Advertisements
Feeds:
Posts
Comments

Archive for the ‘Regenerative Biology and Medicine’ Category


LIVE Key Note Presentations @Biotech Week Boston, October 5, 2016 3:25PM

Boston Convention and Exhibition Center

 

#BiotechWeekBoston

 avivalev-ari@alum.berkeley.edu

@pharma_BI

@AVIVA1950

ANNOUNCEMENT

Leaders in Pharmaceutical Business Intelligence (LPBI) Group, Boston

pharma_bi-background0238

will cover in REAL TIME

Biotech Week Boston, October 5, 2016 @ Boston Convention and Exhibition Center

 

In Attendance, streaming LIVE using Social Media

Aviva Lev-Ari, PhD, RN

Editor-in-Chief

http://pharmaceuticalintelligence.com

 

Key Sessions

Arnold I. Caplan, Ph.D.

Adult Mesenchymal Stem Cells: The New Medicine

Case Western Reserve University

  • HSC – hematopoietic Stem Cells
  • MSC – mesengenic in vitro not in Tissue Engineering – can be derived from multiple tissue sources
  1. all MSC are Pericytes cells on capillaries and microvessels – CD34 CD146
  2. during injury – pericyte – differentiates – sentinal fro damage and innate tissue regenerations, sense environment
  3. MSC = Medicinal Signaling Cell – the injury specific – 655 Clinical Trials – KIDNEY AND OTHER ORGAN TRANSPLANTATION
  4. hCAP-18/LL37 is secreted by hMSCs
  5. LL37 is in Maternal milk and prevent infection in neonatals
  6. Lipoaspirate
  7. immunomodulatory and Trophic activity
  8. Universal Stem Cell Niche pMSC, Niche
  9. MSCs in cancer metastasis: BRCA, prostate and Melanoma (binding protein) – pericyte pull
  10. http://www.ctte

David DiGiusto, Ph.D.

Building a Sustainable Academic Engine for Feeding De-Risked Assets into the Biopharma Pipeline

Stanford Health Care / Stanford School of Medicine, Cell and Gene Medicine

  1. viral vector
  2. bacterial culture
  3. standard tissue
  • Risks of developing Academic Cell Therapy Products
  • Center for Definitive and Curative Medicine – Regenerative Medicine
  1. translational Staging Score: min 6 points
  2. Scientific Review and Prioritization: 0-9 points
  • Lentiviral vector mediated FOXP3 converts T effector cells into T regulatory cells – CD4 LV-FOXP3
  • CD34 CD90+ blood stem cells: ADM of alpha CD117mAB 0 transplant HSC
  • Cancer Immunotherapy: CD19/CD@@: Leukemia
  • cell transduction
  • Vector Production
  • Plasmid Production
  • Skin DEBnb- low keratinocyte stem cells – neoantigen
  • COmbining CRISPR/Cas9 and rAAV6 mediates High Targeting Efficiencies into peripheral blood CD34+ +HSPCs
  1. Cutting: mRNA and RNP
  2. recombination – AAV only, mRNA : Total population vs Sorted GFP(high)  – get 95% due to Gene correction
  3. Genome editing:
  • starting DNA
  • Edited DNA
  • Edited Protein

4. Closed systems, scalable technology, reagents and automation, clean reagents

5. Invest in the Delivery Channel: Basic Research, Product development, clinical research

6. Transplantation Blood and marrow

7. Cell pharmacy

8. Translational Research : Funding Mix Commercialization, licensing and commercial

9. Infrastructure utilized by every stage of development is different

Advertisements

Read Full Post »


LIVE 9/20 2PM to 5:30PM New Viruses for Therapeutic Gene Delivery at CHI’s 14th Discovery On Target, 9/19 – 9/22/2016, Westin Boston Waterfront, Boston

http://www.discoveryontarget.com/

http://www.discoveryontarget.com/crispr-therapies/

Leaders in Pharmaceutical Business Intelligence (LPBI) Group is a

Media Partner of CHI for CHI’s 14th Annual Discovery on Target taking place September 19 – 22, 2016 in Boston.

In Attendance, streaming LIVE using Social Media

Aviva Lev-Ari, PhD, RN

Editor-in-Chief

http://pharmaceuticalintelligence.com

#BostonDOT16

@BostonDOT

 

COMMENTS BY Stephen J Williams, PhD

Gene Therapy Breakthroughs

New Strategies for Better Specificity and Delivery

 

2:05  Chairman’s Remarks

Joseph Gold, Ph.D. Director Manufacturing Center for Biomedicine and Genetics, Beckman Research Institute City of Hope

 

  • CBG (center for biomedicine and Genetics) 20000 sq feet
  • CTPC (center therapy production) mainly CART
  • CBG 16 years operation do all stem cells and >400 products
  • New stem cell Beta cell progenitor
  • Do oncolytic VSV
  • CTPC is investigator driven CART islet cells,
  • Like to do novel work so work with CIRM
  • Banking of modified stem cells
  • Adherent scale out limitations: cost,inefficient; solution can be suspension
  • Establish hESC; plate on CELLstart > Accutase>StemPRO SFM>differentiation process; defined reagents — they use this for cardiomyocyte differentiation: they are functional (inotropy, chronotropy response to isoproterenol) can freeze back cells
  • Create a bank of intermediate cells and when you need it for surgery they will put on their matrix, enrich, expand and ship out
  • Allogeneic cells: project where take allogeneic neural stem cells to deliver a chemotherapy payload as they like to migrate to brain tumors
  • Allogeneic cells: for ALS modified to express GDNF
  • HIV resistance with engineered CCR5 negative blood stem cells
  • Release assay considerations: viability, sterility, if cryopreserved then can determine identity, viral insertions, VSV-G copy number, endotoxin and potency (FDA is wanting phase I potency assays) for CART potency is % transduced
  • Good in vivo activity of the neural stem cells loaded with chemotherapeutic

 

ALS  

  • If deliver GDNF to muscle  using genetically modified myoblasts
  • Best to use fetal stem cells – less issues

 

Canavan disease: progressive fatal neurologic disorder that begins in infancy and don’t make it past teenage years

  • Rossbach is taking autologous cells reprogramming generating iPS cells and then modifying by CRISPR but the CRISPR issues of off target effects persist as well the time required for process and verification; also don’t want to use a selectable marker and put in patients; so you can differentiate the cells and hit them with a lentiviral vector system

 

They have been named a PACT Center Production Assistance for Cell Therapy where you can apply for a project grant.  Applicable for startups up to larger mature companies

www.pactgroup.net

 

They do a standard panel of tests for viral infections.

They work with investigators or companies at all stages of manufacturing processes.

 

@BeckmanInst

@cityofhope

 

2:15 Large-Scale Production of Cell Therapies for Regenerative Medicine

Joseph Gold, Ph.D. Director Manufacturing Center for Biomedicine and Genetics, Beckman Research Institute

 

2:45  Directed Evolution of New Viruses for Therapeutic Gene Delivery

David Schaffer, Ph.D.  Professor of Chemical and Biomolecular Engineering, BioEngineering, Molecular and Cell Biology and Neuroscience;

 

AAV is very safe as many people already infected with it

 

  • Spark (Leber’s cogenital anaurosis
  • Hemophilia B
  • Lipoprotein lipase deficiency
  • Spinal muscular atrophy
  • Challenges: are we just getting the ‘low hanging fruit’ eg Spark therapy must be injected after retinal therapy, hemo B needs to be given at high doses
  • Their theory was AAV had been evolving for its own purposes so hence the limitations of AAV;
  • Utilized 25 different techniques to generate variants of AAV in a library then packaged (each will have its own barcode)
  • Broad platform technology: retina, lung, brain and spinal cord

Retinal:  AAV may be too large to get through layers of the eye, problems; subretinal injections and damage or retinal detachment.  Then they used their whole library in an in-vivo screen (as hard to recapitulate the multi cell layers of the retina).  

 

Cystic Fibrosis

  • AAV2H22 variant worked very well to supply the CFTR gene in pig model of cystic fibrosis and increases chloride transport and reduce bacterial load
  • Then found pig variant AAV did not work well on human tissue so designed a human variant and worked well in human tissues
  • The variant AAV2.5T surrounds sialic acid binding pockets and increases binding and endocytosis

 

Brain and Spinal Cord:  Sanfilippo B trial  8 holes drilled into skull followed by 16 AAV injections

 

  • Injected a generated AAV variant (by evolution process) : engineered AAV2 is 100 fold better getting through blood brain barrier… novel variant undergoes retrograde transport to cortex ; made a cas9 to remove a tdTomato gene overexpressed in mouse and found 90% knockdown
  • Also interesting point: the porcine variant did not work in human and the human variant did not work in porcine.  Implication for FDA safety and efficacy testing must do in monkeys

They started a spinout 4D Molecular Therapeutics

 

4:25 Lentiviral Vectors for Gene Therapy

 

Munapaty Swani, Ph.D. Texas Tech Health Science Center

 

  • Can express multiple shRNA under a separate promoter but toxic so if expressed in miRNA backbone could be safer under a pol II
  • How much of flanking sequence is needed?
  • 30 nt flanking sequence is enough for Drosha processing
  • Constructed 1 to 7 shRNA-miR targeting CCR5 and 6 viral genes; all constructs were functional
  • Problem with pol ii promoter
  • These 7 shRNA miRNA protect against HIV entry if against CCR5 and the 7 viral elements
  • Used the non-integrating lentivirus for transient to see if infect T cells or not versus integrating lentivirus ; results non-integrating lentivirus did not infect t cells so safer to use
  • CCR5 disruption reduced HIV infection in T cells in vitro;
  • ZFN treatment of HIV+ PBMC prevents activation of HIV
  • Encapsulted CAS9 within LV; cas9 protein is incorporated within LV and is functional
  • First transduce then come in with the Cas9 so made all in one lentivirus with Cas9 and an sgRNA expression vector *******
  • This shows that it is possible to put all in a nanoparticle based lentivirus and an all in one may make it easier and safer (supposedly)

 

4:55 AAV Capsid Design

Miguel Seria Esteves, PhD Associate Professor Neurology, Gene Therapy Center University of Massachusetts Medical School

 

-AAV replication dependent no known human disease with native AAV

  •  Multiple barriers to get across blood brain barrier
  • AAV9 preferentially target neonatal neurons and adult astrocytes
  • Multiple capsids can be used for AAV9 infection in brain but not complete
  • Can we design better capsids to give it better tropic properties and better penetration to blood brain barrier
  • Using a polyalanine in the 5’ end of the caspid was most efficeint
  • Increases gene transfer efficiency especially IN SELECT CELL TYPES; Glial transduction and increased in striatum: increase is structure specific so little in thalamus but good in cerebellum and spinal cord
  • AAV9 tranduces also in peripheral tissues with or without modified capsid

 

Huntington’s Disease

  • Polyglutamate disease polyy glu on huntingtin protein
  • They get a 40 to 50% reduction of huntingtin but not significant between capsid design
  • They did a directed evolution of AAV capsid and generated capsid gene delivery diversity: DNA shuffling and in vivo selection
  • AAV-B1 is a new tropic capsid showing transduction of different structures
  • Five fold reduction in tropism to the liver but massive increases in muscle and beta exocrine cells and lung
  • Presence of neutralizing antibodies is a problem with AAV therapy
  • In conclusion unknown mechanisms by whivh a highly hydrophobic string of 19 alanines modifies the CHS tropism of AAV9 kvariants
  • Chimeric capsids identified from in vivo screen can reveal interesting patterns of tropism

