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Background: Squamous cell carcinoma (SCC) of presumed lung origin (PLO) is now the second most frequent histologic subtype of non-small cell carcinoma after adenocarcinoma. The use of clinic-genomic correlation provided by comprehensive genomic profiling (CGP) can revise clinicopathologic diagnoses of presumed primary lung SCC (PLO-SCC) to diagnoses of metastatic SCC of cutaneous origin (C-SCC)., Design: A total of 10 146 samples of clinically advanced PLO-SCC (84% known Stage IV) passed QC metrics and were designated as PLO-SCCs by review of test requisition forms, clinical notes, and pathology reports. One thousand seven hundred sixty-one cases of known primary C-SCC were also included in this study. All samples underwent hybrid capture-based CGP (Foundation Medicine, Inc.) using a targeted gene panel to evaluate all classes of genomic alterations (GA), determine MSI, TMB, and genomic ancestry status. The mutational signature (MS) of each case was called by the decomposition method using reference signatures in the COSMIC database. PD-L1 tumor cell expression was determined by IHC (22C3; Dako). All results were compared using the Fisher exact method with the false discovery rate corrected with a Benjamini-Hochberg adjustment., Results: A total of 253 of 10 146 (2.5%) PLO-SCC cases featured a UV+ MS; 812 of 1761 C-SCC (46.1%) that also featured a UV radiation exposure MS (UV+) were also included in this study. PLO-SCC UV+ cases used for sequencing included tissue samples from the lung (162), lymph node (34), soft tissue (33), liver (8), head and neck (7), brain (5), and skin thought to be metastatic sites from primary lung SCC (4). The PLO-SCC UV+ patients were 78.7% male and had a median age of 72 years, which was younger and more frequently male gender than both the C-SCC UV+ and C-SCC UV- patients (p < 0.0001). Both the PLO-SCC UV+ and C-SCC UV+ featured greater GA per tumor than the PLO-SCC UV- cases (p < 0.0001). In the PLO-SCC UV- cases, tobacco exposure and APOBEC were the most frequent MSs. For the biomarkers associated with immune checkpoint inhibitor efficacy, when compared with the PLO-SCC UV- cases, the PLO-SCC UV+ cases featured more cases with TMB ≥10 mutations/Mb (88.5% vs. 36.5%; p < 0.0001) and ≥20 mutations/Mb (66.8% vs. 6.8%; p < 0.0001) and a trend for less frequent positive PD-L1 (≥50% TPS) IHC staining (30.2% vs. 39.6%; p = 0.062). Compared to PLO-SCC UV- cases, PLO-SCC UV+ and C-SCC UV+ cases were more likely to harbor clinically-actionable GA in PTCH1 and NOTCH1/2 (p < 0.0001) and less likely to harbor clinically-actionable GA in KRAS, PIK3CA, and PTEN (p < 0.0001). The frequency of PTCH1 GA in PLO-SCC UV+ (32% vs. 0.9% in PLO-SCC UV-) suggested that PLO-SCC UV+ may include a mixture of C-SCC and cutaneous basal cell carcinomas (C-BCC) with squamous differentiation., Conclusions: When cases of PLO-SCC undergo CGP, a small 2.5% subset of cases that featured a UV MS emerge that indicates that these tumors may actually represent metastatic cutaneous SCC or BCC with squamous differentiation. Given the significant treatment and clinical impact associated with the resolution of the true diagnosis of these cases, the use of genomic sequencing in PLO-SCC may be clinically beneficial., (© 2024 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Importance: Biliary tract cancers (BTCs) contain several actionable molecular alterations, including FGFR2, IDH1, ERBB2 (formerly HER2), and KRAS. KRAS allelic variants are found in 20% to 30% of BTCs, and multiple KRAS inhibitors are currently under clinical investigation., Objectives: To describe the genomic landscape, co-sequence variations, immunophenotype, genomic ancestry, and survival outcomes of KRAS-mutated BTCs and to calculate the median overall survival (mOS) for the most common allelic variants., Design, Setting, and Participants: This retrospective, multicenter, pooled cohort study obtained clinical and next-generation sequencing data from multiple databases between January 1, 2017, and December 31, 2022. These databases included Princess Margaret Cancer Centre, MD Anderson Cancer Center, Foundation Medicine, American Association for Cancer Research Project GENIE, and cBioPortal for Cancer Genomics. The cohort comprised patients with BTCs who underwent genomic testing., Main Outcome and Measure: The main outcome was mOS, defined as date of diagnosis to date of death, which was measured in months., Results: A total of 7457 patients (n = 3773 males [50.6%]; mean [SD] age, 63 [5] years) with BTCs and genomic testing were included. Of these patients, 5813 had clinical outcome data available, in whom 1000 KRAS-mutated BTCs were identified. KRAS allelic variants were highly prevalent in perihilar cholangiocarcinoma (28.6%) and extrahepatic cholangiocarcinoma (36.1%). Thirty-six KRAS allelic variants were identified, and the prevalence rates in descending order were G12D (41%), G12V (23%), and Q61H (8%). The variant G12D had the highest mOS of 25.1 (95% CI, 22.0-33.0) months compared with 22.8 (95% CI, 19.6-31.4) months for Q61H and 17.8 (95% CI, 16.3-23.1) months for G12V variants. The majority of KRAS-mutated BTCs (98.9%) were not microsatellite instability-high and had low tumor mutational burden (ranging from a median [IQR] of 1.2 (1.2-2.5) to a mean [SD] of 3.3 [1.3]). Immune profiling through RNA sequencing of KRAS and NRAS-mutated samples showed a pattern toward a more immune-inflamed microenvironment with higher M1 macrophage activation (0.16 vs 0.12; P = .047) and interferon-γ expression compared with wild-type tumors. The G12D variant remained the most common KRAS allelic variant in all patient ancestries. Patients with admixed American ancestry had the highest proportion of G12D variant (45.0%)., Conclusions and Relevance: This cohort study found that KRAS allelic variants were relatively common and may be potentially actionable genomic alterations in patients with BTCs, especially perihilar cholangiocarcinoma and extrahepatic cholangiocarcinoma. The findings add to the growing data on genomic and immune landscapes of KRAS allelic variants in BTCs and are potentially of value to the planning of specific therapies for this heterogeneous patient group.

