Tumor molecular profiling is a fundamental component of precision oncology, enabling the identification of genomic alterations in genes and pathways that can be targeted therapeutically. The existence of recurrent targetable alterations across distinct histologically-defined tumor types, coupled with an expanding portfolio of molecularly-targeted therapies, demands flexible and comprehensive approaches to profile clinically significant genes across the full spectrum of cancers. We established a large-scale, prospective clinical sequencing initiative utilizing a comprehensive assay, MSK-IMPACT, through which we have compiled matched tumor and normal sequence data from a unique cohort of more than 10,000 patients with advanced cancer and available pathological and clinical annotations. Using these data, we identified clinically relevant somatic mutations, novel non-coding alterations, and mutational signatures that were shared among common and rare tumor types. Patients were enrolled on genomically matched clinical trials at a rate of 11%. To enable discovery of novel biomarkers and deeper investigation into rare alterations and tumor types, all results are publicly accessible.

We report on 16 patients with relapsed or refractory B cell acute lymphoblastic leukemia (B-ALL) that we treated with autologous T cells expressing the 19-28z chimeric antigen receptor (CAR) specific to the CD19 antigen. The overall complete response rate was 88%, which allowed us to transition most of these patients to a standard-of-care allogeneic hematopoietic stem cell transplant (allo-SCT). This therapy was as effective in high-risk patients with Philadelphia chromosome–positive (Ph+) disease as in those with relapsed disease after previous allo-SCT. Through systematic analysis of clinical data and serum cytokine levels over the first 21 days after T cell infusion, we have defined diagnostic criteria for a severe cytokine release syndrome (sCRS), with the goal of better identifying the subset of patients who will likely require therapeutic intervention with corticosteroids or interleukin-6 receptor blockade to curb the sCRS. Additionally, we found that serum C-reactive protein, a readily available laboratory study, can serve as a reliable indicator for the severity of the CRS. Together, our data provide strong support for conducting a multicenter phase 2 study to further evaluate 19-28z CAR T cells in B-ALL and a road map for patient management at centers now contemplating the use of CAR T cell therapy.

Purpose All patients with EGFR mutant lung cancers eventually develop acquired resistance to EGFR tyrosine kinase inhibitors (TKIs). Smaller series have identified various mechanisms of resistance, but systematic evaluation of a large number of patients to definitively establish the frequency of various mechanisms has not been performed. Experimental Design Patients with lung adenocarcinomas and acquired resistance to erlotinib or gefitinib enrolled onto a prospective biopsy protocol and underwent a re-biopsy after the development of acquired resistance. Histology was reviewed. Samples underwent genotyping for mutations in EGFR, AKT1, BRAF, ERBB2, KRAS, MEK1, NRAS and PIK3CA, and FISH for MET and HER2. Results Adequate tumor samples for molecular analysis were obtained in 155 patients. Ninety-eight had second-site EGFR T790M mutations (63%, 95% CI 55-70%) and four had small cell transformation (3%, 95% CI 0-6%). MET amplification was seen in 4/75 (5%, 95% CI 1-13%). HER2 amplification was seen in 3/24 (13%, 95% CI 3-32%). We did not detect any acquired mutations in PIK3CA, AKT1, BRAF, ERBB2, KRAS, MEK1, or NRAS. (0/88, 0%, 95% CI 0-4%). Overlap among mechanisms of acquired resistance was seen in 4%. Conclusions This is the largest series reporting mechanisms of acquired resistance to EGFR TKI therapy. We identified EGFR T790M as the most common mechanism of acquired resistance, while MET amplification, HER2 amplification, and small cell histologic transformation occur less frequently. More comprehensive methods to characterize molecular alterations in this setting are needed to improve our understanding of acquired resistance to EGFR TKIs.

- The 2013 guideline was largely reaffirmed with updated recommendations to allow testing of cytology samples, require improved assay sensitivity, and recommend against the use of immunohistochemistry for EGFR testing. Key new recommendations include ROS1 testing for all adenocarcinoma patients; the inclusion of additional genes ( ERBB2, MET, BRAF, KRAS, and RET) for laboratories that perform next-generation sequencing panels; immunohistochemistry as an alternative to fluorescence in situ hybridization for ALK and/or ROS1 testing; use of 5% sensitivity assays for EGFR T790M mutations in patients with secondary resistance to EGFR inhibitors; and the use of cell-free DNA to "rule in" targetable mutations when tissue is limited or hard to obtain.