12:45 PM Screening with shRNA and CRISPR

Ryan Raver, PhD Global Product Manager, Functional Genomics, MilliporeSigma

  • KO – Knock Out
  • KD – Knock Down
  1. RNAi -KD
  2. CRISPR-Cas9 – KO

NEW STRATEGIES FOR BETTER SPECIFICITY AND DELIVERY

2:05 Chairperson’s Remarks

Joseph Gold, Center for Biomedicine and Genetics Beckman Research Institute, City of Hope

2:15 Large Scale Production of cell Therapies for Regenerative Medicine: COmbination Cell and Gene Therapy products

Joseph Gold, Center for Biomedicine and Genetics Beckman Research Institute, City of Hope

  • Biological & Cellular GMP manufacturing Core at COH
  • Establishing scalable hESC suspension Culture
  • Optimized small molecule concentration, induction timing, stirring rates
  1. Almost Xeno free
  2. defined
  3. very good reproducibility
  4. high purity and yield:
  • Immuno-staining,
  • FACS – cTnT, sMHC, Alpha-actinin
  • Cryopreservation, Multi-electrode Array (MEA)
  • hESC-RPE monolayer on synthetic substrate

Combination cell/gene therapy products at COH

  1. CAR T CCR5-inactivated CD34+ HSPC – Target: AIDS
  • adoptive immunotherapy using CAR-Engineering T cells – glioblastoma

2. HIV resistance with engineering CCR5-negative blood stem cells: gene KO by ZFNs

  • assay considerations 0 If cryopreservation : Identify, viral insertion, endotoxin, residual beads Potency: CAR T- % transduced cells, CCR5-?-CD34 cells: HIV resistance

3. Glioblastoma

4.  In vivo activity of transduced NSCs: Assay consideration – viability, sterility, mycobatom,

5.  ALS: degeneration of neurones

  • Embryonic stem cells
  • Fetal neural stem cells
  • Adult stem cells – human Proginetor neural cells

6.  Canavan Disease – Y.Shi – ASPA gene mutations cause Canavan disease

Strategy: autologous iPSC-derived, modified neural progenitors: DIferentiate to neural progenitors

Gene therapy: Correction (Off-taregt effects, correct individual cell lines)  or Over expression (Copy number, selection, transduce iPSC)

  • release assay consideration
  • identity – markers and HLA, contaminants, Potency: in vivo efficacy modified autologous
  • production Assistance for Cell therapy
  • cell therapy manufacturing development
  • roles of Biobanks

 

2:45 Directed Evolution of New Viruses for Therapeutic Gene Delivery

David Schaffer, Ph.D., Professor of Chemical and Biomolecular Engineering, Bioengineering, Molecular and Cell Biology, and Neuroscience; Director, Berkeley Stem Cell Center, University of California, Berkeley

Adeno-associated viral (AAV) vectors have been increasingly successful in clinical trials; however, viruses face many delivery barriers that limit their efficacy for most disease targets. We have developed directed vector evolution – the iterative genetic diversification of a viral genome and functional selection for desired properties – to engineer novel, optimized AAV vectors for efficient, selective delivery for a range of tissue and disease targets.

  • DNA (gene therapy and editing) >> RNAi (antisense) >> Protein: Small molecules and monoclonal antibodies
  • Dlivery
  • Adeno Associated Viral Vectors Adenoviral helper genes
  • AAV2, efficacy in Leber’s Congenital samaurosis (AAV)
  • Spinal muscular, lipolrotein lipase deficiency (AAV)

Gene Delivery

  1. neutralization of pre-esisting antibodies
  2. target tissue – deep penetration
  3. Inefficient transduction to target cells
  4. target specific cells

Fitness as Therapy – virus evolutionary: Tropism and immunity

  • AAV directed Vector evolution : Input /sequence >> Diversity/generation >> Packaging
  • Retinoschisis Model – Eye therapeutic gene to protect vision – AAV – transduction of retinal cell, vitreas in Human different scale — eventually Macualr degeneration Therapy for: Photo receptor is the target for therapy
  • Dog: Fundus Imaging of Engineered AAV: Variant Expression GFP Pool Carrying

Lung- Cystic Fibrosis (mucus production amplifies (enhanced transduction) due to MAC3 translation error) – Gene Therapy – cilia function to restore ability to clear mucos: Variant evolved on Human airway epithelia – AA  particles

  1. AAV2>>> AAV%>> T mutation
  • efficacy must be improved

Brain and Spinal Cord

  1. Scull – 8 drills followed by 16 AAV injections
  2. Spinal cord: injuction in muscle

Synthetic version of AAV — Engineered AAV for enhanced Retrograde Transport

  • AAV2-retrograded transport – Noval variant Undergoes Transport along multiple Projections
  • Cas9 – Retrograde Delivery of Cas9 to cortex of mouse – KD –

Summary

  • Virus as gene delivery mechanism
  • designer AAV variants

 

3:15 Sponsored Presentation (Opportunity Available)

3:45 Refreshment Break in the Exhibit Hall with Poster Viewing and Poster Competition Winner Announced

4:25 Lentiviral Vectors for Gene Therapy

Manjunath N. Swamy, MD, Professor of Biomedical Sciences and Co-DIrector of the Center of Emphasis in Infectious Diseases. Paul L Foster School of Medicine, Texas Tech University Health Science Center

  • RNAi targets for HIV
  • Expression of multiple shRNAs
  • COnstruction od 7 shRNA-miR targeting CCR5 and 6 viral genes – protect against both 5 ans x4 tropic HIV-1
  • shRNA expression does not decrease with distance from promoter
  • Lentiviral vector to express ZFNs: HIV envelop – ZFN-mediated CCR5 gene editing in Primary T cells
  • ZFN treatment of HIV+PBMC prevents activation of HIV
  • Strategy to encapsulate Cas9 Protein wihtin LV : AIm to deliver Cas9 protein deliver sgRNA expression vector. A Lentiviral
  • Gene editing by all-in-one Lentivirus = to prepack

 

4:55 AAV Caspid Engineering

Miguel Sena Esteves, Ph.D., Associate Professor, Department of Neurology, Gene Therapy Center, University of Massachusetts Medical School

Adeno-associated virus vectors have become the leading platform for development of in vivo gene therapies for neurological diseases. We have developed new AAV vectors for widespread gene delivery to the CNS through vascular infusion in adult animals through peptide grafting and in vivo library selection. These new neurotropic AAVs have achieved CNS-wide silencing of gene expression using gene-specific microRNAs.

  • Adeno-associated virus – Paravovirus family
  • Recombinant AAV vectors carry gene expression cassette of choice flanked by two
  • CNS – route of gene delivery
  • Crossing BBB
  • Systemic delivery of AAV9 vectors: IV
  • Peptide grafting – in vivo selection of novel CNS: DIstribution of GFP transduced cells – robust neuronal transducture with transduction AAV-AS
  • motor cortex, Straiatum, thalamus, motor cortex, ventral horn of spinal cord, cerebelum, liver, muscle, oculomotor nerve, nucleus of oculomotor nerve

Huntington’s Disease > 40 CAG vs <26 CAG in normal – Peptide grafting of AAV vectors for CNS

  • DNA shuffling and in vivo selection: Brain vs Liver

Next generation of AAV vectors for CNS 

  • Liver, pancreas, lung — same pattern

neural transduction after vascular delivery

  • AAV-B1 caspid vs AAV8 – 19 amino acids

Conclusion

New capsids with improved CNS tropism

19 alanines modifies the CNS tropism of AAV9 variants

Chimeric caspids identified from in vivo screens

 

5:25 Welcome Reception in the Exhibit Hall with Poster Viewing

 

Read Full Post »


cvd-series-a-volume-iv-cover

Series A: e-Books on Cardiovascular Diseases

Series A Content Consultant: Justin D Pearlman, MD, PhD, FACC

VOLUME FOUR

Regenerative and Translational Medicine

The Therapeutic Promise for

Cardiovascular Diseases

  • on Amazon since 12/26/2015

http://www.amazon.com/dp/B019UM909A

 

by  

Larry H Bernstein, MD, FCAP, Senior Editor, Author and Curator

and

Aviva Lev-Ari, PhD, RN, Editor and Curator

 

Part One:

Cardiovascular Diseases,Translational Medicine (TM) and Post TM

Introduction to Part 1: Cardiovascular Diseases,Translational Medicine (TM) and Post TM

Chapter 1: Translational Medicine Concepts

1.0 Post-Translational Modification of Proteins

1.1 Identifying Translational Science within the Triangle of Biomedicine

1.2 State of Cardiology on Wall Stress, Ventricular Workload and Myocardial Contractile Reserve: Aspects of Translational Medicine (TM)

1.3 Risk of Bias in Translational Science

1.4 Biosimilars: Intellectual Property Creation and Protection by Pioneer and by Biosimilar Manufacturers

Chapter 2: Causes and the Etiology of Cardiovascular Diseases: Translational Approaches for Cardiothoracic Medicine

2.1 Genomics

2.1.1 Genomics-Based Classification

2.1.2  Targeting Untargetable Proto-Oncogenes

2.1.3  Searchable Genome for Drug Development

2.1.4 Zebrafish Study Tool

2.1.5  International Human Genome Sequencing Consortium (2004) Finishing the euchromatic sequence of the human genome.

2.2  Proteomics

2.2.1 The Role of Tight Junction Proteins in Water and Electrolyte Transport

2.2.2 Selective Ion Conduction

2.2.3 Translational Research on the Mechanism of Water and Electrolyte Movements into the Cell

2.2.4 Inhibition of the Cardiomyocyte-Specific Kinase TNNI3K ­ Oxidative Stress

2.2.5 Oxidized Calcium Calmodulin Kinase and Atrial Fibrillation

2.2.6 S-Nitrosylation in Cardiac Ischemia and Acute Coronary Syndrome

2.2.7 Acetylation and Deacetylation

2.2.8 Nitric Oxide Synthase Inhibitors (NOS-I) 

2.3 Cardiac and Vascular Signaling

2.3.1 The Centrality of Ca(2+) Signaling and Cytoskeleton Involving Calmodulin Kinases and Ryanodine Receptors in Cardiac Failure, Arterial Smooth Muscle, Post-ischemic Arrhythmia, Similarities and Differences, and Pharmaceutical Targets

2.3.2 Leptin Signaling in Mediating the Cardiac Hypertrophy associated with Obesity

2.3.3 Triggering of Plaque Disruption and Arterial Thrombosis

2.3.4 Sensors and Signaling in Oxidative Stress

2.3.5 Resistance to Receptor of Tyrosine Kinase

2.3.6  S-nitrosylation signaling in cell biology.

2.4  Platelet Endothelial Interaction

2.4.1 Platelets in Translational Research ­ 1

2.4.2 Platelets in Translational Research ­ 2: Discovery of Potential Anti-platelet Targets

2.4.3 The Final Considerations of the Role of Platelets and Platelet Endothelial Reactions in Atherosclerosis and Novel Treatments

2.4.4 Endothelial Function and Cardiovascular Disease
Larry H Bernstein, MD, FCAP

2.5 Post-translational modifications (PTMs)

2.5.1 Post-Translational Modifications

2.5.2.  Analysis of S-nitrosylated Proteins

2.5.3  Mechanisms of Disease: Signal Transduction: Akt Phosphorylates HK-II at Thr-473 and Increases Mitochondrial HK-II Association to Protect Cardiomyocytes

2.5.4  Acetylation and Deacetylation of non-Histone Proteins

2.5.5  Study Finds Low Methylation Regions Prone to Structural Mutation

2.6 Epigenetics and lncRNAs

2.6.1 The Magic of the Pandora’s Box : Epigenetics and Stemness with Long non-coding RNAs (lincRNA)

2.6.2 The SILENCE of the Lambs” Introducing The Power of Uncoded RNA

2.6.3 Long Noncoding RNA Network regulates PTEN Transcription

2.6.4 How mobile elements in “Junk” DNA promote cancer. Part 1: Transposon-mediated tumorigenesis.

2.6.5 Transposon-mediated Gene Therapy improves Pulmonary Hemodynamics and attenuates Right Ventricular Hypertrophy: eNOS gene therapy reduces Pulmonary vascular remodeling and Arterial wall hyperplasia