Purpose: Isocitrate dehydrogenase ( IDH ) 1 / 2 genomic alterations (GA) occur in 20% of intrahepatic cholangiocarcinoma (iCCA); however, the immunogenomic landscape of IDH1-/2- mutated iCCA is largely unknown., Methods: Comprehensive genomic profiling (CGP) was performed on 3,067 cases of advanced iCCA. Tumor mutational burden (TMB), PD-L1 expression (Dako 22C3), microsatellite instability (MSI), and genomic loss of heterozygosity (gLOH) as a surrogate marker for homologous recombination deficiency were examined. RNA sequencing of 73 patient samples was analyzed for differences in stromal/immune cell infiltration, immune marker expression, and T-cell inflammation. Tissue microarray arrays were subjected to multiplex immunohistochemistry and colocalization analysis in 100 surgical samples. Retrospective clinical data were collected for 501 patients with cholangiocarcinoma to examine median overall survival (mOS) in IDH1/2 + versus IDH wt., Results: Of 3,067 iCCA cases subjected to CGP, 426 (14%) were IDH1 + and 125 (4%) were IDH2+ . IDH1 GA included R132C (69%) and R132L/G/S/H/F (16%/7%/4%/3%/<1%). IDH2 GA occurred at R172 (94.4%) and R140 (6.6%). No significant difference was seen in median gLOH between IDH1 + versus IDH wt iCCA ( P = .37), although patterns of comutations differed. MSI-High ( P = .009), TMB ≥10 mut/Mb ( P < .0001), and PD-L1 positivity were lower in IDH1/2+ versus IDHwt iCCA. Resting natural killer cell population, CD70, and programmed cell death 1 expression were significantly higher in non- IDH 1-mutated cases, whereas V-set domain containing T-cell activation inhibitor 1 (B7-H4) expression was significantly higher in IDH1 +. No significant difference in mOS was observed between IDH1/2 + versus IDH wt patients., Conclusion: Significant differences in GA and immune biomarkers are noted between IDH1 / 2 + and IDH wt iCCA. IDH1-/2 -mutated tumors appear immunologically cold without gLOH. These immunogenomic data provide insight for precision targeting of iCCA with IDH alterations.

The micropapillary subtype of urothelial carcinoma (MPUC) of the bladder is a very aggressive histological variant of urothelial bladder cancer (UBC). A high frequency of MPUC contains activating mutations in the extracellular domain (ECD) of ERBB2. We sought to further characterize ERBB2 ECD-mutated MPUC to identify additional genomic alterations that have been associated with tumor progression and therapeutic response. In total, 5,485 cases of archived formalin-fixed, paraffin-embedded UBC underwent comprehensive genomic profiling to identify ERBB2 ECD-mutated MPUC and evaluate the frequencies of genomic co-alterations. We identified 219 cases of UBC with ERBB2 ECD mutations (74% S310F and 26% S310Y), of which 63 (28.8%) were MPUC. Genomic analysis revealed that TERT, TP53, and ARID1A were the most common co-altered genes in ERBB2-mutant MPUC (82.5%, 58.7%, and 39.7%, respectively) and did not differ from ERBB2-mutant non-MPUC (86.5%, 51.9%, and 35.3%). The main differences between ERBB2 ECD-mutated MPUC compared with non-MPUC were KMT2D, RB1, and MTAP alterations. KMT2D and RB1 are tumor-suppressor genes. KMT2D frequency was significantly decreased in ERBB2 ECD-mutated MPUC (6.3%) in contrast to non-MPUC (27.6%; P < .001). RB1 mutations were more frequent in ERBB2 ECD-mutated MPUC (33.3%) than in non-MPUC (17.3%; P = .012). Finally, MTAP loss, an emerging biomarker for new synthetic lethality-based anticancer drugs, was less frequent in ERBB2 ECD-mutated MPUC (11.1%) than in non-MPUC (26.9%; P = .018). Characterizing the genomic landscape of MPUC may not only improve our fundamental knowledge about this aggressive morphological variant of UBC but also has the potential to identify possible prognostic and predictive biomarkers that may drive tumor progression and dictate treatment response to therapeutic approaches., (Copyright © 2024 United States & Canadian Academy of Pathology. Published by Elsevier Inc. All rights reserved.)

Purpose: Approximately 5% of breast cancers each year are diagnosed in young women < 40 years who tend to have worse clinical outcomes. We compared genomic alterations using comprehensive genomic profiling (CGP) of tumor tissue among very young women (< 30 years) and young women (30-39 years) compared to women ≥ 40 years at diagnosis., Methods: 2049 advanced breast cancer cases were submitted to Foundation Medicine within a 22-month window for CGP. Hybrid-capture based CGP was performed to evaluate all classes of genomic alterations. Tumor mutational burden was determined on at least 0.8 Mbp of sequenced DNA and microsatellite instability was determined on at least 95 loci. Immunocyte PD-L1 expression was determined by immunohistochemistry., Results: Of the total cases, 28 (1.37%) were < 30 years, 159 (7.76%) were 30-39 years, and 1862 (90.87%) were ≥ 40 at time of diagnosis. Breast tumors were less likely to be estrogen receptor positive in younger women (54% of < 30 years, p > 0.05; 60% of 30-39 years, p < 0.001; 69.4% of ≥ 40 years) and more likely to be triple negative (43%, p = 0.05; 33%, p = 0.05; 26.1% respectively). Young women had higher rates of BRCA1 mutations (17.9% <30 years, p < 0.001; 10.1% 30-39 years, p < 0.001; 2.6% ≥40 years), but lower rates of CDH1 (7.1% <30 years, p > 0.05; 5.0% 30-39 years, p < 0.001; 15.4% ≥40 years) and PIK3CA mutations (17.9% <30 years, p = 0.02; 17.6% 30-39 years, p < 0.001; 40.0% ≥40 years)., Conclusion: Our findings contribute to the growing literature demonstrating unique genetic profiles among young women diagnosed with breast cancer, compared to older women., (© 2023. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Objective: To evaluate the predictive capability of the pre- and post-pembrolizumab Vesical Imaging-Reporting and Data System (VI-RADS) to identify ypT0N0 or ypT≤1N0 response in muscle-invasive bladder cancer (MIBC) within the PURE-01 trial (ClinicalTrials.gov identifier: NCT02736266)., Patients and Methods: Patients were staged with bladder multiparametric magnetic resonance imaging (mpMRI) before and after treatment (three cycles of pembrolizumab) prior to radical cystectomy (RC). Logistic regression models were used to analyse the pre- and post- pembrolizumab VI-RADS against ypT≤1N0 and ypT0N0 response. The VI-RADS scores were dichotomised between 0 and 3 (0 = no evidence of disease) and 4-5. Event-free survival (EFS) and overall survival (OS) analyses were performed. Comprehensive genomic profiling and transcriptome-wide expression profiling data were matched with the VI-RADS scores., Results: In total, 110 patients underwent centrally reviewed scans (N = 220 mpMRI), treated between February 2017 and July 2020. Both pre- and post-pembrolizumab VI-RADS 0-3 scores were the only significant covariates that predicted the ypT≤1N0 endpoint in multivariable analyses, and the strongest effect was seen with post-pembrolizumab VI-RADS 0-3 predicting the ypT≤1N0 response (P < 0.001). The area under the curve for this model was 0.90. Post-pembrolizumab VI-RADS 0-3 also predicted a longer EFS (P < 0.001) and OS (P = 0.044). The scores of several gene signatures from baseline tumours differed between the pre-pembrolizumab VI-RADS 0-3 and 4-5 categories., Conclusion: Post-pembrolizumab VI-RADS scores are strongly associated with pathological downstaging and survival. VI-RADS scores were also characterised by distinct biomarker features. These results indicate that the VI-RADS is emerging as an important tool for designing next-generation trials for MIBC., (© 2023 BJU International.)

