Author: Tilda Barliya PhD
Metastasis, the spread of cancer cells from a primary tumour to seed secondary tumours in distant sites, is one of the greatest challenges in cancer treatment today. For many patients, by the time cancer is detected, metastasis has already occurred. Over 80% of patients diagnosed with lung cancer, for example, present with metastatic disease. Few patients with metastatic cancer are cured by surgical intervention, and other treatment modalities are limited. Across all cancer types, only one in five patients diagnosed with metastatic cancer will survive more than 5 years. (1,2).
Metastatic Cancer
- Metastatic cancer is cancer that has spread from the place where it first started to another place in the body.
- Metastatic cancer has the same name and same type of cancer cells as the original cancer.
- The most common sites of cancer metastasis are the lungs, bones, and liver.
- Treatment for metastatic cancer usually depends on the type of cancer and the size, location, and number of metastatic tumors.
How do cancer cells spread (3)
- Local invasion: Cancer cells invade nearby normal tissue.
- Intravasation: Cancer cells invade and move through the walls of nearby lymph vessels or blood vessels.
- Circulation: Cancer cells move through the lymphatic system and the bloodstream to other parts of the body.
The ability of a cancer cell to metastasize successfully depends on its individual properties; the properties of the noncancerous cells, including immune system cells, present at the original location; and the properties of the cells it encounters in the lymphatic system or the bloodstream and at the final destination in another part of the body. Not all cancer cells, by themselves, have the ability to metastasize. In addition, the noncancerous cells at the original location may be able to block cancer cell metastasis. Furthermore, successfully reaching another location in the body does not guarantee that a metastatic tumor will form. Metastatic cancer cells can lie dormant (not grow) at a distant site for many years before they begin to grow again, if at all.
Although cancer therapies are improving, many drugs are not reaching the sites of metastases, and doubt remains over the efficacy of those that do. Methods that are effective for treating large, well-vascularized tumours may be inadequate when dealing with small clusters of disseminated malignant cells.
We expect that the expanding capabilities of nanotechnology, especially in targeting, detection and particle trafficking, will enable novel approaches to treat cancers even after metastatic dissemination.
Lymph nodes, which are linked by lymphatic vessels, are distributed throughout the body and have an integral role in the immune response. Dissemination of cancer cells through the lymph network is thought to be an important route for metastatic spread. Tumor proximal lymph nodes are often the first site of metastases, and the presence of lymph node metastases signifies further metastatic spread and poor patient survival.
As such, lymph nodes have been targeted using cell-based nanotechnologies
Lymph nodes are small, bean-shaped organs that act as filters along the lymph fluid channels. As lymph fluid leaves the organ (such as breast, lung etc) and eventually goes back into the bloodstream, the lymph nodes try to catch and trap cancer cells before they reach other parts of the body. Having cancer cells in the lymph nodes suggests an increased risk of the cancer spreading. It is thus very important to evaluate the involvement of lymph nodes when choosing the best possible treatment for the patient.
Although current mapping methods are available such as CT and MRI scans, PET scan, Endobronchial Ultrasound, Mediastinoscopy and lymph node biopsy, sentinel lymph node (SLN) mapping and nodal treatment in lung cancer remain inadequate for routine clinical use.
Certain characteristics are associated with preferential (but not exclusive) nanoparticle trafficking to lymph nodes following intravenous administration.
Targeting is often an indirect process, as receptors on the surface of leukocytes bind nanoparticles and transfer them to lymph nodes as part of a normal immune response. Several strategies have been used to enhance nanoparticle uptake by leukocytes in circulation. Coating iron-oxide nanoparticles with carbohydrates, such as dextran, results in the increased accumulation of these nanoparticles in lymph nodes. Conjugating peptides and antibodies, such as immunoglobulin G (IgG), to the particle surface also increases their accumulation in the lymphatic network. In general, negatively charged particles are taken up at faster rates than positively charged or uncharged particles. Conversely, ‘stealth’ polymers, such as polyethylene glycol (PEG), on the surface of nanoparticles, can inhibit uptake by leukocytes, thereby reducing accumulation in the lymph nodes.
