotherapy, BRAF, Chemotherapy, Clinical Trials, Dacarbazine, Eastern Cooperative Oncology Group, ECOG, FDA, gp100, IFN-gamma, IFN-α-2b, IL-2, Immunotherapy, Interferon gamma, Interleukin-2, Ipilimumab, Jak3, MAPK, Melanoma, metastatic, OS, overall survival, PFS, progression-free survival, Proleukin, response rate, RR, Temozolomide, Therapy, vaccine, VEGFR, Vemurafenib, Yervoy on March 8, 2013| 5 Comments »

In focus: Melanoma therapeutics

Author and Curator: Ritu Saxena, Ph.D.

In the last post of Melanoma titled “In focus: Melanoma Genetics”, I discussed the clinical characteristics and the genetics involved in Melanoma. This post would discuss melanoma therapeutics, both current and novel.

According to the American Cancer Society, more than 76,000 new cases and more than 9100 deaths from melanoma were reported in the United States in 2012[1] Melanoma develops from the malignant transformation of melanocytes, the pigment-producing cells that reside in the basal epidermal layer in human skin. Although most melanomas arise in the skin, they may also arise from mucosal surfaces or at other sites to which neural crest cells migrate.

Melanoma therapeutics

Surgical treatment of cutaneous melanoma employs specific surgical margins depending on the depth of invasion of the tumor and there are specific surgical treatment guidelines of primary, nodal, and metastatic melanoma that surgeons adhere to while treatment. Melanoma researchers have been focusing on developing adjuvant therapies for that would increase the survival post-surgery.

Chemotherapy

Among traditional chemotherapeutic agents, only dacarbazine is FDA approved for the treatment of advanced melanoma (Eggermont AM and Kirkwood JM, Eur J Cancer, Aug 2004;40(12):1825-36). Dacarbazine is a triazene derivative and alkylates and cross-links DNA during all phases of the cell cycle, resulting in disruption of DNA function, cell cycle arrest, and apoptosis. Currently, 17 clinical trials are underway to test the efficacy and effectiveness of dacarbazine against melanoma as either a single agent or in combination chemotherapy regimens with other anti-cancer chemotherapeutic agents such as cisplatin, paclitaxel. Temozolomide is a triazene analog of dacarbazine and is approved for the treatment of malignant gliomas. At physiologic pH, it is converted to a short-lived active cytotoxic compound, monomethyl triazeno imidazole carboxamide (MTIC). MTIC methylates DNA at the O6 and N7 positions of guanine, resulting in inhibition of DNA replication. Unlike dacarbazine, which is metabolized to MITC only in the liver, temozolomide is metabolized to MITC at all sites. Temozolomide is administered orally and penetrates well into the central nervous system. Temozolomide is being tested in many combination regimens for patients with melanoma metastatic to the brain (Douglas JG and Margolin K, Semin Oncol, Oct 2002;29(5):518-24).

Immunotherapy

Melanoma and the immune system are closely related. Hence, immunotherapy has been explored in the treatment of the disease. The two most widely investigated immunotherapy drugs for melanoma are Interferon (IFN)-alpha and Interleukin-2 (IL-2).

The role of IFNalpha-2b in the adjuvant therapy of patients with localized melanoma at high risk for relapse was established by the results of three large randomized trials conducted by the US Intergroup; all three trials demonstrated an improvement in relapse-free survival and two in overall survival. One of these trials, a large randomized multicenter trial performed by the Eastern Cooperative Oncology Group (ECOG), in high-risk melanoma patients showed significant improvements in relapse-free and overall survival with adjuvant IFN-α-2b therapy, compared with standard observation (ECOG 1684). The results of the study led to FDA approval of IFN-α-2b for treatment of melanoma. This study was performed on patients with deep primary tumors without lymph node involvement and node-positive melanomas. In other studies, little antitumor activity has been demonstrated in IFN-α-2b–treated metastatic stage IV melanoma.

