VA National Precision Oncology Program

In partnership with the current DNA sequencing contractor, NPOP provides access to a second gene panel for hematologic malignancies or sarcomas, though neither of these classes of malignancies currently have clear indications for routine NGS multigene panel testing. Given the low rate of finding a gene mutation that would change therapy that could not be found with smaller, less expensive gene panels, NPOP requires prior approval for the use of this panel.

Finally, since early 2019, programmed deathligand 1 (PD-L1) immunohistochemistry analysis is available through NPOP in association with NGS testing of the same sample for those solid tumors with US Food and Drug Administration (FDA)-approved indications that include a PD-L1 companion diagnostic. This service was added to facilitate concurrent testing of PD-L1 and DNA sequencing, which speeds availability of molecular data to the health care provider and veteran.

Determining Clinical Significance

The complexity of tumor NGS gene panel test results is far greater than frequently ordered laboratory or molecular testing due to the near infinite number of possible results and varying degrees of consensus of the significance of the results for therapeutic decision making. That complexity is reflected in the length of the test reports, which are often ≥ 20 pages. Starting from the gene variants identified by the DNA sequencing variant-caller bioinformatics pipeline, there is a 2-step process, referred to as annotation, to interpret the clinical significance that is repeated for each variant.

The first step is to assign a pathogenicity value, also known as oncogenicity, using a 5-point Likert scale from pathogenic to benign with variant of unknown significance (VUS) in the middle of the scale. Only variants that are pathogenic or likely pathogenic are considered further. A VUS is usually communicated to the health care provider but should generally not be acted on, while benign and likely benign variants may or may not be included in the report and should never be acted on. NPOP examined the concordance of pathogenicity calls among 3 annotation services: N-of-One/QCI Precision Insights (qiagen.com), IBM Watson for Genomics (WfG), and OncoKB (www.oncokb.org). ⁶ There was moderate-to-poor concordance, indicating lack of consensus about whether a significant fraction of observed gene variants contributes to the patient’s cancer. This variability likely arises due to differences in algorithms and criteria used to assess pathogenicity.The second step of annotation is assignment of the actionability of the variant, using a level of evidence (LoE) scale from 1 (on-label indication) to 4 (absence of clinical evidence; ie, only preclinical or theoretical evidence). Initially, NPOP used an adaptation of the LoE scales from WfG and OncoKB but now mostly uses the recently revised OncoKB LoE. Actionability also includes prediction of resistance to a treatment (LoE level R1 and R2). An example of a resistance gene variant is a KRAS mutation in colorectal cancer, which predicts lack of clinical benefit from anti- EGFR antibodies. It is important to note that a determination of actionability requires 3 inputs: gene, variant, and tumor type. A BRAF V600E mutation in melanoma has different medications with level 1 LoE than does the same mutation in colorectal cancer, for example.