Purpose Treatment of advanced non-small-cell lung cancer with immune checkpoint inhibitors (ICIs) is characterized by durable responses and improved survival in a subset of patients. Clinically available tools to optimize use of ICIs and understand the molecular determinants of response are needed. Targeted next-generation sequencing (NGS) is increasingly routine, but its role in identifying predictors of response to ICIs is not known. Methods Detailed clinical annotation and response data were collected for patients with advanced non-small-cell lung cancer treated with anti-programmed death-1 or anti-programmed death-ligand 1 [anti-programmed cell death (PD)-1] therapy and profiled by targeted NGS (MSK-IMPACT; n = 240). Efficacy was assessed by Response Evaluation Criteria in Solid Tumors (RECIST) version 1.1, and durable clinical benefit (DCB) was defined as partial response/stable disease that lasted > 6 months. Tumor mutation burden (TMB), fraction of copy number-altered genome, and gene alterations were compared among patients with DCB and no durable benefit (NDB). Whole-exome sequencing (WES) was performed for 49 patients to compare quantification of TMB by targeted NGS versus WES. Results Estimates of TMB by targeted NGS correlated well with WES (ρ = 0.86; P < .001). TMB was greater in patients with DCB than with NDB ( P = .006). DCB was more common, and progression-free survival was longer in patients at increasing thresholds above versus below the 50th percentile of TMB (38.6% v 25.1%; P < .001; hazard ratio, 1.38; P = .024). The fraction of copy number-altered genome was highest in those with NDB. Variants in EGFR and STK11 associated with a lack of benefit. TMB and PD-L1 expression were independent variables, and a composite of TMB plus PD-L1 further enriched for benefit to ICIs. Conclusion Targeted NGS accurately estimates TMB and elevated TMB further improved likelihood of benefit to ICIs. TMB did not correlate with PD-L1 expression; both variables had similar predictive capacity. The incorporation of both TMB and PD-L1 expression into multivariable predictive models should result in greater predictive power.

Human NSCLCs with activating mutations in EGFR frequently respond to treatment with EGFR tyrosine kinase inhibitors (TKIs) such as erlotinib but responses are not durable as tumors acquire resistance. Secondary mutations in EGFR (T790M) or upregulation of the MET kinase are found in over 50% of resistant tumors. Here, we report increased activation of AXL and evidence of epithelial-to-mesenchymal transition (EMT) in multiple in vitro and in vivo EGFR-mutant lung cancer models with erlotinib acquired resistance in the absence of EGFR T790M or MET activation. Genetic or pharmacologic inhibition of AXL restored sensitivity to erlotinib in these tumor models. Increased expression of AXL, and in some cases its ligand GAS6, was found in EGFR-mutant lung cancers obtained from patients with EGFR TKI acquired resistance. These data identify AXL as a promising therapeutic target whose inhibition could prevent or overcome EGFR TKI acquired resistance in EGFR-mutant lung cancer patients.

The identification of specific genetic alterations as key oncogenic drivers and the development of targeted therapies are together transforming clinical oncology and creating a pressing need for increased breadth and throughput of clinical genotyping. Next-generation sequencing assays allow the efficient and unbiased detection of clinically actionable mutations. To enable precision oncology in patients with solid tumors, we developed Memorial Sloan Kettering-Integrated Mutation Profiling of Actionable Cancer Targets (MSK-IMPACT), a hybridization capture-based next-generation sequencing assay for targeted deep sequencing of all exons and selected introns of 341 key cancer genes in formalin-fixed, paraffin-embedded tumors. Barcoded libraries from patient-matched tumor and normal samples were captured, sequenced, and subjected to a custom analysis pipeline to identify somatic mutations. Sensitivity, specificity, reproducibility of MSK-IMPACT were assessed through extensive analytical validation. We tested 284 tumor samples with previously known point mutations and insertions/deletions in 47 exons of 19 cancer genes. All known variants were accurately detected, and there was high reproducibility of inter- and intrarun replicates. The detection limit for low-frequency variants was approximately 2% for hotspot mutations and 5% for nonhotspot mutations. Copy number alterations and structural rearrangements were also reliably detected. MSK-IMPACT profiles oncogenic DNA alterations in clinical solid tumor samples with high accuracy and sensitivity. Paired analysis of tumors and patient-matched normal samples enables unambiguous detection of somatic mutations to guide treatment decisions.

Summary Clonal hematopoiesis (CH), as evidenced by recurrent somatic mutations in leukemia-associated genes, commonly occurs among aging human hematopoietic stem cells. We analyzed deep coverage, targeted next-generation sequencing (NGS) data of paired tumor and blood samples from 8,810 individuals to assess the frequency and clinical relevance of CH in patients with non-hematologic malignancies. We identified CH in 25% of cancer patients, with 4.5% harboring presumptive leukemia driver mutations (CH-PD). CH was associated with increased age, prior radiation therapy, and tobacco use. PPM1D and TP53 mutations were associated with prior exposure to chemotherapy. CH and CH-PD led to an increased incidence for subsequent hematologic cancers, and CH-PD was associated with shorter patient survival. These data suggest CH occurs in an age-dependent manner and specific perturbations can enhance fitness of clonal hematopoietic stem cells, which can impact outcome through progression to hematologic malignancies and through cell non-autonomous effects on solid tumor biology.