Purpose: BRAF mutations are rare in biliary tract cancers (BTC), but are of interest given the recent developments in targeted therapy for BTC. We investigated the clinical outcomes in a cohort of BRAF-mutant advanced BTC treated with first-line chemotherapy. Furthermore, we investigated the genomic landscape of BRAF class I, II, and III mutations in the intrahepatic cholangiocarcinoma (iCCA) subgroup of BTC., Experimental Design: We analyzed two nonoverlapping cohorts. We examined the genomic landscape of BRAF-mutated iCCA in a "genomic cohort" [187 class I, 82 class II, 113 class III BRAF mutants and 8,026 wildtype (WT)]. We also analyzed median progression-free survival (PFS) and overall survival (OS) on first-line chemotherapy in a separate multi-institutional "clinical cohort" of patients with BTC (including iCCA and extrahepatic cholangiocarcinoma (eCCA) and gallbladder cancer; 41 class I, 32 class II+III BRAF mutants and 1,042 WT)., Results: In the entire BTC clinical cohort, the median PFS was shorter for class I [HR, 2.11 (P < 0.001)] and class II+III [HR, 1.72 (P = 0.007)] as compared with BRAF WT. OS was also shorter in class I [HR, 2.04 (P = 0.011)] and class II+III [HR, 1.86 (P = 0.002)] as compared with BRAF WT. In the iCCA subgroup, class I alterations were mutually exclusive with FGFR2, IDH1/2, ERBB2, and KRAS mutations. Class II+III mutations appear to be mutually exclusive with FGFR2 and KRAS., Conclusions: In BTC, all classes of BRAF mutations are associated with a worse prognosis. BRAF mutations occur in 5% of iCCA subgroup and may be mutually exclusive with other targetable mutations., (©2023 American Association for Cancer Research.)

Background: Despite the low rate of urothelial carcinoma of the bladder (UCB) in patients of South Asian (SAS) and East Asian (EAS) descent, they make up a significant portion of the cases worldwide. Nevertheless, these patients are largely under-represented in clinical trials. We queried whether UCB arising in patients with SAS and EAS ancestry would have unique genomic features compared to the global cohort., Methods: Formalin-fixed, paraffin-embedded tissue was obtained for 8728 patients with advanced UCB. DNA was extracted and comprehensive genomic profiling was performed. Ancestry was classified using a proprietary calculation algorithm. Genomic alterations (GAs) were determined using a 324-gene hybrid-capture-based method which also calculates tumor mutational burden (TMB) and determines microsatellite status (MSI)., Results: Of the cohort, 7447 (85.3%) were EUR, 541 (6.2%) were AFR, 461 (5.3%) were of AMR, 74 (0.85%) were SAS, and 205 (2.3%) were EAS. When compared with EUR, TERT GAs were less frequent in SAS (58.1% vs. 73.6%; P = .06). When compared with non-SAS, SAS had less frequent GAs in FGFR3 (9.5% vs. 18.5%, P = .25). TERT promoter mutations were significantly less frequent in EAS compared to non-EAS (54.1% vs. 72.9%; P < .001). When compared with the non-EAS, PIK3CA alterations were significantly less common in EAS (12.7% vs. 22.1%, P = .005). The mean TMB was significantly lower in EAS vs. non-EAS (8.53 vs. 10.02; P = .05)., Conclusions: The results from this comprehensive genomic analysis of UCB provide important insight into the possible differences in the genomic landscape in a population level. These hypothesis-generating findings require external validation and should support the inclusion of more diverse patient populations in clinical trials., (© The Author(s) 2023. Published by Oxford University Press.)

Genomic alterations (GA) in NF2 tumor-suppressor gene have been associated with aggressive behavior in kidney tumors. We used comprehensive genomic profiling (CGP) to evaluate the frequencies of NF2 GA in histologic subtypes of kidney tumors and co-occurring GA in other genes and biomarkers. Advanced kidney tumors included 1875 clear cell (ccRCC), 405 papillary (pRCC), 108 chromophobe (chRCC), 171 sarcomatoid (sRCC), 61 collecting duct (cdRCC), 49 medullary (mRCC), 134 unclassified (uRCC), 906 urothelial carcinoma of renal pelvis (UC), and 147 Wilms tumors underwent hybrid-capture based CGP to evaluate all classes of GA. 192 (4.9%) of kidney tumors featured NF2 GA which were predominantly structural variant mutations (89%), followed by copy number alterations (9%). Gender and age were similar between NF2-mutant (NF2mut) and NF2-wild type (NF2wt) cohorts with male preponderance. NF2 GA frequency was highest in cdRCC (30%), sRCC (21%), uRCC (15%), and pRCC (12%) while lowest in ccRCC (3%), UC (3%) Wilms tumor (1%), and chRCC (0%). NF2 mutational status was associated with loss of Ch 22 (P < .001). NF2mut RCC harbored co-occurring GA including CDKN2A, CDKN2B, SETD2, and BAP1. VHL, PBRM1, PTEN, and FGFR3 GA were significantly more frequent in NF2wt than in NF2mut tumors. MTOR pathway GAs were uncommon in NF2mut tumors. No NF2 mutated RCC featured MSI-high or high TMB. sRCC was associated with high PD-L1 expression. PD-L1 SP142 tumoral (P = .04) and immune cells (P = .013) were more frequent in NF2mut as compared to NF2wt group. Among histologic subtypes of RCC, cdRCC, sRCC, pRCC, and uRCC are enriched in NF2 GA. Co-occurrent GA in CDKN2A/B, SETD2, and BAP1 may represent potential therapeutic targets. Higher level of PD-L1 expression in NF2mut cohort suggests that these tumors might be sensitive to immune checkpoint inhibitor therapies., (© The Author(s) 2023. Published by Oxford University Press.)

