Genetic Engineering & Biotechnology News (GEN) is the world's most widely read biotech publication. It provides the R&D; community with critical information on the tools, technologies, and trends that drive the biotech industry.
Issue link: http://gen.epubxp.com/i/706233
Genetic Engineering & Biotechnology News | GENengnews.com | AUGUST 2016 | 33 achieved significant advances in both sample preparation and informatics analysis." Dr. Vasmatzis brought together a unique combination of technologies to identify large genomic rearrangements and translocations in liquid and solid tumors. The process begins with laser capture microdissection (LCM) to efficiently and accurately extract pure popula- tions of tumor cells. The cells are applied di- rectly into whole-genome amplification pro- tocol, followed by mate-pair (MP) sequencing on Illumina instrumentation. In MP sequencing, the library preparation yields short inserts containing two adjacent fragments that were previously separated by 2–5 kB from each other in the genome. Combining data generated from MP library sequencing with that from short-insert paired- end reads provides a powerful ability to iden- tify complex genomic rearrangements. "The MP approach is a rapid and eco- nomic way of sequencing the whole genome for such abnormalities," asserts Dr. Vas- matzis. "Our Center has already completed over 1,500 tumor genomes." Later this year, the Mayo Clinic plans to offer this test at their CLIA-certified lab. At first, it will support constitutional genetic analysis. Eventually, it will substitute for fluo- rescence in situ hybridization (FISH) and cy- togenetic tests in the analysis of hematologic malignancies and solid tumors. Dr. Vasmatzis describes a typical clinical scenario where this technology may pro- vide valuable information for clinicians: "It is critical to determine whether two distant nodules are genetically related. If they are re- lated, it may indicate an aggressive late-stage tumor, but if they are not related, the disease may still be at stage I, making patient eligible for a potentially curable surgery." Using just a few cells from the fine needle biopsy aspirations, the team is able to detect somatic translocations that are unique for each tumor clone. An added benefit is poten- tial discovery of targetable rearrangements. This protocol may find a wider application in a nationwide screening of individuals with increased cancer risk, such as smokers. "It is still somewhat unclear how the trough of in- formation that our test generates could be pre- sented to physicians," allows Dr. Vasmatzis. "We have been working on visualization strat- egies to make this complex information more readily understood and actionable." Fluid Cancer Monitoring Most patients with metastatic prostate cancer respond well to androgen deprivation therapy. However, many relapse and develop castration-resistant prostate cancer (CRPC). "Novel therapies targeting the androgen receptor (AR) demonstrate significant surviv- al benefit," says Delila Gasi-Tandefelt, Ph.D., Marie Curie Research Fellow, the Institute of Cancer Research (London). "However, 30% of patients do not respond at all, and among those who respond to therapy, invariably all develop resistance." The Institute's laboratory studies the origins and progression of treatment resistance. Mo- lecular characterization of tumors taking serial biopsies could be impractical considering that most of CRPC metastasis develops in bones. In addition, heterogeneity of tumors renders com- prehensive sampling rather challenging. Liquid biopsies present a minimally inva- sive way for longitudinal tracking of tumor molecular makeup. As part of the Institute's treatment resistance team, Dr. Gasi-Tande- felt evaluated the liquid biopsy approach by conducting a comprehensive comparison of cancer tissues and circulating cell-free DNA from the same patients. She showed that tu- mor DNA found in blood is representative of the entire tumor burden. By following mutations in anchor genes, the team characterized the clonality of metastatic disease. The analysis suggested emergence of distinct mechanisms of resistance in different clones that arose or disappeared under treat- ment selection pressure. Sequential monitoring by liquid biopsies may ensure early discontinu- ation of therapies that drive resistance. "Our future research will further focus on evaluation of clonal response to targeted drug therapies," informs Dr. Gasi-Tandefelt. "Tu- mors are very dynamic, especially under selec- tive pressure. Thus, stratifying patients on the basis of past tumor biopsies is problematic." NGS analysis of genetic AR aberrations in CRPC patients, before and after the start of abiraterone treatment, suggested the impor- tant role of somatic mutations and AR gain in resistance. The point mutations could be ob- served months before any clinical manifesta- tion. This data is at the heart of a prospective clinical trial led by Gerhardt Attard, M.D., Ph.D., a senior researcher at the institute. At present, it appears that NGS could inform on the choice of treatment based on the type of the genomic aberration. The next frontier, Dr. Gasi-Tandefelt predicts, will be expansion from the targeted gene panels to whole-genome/whole-exome sequencing, gar- nering even more information for intelligent cancer treatments and single-cell analysis to enable detection of rare resistant clones. Strengthening Quality Control "NGS is revolutionizing our approach to cancer therapeutics," says Francine de Abreu, Ph.D., a genomic analyst at Dartmouth Col- lege. "It is essential for CLIA-certified labora- tories to implement quality control (QC) pro- grams to ensure accuracy and reproducibility of sequencing results. "At present, every CLIA laboratory has its own QC practices. We have established a comprehensive six-step QC process that en- sures accurate sequencing results using forma- lin-fixed paraffin embedded (FFPE) tissues." The workflow is based on the College of American Pathologists (CAP) framework developed over the past few years. It concep- tualizes the overall NGS test process as com- posed of two major analytical components: a "wet bench" component and a "dry bench," or bioinformatics component. Visualize Cellular Responses CELLESTIAL ® Fluorescent Probes www.enzolifesciences.com © 2016 Enzo Life Sciences High-Specifi city Next Generation Fluorescent Probes Our CELLESTIAL ® portfolio of fl uorescent probes and assay kits for cellular analysis provides a complete set of tools for monitoring cell viability, portfolio of fl uorescent probes and assay kits for cellular analysis provides a complete set of tools for monitoring cell viability, portfolio of fl uorescent probes and assay kits for cellular analysis provides a complete set of tools for monitoring cell viability, proliferation, death, oxidative stress and toxicology on fl ow cytometry, microscopy and microplate platforms. Our assays and probes are optimized proliferation, death, oxidative stress and toxicology on fl ow cytometry, microscopy and microplate platforms. Our assays and probes are optimized for the most demanding imaging applications, where consistency and reproducibility are essential. CYTO-ID ® Autophagy Detection Kit A no-transfection assay for monitoring autophagy Fluorescent Microscope Superoxide ROS ROS-ID ª Total ROS/Superoxide Detection Kit Accurately profi le Total ROS and Superoxide with dual-readout assay Fluorescent Microplate Reader NUCLEAR -ID ® Red DNA Stain Brighter cell permeable DNA stain Flow Cytometer CONTROL G 2 /G 1 G 2 /M S scientists enabling scientists. TM ¨ TRANSLATIONAL MEDICINE See NGS on page 34