Rankin, Andrew

Purpose Molecular diagnostic testing has become an integral part of the evaluation of patients with metastatic colorectal cancer (CRC). Expanded mutational testing, such as next-generation sequencing (NGS), often identifies mutations with unclear clinical or prognostic implications. One such example is BRAF mutations that occur outside of codon 600 (non-V600 BRAF mutations). Methods We conducted this multicenter, retrospective cohort study to characterize the clinical, pathologic, and survival implications of non-V600 BRAF mutations in metastatic CRC. We pooled patients in whom non-V600 BRAF mutations were identified from NGS databases at three large molecular genetics reference laboratories. Results A total of 9,643 patients with metastatic CRC underwent NGS testing. We identified 208 patients with non-V600 BRAF mutations, which occurred in 2.2% of all patients tested and accounted for 22% of all BRAF mutations identified. Cancers with non-V600 BRAF mutations, compared with cancers with V600E BRAF (V600E BRAF) mutations, were found in patients who were significantly younger (58 v 68 years, respectively), fewer female patients (46% v 65%, respectively), and patients who had fewer high-grade tumors (13% v 64%, respectively) or right-sided primary tumors (36% v 81%, respectively). Median overall survival was significantly longer in patients with non-V600 BRAF-mutant metastatic CRC compared with those with both V600E BRAF-mutant and wild-type BRAF metastatic CRC (60.7 v 11.4 v 43.0 months, respectively; P < .001). In multivariable analysis, non-V600 BRAF mutation was independently associated with improved overall survival (hazard ratio, 0.18; P < .001). Conclusion Non-V600 BRAF mutations occur in approximately 2.2% of patients with metastatic CRC and define a clinically distinct subtype of CRC with an excellent prognosis.

497 Background: As activating RAS mutations have been shown to predict lack of benefit from anti-EGFR therapies in advanced CRC, NCCN guidelines recommend testing for KRAS exon 2 and non-exon 2 mutations; however, these alterations are thought to explain only a subset of de novo resistance to targeted therapy. In a large set of CRC assayed with comprehensive genomic profiling (CGP) in the course of clinical care, we assessed the frequency of less common KRAS short insertions that may predict failure of anti-EGFR therapy. Methods: 4,422 CRC cases were assayed with CGP performed on hybridization-captured, adaptor ligation based libraries to a mean coverage depth of > 650X for at least 236 cancer-related genes plus 47 introns from 19 genes frequently rearranged in cancer. All classes of genomic alterations (GA) were identified, including base pair substitutions, insertions/deletions, copy number alterations, and rearrangements. Pertinent available prior molecular testing results and clinical history was reviewed for selected cases. Results: Out of 4,422 CRC cases analyzed, KRAS short variants were identified in 50.9% of cases. A majority of cases contained a KRAS alteration at either codon G12 (35.5%) or G13 (9.1%), while alterations at codons Q61 or A146 were identified in 2.2% and 3.1% of cases, respectively. KRAS insertions were identified in 8 ( < 0.5%) cases. All KRAS insertions identified fell within codons 9-13, and 6/8 cases harbored V9_G10 insertions. Out of 6 patients with prior KRAS testing results available, 5 (83%) were negative by previous testing. Conclusions: CGP identifies KRAS insertions within or adjacent to hotspot regions in CRC cases which have previously tested negative for KRAS mutations. Given the importance of KRAS alterations in predicting lack of response to anti-EGFR therapies in CRC, accurate detection of these alterations in the course of clinical care is essential for effective treatment. CGP offers the possibility of identifying KRAS insertions that may impact efficacy of anti-EGFR targeted therapy and should be considered when previous focused testing for KRAS mutations is negative.

Abstract Introduction. A KRAS mutation represented the first genomic biomarker to predict lack of benefit from anti-epidermal growth factor receptor (EGFR) antibody therapy in advanced colorectal cancer (CRC). Expanded RAS testing has further refined the treatment approach, but understanding of genomic alterations underlying primary and acquired resistance is limited and further study is needed. Materials and Methods. We prospectively analyzed 4,422 clinical samples from patients with advanced CRC, using hybrid-capture based comprehensive genomic profiling (CGP) at the request of the individual treating physicians. Comparison with prior molecular testing results, when available, was performed to assess concordance. Results. We identified a RAS/RAF pathway mutation or amplification in 62% of cases, including samples harboring KRAS mutations outside of the codon 12/13 hotspot region in 6.4% of cases. Among cases with KRAS non-codon 12/13 alterations for which prior test results were available, 79 of 90 (88%) were not identified by focused testing. Of 1,644 RAS/RAF wild-type cases analyzed by CGP, 31% harbored a genomic alteration (GA) associated with resistance to anti-EGFR therapy in advanced CRC including mutations in PIK3CA, PTEN, EGFR, and ERBB2. We also identified other targetable GA, including novel kinase fusions, receptor tyrosine kinase amplification, activating point mutations, as well as microsatellite instability. Conclusion. Extended genomic profiling reliably detects alterations associated with lack of benefit to anti-EGFR therapy in advanced CRC, while simultaneously identifying alterations potentially important in guiding treatment. The use of CGP during the course of clinical care allows for the refined selection of appropriate targeted therapies and clinical trials, increasing the chance of clinical benefit and avoiding therapeutic futility.