Recombinant IL-2 showed an overall response rate of 15-20% in metastatic melanoma and was capable of producing complete and durable remissions in about 6% of patients treated. Based upon these data, the US FDA has approved the use of high-dose IL-2 for the therapy of patients with metastatic melanoma. Aldesleukin (Brand name: Proleukin) is a recombinant analog of the endogenous cytokine interleukin-2 (IL-2). It binds to and activates the IL-2 receptor (IL-2R), followed by heterodimerization of the IL-2R beta and gamma(c) cytoplasmic chains; activation of Jak3; and phosphorylation of tyrosine residues on the IL-2R beta chain, resulting in an activated receptor complex (NCI). The activated complex recruits several signaling molecules that act as substrates for regulatory enzymes associated with the complex. It is administered intravenously and stimulates lymphokine-activating killer (LAK) cells, natural killer (NK) cells and the production of cytokines such as gamma interferon (nm|OK). Several clinical trials are currently underway using Aldesleukin to determine the efficacy of combination treatment in melanoma patients.

Another anti-cancer immunotherapeuty-based mechanism involved inhibition of inhibitory signal of cytotoxic T lymphocyte-associated antigen 4 (CTLA-4), a molecule on T-cells that plays a critical role in regulating natural immune responses. Ipilimumab (Brand name: Yervoy) was by FDA for melanoma treatment. It is a human monoclonal antibody (MAb) T-cell potentiator that specifically blocks CTLA-4. It is approved for inoperable advanced (Stage III) or metastatic (Stage IV) melanoma in newly diagnosed or previously treated patients (nm|OK). The approval, March 25, 2011, was based on a randomized (3:1:1) double-blind double-dummy clinical trial (MDX010-20) in patients with unresectable or metastatic melanoma who had received at least one prior systemic treatment for melanoma. Patients were randomly assigned to receive either ipilimumab, 3 mg/kg intravenously, in combination with the tumor vaccine (n=403); ipilimumab plus vaccine placebo (n=137); or tumor vaccine with placebo (n=136). Patients treated with ipilimumab alone had a median overall survival (OS) of 10 months while those treated with tumor vaccine had a median OS of 6 months. The trial also demonstrated a statistically significant improvement in OS for patients treated with the combination of ipilimumab plus tumor vaccine compared with patients treated with tumor vaccine alone. For more information on the trial, check the clinical trials website, www.clinicaltrials.gov

Signaling pathway inhibitors

Approximately 90% of BRAF gene mutations involve valine (V) to glutamic acid (E) mutation at number 600 residue (V600E). The resulting oncogene product, BRAF (V600E) kinase is highly active and exhibits elevated MAPK pathway. The BRAF(V600E) gene mutation occurs in approximately 60% of melanomas indicating that it could be therapeutically relevant. Vemurafenib (Brand name: Zelboraf) is a novel small-molecule inhibitor of BRAF (V600E) kinase. It selectively binds to the ATP-binding site and inhibits the activity of BRAF (V600E) kinase. Vemurafebib inhibits over active MAPK pathway by inhibiting the mutated BRAF kinase, thereby reducing tumor cell proliferation (NCI). Encouraging results of phase III randomized, open-label, multicenter trial were reported recently at the 2011 ASCO meeting (Chapman PB, et al, ASCO 2011, Abstract LBA4). The trial compared the novel BRAF inhibitor vemurafenib with dacarbazine in patients with BRAF-mutated melanoma. Previously untreated, unresectable stage IIIC or stage IV melanoma that tested positive for BRAF mutation were randomized (1:1) to vemurafenib or dacarbazine. The response rate (RR) was significantly high (48.4%) in vemurafenib treated patients as compared to 5.5% in dacarbazine among the 65% of patients evaluable for RR to date. In addition, vemurafenib was associated with significantly improved OS and PFS compared to dacarbazine in patients with previously untreated BRAF (V600E) mutation bearing patients with metastatic melanoma.

