Chairman and Director Chief of Lymphoma Director, Clinical and Translational Cancer Research

		John Theurer Cancer Center at Hackensack University Medical Center

Medicine has transformed at a dizzying pace over the last decade, much of it fueled by rapid advances in genomics and high throughput technology, particularly next-generation sequencing (NGS). While the first human genome took almost 7 years to complete at a cost of $3 billion, the same genome can now be sequenced in less than 2 weeks at a cost that will soon be under $1000.

 Since cancer is fundamentally a disorder of DNA, there has been growing angst amongst some anatomic pathologists that their traditional roles as diagnosticians will succumb to an army of DNA sequencers—the concern is that if tissue is collected by surgeons and interventional radiologists and directly sequenced, pathologists will be bypassed entirely.

These fears are unfounded. In fact, as high-throughput technology becomes more widely available, it will be integrated as a tool for pathologists and clinicians working together for better patient stratification and improved outcomes.

As Dr Juan Rosai, one of the greatest surgical pathologists of our time, has written, the amount of information gleaned from a single H&E slide is “staggering.” He argues that “the morphologic appearance of a tumor as seen in an H&E slide represents the grand synthesis of thousands of genes working in concert and sometimes in opposition, and there is probably not a single gene that plays an important role in the neoplastic process whose expression is not manifested in one way or another in a morphologic change that can be detected by those with the training and ability to do it.”

Traditional high-throughput molecular analysis, such as DNA sequencing or microarrays, starts by grinding up the tissue into a “mix,” which obscures the spatial components of the disease. Next-generation sequencing definitely benefits from a pathologist choosing the most appropriate tissue for testing and, in some cases, should use microdissection.

Depending on the chemistry employed by the NGS analyzer, the length of read, and the type of read (uni- vs bidirectional), NGS can have significant base-call error rates, leading to background noise.

 In specimens with low concentrations of tumor, this “noise” can mask low-level changes in DNA sequences. Accurate identification of the tumor tissue-of-origin and subtype classification is also a prerequisite for proper interpretation of molecular results.

The most common example is lung cancer, in which adenocarcinoma and small cell lung carcinoma (SCLC) are known to be biologically distinct. The finding of an EGFR mutation in a lung adenocarcinoma does confer increased sensitivity to EGFR tyrosine kinase inhibitors, while the same mutation has no (defined) such benefit in SCLC.

Similarly, while a BRAF V600E mutation may predict increased sensitivity to vemurafenib in melanoma, the same mutation abrogates the favorable outcome of anti-EGFR monoclonal antibodies in colorectal cancers that are wild-type KRAS.

 Clearly, the significance of any mutation must be interpreted in the context of morphology.

Finally, the presence of a mutation or an atypical molecular result may not be an indicator of malignancy. Multiple studies have shown that highly sensitive PCR techniques can detect the presence of a BCR-ABL transcript in up to 10% of healthy individuals who have no evidence of chronic myelogenous leukemia.

 Other studies have identified IGH/BCL2 translocations, the most common cytogenetic finding in follicular lymphoma, in up to 50% of peripheral blood of otherwise normal individuals.

 Such findings may become increasing common as new techniques become available to analyze minute quantities of cell-free DNA in plasma. Pathologists will be required to interpret and integrate these results as part of the overall diagnostic work-up.

The continued progress of cancer cell biology and rapidly advancing molecular techniques will continue to help define more precise subentities. The complexity of the results needs to be put in perspective of the overall diagnosis for its clinical significance. A few decades ago pathology diagnosis was based essentially on morphology, then came immunohistochemistry and flow cytometry, which changed the field and did not replace morphology. As expected, these technologies were progressively integrated into the full pathology analysis. No one nowadays would sign a report without IHC markers and/or flow cytometry results to confirm clonality or give prognostic information. Additional emerging technologies, such as digitalization of tissue, will help through automation both processing speed and quantification of components involved in morphology analysis.

The origins of pathology (Greek, pathologia: “study of emotions or suffering”) could perhaps date back to ancient physicians struggling through observations and philosophy while trying to unravel the mysteries of the human body and its afflictions. Pathologists in the molecular era, I would argue, will become critical in teaching and further guiding clinicians on how to interpret molecular findings and, as such, will be a central to the effort of precision medicine.

			National Human Genome Research Institute. “About About NHGRI: A Brief History and Timeline.” http://www.genome.gov/10001763. Accessed March 17, 2015.

			Rosai, J. The continuing role of morphology in the molecular age. 

			Wall, JD et al. Estimating genotype error rates from high-coverage next- generation sequence data. 

			Liu, L et al. Comparison of next-generation sequencing systems. 

			Cagle PT, Myers J. Precision medicine for lung cancer: role of the surgical pathologist. 

			Mao C et al. BRAF V600E mutation and resistance to anti-EGFR monoclonal antibodies in patients with metastatic colorectal cancer: a meta-analysis. 

			Ismail SI et al. Incidence of bcr-abl fusion transcripts in healthy indi- viduals. Mol Med Rep. 2014;9(4):1271-1276.

			Paltiel O et al. Translocation t(14;18) in healthy individuals: preliminary study of its association with family history and agricultural expo- sure. 

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