Activating point mutations in Anaplastic Lymphoma Kinase (ALK) have positioned ALK as the only mutated oncogene tractable for targeted therapy in neuroblastoma. Cells with these mutations respond to lorlatinib in pre-clinical studies, providing the rationale for a first-in-child Phase 1 trial (NCT03107988) in patients with ALK-driven neuroblastoma. To track evolutionary dynamics and heterogeneity of tumors, and to detect early emergence of lorlatinib resistance, we collected serial circulating tumor DNA samples from patients enrolled on this trial. Here we report the discovery of off-target resistance mutations in 11 patients (27%), predominantly in the RAS-MAPK pathway. We also identify newly acquired secondary compound ALK mutations in 6 (15%) patients, all acquired at disease progression. Functional cellular and biochemical assays and computational studies elucidate lorlatinib resistance mechanisms. Our results establish the clinical utility of serial circulating tumor DNA sampling to track response and progression and to discover acquired resistance mechanisms that can be leveraged to develop therapeutic strategies to overcome lorlatinib resistance., (© 2023. The Author(s).)

Introduction: New treatment strategies for advanced non-small-cell lung carcinoma (NSCLC) include synthetic lethality targets focused on protein arginine methyl transferases such as PRMT5 that exploit the impact of genomic loss of methylthioadenosine phosphorylase (MTAP)., Methods: Twenty nine thousand three hundred seventy nine advanced NSCLC cases underwent hybrid-capture based comprehensive genomic profiling between June 1, 2018 and May 31, 2020. PD-L1 expression was determined by immunohistochemistry (Dako 22C3 PharmDx assay)., Results: 13.4% (3928/29,379) NSCLC cases exhibited MTAP loss distributed in adenocarcinoma (59%), squamous cell carcinoma (22%), NSCLC not otherwise specified (16%), and 1% each for large-cell neuroendocrine, sarcomatoid, and adenosquamous carcinoma. Statistically significant differences in mitogenic driver alterations included more KRAS G12C mutations in MTAP-intact versus MTAP-lost (12% vs. 10%, p = 0.0003) and fewer EGFR short variant mutations in MTAP-intact versus MTAP-lost NSCLC (10% vs. 13%, p < 0.0001). Statistically significant differences in currently untargetable genomic alterations included higher frequencies of TP53 (70% vs. 63%, p < 0.0001) and RB1 inactivation (10% vs. 2%, p < 0.0001) in MTAP-intact compared to MTAP-lost NSCLC. SMARCA4 inactivation (7% vs. 10%, p < 0.0001) was less frequent in MTAP-intact versus MTAP-lost NSCLC. Alterations in ERBB2, MET, ALK, ROS1, and NTRK1 did not significantly differ between the two groups. Predictors of immunotherapy efficacy were higher in MTAP-intact versus MTAP-lost NSCLC including tumor mutational burden (9.4 vs. 8.6 mut/Mb, p = 0.001) and low (30% vs. 28%, p = 0.01) and high PD-L1 (32% vs. 30%, p = 0.01) expression. Alterations in biomarkers potentially predictive of immune checkpoint inhibitor resistance (STK11, KEAP1, and MDM2) were similar in the two groups., Conclusions: MTAP loss occurs in 13% of NSCLC, supporting the development of targeted therapies to exploit PRMT5 hyper-dependence. MTAP loss is accompanied by small differences in targeted and immunotherapy options which may impact future combination strategies., (© 2022 The Authors. Cancer Medicine published by John Wiley & Sons Ltd.)

Pathological and genomic profiling have transformed breast cancer care by matching patients to targeted treatments. However, tumors evolve and evade therapeutic interventions often through the acquisition of genomic mutations. Here we examine patients profiled with tissue (TBx) and liquid biopsy (LBx) as part of routine clinical care, to characterize the tumor evolutionary landscape and identify potential vulnerabilities in the relapsed setting. Real-world evidence demonstrates that LBx is utilized later in care and identifies associations with intervening therapy. While driver events are frequently shared, acquired LBx alterations are detected in a majority of patients, with the highest frequency in ER+ disease and in patients with longer biopsy intervals. Acquired mutations are often polyclonal and present at lower allelic fractions, suggesting multi-clonal convergent evolution. In addition to well-characterized resistance mutations (e.g., ESR1, NF1, RB1, ERBB2), we observe a diversity of rarer but potentially targetable mutations (e.g., PIK3CA, HRAS/NRAS/KRAS, FGFR1/2/3, BRAF) and fusions (e.g., FGFR1/2, ERBB2, RET), as well as BRCA1/2 reversions through a variety of mechanisms, including splice alterations and structural deletions. This study provides insights on treatment and selection-driven tumor evolution and identifies potential combinatorial treatment options in advanced breast cancer., (© 2022. The Author(s).)

Background: Carcinoma of the urethra (UrthCa) is an uncommon Genitourinary (GU) malignancy that can progress to advanced metastatic disease., Methods: One hundred twenty-seven metastatic UrthCa underwent hybrid capture-based comprehensive genomic profiling to evaluate all classes of genomic alterations (GA). Tumor mutational burden was determined on up to 1.1 Mbp of sequenced DNA, and microsatellite instability was determined on 114 loci. PD-L1 expression was determined by IHC (Dako 22C3)., Results: Forty-nine (39%) urothelial (UrthUC), 31 (24%) squamous (UrthSCC), 24 (19%) adenocarcinomas NOS (UrthAC), and 12 (9%) clear cell (UrthCC) were evaluated. UrthUC and UrthSCC are more common in men; UrthAC and UrthCC are more common in women. Ages were similar in all 4 groups. GA in PIK3CA were the most frequent potentially targetable GA; mTOR pathway GA in PTEN were also identified. GA in other potentially targetable genes were also identified including ERBB2 (6% in UrthUC, 3% in UrthSCC, and 12% in UrthAC), FGFR1-3 (3% in UrthSCC), BRAF (3% in UrthAC), PTCH1 (8% in UrthCC), and MET (8% in UrthCC). Possibly reflecting their higher GA/tumor status, potential for immunotherapy benefit associated with higher tumor mutational burden and PD-L1 staining levels were seen in UrthUC and UrthSCC compared to UrthAC and UrthCC. Microsatellite instability high status was absent throughout., Conclusions: Comprehensive genomic profiling reveals GA that may be predictive of both targeted and immunotherapy benefit in patients with advanced UrthCa and that could potentially be used in future adjuvant, neoadjuvant, and metastatic disease trials., (Copyright © 2020. Published by Elsevier Inc.)