Lymph node targeting may be achieved by other routes of administration. Tsuda and co-workers reported that non-cationic particles with a size range of 6–34nm, when introduced to the lungs (intrapulmonary administration), are trafficked rapidly (<1 hour) to local lymph nodes. Administering particles <80 nm in size subcutaneously also results in trafficking to lymph nodes. Interestingly, some studies have indicated that non-pegylated particles exhibit enhanced accumulation in the lymphatics and that pegylated particles tend to appear in the circulation several hours after administration.
Over the last twenty years, sentinel lymph node (SLN) imaging has revolutionized the treatment of several malignancies, such has melanoma and breast cancer, and has the potential to drastically improve treatment in other malignancies, including lung cancer. Several attempts at developing an easy, reliable, and effective method for SLN mapping in lung cancer have been unsuccessful due to unique difficulties inherent to the lung and to operating in the thoracic cavity.
An inexpensive method offering rapid, intraoperative identification of SLNs, with minimal risk to both patient and provider, would allow for improved staging in patients. This, in turn, would permit better selection of patients for adjuvant therapy, thus reducing morbidity in those patients for whom adjuvant treatment is inappropriate, and ensuring that those who need this added therapy actually receive it. (http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3109504/)
Current methods for SLN identification involve the use of radioactivity-guided mapping with technetium-99m sulfur colloid and/or visual mapping using vital blue dyes. Unfortunately these methods can be inadequate for SLN mapping in non-small cell lung cancer (NSCLC) The use of vital blue dyes is limited in vivo by poor visibility, particularly in the presence of anthracotic mediastinal nodes, thereby decreasing the signal-to-background ratio (SBR) that enables nodal detection. Similarly, results with technetium-99m sulfur colloid have been mixed when used in the thoracic cavity, where hilar structures and aberrant patterns of lymphatic drainage make detection more difficult.
Although Nomori et al. have reported an 83% nodal identification rate following a preoperative injection of technetium-99 colloid, there is an associated increased risk of pneumothorax and bleeding with this method. Further, the recently completed CALGB 140203 multicenter Phase 2 trial investigating the use of intraoperative technetium-99m colloid found an identification rate of only 51% with this technique. Clearly a technology with greater accuracy, improved SBR, and less potential risk to surgeon and patient would be welcome in the field of thoracic oncology.
Near-infrared (NIR) fluorescence imaging has the potential to meet this difficult challenge.
Near-Infrared Light
NIR light is defined as that within the wavelength range of 700 to 1000 nm. Although NIR light is invisible to the naked eye, it can be thought of as “redder” than UV and visible light.
- Absorption, scatter, and autofluorescence are all significantly reduced at redder wavelengths. For instance, Hemoglobin, water, lipids, and other endogenous chromophores, such as melanin, have their lowest absorption within the NIR spectrum, which permits increased photon depth penetration into tissues
- In addition, imaging can also be affected by photon scatter, which describes the reflection and/or deflection of light when it interacts with tissue. Scatter, on an absolute scale, is often ten-times higher than absorption. However, the two major types of scatter, Mie and Rayleigh, are both reduced in the NIR, making the use of NIR wavelengths especially important for the reduction of photon attenuation.
- living tissue has extremely high “autofluorescence” in the UV and visible wavelength ranges due to endogenous fluorophores, such as NADH and the porphyrins. Therefore, UV/visible fluorescence imaging of the intestines, bladder, and gallbladder is essentially precluded. However, in the NIR spectrum, autofluorescence is extremely low, providing the black imaging background necessary for optimal detection of a NIR fluorophore within the surgical field
- Additionally, optical imaging techniques, such as NIR fluorescence, eliminate the need for ionizing radiation. This, combined with the availability of a NIR fluorophore already FDA-approved for other indications and having extremely low toxicity (discussed below), make this a potentially safe imaging modality.
The main disadvantage is that it’s invisible to the human eye, requiring special imaging-systems to “see” the NIR fluorescence.