Biochemotherapy

Biochemothreapy combine traditional chemotherapy with immunotherapies, such as IL-2 and IFN-α-2b. These combination therapies seemed promising in phase II trials, however, seven large studies failed to show statistically significant increased overall survival rates for various biochemotherapy regimens in patients with stage IV metastasis (Margolin KA, et al, Cancer, 1 Aug 2004;101(3):435-8). Owing to inconsistent results of the available studies with regard to benefit including RR, OS and progression time, and consistently high toxicity rates, clinical practice guideline do not recommend biochemotherapy for the treatment of metastatic melanoma (Verma S, et al, Curr Oncol, April 2008; 15(2): 85–89).

Vaccines

The use of therapeutic vaccines is an ongoing area of research, and clinical trials of several types of vaccines (whole cell, carbohydrate, peptide) are being conducted in patients with intermediate and late-stage melanoma. Vaccines are also being tested in patients with metastatic melanoma to determine their immune effects and to define their activity in combination with other immunotherapeutic agents such as IL-2 or IFNalpha (Agarwala S, Am J Clin Dermatol, 2003;4(5):333-46). In fact, recently investigators at the Indiana University Health Goshen Center for Cancer Care (Goshen, IN) conducted a randomized, multicenter phase III trial involving 185 patients with stage IV or locally advanced stage III cutaneous melanoma. The patients were assigned into treatment groups with IL-2 alone or with vaccine (gp100) followed by IL-2. The vaccine-IL-2 group had a significantly improved OR as compared to the IL-2-only group (16% Vs. 6%) and longer progression free survival (2.2 months Vs. 1.6 months). The median overall survival was also longer in the vaccine-interleukin-2 group than in the interleukin-2-only group (17.8 months Vs. 11.1 months). Thus, a combination of vaccine and immunotherapy showed a better response rate and longer progression-free survival than with interleukin-2 alone in patients with advanced melanoma (Schwartzentruber DJ, et al, N Engl J Med, 2 Jun 2011;364(22):2119-27).

Which Treatment When?

Earlier, there were essentially two main options for patients suffering from advanced melanoma, dacarbazine and IL-2. Dacarbazine, a chemotherapeutic agent produces modest improvements in survival or symptomatic benefits in most patients. Interleukin-2 -based drugs, on the other hand, induce long-term remissions in a small group of patients but are highly toxic. Recently, FDA approved ipilimumab and vemurafenib for patients with metastatic melanoma. Apart from these, therapies are also aiming at starving the tumor by inhibiting angiogenesis or depleting nutrients essential for cancer growth. Of the antiangiogenic compounds, VEGFR inhibitors SU5416 and AG-013736 demonstrated broad-spectrum antitumor activity in mice bearing xenografts of human cancer cell lines originating from various tissues, including melanoma. In addition, several trials are currently underway to test the efficacy of the drugs in combination. In the future, personalized medicine-based recommendations of novel and existing drugs for melanoma patients might be the way to go.

Reference:

Related articles on Melanoma on this Open Access Online Scientific Journal:

In focus: Melanoma Genetics Curator- Ritu Saxena, Ph.D.
Thymosin alpha1 and melanoma Author/Editor- Tilda Barliya, Ph.D.
A New Therapy for Melanoma Reporter- Larry H Bernstein, M.D.
Melanoma: Molecule in Immune System Could Help Treat Dangerous Skin Cancer Reporter: Prabodh Kandala, Ph.D.
Why Braf inhibitors fail to treat melanoma. Reporter: Prabodh Kandala, Ph.D.