2.6.6 Junk DNA codes for valuable miRNAs: non-coding DNA controls Diabetes

2.6.7 Targeted Nucleases

2.6.8 Late Onset of Alzheimer’s Disease and One-carbon Metabolism
Dr. Sudipta Saha

2.6.9 Amyloidosis with Cardiomyopathy

2.6.10 Long non-coding RNAs: Molecular Regulators of Cell Fate

2.7 Metabolomics

2.7.1 Expanding the Genetic Alphabet and Linking the Genome to the Metabolome

2.7.2 How Methionine Imbalance with Sulfur-Insufficiency Leads to Hyperhomocysteinemia

2.7.3 A Second Look at the Transthyretin Nutrition Inflammatory Conundrum

2.7.4 Transthyretin and Lean Body Mass in Stable and Stressed State

2.7.5 Hyperhomocysteinemia interaction with Protein C and Increased Thrombotic Risk

2.7.6 Telling NO to Cardiac Risk

2.8 Mitochondria and Oxidative Stress

2.8.1 Reversal of Cardiac Mitochondrial Dysfunction

2.8.2 Calcium Signaling, Cardiac Mitochondria and Metabolic Syndrome

2.8.3. Mitochondrial Dysfunction and Cardiac Disorders

2.8.4 Mitochondrial Metabolism and Cardiac Function

2.8.5 Mitochondria and Cardiovascular Disease: A Tribute to Richard Bing

2.8.6 MIT Scientists on Proteomics: All the Proteins in the Mitochondrial Matrix Identified

2.8.7 Mitochondrial Dynamics and Cardiovascular Diseases

2.8.8 Mitochondrial Damage and Repair under Oxidative Stress

2.8.9 Nitric Oxide has a Ubiquitous Role in the Regulation of Glycolysis -with a Concomitant Influence on Mitochondrial Function

2.8.10 Mitochondrial Mechanisms of Disease in Diabetes Mellitus

2.8.11 Mitochondria Dysfunction and Cardiovascular Disease – Mitochondria: More than just the “Powerhouse of the Cell”

Chapter 3: Risks and Biomarkers for Diagnosis and Prognosis in Translational Cardiothoracic Medicine

3.1 Biomarkers. Diagnosis and Management: Biomarkers. Present and Future.

3.2 Landscape of Cardiac Biomarkers for Improved Clinical Utilization

3.3 Achieving Automation in Serology: A New Frontier in Best

3.4 Accurate Identification and Treatment of Emergent Cardiac Events

3.5 Prognostic Marker Importance of Troponin I in Acute Decompensated Heart Failure (ADHF)

3.6 High-Sensitivity Cardiac Troponin Assays Preparing the United States for High-Sensitivity Cardiac Troponin Assays

3.7 Voices from the Cleveland Clinic On Circulating apoA1: A Biomarker for a Proatherogenic Process in the Artery Wall

3.8 Triggering of Plaque Disruption and Arterial Thrombosis

3.9 Relationship between Adiposity and High Fructose Intake Revealed

3.10 The Cardio-Renal Syndrome (CRS) in Heart Failure (HF)

3.11 Aneuploidy and Carcinogenesis

3.12 “Sudden Cardiac Death,” SudD is in Ferrer inCode’s Suite of Cardiovascular Genetic Tests to be Commercialized in the US

Chapter 4: Therapeutic Aspects in Translational Cardiothoracic Medicine

4.1 Molecular and Cellular Cardiology

4.1.1 αllbβ3 Antagonists As An Example of Translational Medicine Therapeutics

4.1.2 Three-Dimensional Fibroblast Matrix Improves Left Ventricular Function post MI

4.1.3 Biomaterials Technology: Models of Tissue Engineering for Reperfusion and Implantable Devices for Revascularization

4.1.4 CELLWAVE Randomized Clinical Trial: Modest improvement in LVEF at 4 months ­ “Shock wave­facilitated intracoronary administration of BMCs” vs “Shock wave treatment alone”

4.1.5 Prostacyclin and Nitric Oxide: Adventures in vascular biology –  a tale of two mediators

4.1.6 Cardiac Contractility & Myocardium Performance: Ventricular Arrhythmiasand Non-ischemic Heart Failure – Therapeutic Implications for Cardiomyocyte Ryanopathy

4.1.7 Publications on Heart Failure by Prof. William Gregory Stevenson, M.D., BWH

4.2 Interventional Cardiology and Cardiac Surgery – Mechanical Circulatory Support and Vascular Repair

4.2.1 Mechanical Circulatory Support System, LVAD, RVAD, Biventricular as a Bridge to Heart Transplantation or as “Destination Therapy”: Options for Patients in Advanced Heart Failure

4.2.2 Heart Transplantation: NHLBI’s Ten Year Strategic Research Plan to Achieving Evidence-based Outcomes

4.2.3 Improved Results for Treatment of Persistent type 2 Endoleak after Endovascular Aneurysm Repair: Onyx Glue Embolization

4.2.4 Carotid Endarterectomy (CEA) vs. Carotid Artery Stenting (CAS): Comparison of CMMS high-risk criteria on the Outcomes after Surgery: Analysis of the Society for Vascular Surgery (SVS) Vascular Registry Data

4.2.5 Effect of Hospital Characteristics on Outcomes of Endovascular Repair of Descending Aortic Aneurysms in US Medicare Population

4.2.6 Hypertension and Vascular Compliance: 2013 Thought Frontier – An Arterial Elasticity Focus

4.2.7 Preventive Medicine Philosophy: Excercise vs. Drug, IF More of the First THEN Less of the Second

4.2.8 Cardio-oncology and Onco-Cardiology Programs: Treatments for Cancer Patients with a History of Cardiovascular Disease

Summary to Part One

 

Part Two:

Cardiovascular Diseases and Regenerative Medicine

Introduction to Part Two

Chapter 1: Stem Cells in Cardiovascular Diseases

1.1 Regeneration: Cardiac System (cardiomyogenesis) and Vasculature (angiogenesis)

1.2 Notable Contributions to Regenerative Cardiology by Richard T. Lee (Lee’s Lab, Part I)

1.3 Contributions to Cardiomyocyte Interactions and Signaling (Lee’s Lab, Part II)

1.4 Jmjd3 and Cardiovascular Differentiation of Embryonic Stem Cells

1.5 Stem Cell Therapy for Coronary Artery Disease (CAD)

1.6 Intracoronary Transplantation of Progenitor Cells after Acute MI

1.7 Progenitor Cell Transplant for MI and Cardiogenesis (Part 1)

1.8 Source of Stem Cells to Ameliorate Damage Myocardium (Part 2)

1.9 Neoangiogenic Effect of Grafting an Acellular 3-Dimensional Collagen Scaffold Onto Myocardium (Part 3)

1.10 Transplantation of Modified Human Adipose Derived Stromal Cells Expressing VEGF165

1.11 Three-Dimensional Fibroblast Matrix Improves Left Ventricular Function Post MI

Chapter 2: Regenerative Cell and Molecular Biology

2.1 Circulating Endothelial Progenitors Cells (cEPCs) as Biomarkers

2.2 Stem Cell Research — The Frontier at the Technion in Israel

2.3 Blood vessel-generating stem cells discovered

2.4 Heart Renewal by pre-existing Cardiomyocytes: Source of New Heart Cell Growth Discovered

2.5 The Heart: Vasculature Protection – A Concept-based Pharmacological Therapy including THYMOSIN

2.6 Innovations in Bio instrumentation for Measurement of Circulating Progenetor Endothelial Cells in Human Blood.

2.7 Endothelial Differentiation and Morphogenesis of Cardiac Precursor

Chapter 3: Therapeutics Levels In Molecular Cardiology

3.1 Secrets of Your Cells: Discovering Your Body’s Inner Intelligence (Sounds True, on sale May 1, 2013) by Sondra Barrett

3.2 Human Embryonic-Derived Cardiac Progenitor Cells for Myocardial Repair

3.3 Repair using iPPCs or Stem Cells

3.3.1 Reprogramming cell in Tissue Repair

3.3.2 Heart patients’ skin cells turned into healthy heart muscle cells

3.4 Arteriogenesis and Cardiac Repair: Two Biomaterials – Injectable Thymosin beta4 and Myocardial Matrix Hydrogel 

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

3.6 Calcium Cycling (ATPase Pump) in Cardiac Gene Therapy: Inhalable Gene Therapy for Pulmonary Arterial Hypertension and Percutaneous Intra-coronary Artery Infusion for Heart Failure: Contributions by Roger J. Hajjar, MD

Chapter 4: Research Proposals for Endogenous Augmentation of circulating Endothelial Progenitor Cells (cEPCs)

4.1 Peroxisome proliferator-activated receptor (PPAR-gamma) Receptors Activation: PPARγ transrepression for Angiogenesis in Cardiovascular Disease and PPARγ transactivation for Treatment of Diabetes

4.2 Clinical Trials Results for Endothelin System: Pathophysiological role in Chronic Heart Failure, Acute Coronary Syndromes and MI – Marker of Disease Severity or Genetic Determination?

4.3 Endothelin Receptors in Cardiovascular Diseases: The Role of eNOS Stimulation

4.4 Inhibition of ET-1, ETA and ETA-ETB, Induction of NO production, stimulation of eNOS and Treatment Regime with PPAR-gamma agonists (TZD): cEPCs Endogenous Augmentation for Cardiovascular Risk Reduction – A Bibliography

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

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

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

4.8 Cardiovascular Disease (CVD) and the Role of agent alternatives in endothelial Nitric Oxide Synthase (eNOS) Activation and Nitric Oxide Production

4.9 Resident-cell-based Therapy in Human Ischaemic Heart Disease: Evolution in the PROMISE of Thymosin beta4 for Cardiac Repair

4.10 Macrovascular Disease – Therapeutic Potential of cEPCs: Reduction Methods for CV Risk

4.11 Bystolic’s generic Nebivolol – positive effect on circulating Endothelial Proginetor Cells endogenous augmentation

4.12 Heart Vasculature – Regeneration and Protection of Coronary Artery Endothelium and Smooth Muscle: A Concept-based Pharmacological Therapy of a Combination Three Drug Regimen including THYMOSIN

Summary to Part Two

Epilogue to Volume Four

Read Full Post »


Agenda for Frontiers in Cancer Immunotherapy, February 27 – 28, 2017, The New York Academy of Sciences

Reporter: Aviva Lev-Ari, PhD, RN

 

  •  Frontiers in Cancer Immunotherapy

    February 27 – 28, 2017
    The New York Academy of Sciences

    Presented by The Mushett Family Foundation, Science, Science Immunology, Science Translational Medicine, and the New York Academy of Sciences

    Register Now

    Agenda

    * Presentation titles and times are subject to change.