Currently there are three intraoperative NIR imaging systems in various stages of development:
- The SPY system™ (Novadaq, Canada) – utilizes laser light excitation in order to obtain fluorescent images. The Spy™ system has been studied for imaging patency of vascular anastamoses following CABG and organ transplantation
- The Photodynamic Eye™(Hamamatsu, Japan) – is presently available only in Japan
- The Fluorescence-Assisted Resection and Exploration (FLARE™) system ()- developed by the authors’ laboratory utilizes NIR light-emitting diode (LED) excitation, eliminating the need for a potentially harmful laser. Additionally, the FLARE™system has the advantage of being able to provide simultaneous color imaging, NIR fluorescence imaging, and color-NIR merged images, allowing the surgeon to simultaneously visualize invisible NIR fluorescence images within the context of surgical anatomy.
Near-Infrared Fluorescent Nanoparticle Contrast Agents
The ideal contrast agent for SLN mapping would be anionic and within 10–50 nm in size in order to facilitate rapid uptake into lymphatic vessels with optimal retention within the SLN.
Due to the lack of endogenous NIR tissue fluorescence, exogenous contrast agents must be administered for in vivo studies. The most important contrast agents that emit within the NIR spectrum are the heptamethine cyanines fluorophores, of which indocyanine green (ICG) is the most widely used, and fluorescent semiconductor nanocrystals, also known as quantum dots (QDs).
- ICG is an extremely safe NIR fluorophore, with its only known toxicity being rare anaphylaxis. The dye was FDA approved in 1958 for systemic administration for indicator-dilution studies including measurements of cardiac output and hepatic function. Additionally, it is commonly used in ophthalmic angiography. When given intravenously, ICG is rapidly bound to plasma albumin and cleared from the blood via the biliary system. Peak absorption and emission of ICG occur at 780 nm and 830 nm respectively, within the window where in vivo tissue absorption is at its minimum. ICG has a relatively neutral charge, has a hydrodynamic diameter of only 1.2 nm, and is relatively hydrophobic. Unfortunately, this results in rapid transport out of the SLN and relatively low fluorescence yield, thereby decreasing its efficacy in mapping techniques. However, noncovalent adsorption of ICG to human serum albumin (HSA), as occurs within plasma, results in an anionic nanoparticle with a diameter of 7.3 nm and a three-fold increase in fluorescence yield markedly improving its utility in SLN mapping.
- QDs consist of an inorganic heavy metal core and shell which emit within the NIR spectrum. This structure is then surrounded by a hydrophilic organic coating which facilitates aqeuous solubility and lymphatic distrubtion. QDs have been extensively studied and are ideal for SLN mapping as their hydrodynamic diameter can be customized to the appropriate size within a narrow distribution (15–20 nm), they can be engineered to have an anionic surface charge, and exhibit an extremely high SBRs with significant photostability. Unfortunately, safety concerns due to the presence of heavy metals within the QDs so far have precluded clinical application
Human Clinical Trials and NIR SLN mapping
Several studies have investigated the clinical use of indocyanine green without adsorption to HSA for NIR fluorescence-guided SLN mapping in breast and gastric cancer with good success (9-13).
Kitai et al. first examined this technique in 2005 in breast cancer patients, and was able to identify a SLN node in 17 of 18 patients using NIR fluorescence rather than the visible green color of ICG (9). Sevick-Muraca et al. reported similar results using significantly lower microdoses of ICG (10 – 100 μg), successfully identifying the SLN in 8 of 9 patients (11). Similar to these subcutaneous studies, 56 patients with gastric cancer underwent endoscopic ICG injection into the submucosa around the tumor 1 to 3 days preoperatively or injection directly into the subserosa intraoperatively with identification of the SLN in 54 patients (13).