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Genomics & Genetics of Cardiovascular Disease Diagnoses: A Literature Survey of AHA’s Circulation Cardiovascular Genetics, 3/2010 – 3/2013

Posted in Atherogenic Processes & Pathology, Biological Networks, Gene Regulation and Evolution, Biomarkers & Medical Diagnostics, Cardiovascular Pharmacogenomics, Chemical Genetics, Computational Biology/Systems and Bioinformatics, Disease Biology, Small Molecules in Development of Therapeutic Drugs, FDA Regulatory Affairs, Frontiers in Cardiology and Cardiovascular Disorders, Genome Biology, Genomic Testing: Methodology for Diagnosis, HTN, Liver & Digestive Diseases Research, Metabolomics, Molecular Genetics & Pharmaceutical, Origins of Cardiovascular Disease, Personalized and Precision Medicine & Genomic Research, Pharmaceutical Analytics, Pharmacogenomics, Population Health Management, Genetics & Pharmaceutical, Proteomics, Regulated Clinical Trials: Design, Methods, Components and IRB related issues, Statistical Methods for Research Evaluation, Systemic Inflammatory Response Related Disorders, Technology Transfer: Biotech and Pharmaceutical, tagged Cardiovascular disease, Carlos Bustamante, CVD, genetics, Genome-wide association study, GWAS, Hyperlipidemia, Larry H. Bernstein, pharmacogenomics, Single-nucleotide polymorphism, Stanford University, Stanford University School of Medicine on March 7, 2013| 25 Comments »

Genomics & Genetics of Cardiovascular Disease Diagnoses: A Literature Survey of AHA’s Circulation Cardiovascular Genetics, 3/2010 – 3/2013

Curators: Aviva Lev-Ari, PhD, RN and Larry H. Bernstein, MD, FCAP

348 articles that appeared in AHA’s Circulation Cardiovascular Genetics, 3/2010 – 3/2013 were classified by the curators of this article into the following TEN categories. The first 9, represent DIAGNOSES of cardiovascular diseases, the last, deals with Pharmacogenomics.

The Cardiovascular Diagnoses that were covered in the period of 3/2010 – 3/2013, include the following:

Preventative Cardiology

MicroRNA in Serum as Bimarker for Cardiovascular Pathologies: acute myocardial infarction, viral myocarditis, diastolic dysfunction, and acute heart failure

Genetic Determinants of Potassium Sensitivity and Hypertension

Heart and Aging Research in Genomic Epidemiology: 1700 MIs and 2300 coronary heart disease events among about 29 000 eligible patients

Genomics of Ventricular arrhythmias, A-Fib, Right Ventricular Dysplasia, Cardiomyopathy

Genetics of CVD and Hyperlipidemia, Hyper Cholesterolemia, Metabolic Syndrome

Genetics and Vascular Pathologies and Platelet Aggregation, Cardiac Troponin T in Serum

Genomics and Valvular Disease

Heredity of Cardiovascular Disorders Inheritance

Pharmacogenomics

Introductions

Larry H. Bernstein, MD, FCAP

The curation of this large amount of material in 10 categories begins with a first chapter on preventative cardiology, which has had much public attention for the last decade. Much of the concern with preventive cardiology has emphasized diet and exercise. There is much to be said about this in articles not yet written. However, there are several decades of research on the amino acid composition of foods, and the essential fatty acids, that indicates an essential balance between proinflammatory and antiinflammatory fatty acids in polyunsaturated fatty acids, and of the harmful effects of saturated fats. There is also much to be said of essential amino acids, and in particular, those essential for methylation processes, and sulfur metabolism.

The next eight chapters are all concerned with genomics in cardiovascular disease. This is in no small part a follow up on the completion of the genetic code in 2003, a seminal event. Let us look at these in clusters.

[1] microRNA in serum is now considered for a biomarker for cardiovascular disease. It can be measured at very low levels, but we don’t yet know where it fits. It might be more revealing once we understand the adaptive mechanism in development of congestive heart failure, renal hypertension, and post-genomic events.

[2] It appears to me that potassium sensitivity and hypertension approached from the genomic side is more complicate. Why is that? The kidney excretes a sodium load and in metabolic acidosis, the serum potassium rises with a metabolic acidemia that can’t be compensated by the respiratory loss of CO2 through the carbonic anhydrase mechanism.