    Day 1: February 27, 2017

    7:45 AM Breakfast and Registration
    8:30 AM Introduction and Welcome Remarks
    Representative, Mushett Family Foundation
    Representative, Science, Science Translational Medicine, and Science Immunology
    Representative, The New York Academy of Sciences
    8:55 AM Day 1 Keynote Address:
    Immune Checkpoint Blockade in Cancer Therapy: New Insights, Opportunities and Prospects for A Cure
    James P. Allison, PhD, The University of Texas MD Anderson Cancer Center

    Session 1: Assessment of Current Therapeutic Approaches in Cancer Immunotherapy: Successes and Challenges

    9:40 AM Title to Be Announced
    Phillip D. Greenberg, MD, University of Washington
    10:05 AM In situ Vaccination for the Treatment of Cancer
    Nina Bhardwaj, MD, PhD, Mount Sinai School of Medicine
    10:30 AM Treating the Tumor and Treating the Host
    Ronald Levy, MD, Stanford University
    10:55 AM Networking Coffee Break

    Session 2: Evaluation of Combination Therapy Strategies to Improve Clinical Outcomes

    11:25 AM Title to Be Announced
    Jedd D. Wolchok, MD, Memorial Sloan Kettering Cancer Center
    11:50 AM The Promise of Epigenetic Therapies for Enhancing the Efficacy of Immune Checkpoint Therapy
    Stephen B. Baylin, MD, Johns Hopkins University
    12:15 PM HPV Theraputic Vacccines: Where Are We Now and Where Are We Going?
    Cornelia (Connie) Liu Trimble, MD, Johns Hopkins University
    12:40 PM Networking Lunch and Poster Viewing
    12:55 PM Underrepresented Minorities, Women, and Early Career Investigator Career Development Workshop and Lunch Running Concurrently with the Networking Lunch

    For Graduate Students, Post-doctoral Fellows, and Junior Faculty

    Editor’s Guide to Writing and Publishing Your Paper

    Angela Colmone, PhD, Senior Editor, Science Translational Medicine
    Kristen Mueller, PhD, Senior Editor, Science
    Yevgeniya Nusinovich, MD, PhD, Associate Editor, Science Translational Medicine

    Session 3: Identification of Relevant Prognostic and Predictive Biomarkers for the Development of Immune-Monitoring Strategies

    2:00 PM Inflammation and Cancer: Fueling Response and Resistance of Immunotherapies
    Lisa Coussens, PhD, Oregon Health Sciences University
    2:25 PM High-throughput Single-Cell Analysis for T-Cell Receptor Ligands and Sequences
    Mark Davis, PhD, Stanford University
    2:50 PM Response and Resistance to PD-1 Blockade Therapy
    Antoni Ribas, MD, PhD, University of California Los Angeles
    3:15 PM Networking Coffee Break

    Session 4: Hot Topic Short Talks

    3:40 PM Short Talk Selected from Submitted Abstracts
    3:55 PM Short Talk Selected from Submitted Abstracts

    Session 5: Development of Strategies to Overcome Immune Tolerance

    4:10 PM Title to Be Announced
    Thomas Gajewski, MD, PhD, University of Chicago
    4:35 PM Molecular and Epigenetic Programs Underlying CD8 T Cell Dysfunction in Solid Tumors
    Andrea Schietinger, PhD, Memorial Sloan Kettering Cancer Center
    5:00 PM Targeting FoxP3+ T-cells in Cancers; Friends or Foes?
    Hiroyoshi Nishikawa, MD, PhD, National Cancer Center Hospital, Japan
    5:25 PM Panel Discussion
    5:50 PM Closing Remarks
    6:00 PM Poster Session 1 and Networking Reception
    7:00 PM Day 1 Adjourns

    Day 2: February 28, 2017

    8:00 AM Breakfast and Registration
    8:00 PM Underrepresented Minorities, Women, and Early Career Investigator Mentoring Breakfast Running Concurrently with the General Breakfast

    Session 6: Clinical Research Break Out Sessions

    9:00 AM Breakout Group 1: How to Manage Toxicities in Cancer Immunotherapy?
    Jedd D. Wolchok, MD, Memorial Sloan Kettering Cancer Center
    Breakout Group 2: How to Incorporate Biomarkers into Early Phase Immunotherapy Trials?
    Jonathan Cebon, MBBS, PhD, FRACP, Olivia Newton-John Cancer and Wellness Centre
    Breakout Group 3: What are the Latest Strategies for Cancer Immunotherapy Development — a Pharmaceutical Industry Perspective?
    Ira Mellman, PhD, Genentech
    10:00 AM Networking Coffee Break

    Session 7: Optimizing Incorporation of Cancer Genomics and Epigenomics into Immunotherapy Research and Clinical Strategies

    10:30 AM Title to Be Announced
    Steven Rosenberg, MD, PhD, National Cancer Institute, U.S. National Institutes of Health
    10:55 AM Title to Be Announced
    Michele Maio, MD, PhD, University Hospital, Sienna, Italy
    11:20 AM Using Cancer-specific Neoantigens to Personalize Cancer Immunotherapy
    Robert Schreiber, PhD, Washington University
    11:45 AM Networking Lunch and Poster Session 2

    Session 8: Targeting T-cells in Cancer Immunotherapy

    1:15 PM Regulatory T-cells and Strategies for Inhibition for Cancer Therapy
    Alexander Rudensky, PhD, Memorial Sloan Kettering Cancer Center
    1:40 PM Use of Engineered Chimeric Antigen Receptors for Leukemia Treatment
    Crystal Mackall, MD, Stanford University
    2:05 PM Title to Be Announced
    Kunle Odunsi, MD, PhD, Roswell Park Cancer Institute
    2:30 PM Networking Coffee Break

    Session 9: Hot Topic Short Talks

    3:00 PM Short Talk Selected from Submitted Abstracts
    3:15 PM Short Talk Selected from Submitted Abstracts

    Session 10: Emerging Technologies in Cancer Immunotherapy

    3:30 PM Title to Be Announced
    Laurence Zitvogel, MD, PhD, Institut National de la Santé et Recherche Médicale
    3:55 PM Emerging Technologies in Cancer Immunotherapy
    Carl June, MD, University of Pennsylvania
    4:20 PM Closing Remarks
    4:30 PM Conference Adjourns

Read Full Post »


GE Healthcare has acquired Biosafe Group SA, a supplier of Integrated Cell Bioprocessing Systems for Cell Therapy and Regenerative Medicine Industry

Reporter: Aviva Lev-Ari, PhD, RN

 

CHALFONT ST GILES, England–(BUSINESS WIRE)–GE Healthcare has acquired Biosafe Group SA, a supplier of integrated cell bioprocessing systems for the rapidly growing cell therapy and regenerative medicine industry for an undisclosed sum. The acquisition of Biosafe expands GE Healthcare’s end-to-end ecosystem of products, solutions and services for our cell therapy customers, and expands GE’s technology reach to a number of new cell and therapy types.

“Together with GE we will have the combination of biological, engineering and industrial capabilities to help accelerate the fields of cell therapy and cellular immunotherapy into the mainstream, benefitting patients globally, and bringing the vision of personalized medicine to reality.”

Tweet this

Cellular therapies are rapidly changing the healthcare landscape by providing life-saving and potentially curative treatments for many of the world’s most challenging diseases, especially cancer. The cell therapy oncology market alone is expected to reach $30 billion by 20301 with more than 600 potentially life-changing therapies in clinical trials at the end of 20152.

Biosafe, headquartered in the Lake Geneva region in Switzerland, with a global presence, has a 20 year track-record in automated cell processing and is a recognized leader in the field with reliable applications in bioprocessing, regenerative medicine and stem cell banking. Its proprietary products offer significant advantages over conventional processing tools, with closed fluid pathways, built-in traceability and single-use consumables. The strong strategic fit and complementary business models of GE Healthcare’s Life Sciences business and Biosafe combined with expanded capabilities in product development and commercial reach, will offer significant customer and ultimately patient benefits.

Kieran Murphy, CEO Life Sciences, GE Healthcare said: “GE is building a world-class set of tools, technologies and services for cell and gene therapy and Biosafe’s expertise and innovative systems will strongly enhance our customer offering. GE and Biosafe share a vision of an integrated approach to helping customers optimize every stage of their process to reduce production risks dramatically and increase access to these remarkable new medicines.”

Claude Fell, Founder and Chairman, Biosafe Group SA, said: “Together with GE we will have the combination of biological, engineering and industrial capabilities to help accelerate the fields of cell therapy and cellular immunotherapy into the mainstream, benefitting patients globally, and bringing the vision of personalized medicine to reality.” Olivier Waridel, Biosafe CEO, who will continue to lead Biosafe within the new integrated GEHC structure, added: “Joining GE Healthcare will give Biosafe an outstanding opportunity to couple its unique cell processing technology with GE Healthcare’s strong, global infrastructure, leading to improved capabilities for our customers and enhanced market penetration.”

GE’s strategy is to work with the industry and partners to develop a digitally-enabled ecosystem of complete tools, solutions and services for cell therapy aimed at accelerating the standardization, collaboration and integration customers need to bring these new therapies into mainstream clinical practice. GE has engaged globally with leaders in the industry, such as Canada’s Center for the Commercialization of Regenerative Medicine, the UK’s Cell and Gene Therapy Catapult, Australia’s Cell Therapy Manufacturing Cooperative Research Centre and leading clinical centers such as UPenn, Karolinska Institute, Memorial Sloan-Kettering and Mayo Clinic.

In 2016, GE has announced further significant investments in the cell therapy and regenerative medicine space. In April, GE Ventures and Mayo Clinic announced the launch of Vitruvian Networks, Inc., an independent platform company committed to accelerating access to cell and gene therapies through advanced, cloud-ready software systems and manufacturing services. In January, GE announced the BridGE@CCRM Cell Therapy Centre of Excellence, a US $31.5 million co-investment with the Canadian Government to promote new technologies for the production of cellular therapies in Toronto.

*END*

About GE Healthcare

GE Healthcare provides transformational medical technologies and services to meet the demand for increased access, enhanced quality and more affordable healthcare around the world. GE (NYSE: GE) works on things that matter – great people and technologies taking on tough challenges. From medical imaging, software & IT, patient monitoring and diagnostics to drug discovery, biopharmaceutical manufacturing technologies and performance improvement solutions, GE Healthcare helps medical professionals deliver great healthcare to their patients. For more information about GE Healthcare, visit our website atwww.gehealthcare.com.

About Biosafe Group SA

Founded in 1997 the Biosafe Group is active in the design, manufacture and marketing of automated cell processing systems. Headquartered in Switzerland and privately-owned, the Biosafe Group operates through regional subsidiaries (Geneva, Houston, Hong-Kong, Shanghai and São Paulo) and is present in more than 50 countries, either directly or through distributors. For more information about Biosafe, visit the website www.biosafe.ch.

1 http://www.centerwatch.com/news-online/2015/10/23/t-cell-immunotherapy-market-may-be-worth-30b-by-2030/

2 Alliance for Regenerative Medicine Q1 Data Report 2016 http://alliancerm.org/page/arm-data-reports#

GE to boost cell therapy tech with Biosafe acquisition

 

Biosafe has been working in the automated cell processing arena and offers proprietary products that include advantages–such as closed fluid pathways, built-in traceability and single-use consumables–over conventional tech.

“GE is building a world-class set of tools, technologies and services for cell and gene therapy and Biosafe’s expertise and innovative systems will strongly enhance our customer offering,” said Kieran Murphy, the CEO of life sciences at GE Healthcare. “GE and Biosafe share a vision of an integrated approach to helping customers optimize every stage of their process to reduce production risks dramatically and increase access to these remarkable new medicines.”

GE explained in the announcement that its strategy is to work with its partners on cell therapy, specifically looking to develop a digitally enabled offering of tools, solutions and services for cell therapy. The aim is to boost up standardization, collaboration and integration for customers so cell therapies can become mainstream in clinical practice, GE noted.

“Joining GE Healthcare will give Biosafe an outstanding opportunity to couple its unique cell processing technology with GE Healthcare’s strong, global infrastructure, leading to improved capabilities for our customers and enhanced market penetration,” said Biosafe CEO Olivier Waridel.