Immune checkpoint inhibitors targeting the PD-1/PD-L1 axis lead to durable clinical responses in subsets of cancer patients across multiple indications, including non-small cell lung cancer (NSCLC), urothelial carcinoma (UC) and renal cell carcinoma (RCC). Herein, we complement PD-L1 immunohistochemistry (IHC) and tumor mutation burden (TMB) with RNA-seq in 366 patients to identify unifying and indication-specific molecular profiles that can predict response to checkpoint blockade across these tumor types. Multiple machine learning approaches failed to identify a baseline transcriptional signature highly predictive of response across these indications. Signatures described previously for immune checkpoint inhibitors also failed to validate. At the pathway level, significant heterogeneity is observed between indications, in particular within the PD-L1 + tumors. mUC and NSCLC are molecularly aligned, with cell cycle and DNA damage repair genes associated with response in PD-L1- tumors. At the gene level, the CDK4/6 inhibitor CDKN2A is identified as a significant transcriptional correlate of response, highlighting the association of non-immune pathways to the outcome of checkpoint blockade. This cross-indication analysis reveals molecular heterogeneity between mUC, NSCLC and RCC tumors, suggesting that indication-specific molecular approaches should be prioritized to formulate treatment strategies.

Introduction: Lung adenocarcinomas in young patients (<40 y) are more likely to harbor targetable genomic alterations. This study aimed to determine whether the prevalence of targetable alterations is greater in young adults with lung carcinoma than in the overall lung cancer population. To reach this rare patient population, a web-based platform was used to recruit and enroll patients remotely., Methods: In this prospective study, patients less than 40 years old at the time of primary lung cancer diagnosis with confirmed lung carcinoma were recruited from four global sites and remotely by means of a website. Genotyping data were collected, if available, or obtained by means of next-generation sequencing using the FoundationOne platform. The prevalence of targetable alterations was quantified across patients with advanced adenocarcinoma., Results: Overall, 133 patients across five continents were included, 41% of whom enrolled online. The mean (SD) age at diagnosis was 34 (5.2) years; 79% had stage IV disease at diagnosis. Among patients with adenocarcinoma (n = 115), 112 entered the study with previous genomic testing results and 86 (77%) had targetable alterations in EGFR , ALK , ROS1 , MET , ERBB2 , or RET . Among those without targetable alterations, 14 received further testing and a targetable alteration was identified in eight (57%)., Conclusions: This study revealed the feasibility of using a web-based platform to recruit young patients with lung cancer and revealed that 94 of 112 (84%) with adenocarcinoma at any stage had targetable genomic alterations. Among patients with stage IV adenocarcinoma, 85% had a targetable alteration, which is higher than historical expectations for the general population., (© 2021 The Authors.)

Based on the approvals of crizotinib and entrectinib by the Food and Drug Administration for the treatment of ROS1 positive nonsmall cell lung cancer (NSCLC), we sought to examine the mutational profile of a variety of solid tumors (excluding sarcomas) with ROS1 fusions that underwent comprehensive genomic profiling. A review of our database was performed to extract all nonsarcoma patients with ROS1 fusions that were discovered by the hybrid capture-based DNA only sequencing assays. We examined the coalterations representing potentially targetable biomarkers, resistance alterations and other alterations in these cases. In addition, we examined the histologic characteristics and protein expression with immunohistochemistry (IHC). From a series of clinically advanced nonsarcoma solid tumors, 356 unique cases with ROS1 fusions included 275 (77.2%) NSCLC and 81 (22.8%) non-NSCLC. Ten novel ROS1 fusions were discovered. Importantly, the NSCLC ROS1 fusion pos tumors had a higher PD-L1 IHC expression positivity when compared to the NSCLC ROS1 fusion neg population (P = .012, Chi-squared). The frequency of known and likely anti-ROS1 targeted therapy resistance genomic alterations in NSCLC was 7.3% (20/275) and in non-NSCLC was 4.9% (4/81). Overall, the coalteration profile of ROS1 fusion pos NSCLC and non-NSCLC was similar with only three genes altered significantly more frequently in non-NSCLC vs NSCLC: TERT, PTEN, APC. In our study, we characterized a large cohort of ROS1 fusion pos NSCLC and non-NSCLC solid tumors and discovered 10 novel ROS1 fusions., (© 2020 UICC.)

RAF family protein kinases signal through the MAPK pathway to orchestrate cellular proliferation, survival, and transformation. Identifying BRAF alterations in pediatric cancers is critically important as therapeutic agents targeting BRAF or MEK may be incorporated into the clinical management of these patients. In this study, we performed comprehensive genomic profiling on 3,633 pediatric cancer samples and identified a cohort of 221 (6.1%) cases with known or novel alterations in BRAF or RAF1 detected in extracranial solid tumors, brain tumors, or hematological malignancies. Eighty percent (176/221) of these tumors had a known-activating short variant (98, 55.7%), fusion (72, 40.9%), or insertion/deletion (6, 3.4%). Among BRAF altered cancers, the most common tumor types were brain tumors (74.4%), solid tumors (10.8%), hematological malignancies (9.1%), sarcomas (3.4%), and extracranial embryonal tumors (2.3%). RAF1 fusions containing intact RAF1 kinase domain (encoded by exons 10-17) were identified in seven tumors, including two novel fusions TMF1-RAF1 and SOX6-RAF1. Additionally, we highlight a subset of patients with brain tumor with positive clinical response to BRAF inhibitors, demonstrating the rationale for incorporating precision medicine into pediatric oncology. IMPLICATIONS FOR PRACTICE: Precision medicine has not yet gained a strong foothold in pediatric cancers. This study describes the landscape of BRAF and RAF1 genomic alterations across a diverse spectrum of pediatric cancers, primarily brain tumors, but also encompassing melanoma, sarcoma, several types of hematologic malignancy, and others. Given the availability of multiple U.S. Food and Drug Administration-approved BRAF inhibitors, identification of these alterations may assist with treatment decision making, as described here in three cases of pediatric cancer., (© AlphaMed Press 2020.)