[3] Heart and aging research is a rich area for work on the long term post-genomic changes, and it involves a large population base.

[4][5] The genomics of cardiac dysrrhytmias and cardiomyopathies will open new doors into our understanding of the mechanisms of these diseases, and perhaps find therapeutic targets. There has been a large volume of work on lipid synthesis, the role of the liver in generating apolipoproteins, and this has new answers on the way. The most important feature, not readily accepted is the measurement of particles, which has now been done by a monoclonal antibody. Metabolic syndrome brings together adipose tissue metabolism, endocrine and changes in CRP and IL-1.

[6] Vascular pathologies and coagulation, hyperviscosity has had an enormous increase in intensity of research. The concept of plaque rupture to account for all AMIs is being modified, and the high sensitivity cardio-specific troponins have become the most widely use test.

[7] The genomics of valvular disease fits with the increased surgical procedures for valvular disease related to atheroschlerosis and advent of minimally invasive surgical procedures for the reapir and replacement of valves, procedure called TAVR vs. Openhealrt surgery for valve replacement.

[8] Inherited cardiovascular disease is an older family of disorders, going back to Victor McKusik, and also the “Blue Baby” operation, both at Johns Hopkins.

[9] Pharmacogenomics is a vary active field of investigation and has uncovered inter-individual differences in handling Warfarin as a starter.

Preventative Cardiology

Methods in Genetics and Clinical Interpretation Randomized Trial of Personal Genomics for Preventive Cardiology Design and Challenges

Joshua W. Knowles, MD, PhD, Themistocles L. Assimes, MD, PhD, Michaela Kiernan, PhD, Aleksandra Pavlovic, BS, Benjamin A. Goldstein, PhD, Veronica Yank, MD, Michael V. McConnell, MD, Devin Absher, PhD, Carlos Bustamante, PhD, Euan A. Ashley, MD, DPhil and John P.A. Ioannidis, MD, DSc

Author Affiliations

From the Division of Cardiovascular Medicine (J.W.K., T.L.A., A.P., M.V.M., E.A.A.), Stanford Prevention Research Center (M.K., V.Y., J.P.A.I.), Division of General Medical Disciplines (V.Y.), Department of Genetics (C.B.), Department of Health Research and Policy (J.P.A.I.), Stanford University School of Medicine, Stanford, CA; Quantitative Sciences Unit, Stanford University School of Medicine, Palo Alto, CA (B.A.G.); HudsonAlpha Institute for Biotechnology, Huntsville, AL (D.A.); Department of Statistics, Stanford University School of Humanities and Sciences, Stanford, CA (J.P.A.I.).

Correspondence to Joshua W. Knowles, MD, PhD, Stanford University School of Medicine, Division of Cardiovascular Medicine, Falk CVRC, 300 Pasteur Dr, Stanford, CA 94305. E-mail knowlej@stanford.edu

Background

Genome-wide association studies (GWAS) have identified more than 1500 disease-associated single nucleotide polymorphisms (SNPs), including many related to atherosclerotic cardiovascular disease (CVD). Associations have been found for most traditional risk factors (TRFs), including lipids,1^,2 blood pressure/hypertension,3^,4 weight/body mass index,5^,6 smoking behavior,7 and diabetes.8–13 GWAS have also identified susceptibility variants for coronary heart disease (CHD). The first and, so far, strongest of these signals was found in the 9p21.3 locus, where common variants in this region increase the relative risk of CVD by 15% to 30% per risk allele in most race/ethnic groups.13–20 Subsequent large-scale GWAS meta-analyses and replication studies in largely white/European populations have led to the reliable identification of an additional 26 loci conferring susceptibility to CHD,2^,20–23 all with substantially lower effects sizes compared with the 9p21 locus. Many of these CVD susceptibility loci appear to be conferring risk independent of TRFs and thus cannot currently be assessed by surrogate clinical measures (Table 1). Among the 27 independent loci identified in the most recent large meta-analyses of CVD, 21 were reported not to be associated with any of the TRFs.20^,21