SOURCE

http://www.fiercebiotech.com/medical-devices/ge-to-boost-cell-therapy-tech-biosafe-group-sa-acquisition?utm_medium=nl&utm_source=internal&mrkid=993697&mkt_tok=eyJpIjoiWVdabE1EQXhaVEJrWkdaayIsInQiOiI1RzVJMFAwbFNxcCt1eFNhSktLQTZYTFhVUit4UlwvcnBFdFJYNlNvbWlNMG9UYUFJUUc0UmpPckJmS0ZoUXNYNUNScFJPQVwvQnpSY0hubjJxN1dkeDdENXprS0VWRDhtdEhHQlJBXC9JeE9Sdz0ifQ%3D%3D

 

Read Full Post »


The Possibility of rejuvenating ageing bodies with injections of Blood Stem Cells saved from birth or early life: Free of Mutations and have Full-length Telomeres

Reporter: Aviva Lev-Ari, PhD, RN

 

 

Somatic mutations found in the healthy blood compartment of a 115-yr-old woman demonstrate oligoclonal hematopoiesis

  1. Henne Holstege1,10,
  2. Wayne Pfeiffer2,
  3. Daoud Sie3,
  4. Marc Hulsman4,
  5. Thomas J. Nicholas5,
  6. Clarence C. Lee6,
  7. Tristen Ross6,
  8. Jue Lin7,
  9. Mark A. Miller2,
  10. Bauke Ylstra3,
  11. Hanne Meijers-Heijboer1,
  12. Martijn H. Brugman8,
  13. Frank J.T. Staal8,
  14. Gert Holstege9,
  15. Marcel J.T. Reinders4,
  16. Timothy T. Harkins6,
  17. Samuel Levy5 and
  18. Erik A. Sistermans1

+Author Affiliations


  1. 1Department of Clinical Genetics, VU University Medical Center, 1007 MB Amsterdam, The Netherlands;

  2. 2San Diego Supercomputer Center, UCSD, La Jolla, California 92093, USA;

  3. 3Department of Pathology, VU University Medical Center, 1007 MB Amsterdam, The Netherlands;

  4. 4Delft Bioinformatics Laboratory, Delft University of Technology, 2628 CD Delft, The Netherlands;

  5. 5Department of Molecular and Experimental Medicine, Scripps Translational Science Institute, San Diego, California 92037, USA;

  6. 6Advanced Applications Group, Life Technologies, Beverly, Massachusetts 01915, USA;

  7. 7Department of Biochemistry and Biophysics UCSF, San Francisco, California 94143, USA;

  8. 8Department of Immunohematology and Blood Transfusion, Leiden University Medical Center, 2333 ZA Leiden, The Netherlands;

  9. 9Centre for Clinical Research, University of Queensland, Herston, QLD 4006, Australia

Abstract

The somatic mutation burden in healthy white blood cells (WBCs) is not well known. Based on deep whole-genome sequencing, we estimate that approximately 450 somatic mutations accumulated in the nonrepetitive genome within the healthy blood compartment of a 115-yr-old woman. The detected mutations appear to have been harmless passenger mutations: They were enriched in noncoding, AT-rich regions that are not evolutionarily conserved, and they were depleted for genomic elements where mutations might have favorable or adverse effects on cellular fitness, such as regions with actively transcribed genes. The distribution of variant allele frequencies of these mutations suggests that the majority of the peripheral white blood cells were offspring of two related hematopoietic stem cell (HSC) clones. Moreover, telomere lengths of the WBCs were significantly shorter than telomere lengths from other tissues. Together, this suggests that the finite lifespan of HSCs, rather than somatic mutation effects, may lead to hematopoietic clonal evolution at extreme ages.

SOURCE

http://genome.cshlp.org/content/24/5/733

Blood of world’s oldest woman hints at limits of life

By Andy Coghlan

https://www.newscientist.com/article/dn25458-blood-of-worlds-oldest-woman-hints-at-limits-of-life

Read Full Post »


Familial transthyretin amyloid polyneuropathy

Curator: Larry H. Bernstein, MD, FCAP

LPBI

 

First-Ever Evidence that Patisiran Reduces Pathogenic, Misfolded TTR Monomers and Oligomers in FAP Patients

We reported data from our ongoing Phase 2 open-label extension (OLE) study of patisiran, an investigational RNAi therapeutic targeting transthyretin (TTR) for the treatment of TTR-mediated amyloidosis (ATTR amyloidosis) patients with familial amyloidotic polyneuropathy (FAP). Alnylam scientists and collaborators from The Scripps Research Institute and Misfolding Diagnostics, Inc. were able to measure the effects of patisiran on pathogenic, misfolded TTR monomers and oligomers in FAP patients. Results showed a rapid and sustained reduction in serum non-native conformations of TTR (NNTTR) of approximately 90%. Since NNTTR is pathogenic in ATTR amyloidosis and the level of NNTTR reduction correlated with total TTR knockdown, these results provide direct mechanistic evidence supporting the therapeutic hypothesis that TTR knockdown has the potential to result in clinical benefit. Furthermore, complete 12-month data from all 27 patients that enrolled in the patisiran Phase 2 OLE study showed sustained mean maximum reductions in total serum TTR of 91% for over 18 months and a mean 3.1-point decrease in mNIS+7 at 12 months, which compares favorably to an estimated increase in mNIS+7 of 13 to 18 points at 12 months based upon analysis of historical data sets in untreated FAP patients with similar baseline characteristics. Importantly, patisiran administration continues to be generally well tolerated out to 21 months of treatment.

Read our press release

View the non-native TTR poster (480 KB PDF)

View the complete 12-month patisiran Phase 2 OLE data presentation (620 KB PDF)

We are encouraged by these new data that provide continued support for our hypothesis that patisiran has the potential to halt neuropathy progression in patients with FAP. If these results are replicated in a randomized, double-blind, placebo-controlled study, we believe that patisiran could emerge as an important treatment option for patients suffering from this debilitating, progressive and life-threatening disease.

 

Hereditary ATTR Amyloidosis with Polyneuropathy (hATTR-PN)

ATTR amyloidosis is a progressive, life-threatening disease caused by misfolded transthyretin (TTR) proteins that accumulate as amyloid fibrils in multiple organs, but primarily in the peripheral nerves and heart. ATTR amyloidosis can lead to significant morbidity, disability, and mortality. The TTR protein is produced primarily in the liver and is normally a carrier for retinol binding protein – one of the vehicles used to transport vitamin A around the body.  Mutations in the TTR gene cause misfolding of the protein and the formation of amyloid fibrils that typically contain both mutant and wild-type TTR that deposit in tissues such as the peripheral nerves and heart, resulting in intractable peripheral sensory neuropathy, autonomic neuropathy, and/or cardiomyopathy.

Click to Enlarge

 

ATTR represents a major unmet medical need with significant morbidity and mortality. There are over 100 reported TTR mutations; the particular TTR mutation and the site of amyloid deposition determine the clinical manifestations of the disease whether it is predominantly symptoms of neuropathy or cardiomyopathy.

Specifically, hereditary ATTR amyloidosis with polyneuropathy (hATTR-PN), also known as familial amyloidotic polyneuropathy (FAP), is an inherited, progressive disease leading to death within 5 to 15 years. It is due to a mutation in the transthyretin (TTR) gene, which causes misfolded TTR proteins to accumulate as amyloid fibrils predominantly in peripheral nerves and other organs. hATTR-PN can cause sensory, motor, and autonomic dysfunction, resulting in significant disability and death.

It is estimated that hATTR-PN, also known as FAP, affects approximately 10,000 people worldwide.  Patients have a life expectancy of 5 to 15 years from symptom onset, and the only treatment options for early stage disease are liver transplantation and TTR stabilizers such as tafamidis (approved in Europe) and diflunisal.  Unfortunately liver transplantation has limitations, including limited organ availability as well as substantial morbidity and mortality. Furthermore, transplantation eliminates the production of mutant TTR but does not affect wild-type TTR, which can further deposit after transplantation, leading to cardiomyopathy and worsening of neuropathy. There is a significant need for novel therapeutics to treat patients who have inherited mutations in the TTR gene.

Our ATTR program is the lead effort in our Genetic Medicine Strategic Therapeutic Area (STAr) product development and commercialization strategy, which is focused on advancing innovative RNAi therapeutics toward genetically defined targets for the treatment of rare diseases with high unmet medical need.  We are developing patisiran (ALN-TTR02), an intravenously administered RNAi therapeutic, to treat the hATTR-PN form of the disease.

Patisiran for the Treatment hATTR-PN

APOLLO Phase 3 Trial

In 2012, Alnylam entered into an exclusive alliance with Genzyme, a Sanofi company, to develop and commercialize RNAi therapeutics, including patisiran and revusiran, for the treatment of ATTR amyloidosis in Japan and the broader Asian-Pacific region. In early 2014, this relationship was extended as a significantly broader alliance to advance RNAi therapeutics as genetic medicines. Under this new agreement, Alnylam will lead development and commercialization of patisiran in North America and Europe while Genzyme will develop and commercialize the product in the rest of world.

 

Hereditary ATTR Amyloidosis with Cardiomyopathy (hATTR-CM)

ATTR amyloidosis is a progressive, life-threatening disease caused by misfolded transthyretin (TTR) proteins that accumulate as amyloid fibrils in multiple organs, but primarily in the peripheral nerves and heart. ATTR amyloidosis can lead to significant morbidity, disability, and mortality. The TTR protein is produced primarily in the liver and is normally a carrier for retinol binding protein – one of the vehicles used to transport vitamin A around the body.  Mutations in the TTR gene cause misfolding of the protein and the formation of amyloid fibrils that typically contain both mutant and wild-type TTR that deposit in tissues such as the peripheral nerves and heart, resulting in intractable peripheral sensory neuropathy, autonomic neuropathy, and/or cardiomyopathy.

Click to Enlarge                            http://www.alnylam.com/web/assets/tetramer.jpg

ATTR represents a major unmet medical need with significant morbidity and mortality. There are over 100 reported TTR mutations; the particular TTR mutation and the site of amyloid deposition determine the clinical manifestations of the disease, whether it is predominantly symptoms of neuropathy or cardiomyopathy.

Specifically, hereditary ATTR amyloidosis with cardiomyopathy (hATTR-CM), also known as familial amyloidotic cardiomyopathy (FAC), is an inherited, progressive disease leading to death within 2 to 5 years. It is due to a mutation in the transthyretin (TTR) gene, which causes misfolded TTR proteins to accumulate as amyloid fibrils primarily in the heart. Hereditary ATTR amyloidosis with cardiomyopathy can result in heart failure and death.

While the exact numbers are not known, it is estimated hATTR-CM, also known as FAC affects at least 40,000 people worldwide.  hATTR-CM is fatal within 2 to 5 years of diagnosis and treatment is currently limited to supportive care.  Wild-type ATTR amyloidosis (wtATTR amyloidosis), also known as senile systemic amyloidosis, is a nonhereditary, progressive disease leading to death within 2 to 5 years. It is caused by misfolded transthyretin (TTR) proteins that accumulate as amyloid fibrils in the heart. Wild-type ATTR amyloidosis can cause cardiomyopathy and result in heart failure and death. There are no approved therapies for the treatment of hATTR-CM or SSA; hence there is a significant unmet need for novel therapeutics to treat these patients.

Our ATTR program is the lead effort in our Genetic Medicine Strategic Therapeutic Area (STAr) product development and commercialization strategy, which is focused on advancing innovative RNAi therapeutics toward genetically defined targets for the treatment of rare diseases with high unmet medical need.  We are developing revusiran (ALN-TTRsc), a subcutaneously administered RNAi therapeutic for the treatment of hATTR-CM.

