Comparison of tissue procurement approaches for genomic profiling
Evaluation of: Mojica WD, Sykes DE, Conroy J, Gaile D, Fang X, Nowak N: A comparative analysis of two tissue procurement approaches for the genomic profiling of clinical colorectal cancer samples. Int. J. Colorectal Dis. 23(11), 1089–1098 (2008).
– John W Gillespie
Molecular analysis of cells that are directly involved with the disease processes will play a very important role in the development of personalized medicine. In this article Citation[1], the authors described the most common techniques for sample acquisition for analysis, including tissue sections (curls), laser capture microdissection and the recently developed technique, exfoliation and enrichment (EE), based on three criteria: time, cost and maintenance of the integrity of large-molecular-weight DNA. Since EE and curls are rapid, low-cost techniques that preserve high-molecular-weight DNA, they were compared in their study. They did not include laser capture microdissection in the study, owing to its high cost and long time for cell procurement. Since DNA preservation is well known to be compromised in tissue that has been formalin-fixed and paraffin-embedded, only frozen curls were compared with EE. In their study that compared DNA recovery from EE and curls, they used array comparative genomic hybridization. In this study, they detected a region in chromosome 18 that showed the greatest genetic alterations between normal and cancer. Further examination in this region using quantitative PCR for the curls and EE, demonstrated that the presence of noncancer-contaminating cells in the curls biased the genetic data.
Even though time, cost and DNA integrity are very important criteria to assess cell procurement techniques, the availability of antibodies specific for the target cell population will be the ultimate determinant of the usefulness of EE. In the example of colon cancer used in this study, they used the antibody ber-Ep that binds to both normal colonic epithelial and cancer cells. Since the cancerous and normal areas are readily grossly discernible in colon cancer, they were able to obtain cancer cells with very little contamination with normal colonic epithelial cells. It should be emphasized that a variety of cancer types microscopically show several different admixed cell populations with variable lineages, such as inflammatory cells, blood vessels, fibroblasts, smooth muscle and normal epithelial cells. In this situation, if a cancer-specific antibody is not available, laser microdissection Citation[2–4] may be a very useful alternative. On the other hand, if the cells of interest are difficult to target with laser microdissection (e.g., endothelial cells) and a cell-specific antibody is available, EE may represent the only effective technique to procure these cells. Owing to the high-throughput, low cost, preservation of high-molecular-weight DNA, and in some cases, the only option available to procure pure populations of cells, EE represents a major milestone in the ‘toolbox‘ of technologies that will greatly accelerate the development of personalized medicine.
Financial & competing interests disclosure
This project has been funded in whole or in part with federal funds from the National Cancer Institute, National Institutes of Health, under Contract No. HHSN261200800001E. The content of this publication does not necessarily reflect the views or policies of the Department of Health and Human Services, nor does mention of trade names, commercial products, or organizations imply endorsement by the US Government. The authors have no other relevant affiliations or financial involvement with any organization or entity with a financial interest in or financial conflict with the subject matter or materials discussed in the manuscript apart from those disclosed.
No writing assistance was utilized in the production of this manuscript.
EGFR Mutations in Circulating Cancer Cells
Evaluation of: Maheswaran S, Sequist LV, Nagrath S et al.: Detection of mutations in EGFR in circulating lung-cancer cells. N. Engl. J. Med. 359(4), 366–377 (2008).
– Rafael Rosell, Miquel Taron, Cristina Queralt & Jia Wei
The detection of circulating tumor cells (CTCs) in peripheral blood was first described by Ashworth in 1869 (reviewed in Citation[1]). Since then, the analysis of CTCs has developed, and now shows great promise for the monitoring of breast cancer Citation[2,3]. The CellSearch™ assay, used for the isolation and enumeration of CTCs, consists of the use of magnetic bead-conjugated antibodies against epithelial-cell adhesion molecule (EpCAM) Citation[2,3]. The assay is highly accurate and reproducible Citation[4] and has received US FDA clearance Citation[1]; however, it allows for the detection of a relatively small number of CTCs. When breast cancer patients were assessed with the CellSearch assay, those with more than five CTCs per 7.5 ml of whole blood at baseline and first follow-up had significantly shorter progression-free and overall survival than those with less than five Citation[2,3].
However, the clinical validation of this method remains an unmet challenge for other common cancers. Investigators from Massachusetts General Hospital (MA, USA) have developed a microfluidic-based device called the CTC chip that can isolate, quantify and analyze a median of 67 CTCs per ml in almost all tested patients with metastatic cancer, including nonsmall-cell lung cancer (NSCLC), prostate, pancreatic and colorectal cancers, but not from healthy controls Citation[5]. Maheswaran et al. used the CTC chip to capture CTCs from the blood of NSCLC patients, with a median number of 7 Citation[6]. CTCs/mm and EGFR mutations were identified in CTCs from 11 of 12 NSCLC patients (92%). The authors also detected the drug-resistant T790M mutation in CTCs collected from patients with EGFR mutations who had received tyrosine kinase inhibitors. Importantly, serial analysis of CTCs showed that a reduction in the number of captured cells was associated with tumor response, while an increase was related to tumor progression Citation[6].
The field of CTC research could be even more important if gene-expression profiling with this method becomes feasible, in which case CTC evaluation could be used as a real-time biopsy Citation[1]. Intriguingly, in experimental models of lung tumor cells, drug-surviving lung cancer stem cells have been identified following chemotherapy, with high metastatic potential Citation[7]. We can speculate that if CTC evaluation could identify these cancer stem cells among the CTCs, it would become a very useful tool for predicting chemoresistance and designing alternative treatment strategies, including customized therapies.
Financial & competing interests disclosure
The authors have no relevant affiliations or financial involvement with any organization or entity with a financial interest in or financial conflict with the subject matter or materials discussed in the manuscript. This includes employment, consultancies, honoraria, stock ownership or options, expert testimony, grants or patents received or pending, or royalties.
No writing assistance was utilized in the production of this manuscript.
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- Maheswaran S , SequistLV, NagrathS et al.: Detection of mutations in EGFR in circulating lung-cancer cells.N. Engl. J. Med.359 , 366–377 (2008).
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