SOURCE

Circulation: Cardiovascular Genetics 2012; 5: 368-376

doi: 10.1161/ CIRCGENETICS.112.962746

MicroRNA in Serum as Bimarker for Cardiovascular Pathologies: acute myocardial infarction, viral myocarditis, diastolic dysfunction, and acute heart failure

Increased MicroRNA-1 and MicroRNA-133a Levels in Serum of Patients With Cardiovascular Disease Indicate Myocardial Damage

Yasuhide Kuwabara, MD, Koh Ono, MD, PhD, Takahiro Horie, MD, PhD, Hitoo Nishi, MD, PhD, Kazuya Nagao, MD, PhD, Minako Kinoshita, MD, PhD, Shin Watanabe, MD, PhD, Osamu Baba, MD, Yoji Kojima, MD, PhD, Satoshi Shizuta, MD, Masao Imai, MD, Toshihiro Tamura, MD, Toru Kita, MD, PhD and Takeshi Kimura, MD, PhD

Author Affiliations

From the Department of Cardiovascular Medicine, Graduate School of Medicine, Kyoto University, Kyoto, Japan (Y. Kuwabara, K.O., T.H., H.N., K.N., M.K., S.W., O.B., Y. Kojima, S.S., M.I., T.T., T. Kimura); and Kobe City Medical Center General Hospital, Kobe, Japan (T. Kita).

Correspondence to Koh Ono, MD, PhD, Department of Cardiovascular Medicine, Graduate School of Medicine, Kyoto University, 54 Shogoin-kawahara-cho, Sakyo-ku, Kyoto, Japan 606-8507. E-mail kohono@kuhp.kyoto-u.ac.jp

Abstract

Background—Recently, elevation of circulating muscle-specific microRNA (miRNA) levels has been reported in patients with acute myocardial infarction. However, it is still unclear from which part of the myocardium or under what conditions miRNAs are released into circulating blood. The purpose of this study was to identify the source of elevated levels of circulating miRNAs and their function in cardiovascular diseases.

Conclusions—These results suggest that elevated levels of circulating miR-133a in patients with cardiovascular diseases originate mainly from the injured myocardium. Circulating miR-133a can be used as a marker for cardiomyocyte death, and it may have functions in cardiovascular diseases.

SOURCE:

Circulation: Cardiovascular Genetics. 2011; 4: 446-454

Published online before print June 2, 2011,

doi: 10.1161/ CIRCGENETICS.110.958975

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Introductions

Preventative Cardiology

Methods in Genetics and Clinical Interpretation Randomized Trial of Personal Genomics for Preventive Cardiology Design and Challenges

MicroRNA in Serum as Bimarker for Cardiovascular Pathologies: acute myocardial infarction, viral myocarditis, diastolic dysfunction, and acute heart failure

Increased MicroRNA-1 and MicroRNA-133a Levels in Serum of Patients With Cardiovascular Disease Indicate Myocardial Damage

Selective Articles citing this article

Circulating MicroRNA-208b and MicroRNA-499 Reflect Myocardial Damage in Cardiovascular Disease

Selective Articles citing this article

Genetic Determinants of Potassium Sensitivity and Hypertension

Integrated Computational and Experimental Analysis of the Neuroendocrine Transcriptome in Genetic Hypertension Identifies Novel Control Points for the Cardiometabolic Syndrome

Genome-Wide Linkage and Positional Candidate Gene Study of Blood Pressure Response to Dietary Potassium Intervention

Genome-Wide Association Study of Cardiac Structure and Systolic Function in African Americans

Heart and Aging Research in Genomic Epidemiology: 1700 MIs and 2300 coronary heart disease events among about 29 000 eligible patients