Revusiran for the Treatment of hATTR-CM

ENDEAVOUR Phase 3 Trial

In 2012, Alnylam entered into an exclusive alliance with Genzyme, a Sanofi company, to develop and commercialize RNAi therapeutics, including patisiran and revusiran, for the treatment of ATTR amyloidosis in Japan and the broader Asian-Pacific region. In early 2014, this relationship was extended as a broader alliance to advance RNAi therapeutics as genetic medicines. Under this new agreement, Alnylam and Genzyme have agreed to co-develop and co-commercialize revusiran in North America and Europe, with Genzyme developing and commercializing the product in the rest of world. This broadened relationship on revusiran is aimed at expanding and accelerating the product’s global value.

Pre-Clinical Data and Advancement of ALN-TTRsc02 for Transthyretin-Mediated Amyloidosis

We presented pre-clinical data with ALN-TTRsc02, an investigational RNAi therapeutic targeting transthyretin (TTR) for the treatment of TTR-mediated amyloidosis (ATTR amyloidosis).  In pre-clinical studies, including those in non-human primates (NHPs), ALN-TTRsc02 achieved potent and highly durable knockdown of serum TTR of up to 99% with multi-month durability achieved after just a single dose, supportive of a potentially once quarterly dose regimen. Results from studies comparing TTR knockdown activity of ALN-TTRsc02 to that of revusiran showed that ALN-TTRsc02 has a markedly superior TTR knockdown profile.  Further, in initial rat toxicology studies, ALN-TTRsc02 was found to be generally well tolerated with no significant adverse events at doses as high as 100 mg/kg.

Read our press release

View the presentation

http://www.alnylam.com/product-pipeline/hereditary-attr-amyloidosis-with-cardiomyopathy/

 

Emerging Therapies for Transthyretin Cardiac Amyloidosis Could Herald a New Era for the Treatment of HFPEF

Oct 14, 2015   |  Adam Castano, MDDavid Narotsky, MDMathew S. Maurer, MD, FACC

http://www.acc.org/latest-in-cardiology/articles/2015/10/13/08/35/emerging-therapies-for-transthyretin-cardiac-amyloidosis#sthash.9xzc0rIe.dpuf

Heart failure with a preserved ejection fraction (HFPEF) is a clinical syndrome that has no pharmacologic therapies approved for this use to date. In light of failed medicines, cardiologists have refocused treatment strategies based on the theory that HFPEF is a heterogeneous clinical syndrome with different etiologies. Classification of HFPEF according to etiologic subtype may, therefore, identify cohorts with treatable pathophysiologic mechanisms and may ultimately pave the way forward for developing meaningful HFPEF therapies.1

A wealth of data now indicates that amyloid infiltration is an important mechanism underlying HFPEF. Inherited mutations in transthyretin cardiac amyloidosis (ATTRm) or the aging process in wild-type disease (ATTRwt) cause destabilization of the transthyretin (TTR) protein into monomers or oligomers, which aggregate into amyloid fibrils. These insoluble fibrils accumulate in the myocardium and result in diastolic dysfunction, restrictive cardiomyopathy, and eventual congestive heart failure (Figure 1). In an autopsy study of HFPEF patients, almost 20% without antemortem suspicion of amyloid had left ventricular (LV) TTR amyloid deposition.2 Even more resounding evidence for the contribution of TTR amyloid to HFPEF was a study in which 120 hospitalized HFPEF patients with LV wall thickness ≥12 mm underwent technetium-99m 3,3-diphosphono-1,2-propranodicarboxylic acid (99mTc-DPD) cardiac imaging,3,4 a bone isotope known to have high sensitivity and specificity for diagnosing TTR cardiac amyloidosis.5,6 Moderate-to-severe myocardial uptake indicative of TTR cardiac amyloid deposition was detected in 13.3% of HFPEF patients who did not have TTR gene mutations. Therefore, TTR cardiac amyloid deposition, especially in older adults, is not rare, can be easily identified, and may contribute to the underlying pathophysiology of HFPEF.

Figure 1

As no U.S. Food and Drug Administration-approved drugs are currently available for the treatment of HFPEF or TTR cardiac amyloidosis, the development of medications that attenuate or prevent TTR-mediated organ toxicity has emerged as an important therapeutic goal. Over the past decade, a host of therapies and therapeutic drug classes have emerged in clinical trials (Table 1), and these may herald a new direction for treating HFPEF secondary to TTR amyloid.

Table 1

TTR Silencers (siRNA and Antisense Oligonucleotides)

siRNA

Ribonucleic acid interference (RNAi) has surfaced as an endogenous cellular mechanism for controlling gene expression. Small interfering RNAs (siRNAs) delivered into cells can disrupt the production of target proteins.7,8 A formulation of lipid nanoparticle and triantennary N-acetylgalactosamine (GalNAc) conjugate that delivers siRNAs to hepatocytes is currently in clinical trials.9 Prior research demonstrated these GalNAc-siRNA conjugates result in robust and durable knockdown of a variety of hepatocyte targets across multiple species and appear to be well suited for suppression of TTR gene expression and subsequent TTR protein production.

The TTR siRNA conjugated to GalNAc, ALN-TTRSc, is now under active investigation as a subcutaneous injection in phase 3 clinical trials in patients with TTR cardiac amyloidosis.10 Prior phase 2 results demonstrated that ALN-TTRSc was generally well tolerated in patients with significant TTR disease burden and that it reduced both wild-type and mutant TTR gene expression by a mean of 87%. Harnessing RNAi technology appears to hold great promise for treating patients with TTR cardiac amyloidosis. The ability of ALN-TTRSc to lower both wild-type and mutant proteins may provide a major advantage over liver transplantation, which affects the production of only mutant protein and is further limited by donor shortage, cost, and need for immunosuppression.

Antisense Oligonucleotides

Antisense oligonucleotides (ASOs) are under clinical investigation for their ability to inhibit hepatic expression of amyloidogenic TTR protein. Currently, the ASO compound, ISIS-TTRRx, is under investigation in a phase 3 multicenter, randomized, double-blind, placebo-controlled clinical trial in patients with familial amyloid polyneuropathy (FAP).11 The primary objective is to evaluate its efficacy as measured by change in neuropathy from baseline relative to placebo. Secondary measures will evaluate quality of life (QOL), modified body mass index (mBMI) by albumin, and pharmacodynamic effects on retinol binding protein. Exploratory objectives in a subset of patients with LV wall thickness ≥13 mm without a history of persistent hypertension will examine echocardiographic parameters, N-terminal pro–B-type natriuretic peptide (NT-proBNP), and polyneuropathy disability score relative to placebo. These data will facilitate analysis of the effect of antisense oligonucleotide-mediated TTR suppression on the TTR cardiac phenotype with a phase 3 trial anticipated to begin enrollment in 2016.

TTR Stabilizers (Diflunisal, Tafamidis)

Diflunisal

Several TTR-stabilizing agents are in various stages of clinical trials. Diflunisal, a traditionally used and generically available nonsteroidal anti-inflammatory drug (NSAID), binds and stabilizes familial TTR variants against acid-mediated fibril formation in vitro and is now in human clinical trials.12,13 The use of diflunisal in patients with TTR cardiac amyloidosis is controversial given complication of chronic inhibition of cyclooxygenase (COX) enzymes, including gastrointestinal bleeding, renal dysfunction, fluid retention, and hypertension that may precipitate or exacerbate heart failure in vulnerable individuals.14-17 In TTR cardiac amyloidosis, an open-label cohort study suggested that low-dose diflunisal with careful monitoring along with a prophylactic proton pump inhibitor could be safely administered to compensated patients.18 An association was observed, however, between chronic diflunisal use and adverse changes in renal function suggesting that advanced kidney disease may be prohibitive in diflunisal therapy.In FAP patients with peripheral or autonomic neuropathy randomized to diflunisal or placebo, diflunisal slowed progression of neurologic impairment and preserved QOL over two years of follow-up.19 Echocardiography demonstrated cardiac involvement in approximately 50% of patients.20 Longer-term safety and efficacy data over an average 38 ± 31 months in 40 Japanese patients with hereditary ATTR amyloidosis who were not candidates for liver transplantation showed that diflunisal was mostly well tolerated.12 The authors cautioned the need for attentive monitoring of renal function and blood cell counts. Larger multicenter collaborations are needed to determine diflunisal’s true efficacy in HFPEF patients with TTR cardiac amyloidosis.

Tafamidis

Tafamidis is under active investigation as a novel compound that binds to the thyroxine-binding sites of the TTR tetramer, inhibiting its dissociation into monomers and blocking the rate-limiting step in the TTR amyloidogenesis cascade.21 The TTR compound was shown in an 18-month double-blind, placebo-controlled trial to slow progression of neurologic symptoms in patients with early-stage ATTRm due to the V30M mutation.22 When focusing on cardiomyopathy in a phase 2, open-label trial, tafamidis also appeared to effectively stabilize TTR tetramers in non-V30M variants, wild-type and V122I, as well as biochemical and echocardiographic parameters.23,24 Preliminary data suggests that clinically stabilized patients had shorter disease duration, lower cardiac biomarkers, less myocardial thickening, and higher EF than those who were not stabilized, suggesting early institution of therapy may be beneficial. A phase 3 trial has completed enrollment and will evaluate the efficacy, safety, and tolerability of tafamidis 20 or 80 mg orally vs. placebo.25 This will contribute to long-term safety and efficacy data needed to determine the therapeutic effects of tafamidis among ATTRm variants.

Amyloid Degraders (Doxycycline/TUDCA and Anti-SAP Antibodies)

Doxycycline/TUDCA

While silencer and stabilizer drugs are aimed at lowering amyloidogenic precursor protein production, they cannot remove already deposited fibrils in an infiltrated heart. Removal of already deposited fibrils by amyloid degraders would be an important therapeutic strategy, particularly in older adults with heavily infiltrated hearts reflected by thick walls, HFPEF, systolic heart failure, and restrictive cardiomyopathy. Combined doxycycline and tauroursodeoxycholic acid (TUDCA) disrupt TTR amyloid fibrils and appeared to have an acceptable safety profile in a small phase 2 open-label study among 20 TTR patients. No serious adverse reactions or clinical progression of cardiac or neuropathic involvement was observed over one year.26 An active phase 2, single-center, open-label, 12-month study will assess primary outcome measures including mBMI, neurologic impairment score, and NT-proBNP.27 Another phase 2 study is examining the tolerability and efficacy of doxycycline/TUDCA over an 18-month period in patients with TTR amyloid cardiomyopathy.28 Additionally, a study in patients with TTR amyloidosis is ongoing to determine the effect of doxycycline alone on neurologic function, cardiac biomarkers, echocardiographic parameters, modified body mass index, and autonomic neuropathy.29

Anti-SAP Antibodies

In order to safely clear established amyloid deposits, the role of the normal, nonfibrillar plasma glycoprotein present in all human amyloid deposits, serum amyloid P component (SAP), needs to be more clearly understood.30 In mice with amyloid AA type deposits, administration of antihuman SAP antibody triggered a potent giant cell reaction that removed massive visceral amyloid deposits without adverse effects.31 In humans with TTR cardiac amyloidosis, anti-SAP antibody treatments could be feasible because the bis-D proline compound, CPHPC, is capable of clearing circulating human SAP, which allow anti-SAP antibodies to reach residual deposited SAP. In a small, open-label, single-dose-escalation, phase 1 trial involving 15 patients with systemic amyloidosis, none of whom had clinical evidence of cardiac amyloidosis, were treated with CPHPC followed by human monoclonal IgG1 anti-SAP antibody.32 No serious adverse events were reported and amyloid deposits were cleared from the liver, kidney, and lymph node. Anti-SAP antibodies hold promise as a potential amyloid therapy because of their potential to target all forms of amyloid deposits across multiple tissue types.