A high tumour mutational burden (hypermutation) is observed in some gliomas 1-5 ; however, the mechanisms by which hypermutation develops and whether it predicts the response to immunotherapy are poorly understood. Here we comprehensively analyse the molecular determinants of mutational burden and signatures in 10,294 gliomas. We delineate two main pathways to hypermutation: a de novo pathway associated with constitutional defects in DNA polymerase and mismatch repair (MMR) genes, and a more common post-treatment pathway, associated with acquired resistance driven by MMR defects in chemotherapy-sensitive gliomas that recur after treatment with the chemotherapy drug temozolomide. Experimentally, the mutational signature of post-treatment hypermutated gliomas was recapitulated by temozolomide-induced damage in cells with MMR deficiency. MMR-deficient gliomas were characterized by a lack of prominent T cell infiltrates, extensive intratumoral heterogeneity, poor patient survival and a low rate of response to PD-1 blockade. Moreover, although bulk analyses did not detect microsatellite instability in MMR-deficient gliomas, single-cell whole-genome sequencing analysis of post-treatment hypermutated glioma cells identified microsatellite mutations. These results show that chemotherapy can drive the acquisition of hypermutated populations without promoting a response to PD-1 blockade and supports the diagnostic use of mutational burden and signatures in cancer.

Purpose: BRCA1 or BRCA2 loss of function results in homologous recombination deficiency (HRD), which is targetable by poly (ADP-ribose) polymerase (PARP) inhibitors and other DNA-damaging agents. In cancers associated with germline BRCA1/2 alterations ( BRCA1/2 -associated cancers: breast, ovarian, pancreatic, prostate), BRCA1/2 alterations result in HRD and are biomarkers for PARP inhibitor use. In other (non- BRCA1/2 -associated) cancer types, the association between BRCA1/2 alteration and HRD is less clear., Methods: A total of 234,154 tumor samples were sequenced by hybrid capture-based comprehensive genomic profiling. Somatic, germline, and zygosity status was determined computationally. BRCA1/2 alterations were classified as predicted germline/somatic and biallelic/monoallelic. Genome-wide loss of heterozygosity (gLOH) was evaluated as a marker of HRD., Results: BRCA1/2 alterations were observed at a 4.7% frequency. BRCA1/2 mutations were predicted germline in 57.4% of BRCA1/2 -associated and 37.2% of non- BRCA1/2 -associated cancers. The fraction of BRCA1/2 -altered cases that were biallelic was 68.7%, with a higher biallelic fraction in BRCA1/2 -associated (89.9%) versus non- BRCA1/2 -associated cancers (43.6%). Differences in tissue distribution of biallelic BRCA1 versus BRCA2 alterations were noted, including a higher rate of biallelic BRCA2 alteration in prostate cancer. Biallelic BRCA1/2 alteration was observed at a 3.2% frequency ( BRCA1/2 -associated cancers, 8.9%; non- BRCA1/2 -associated cancers, 1.3%) and > 1% frequency in at least 13 cancer types. Across cancer types, biallelic BRCA1/2 alteration was associated with increased gLOH versus monoallelic or wild-type BRCA1/2 ; predicted germline or somatic mutations were both associated with elevated gLOH., Conclusion: Biallelic BRCA1/2 alterations were associated with elevated gLOH in diverse cancer types, including those not traditionally associated with BRCA1/2 cancer syndromes. Biomarker development for PARP inhibitors should integrate methods to distinguish biallelic from monoallelic BRCA1/2 status, and biallelic BRCA1/2 alteration should be broadly evaluated across cancer types as a biomarker for underlying HRD and PARP inhibitor sensitivity., Competing Interests: AUTHORS’ DISCLOSURES OF POTENTIAL CONFLICTS OF INTEREST The following represents disclosure information provided by authors of this manuscript. All relationships are considered compensated unless otherwise noted. Relationships are self-held unless noted. I = Immediate Family Member, Inst = My Institution. Relationships may not relate to the subject matter of this manuscript. For more information about ASCO’s conflict of interest policy, please refer to www.asco.org/rwc or ascopubs.org/po/author-center. Open Payments is a public database containing information reported by companies about payments made to US-licensed physicians (Open Payments). We demonstrate that biallelic BRCA1/2 alterations are associated with elevated gLOH across all cancer types evaluated and may therefore represent a therapeutic vulnerability targetable by PARPi. Biomarker development for PARPi should reliably distinguish biallelic/monoallelic BRCA1/2 status, and biallelic BRCA1/2 alteration should be broadly evaluated as a pan-cancer biomarker in PARPi clinical trials. Ethan S. Sokol Employment: Foundation Medicine Stock and Other Ownership Interests: Roche Dean Pavlick Stock and Other Ownership Interests: Roche Garrett M. Frampton Employment: Foundation Medicine Stock and Other Ownership Interests: Roche Jeffrey S. Ross Employment: Foundation Medicine Leadership: Foundation Medicine Stock and Other Ownership Interests: Foundation Medicine Research Funding: Foundation Medicine Jeffrey P. Gregg Consulting or Advisory Role: AstraZeneca, Bristol-Myers Squibb, Roche, Foundation Medicine, Luminex Speakers’ Bureau: AstraZeneca, Foundation Medicine, Bristol-Myers Squibb Primo N. Lara Consulting or Advisory Role: Janssen Pharmaceuticals, Merck, CellMax Life, Pfizer Research Funding: Aragon Pharmaceuticals (Inst), Janssen Biotech (Inst), TRACON Pharma (Inst), Merck (Inst), Pharmacyclics (Inst), Incyte (Inst), Taiho Pharmaceutical (Inst) Steffi Oesterreich Stock and Other Ownership Interests: Ocean Genomics (I) Research Funding: GE, Blueprint Medicine, H3 Biomedicine, AstraZeneca, Illumina Neeraj Agarwal Consulting or Advisory Role: Pfizer, Medivation, Astellas Pharma, Bristol-Myers Squibb, AstraZeneca, Nektar, Eli Lilly, Bayer AG, Foundation Medicine, Pharmacyclics, Exelixis, Janssen Oncology, Merck, Novartis Research Funding: Bayer AG (Inst), Bristol-Myers Squibb (Inst), GlaxoSmithKline (Inst), Takeda Pharmaceuticals (Inst), Novartis (Inst), Pfizer (Inst), BN ImmunoTherapeutics (Inst), Exelixis (Inst), TRACON Pharma (Inst), Rexahn Pharmaceuticals (Inst), Amgen (Inst), AstraZeneca (Inst), Active Biotech (Inst), Bavarian Nordic (Inst), Calithera Biosciences (Inst), Celldex (Inst),Eisai (Inst), Genentech (Inst), Immunomedics (Inst), Janssen Pharmaceuticals (Inst), Merck (Inst), NewLink Genetics (Inst), Prometheus (Inst), Sanofi (Inst) Andrea Necchi Employment: Bayer AG (I) Stock and Other Ownership Interests: Bayer AG (I) Honoraria: Roche, Merck, AstraZeneca, Janssen Pharmaceuticals, Foundation Medicine, Bristol-Myers Squibb Consulting or Advisory Role: Merck Sharp & Dohme, Roche, Bayer AG, AstraZeneca, Clovis Oncology, Janssen Pharmaceuticals, Incyte, Seattle Genetics, Astellas Pharma, Bristol-Myers Squibb, Rainier Therapeutics, GlaxoSmithKline, Ferring Research Funding: Merck Sharp & Dohme (Inst), AstraZeneca (Inst), Ipsen (Inst) Travel, Accommodations, Expenses: Roche, Merck Sharp & Dohme, AstraZeneca, Janssen Pharmaceuticals, Rainier Therapeutics Other Relationship: Bayer AG (I) Vincent A. Miller Employment: Foundation Medicine Leadership: Foundation Medicine, Revolution Medicines Stock and Other Ownership Interests: Foundation Medicine, Mirati Therapeutics, Revolution Medicines Patents, Royalties, Other Intellectual Property: Periodic royalties related to T790M patent awarded to Memorial Sloan Kettering Cancer Center Brian Alexander Employment: Foundation Medicine Leadership: Foundation Medicine Stock and Other Ownership Interests: Roche Research Funding: Eli Lilly (Inst), Puma (Inst), Celgene (Inst) Open Payments Link: https://openpaymentsdata.cms.gov/physician/854258/summary Siraj M. Ali Employment: Foundation Medicine, EQRX, EQRX (I) Leadership: Incysus Stock and Other Ownership Interests: Exelixis, Blueprint Medicines, Agios, Genocea Biosciences Consulting or Advisory Role: Revolution Medicines, Azitra (I), Princepx Tx (I) Patents, Royalties, Other Intellectual Property: Patents through Foundation Medicine, patents through Seres Health on microbiome stuff in non-neoplastic disease (I) Shridar Ganesan Employment: Merck (I) Stock and Other Ownership Interests: Ibris, Inspirata, Merck (I) Consulting or Advisory Role: Inspirata, Novartis, Roche, Foghorn Therapeutics, Foundation Medicine Patents, Royalties, Other Intellectual Property: Two patents for digital imaging that may be licensed to Ibris and Inspirata Travel, Accommodations, Expenses: Inspirata Jon H. Chung Employment: Foundation Medicine Stock and Other Ownership Interests: Foundation Medicine No other potential conflicts of interest were reported.