Recently, Troyan et al. have completed a pilot phase I clinical trial examining the utility of NIR imaging the ICG:HSA nanoparticle fluorophore for SLN mapping/biopsy in breast cancer using the FLARE™system. In this study, 6 patients received both 99mTc-sulfur colloid lymphoscintigraphy along with ICG:HSA at micromolar doses. SLNs were identified in all patients using both methods. In 4 of 6 patients the SLNs identified were the same, while in the remaining two, lymphoscintigraphy identified an additional node in one patient and ICG:HSA identified an additional SLN in the other. Irrespective, this study demonstrates that NIR SLN mapping with low dose ICG:HSA is a viable method for intraoperative SLN identification.
Nanotechnology and Drug Delivery in Lung cancer
We previously explored Lung cancer and nanotechnology aspects as polymer nanotechnology has been an area of significant research over the past decade as polymer nanoparticle drug delivery systems offer several advantages over traditional methods of chemotherapy delivery
see: (15) http://pharmaceuticalintelligence.com/2012/11/08/lung-cancer-nsclc-drug-administration-and-nanotechnology/ (16) http://pharmaceuticalintelligence.com/2012/12/01/diagnosing-lung-cancer-in-exhaled-breath-using-gold-nanoparticles/
As the importance of micrometastatic lymphatic spread of tumor becomes clearer, there has been much interest in the use of nanoparticles for lymphatic drug delivery. The considerable focus on developing an effective method for SLN mapping for lung cancer is indicative of the importance of nodal spread on overall survival.
Our lab is investigating the use of image-guided nanoparticles engineered for lymphatic drug delivery. We have previously described the synthesis of novel, pH-responsive methacrylate nanoparticle systems (14). Following a simple subcutaneous injection of NIR fluorophore-labeled nanoparticles 70 nm in size, we have shown that we can deliver paclitaxel loaded within the particles to regional draining lymph nodes in several organ systems of Yorkshire pigs while simultaneously confirming nodal migration using NIR fluorescent light. Future studies will need to investigate the ability of nanoparticles to treat and prevent nodal metastases in animal cancer models. Additionally, the development of tumor specific nanoparticles will potentially allow for targeting of chemotherapy to small groups of metastatic tumor cells further limiting systemic toxicities by narrowing the delivery of cytotoxic drugs.
Ref:
1. http://www.nature.com.rproxy.tau.ac.il/nrc/journal/v12/n1/pdf/nrc3180.pdf
2. http://www.nature.com/nrc/focus/metastasis/index.html
3. http://www.cancer.gov/cancertopics/factsheet/Sites-Types/metastatic
4. http://www.cancerresearchuk.org/cancer-help/about-cancer/what-is-cancer/body/the-lymphatic-system
6. Khullar O, Frangioni JV and Colson YL. Image-Guided Sentinel Lymph Node Mapping and Nanotechnology-Based Nodal Treatment in Lung Cancer using Invisible Near-Infrared Fluorescent Light. Semi Thorac Cardiovasc Surg 2009 :21 (4); 309-315. http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3109504/
7. Stacker SA, Achen MG, Jussila L, Baldwin ME and Alitalo K. Metastasis: Lymphangiogenesis and cancer metastasis. Nature Reviews Cancer 2002 2, 573-583. http://www.nature.com/nrc/journal/v2/n8/full/nrc863.html
8. Schroeder A., Heller DA., Winslow MM., Dahlman JE., Pratt GW., Langer R., Jacks T and Anderson DG.. Nature Reviews Cancer 2012; 12(1), 39-50. Treating metastatic cancer with nanotechnology. http://www.nature.com.rproxy.tau.ac.il/nrc/journal/v12/n1/pdf/nrc3180.pdf
http://www.nature.com.rproxy.tau.ac.il/nrc/journal/v12/n1/full/nrc3180.html
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11. Sevick-Muraca EM, Sharma R, Rasmussen JC, et al. Imaging of lymph flow in breast cancer patients after microdose administration of a near-infrared fluorophore: feasibility study. Radiology.2008;246:734–741.
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16. http://pharmaceuticalintelligence.com/2012/12/01/diagnosing-lung-cancer-in-exhaled-breath-using-gold-nanoparticles/
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