Mutant or wild-type TTR cardiac amyloidoses are increasingly recognized as a cause of HFPEF. Clinicians need to be aware of this important HFPEF etiology because the diverse array of emerging disease-modifying agents for TTR cardiac amyloidosis in human clinical trials has the potential to herald a new era for the treatment of HFPEF.

References

  1. Maurer MS, Mancini D. HFpEF: is splitting into distinct phenotypes by comorbidities the pathway forward? J Am Coll Cardiol 2014;64:550-2.
  2. Mohammed SF, Mirzoyev SA, Edwards WD, et al. Left ventricular amyloid deposition in patients with heart failure and preserved ejection fraction. JACC Heart Fail 2014;2:113-22.
  3. González-López E, Gallego-Delgado M, Guzzo-Merello G, et al. Wild-type transthyretin amyloidosis as a cause of heart failure with preserved ejection fraction. Eur Heart J 2015.
  4. Castano A, Bokhari S, Maurer MS. Unveiling wild-type transthyretin cardiac amyloidosis as a significant and potentially modifiable cause of heart failure with preserved ejection fraction. Eur Heart J 2015 Jul 28. [Epub ahead of print]
  5. Rapezzi C, Merlini G, Quarta CC, et al. Systemic cardiac amyloidoses: disease profiles and clinical courses of the 3 main types. Circulation 2009;120:1203-12.
  6. Bokhari S, Castano A, Pozniakoff T, Deslisle S, Latif F, Maurer MS. (99m)Tc-pyrophosphate scintigraphy for differentiating light-chain cardiac amyloidosis from the transthyretin-related familial and senile cardiac amyloidoses. Circ Cardiovasc Imaging 2013;6:195-201.
  7. Fire A, Xu S, Montgomery MK, Kostas SA, Driver SE, Mello CC. Potent and specific genetic interference by double-stranded RNA in Caenorhabditis elegans. Nature 1998;391:806-11.
  8. Elbashir SM, Harborth J, Lendeckel W, Yalcin A, Weber K, Tuschl T. Duplexes of 21-nucleotide RNAs mediate RNA interference in cultured mammalian cells. Nature 2001;411:494-8.
  9. Kanasty R, Dorkin JR, Vegas A, Anderson D. Delivery materials for siRNA therapeutics. Nature Mater 2013;12:967-77.
  10. U.S. National Institutes of Health. Phase 2 Study to Evaluate ALN-TTRSC in Patients With Transthyretin (TTR) Cardiac Amyloidosis (ClinicalTrials.gov website). 2014. Available at: https://www.clinicaltrials.gov/ct2/show/NCT01981837. Accessed 8/19/2015.
  11. U.S. National Institutes of Health. Efficacy and Safety of ISIS-TTRRx in Familial Amyloid Polyneuropathy (Clinical Trials.gov Website. 2013. Available at: http://www.clinicaltrials.gov/ct2/show/NCT01737398. Accessed 8/19/2015.
  12. Sekijima Y, Dendle MA, Kelly JW. Orally administered diflunisal stabilizes transthyretin against dissociation required for amyloidogenesis. Amyloid 2006;13:236-49.
  13. Tojo K, Sekijima Y, Kelly JW, Ikeda S. Diflunisal stabilizes familial amyloid polyneuropathy-associated transthyretin variant tetramers in serum against dissociation required for amyloidogenesis. Neurosci Res 2006;56:441-9.
  14. Epstein M. Non-steroidal anti-inflammatory drugs and the continuum of renal dysfunction. J Hypertens Suppl 2002;20:S17-23.
  15. Wallace JL. Pathogenesis of NSAID-induced gastroduodenal mucosal injury. Best Pract Res Clin Gastroenterol 2001;15:691-703.
  16. Mukherjee D, Nissen SE, Topol EJ. Risk of cardiovascular events associated with selective COX-2 inhibitors. JAMA 2001;286:954-9.
  17. Page J, Henry D. Consumption of NSAIDs and the development of congestive heart failure in elderly patients: an underrecognized public health problem. Arch Intern Med 2000;160:777-84.
  18. Castano A, Helmke S, Alvarez J, Delisle S, Maurer MS. Diflunisal for ATTR cardiac amyloidosis. Congest Heart Fail 2012;18:315-9.
  19. Berk JL, Suhr OB, Obici L, et al. Repurposing diflunisal for familial amyloid polyneuropathy: a randomized clinical trial. JAMA 2013;310:2658-67.
  20. Quarta CCF, Solomon RH Suhr SD, et al. The prevalence of cardiac amyloidosis in familial amyloidotic polyneuropathy with predominant neuropathy: The Diflunisal Trial. International Symposium on Amyloidosis 2014:88-9.
  21. Hammarstrom P, Jiang X, Hurshman AR, Powers ET, Kelly JW. Sequence-dependent denaturation energetics: A major determinant in amyloid disease diversity. Proc Natl Acad Sci U S A 2002;99 Suppl 4:16427-32.
  22. Coelho T, Maia LF, Martins da Silva A, et al. Tafamidis for transthyretin familial amyloid polyneuropathy: a randomized, controlled trial. Neurology 2012;79:785-92.
  23. Merlini G, Plante-Bordeneuve V, Judge DP, et al. Effects of tafamidis on transthyretin stabilization and clinical outcomes in patients with non-Val30Met transthyretin amyloidosis. J Cardiovasc Transl Res 2013;6:1011-20.
  24. Maurer MS, Grogan DR, Judge DP, et al. Tafamidis in transthyretin amyloid cardiomyopathy: effects on transthyretin stabilization and clinical outcomes. Circ Heart Fail 2015;8:519-26.
  25. U.S. National Institutes of Health. Safety and Efficacy of Tafamidis in Patients With Transthyretin Cardiomyopathy (ATTR-ACT) (ClinicalTrials.gov website). 2014. Available at: http://www.clinicaltrials.gov/show/NCT01994889. Accessed 8/19/2015.
  26. Obici L, Cortese A, Lozza A, et al. Doxycycline plus tauroursodeoxycholic acid for transthyretin amyloidosis: a phase II study. Amyloid 2012;19 Suppl 1:34-6.
  27. U.S. National Institutes of Health. Safety, Efficacy and Pharmacokinetics of Doxycycline Plus Tauroursodeoxycholic Acid in Transthyretin Amyloidosis (ClinicalTrials.gov website). 2011. Available at: http://www.clinicaltrials.gov/ct2/show/NCT01171859. Accessed 8/19/2015.
  28. U.S. National Institutes of Health. Tolerability and Efficacy of a Combination of Doxycycline and TUDCA in Patients With Transthyretin Amyloid Cardiomyopathy (ClinicalTrials.gov website). 2013. Available at: http://www.clinicaltrials.gov/ct2/show/NCT01855360. Accessed 8/19/2015.
  29. U.S. National Institutes of Health. Safety and Effect of Doxycycline in Patients With Amyloidosis (ClinicalTrials.gov website).2015. Available at: https://clinicaltrials.gov/ct2/show/NCT01677286. Accessed 8/19/2015.
  30. Pepys MB, Dash AC. Isolation of amyloid P component (protein AP) from normal serum as a calcium-dependent binding protein. Lancet 1977;1:1029-31.
  31. Bodin K, Ellmerich S, Kahan MC, et al. Antibodies to human serum amyloid P component eliminate visceral amyloid deposits. Nature 2010;468:93-7.
  32. Richards DB, Cookson LM, Berges AC, et al. Therapeutic Clearance of Amyloid by Antibodies to Serum Amyloid P Component. N Engl J Med 2015;373:1106-14.

 

The Acid-Mediated Denaturation Pathway of Transthyretin Yields a Conformational Intermediate That Can Self-Assemble into Amyloid

Zhihong Lai , Wilfredo Colón , and Jeffery W. Kelly *
Department of Chemistry, Texas A&M University, College Station, Texas 77843-3255
Biochemistry199635 (20), pp 6470–6482   http://dx.doi.org:/10.1021/bi952501g
Publication Date (Web): May 21, 1996  Copyright © 1996 American Chemical Society

Transthyretin (TTR) amyloid fibril formation is observed during partial acid denaturation and while refolding acid-denatured TTR, implying that amyloid fibril formation results from the self-assembly of a conformational intermediate. The acid denaturation pathway of TTR has been studied in detail herein employing a variety of biophysical methods to characterize the intermediate(s) capable of amyloid fibril formation. At physiological concentrations, tetrameric TTR remains associated from pH 7 to pH 5 and is incapable of amyloid fibril formation. Tetrameric TTR dissociates to a monomer in a process that is dependent on both pH and protein concentration below pH 5. The extent of amyloid fibril formation correlates with the concentration of the TTR monomer having an altered, but defined, tertiary structure over the pH range of 5.0−3.9. The inherent Trp fluorescence-monitored denaturation curve of TTR exhibits a plateau over the pH range where amyloid fibril formation is observed (albeit at a higher concentration), implying that a steady-state concentration of the amyloidogenic intermediate with an altered tertiary structure is being detected. Interestingly, 1-anilino-8-naphthalenesulfonate fluorescence is at a minimum at the pH associated with maximal amyloid fibril formation (pH 4.4), implying that the amyloidogenic intermediate does not have a high extent of hydrophobic surface area exposed, consistent with a defined tertiary structure. Transthyretin has two Trp residues in its primary structure, Trp-41 and Trp-79, which are conveniently located far apart in the tertiary structure of TTR. Replacement of each Trp with Phe affords two single Trp containing variants which were used to probe local pH-dependent tertiary structural changes proximal to these chromophores. The pH-dependent fluorescence behavior of the Trp-79-Phe mutant strongly suggests that Trp-41 is located near the site of the tertiary structural rearrangement that occurs in the formation of the monomeric amyloidogenic intermediate, likely involving the C-strand−loop−D-strand region. Upon further acidification of TTR (below pH 4.4), the structurally defined monomeric amyloidogenic intermediate begins to adopt alternative conformations that are not amyloidogenic, ultimately forming an A-state conformation below pH 3 which is also not amyloidogenic. In summary, analytical equilibrium ultracentrifugation, SDS−PAGE, far- and near-UV CD, fluorescence, and light scattering studies suggest that the amyloidogenic intermediate is a monomeric predominantly β-sheet structure having a well-defined tertiary structure.

 

Prevention of Transthyretin Amyloid Disease by Changing Protein Misfolding Energetics

Per Hammarström*, R. Luke Wiseman*, Evan T. Powers, Jeffery W. Kelly   + Author Affiliations

Science  31 Jan 2003; 299(5607):713-716   http://dx.doi.org:/10.1126/science.1079589

Genetic evidence suggests that inhibition of amyloid fibril formation by small molecules should be effective against amyloid diseases. Known amyloid inhibitors appear to function by shifting the aggregation equilibrium away from the amyloid state. Here, we describe a series of transthyretin amyloidosis inhibitors that functioned by increasing the kinetic barrier associated with misfolding, preventing amyloidogenesis by stabilizing the native state. The trans-suppressor mutation, threonine 119 → methionine 119, which is known to ameliorate familial amyloid disease, also functioned through kinetic stabilization, implying that this small-molecule strategy should be effective in treating amyloid diseases.