Background: CDK12 loss-of-function (LOF) genomic alterations are associated with focal tandem duplications (FTDs) in ovarian and prostate cancers. Because these FTDs may produce fusion-induced neoantigens (FINAs), CDK12 alteration is a candidate biomarker for immune checkpoint inhibitor sensitivity. Here we determine the prevalence of CDK12- LOF alterations and their association with FTDs across diverse tumor types., Materials and Methods: A total of 142,133 tumor samples comprising 379 cancer types were sequenced (August 2014 to April 2018) by hybrid capture-based comprehensive genomic profiling (Foundation Medicine, Cambridge, MA) as part of routine clinical care. Results were analyzed for base substitutions, short insertions/deletions, rearrangements, and copy number alterations. CDK12- LOF genomic alterations were assessed for zygosity status and association with FTDs/focal copy number gain., Results: CDK12 genomic alterations were detected in 1.1% of all cases, most frequently in prostate cancer (5.6%), but were also observed at >1% frequency in 11 cancer types. Across multiple cancer types, including prostate, gastric/esophageal, ovarian, breast, and endometrial cancer, the number of FTDs was significantly increased in CDK12- LOF versus CDK12 wild-type cases. Notably, CDK12 -LOF was not consistently associated with a homologous recombination deficiency genomic signature. Quantitative assessment of CDK12- associated FTDs by measurement of single copy number gains identified novel likely deleterious CDK12 kinase-domain mutations in prostate and ovarian cancers., Conclusion: Detection of CDK12- LOF genomic alterations and their association with FTDs in a diverse spectrum of malignancies suggests that immunotherapy approaches targeting FINAs derived from CDK12- associated FTDs may be a broadly applicable strategy that could be explored across cancer types in a tumor-agnostic manner., Implications for Practice: CDK12 inactivation in ovarian and prostate cancer results in the generation of focal tandem duplications, which can cause fusion-induced neoantigens. In prostate cancer, CDK12 alterations have demonstrated promise as a potential predictive biomarker for response to immune checkpoint blockade. This study evaluated genomic profiling data from >142,000 tumors to determine the prevalence of CDK12 loss-of-function genomic alterations across tumor types and demonstrated that CDK12 alterations are associated with the tandem-duplicator phenotype in cancer types other than ovarian and prostate cancer. The association of CDK12 alterations with focal tandem duplications across broad cancer types suggests that CDK12 inactivation warrants further investigation as a pan-cancer biomarker for immunotherapy benefit., Competing Interests: Disclosures of potential conflicts of interest may be found at the end of this article., (© AlphaMed Press 2019.)

Infantile myofibromatosis (IM) is an aggressive neoplasm composed of myofibroblast-like cells in children. Although typically localized, it can also present as multifocal disease, which represents a challenge for effective treatment. IM has previously been linked to activating somatic and germline point mutations in the PDGFRβ tyrosine kinase encoded by the PDGFRB gene. Clinical panel-based targeted tumor sequencing of a tumor from a newborn with multifocal IM revealed a novel PDGFRB rearrangement, which was reported as being of unclear significance. Additional sequencing of cDNA from tumor and germline DNA confirmed a complex somatic/mosaic PDGFRB rearrangement with an apparent partial tandem duplication disrupting the juxtamembrane domain. Ectopic expression of cDNA encoding the mutant form of PDGFRB markedly enhanced cell proliferation of mouse embryo fibroblasts (MEFs) compared to wild-type PDGFRB and conferred tumor-forming capacity on nontumorigenic 10T1/2 fibroblasts. The mutated protein enhanced MAPK activation and retained sensitivity to the PDGFRβ inhibitor imatinib. Our findings reveal a new mechanism by which PDGFRB can be activated in IM, suggest that therapy with tyrosine kinase inhibitors including imatinib may be beneficial, and raise the possibility that this receptor tyrosine kinase might be altered in a similar fashion in additional cases that would similarly present annotation challenges in clinical DNA sequencing analysis pipelines., (© 2019 Hassan et al.; Published by Cold Spring Harbor Laboratory Press.)