Cohorts for Heart and Aging Research in Genomic Epidemiology (CHARGE) Consortium

Genomics of Ventricular arrhythmias, A-Fib, Right Ventricular Dysplasia, Cardiomyopathy

Selective Articles citing this article

Large-Scale Candidate Gene Analysis in Whites and African Americans Identifies IL6R Polymorphism in Relation to Atrial Fibrillation

PITX2c Is Expressed in the Adult Left Atrium, and Reducing Pitx2c Expression Promotes Atrial Fibrillation Inducibility and Complex Changes in Gene Expression

Genetics of CVD and Hyperlipidemia, Hyper Cholesterolemia, Metabolic Syndrome

Genetic Loci Associated With Plasma Concentration of Low-Density Lipoprotein Cholesterol, High-Density Lipoprotein Cholesterol, Triglycerides, Apolipoprotein A1, and Apolipoprotein B Among 6382 White Women in Genome-Wide Analysis With Replication

Integrated Computational and Experimental Analysis of the Neuroendocrine Transcriptome in Genetic Hypertension Identifies Novel Control Points for the Cardiometabolic Syndrome

Associations Between Incident Ischemic Stroke Events and Stroke and Cardiovascular Disease-Related Genome-Wide Association Studies Single Nucleotide Polymorphisms in the Population Architecture Using Genomics and Epidemiology Study

Common Variation in Fatty Acid Genes and Resuscitation From Sudden Cardiac Arrest

Genome-Wide Association Study Pinpoints a New Functional Apolipoprotein B Variant Influencing Oxidized Low-Density Lipoprotein Levels But Not Cardiovascular Events

Genome-Wide Screen for Metabolic Syndrome Susceptibility Loci Reveals Strong Lipid Gene Contribution But No Evidence for Common Genetic Basis for Clustering of Metabolic Syndrome Traits

Genetics and Vascular Pathologies and Platelet Aggregation, Cardiac Troponin T in Serum

TGFβRIIb Mutations Trigger Aortic Aneurysm Pathogenesis by Altering Transforming Growth Factor β2 Signal Transduction

Matrix Metalloproteinase-9 Genotype as a Potential Genetic Marker for Abdominal Aortic Aneurysm

Common Genetic Variation in the 3′–BCL11B Gene Desert Is Associated With Carotid-Femoral Pulse Wave Velocity and Excess Cardiovascular Disease Risk

Genetic Variation in PEAR1 is Associated with Platelet Aggregation and Cardiovascular Outcomes

Association of Genome-Wide Variation With Highly Sensitive Cardiac Troponin-T Levels in European Americans and Blacks

Genomics and Valvular Disease

Supravalvular Aortic Stenosis Elastin Arteriopathy

Genetic Loci for Coronary Calcification and Serum Lipids Relate to Aortic and Carotid Calcification

Joint Associations of 61 Genetic Variants in the Nicotinic Acetylcholine Receptor Genes with Subclinical Atherosclerosis in American Indians

Heredity of Cardiovascular Disorders Inheritance

A Clinical Approach to Common Cardiovascular Disorders When There Is a Family History

The Implications of Inheritance for Clinical Management

Clinical Considerations of Heritable Factors in Common Heart Failure

Pharmacogenomics

Hypertension Susceptibility Loci and Blood Pressure Response to Antihypertensives

Results From the Pharmacogenomic Evaluation of Antihypertensive Responses Study

Genetic Determinants of Statin-Induced Low-Density Lipoprotein Cholesterol Reduction

The Justification for the Use of Statins in Prevention: An Intervention Trial Evaluating Rosuvastatin (JUPITER) Trial

Genetic Variation in the β2 Subunit of the Voltage-Gated Calcium Channel and Pharmacogenetic Association With Adverse Cardiovascular Outcomes in the INternational VErapamil SR-Trandolapril STudy GENEtic Substudy (INVEST-GENES)