 

Rational design of potent human transthyretin amyloid disease inhibitors

Thomas Klabunde1,2, H. Michael Petrassi3, Vibha B. Oza3, Prakash Raman3, Jeffery W. Kelly3 & James C. Sacchettini1

Nature Structural & Molecular Biology 2000; 7: 312 – 321.                http://dx.doi.org:/10.1038/74082

The human amyloid disorders, familial amyloid polyneuropathy, familial amyloid cardiomyopathy and senile systemic amyloidosis, are caused by insoluble transthyretin (TTR) fibrils, which deposit in the peripheral nerves and heart tissue. Several nonsteroidal anti-inflammatory drugs and structurally similar compounds have been found to strongly inhibit the formation of TTR amyloid fibrils in vitro. These include flufenamic acid, diclofenac, flurbiprofen, and resveratrol. Crystal structures of the protein–drug complexes have been determined to allow detailed analyses of the protein–drug interactions that stabilize the native tetrameric conformation of TTR and inhibit the formation of amyloidogenic TTR. Using a structure-based drug design approach ortho-trifluormethylphenyl anthranilic acid and N-(meta-trifluoromethylphenyl) phenoxazine 4,6-dicarboxylic acid have been discovered to be very potent and specific TTR fibril formation inhibitors. This research provides a rationale for a chemotherapeutic approach for the treatment of TTR-associated amyloid diseases.

 

First European consensus for diagnosis, management, and treatment of transthyretin familial amyloid polyneuropathy

Adams, Davida; Suhr, Ole B.b; Hund, Ernstc; Obici, Laurad; Tournev, Ivailoe,f; Campistol, Josep M.g; Slama, Michel S.h; Hazenberg, Bouke P.i; Coelho, Teresaj; from the European Network for TTR-FAP (ATTReuNET)

Current Opin Neurol: Feb 2016; 29 – Issue – p S14–S26      http://dx.doi.org:/10.1097/WCO.0000000000000289

Purpose of review: Early and accurate diagnosis of transthyretin familial amyloid polyneuropathy (TTR-FAP) represents one of the major challenges faced by physicians when caring for patients with idiopathic progressive neuropathy. There is little consensus in diagnostic and management approaches across Europe.

Recent findings: The low prevalence of TTR-FAP across Europe and the high variation in both genotype and phenotypic expression of the disease means that recognizing symptoms can be difficult outside of a specialized diagnostic environment. The resulting delay in diagnosis and the possibility of misdiagnosis can misguide clinical decision-making and negatively impact subsequent treatment approaches and outcomes.

Summary: This review summarizes the findings from two meetings of the European Network for TTR-FAP (ATTReuNET). This is an emerging group comprising representatives from 10 European countries with expertise in the diagnosis and management of TTR-FAP, including nine National Reference Centres. The current review presents management strategies and a consensus on the gold standard for diagnosis of TTR-FAP as well as a structured approach to ongoing multidisciplinary care for the patient. Greater communication, not just between members of an individual patient’s treatment team, but also between regional and national centres of expertise, is the key to the effective management of TTR-FAP.

http://images.journals.lww.com/co-neurology/Original.00019052-201602001-00003.FF1.jpeg

Transthyretin familial amyloid polyneuropathy (TTR-FAP) is a highly debilitating and irreversible neurological disorder presenting symptoms of progressive sensorimotor and autonomic neuropathy [1▪,2▪,3]. TTR-FAP is caused by misfolding of the transthyretin (TTR) protein leading to protein aggregation and the formation of amyloid fibrils and, ultimately, to amyloidosis (commonly in the peripheral and autonomic nervous system and the heart) [4,5]. TTR-FAP usually proves fatal within 7–12 years from the onset of symptoms, most often due to cardiac dysfunction, infection, or cachexia [6,7▪▪].

The prevalence and disease presentation of TTR-FAP vary widely within Europe. In endemic regions (northern Portugal, Sweden, Cyprus, and Majorca), patients tend to present with a distinct genotype in large concentrations, predominantly a Val30Met substitution in the TTR gene [8–10]. In other areas of Europe, the genetic footprint of TTR-FAP is more varied, with less typical phenotypic expression [6,11]. For these sporadic or scattered cases, a lack of awareness among physicians of variable clinical features and limited access to diagnostic tools (i.e., pathological studies and genetic screening) can contribute to high rates of misdiagnosis and poorer patient outcomes [1▪,11]. In general, early and late-onset variants of TTR-FAP, found within endemic and nonendemic regions, present several additional diagnostic challenges [11,12,13▪,14].

Delay in the time to diagnosis is a major obstacle to the optimal management of TTR-FAP. With the exception of those with a clearly diagnosed familial history of FAP, patients still invariably wait several years between the emergence of first clinical signs and accurate diagnosis [6,11,14]. The timely initiation of appropriate treatment is particularly pertinent, given the rapidity and irreversibility with which TTR-FAP can progress if left unchecked, as well as the limited effectiveness of available treatments during the later stages of the disease [14]. This review aims to consolidate the existing literature and present an update of the best practices in the management of TTR-FAP in Europe. A summary of the methods used to achieve a TTR-FAP diagnosis is presented, as well as a review of available treatments and recommendations for treatment according to disease status.

Patients with TTR-FAP can present with a range of symptoms [11], and care should be taken to acquire a thorough clinical history of the patient as well as a family history of genetic disease. Delay in diagnosis is most pronounced in areas where TTR-FAP is not endemic or when there is no positive family history [1▪]. TTR-FAP and TTR-familial amyloid cardiomyopathy (TTR-FAC) are the two prototypic clinical disease manifestations of a broader disease spectrum caused by an underlying hereditary ATTR amyloidosis [19]. In TTR-FAP, the disease manifestation of neuropathy is most prominent and definitive for diagnosis, whereas cardiomyopathy often suggests TTR-FAC. However, this distinction is often superficial because cardiomyopathy, autonomic neuropathy, vitreous opacities, kidney disease, and meningeal involvement all may be present with varying severity for each patient with TTR-FAP.

Among early onset TTR-FAP with usually positive family history, symptoms of polyneuropathy present early in the disease process and usually predominate throughout the progression of the disease, making neurological testing an important diagnostic aid [14]. Careful clinical examination (e.g., electromyography with nerve conduction studies and sympathetic skin response, quantitative sensation test, quantitative autonomic test) can be used to detect, characterize, and scale the severity of neuropathic abnormalities involving small and large nerve fibres [10]. Although a patient cannot be diagnosed definitively with TTR-FAP on the basis of clinical presentation alone, symptoms suggesting the early signs of peripheral neuropathy, autonomic dysfunction, and cardiac conduction disorders or infiltrative cardiomyopathy are all indicators that further TTR-FAP diagnostic investigation is warranted. Late-onset TTR-FAP often presents as sporadic cases with distinct clinical features (e.g., milder autonomic dysfunction) and can be more difficult to diagnose than early-onset TTR-FAP (Table 2) [1▪,11,12,13▪,14,20].

http://images.journals.lww.com/co-neurology/LargeThumb.00019052-201602001-00003.TT2.jpeg

Genetic testing is carried out to allow detection of specific amyloidogenic TTR mutations (Table 1), using varied techniques depending on the expertise and facilities available in each country (Table S2, http://links.lww.com/CONR/A39). A targeted approach to detect a specific mutation can be used for cases belonging to families with previous diagnosis. In index cases of either endemic and nonendemic regions that do not have a family history of disease, are difficult to confirm, and have atypical symptoms, TTR gene sequencing is required for the detection of both predicted and new amyloidogenic mutations [26,27].

Following diagnosis, the neuropathy stage and systemic extension of the disease should be determined in order to guide the next course of treatment (Table 4) [3,30,31]. The three stages of TTR-FAP severity are graded according to a patient’s walking disability and degree of assistance required [30]. Systemic assessment, especially of the heart, eyes, and kidney, is also essential to ensure all aspects of potential impact of the disease can be detected [10].

Table 4

http://images.journals.lww.com/co-neurology/LargeThumb.00019052-201602001-00003.TT4.jpeg

Image Tools

The goals of cardiac investigations are to detect serious conduction disorders with the risk of sudden death and infiltrative cardiomyopathy. Electrocardiograms (ECG), Holter-ECG, and intracardiac electrophysiology study are helpful to detect conduction disorders. Echocardiograms, cardiac magnetic resonance imaging, scintigraphy with bone tracers, and biomarkers (e.g., brain natriuretic peptide, troponin) can all help to diagnose infiltrative cardiomyopathy[10]. An early detection of cardiac abnormalities has obvious benefits to the patient, given that the prophylactic implantation of pacemakers was found to prevent 25% of major cardiac events in TTR-FAP patients followed up over an average of 4 years [32▪▪]. Assessment of cardiac denervation with 123-iodine meta-iodobenzylguanidine is a powerful prognostic marker in patients diagnosed with FAP [33].

…..

Tafamidis

Tafamidis is a first-in-class therapy that slows the progression of TTR amyloidogenesis by stabilizing the mutant TTR tetramer, thereby preventing its dissociation into monomers and amyloidogenic and toxic intermediates [55,56]. Tafamidis is currently indicated in Europe for the treatment of TTR amyloidosis in adult patients with stage I symptomatic polyneuropathy to delay peripheral neurological impairment [57].

In an 18-month, double-blind, placebo-controlled study of patients with early-onset Val30Met TTR-FAP, tafamidis was associated with a 52% lower reduction in neurological deterioration (P = 0.027), a preservation of nerve function, and TTR stabilization versus placebo [58▪▪]. However, only numerical differences were found for the coprimary endpoints of neuropathy impairment [neuropathy impairment score in the lower limb (NIS-LL) responder rates of 45.3% tafamidis vs 29.5% placebo; P = 0.068] and quality of life scores [58▪▪]. A 12-month, open-label extension study showed that the reduced rates of neurological deterioration associated with tafamidis were sustained over 30 months, with earlier initiation of tafamidis linking to better patient outcomes (P = 0.0435) [59▪]. The disease-slowing effects of tafamidis may be dependent on the early initiation of treatment. In an open-label study with Val30Met TTR-FAP patients with late-onset and advanced disease (NIS-LL score >10, mean age 56.4 years), NIS-LL and disability scores showed disease progression despite 12 months of treatment with tafamidis, marked by a worsening of neuropathy stage in 20% and the onset of orthostatic hypotension in 22% of patients at follow-up [60▪].

Tafamidis is not only effective in patients exhibiting the Val30Met mutation; it also has proven efficacy, in terms of TTR stabilization, in non-Val30Met patients over 12 months [61]. Although tafamidis has demonstrated safe use in patients with TTR-FAP, care should be exercised when prescribing to those with existing digestive problems (e.g., diarrhoea, faecal incontinence) [60▪].

Back to Top | Article Outline

Diflunisal

Diflunisal is a nonsteroidal anti-inflammatory drug (NSAID) that, similar to tafamidis, slows the rate of amyloidogenesis by preventing the dissociation, misfolding, and misassembly of the mutated TTR tetramer [62,63]. Off-label use has been reported for patients with stage I and II disease, although diflunisal is not currently licensed for the treatment of TTR-FAP.

Evidence for the clinical effectiveness of diflunisal in TTR-FAP derives from a placebo-controlled, double-blind, 24-month study in 130 patients with clinically detectable peripheral or autonomic neuropathy[64▪]. The deterioration in NIS scores was significantly more pronounced in patients receiving placebo compared with those taking diflunisal (P = 0.001), and physical quality of life measures showed significant improvement among diflunisal-treated patients (P = 0.001). Notable during this study was the high rate of attrition in the placebo group, with 50% more placebo-treated patients dropping out of this 2-year study as a result of disease progression, advanced stage of the disease, and varied mutations.

One retrospective analysis of off-label use of diflunisal in patients with TTR-FAP reported treatment discontinuation in 57% of patients because of adverse events that were largely gastrointestinal [65]. Conclusions on the safety of diflunisal in TTR-FAP will depend on further investigations on the impact of known cardiovascular and renal side-effects associated with the NSAID drug class [66,67].

 

 

 

 

Read Full Post »

Older Posts »