Background: Liquid biopsy offers the ability to non-invasively analyze the genome of a tumor through circulating tumor DNA (ctDNA) to identify targetable and prognostic genomic alterations. Few studies have rigorously analyzed ctDNA results and determined the fidelity with which they recapitulate the genomics of a sequenced tissue sample obtained from the same tumor. The clinical utility study (CUS) for the FoundationACT™ ctDNA assay (Foundation Medicine, Cambridge, MA, USA; NCT02620527) is a multi-center prospective clinical study for multiple solid tumor types to compare genomic profiling of paired tissue and blood samples from the same patient. In this subset of the study, paired specimens from 96 patients with colorectal cancer (CRC) were analyzed with comprehensive genomic profiling (CGP) of the tumor tissue sample (FoundationOne ® ) and blood sample (FoundationACT™)., Methods: Both samples underwent CGP using the hybrid capture-based Illumina Hi-Seq technology. Maximum somatic allele frequency (MSAF) was used to estimate the fraction of ctDNA in the sample. The set of genes and targeted regions common to both tumor and liquid were compared for each subject., Results: Among these patients, 61% were male; 74% had clinical stage IV disease, 19% had clinical stage III disease, and 7% had clinical stage II disease. Time between the tissue biopsy and liquid biopsy (range, 0 - 709 days) had a significant impact on the positive percent agreement (PPA) between the two assays. Eighty percent of cases had evidence of ctDNA in the blood (MSAF >0). For all cases with MSAF >0, 171 base substitutions and insertions/deletions (indels) were identified in the tumor, and 79% (PPA) of these identical alterations were also identified in matched ctDNA samples; PPA increased to 87% for cases <270 days between the tissue and liquid biopsy, 95% for <90 days, and 100% PPA for <30 days. All known and likely short variants in KRAS, NRAS , and BRAF were analyzed independently as testing of these genes is recommended by the National Comprehensive Cancer Network (NCCN) for patients with CRC and have therapeutic implications. For NCCN genes, PPA was 80% for all time points for short variants; PPA increased to 90% for cases <270 days between the tissue and liquid biopsy. There was high concordance for KRAS G12X between tissue and liquid: overall percent agreement (97%), PPA (93%), negative percent agreement (NPA) (100%), positive predictive value (PPV) (100%), and negative predictive value (NPV) (96%) for the <270 day cohort., Conclusions: In cases where tumor tissue profiling is not possible, these results provide compelling evidence that genomic profiling of ctDNA in late stage CRC shows a high concordance with tumor tissue sequencing results and can be used to identify most clinically relevant alterations capable of guiding therapy for these patients., Competing Interests: Conflicts of Interest: G Li, D Pavlick, JH Chung, J He, M Cooke, J Hughes, SM Ali, M Nahas, VA Miller, P Stephens, B Forcier, JS Ross, AB Schrock are employees of Foundation Medicine, Inc. JP Gregg is a consultant/advisor and external speaker for FMI and on Speaker Bureau for AstraZeneca and BMS. The other authors have no conflicts of interest to declare., (2019 Journal of Gastrointestinal Oncology. All rights reserved.)

Introduction: Genomic profiling informs selection of matched targeted therapies as part of routine clinical care in NSCLC. Tissue biopsy is the criterion standard; however, genomic profiling of blood-derived circulating tumor DNA (ctDNA) has emerged as a minimally invasive alternative., Methods: Hybrid capture-based genomic profiling of 62 genes was performed on blood-based ctDNA from 1552 patients with NSCLC., Results: Evidence of ctDNA was detected in 80% of samples, and in 86% of these cases, at least one reportable genomic alteration (GA) was detected. Frequently altered genes were tumor protein p53 gene (TP53) (59%), EGFR (25%), and KRAS (17%). Comparative analysis with a tissue genomic database (N = 21,500) showed similar frequencies of GAs per gene, although KRAS mutation and EGFR T790M were more frequent in tissue and ctDNA, respectively (both p < 0.0001), likely reflecting the use of liquid versus tissue biopsy after relapse during targeted therapy. In temporally matched ctDNA and tissue samples from 33 patients with evidence of ctDNA in their blood, 64% of GAs detected in tissue were also detected in ctDNA, including 78% of short variants (58 of 74) and 100% of rearrangements (four of four), but only 16% of amplifications (four of 25)., Conclusions: Genomic profiling of ctDNA detected clinically relevant GAs in a significant subset of NSCLC cases. Most alterations detected in matched tissue were also detected in ctDNA. These results suggest the utility of ctDNA testing in advanced NSCLC as a complementary approach to tissue testing. Blood-based ctDNA testing may be particularly useful at the time of progression during targeted therapy., (Copyright © 2018 International Association for the Study of Lung Cancer. Published by Elsevier Inc. All rights reserved.)

Background: This retrospective study was undertaken to determine if the plasma circulating tumor DNA (ctDNA) level and tumor biological features in patients with advanced solid tumors affected the detection of genomic alterations (GAs) by a plasma ctDNA assay., Method: Cell-free DNA (cfDNA) extracted from frozen plasma (N = 35) or fresh whole blood (N = 90) samples were subjected to a 62-gene hybrid capture-based next-generation sequencing assay FoundationACT. Concordance was analyzed for 51 matched FoundationACT and FoundationOne (tissue) cases. The maximum somatic allele frequency (MSAF) was used to estimate the amount of tumor fraction of cfDNA in each sample. The detection of GAs was correlated with the amount of cfDNA, MSAF, total tumor anatomic burden (dimensional sum), and total tumor metabolic burden (SUVmax sum) of the largest ten tumor lesions on PET/CT scans., Results: FoundationACT detected GAs in 69 of 81 (85%) cases with MSAF > 0. Forty-two of 51 (82%) cases had ≥ 1 concordance GAs matched with FoundationOne, and 22 (52%) matched to the National Comprehensive Cancer Network (NCCN)-recommended molecular targets. FoundationACT also detected 8 unique molecular targets, which changed the therapy in 7 (88%) patients who did not have tumor rebiopsy or sufficient tumor DNA for genomic profiling assay. In all samples (N = 81), GAs were detected in plasma cfDNA from cancer patients with high MSAF quantity (P = 0.0006) or high tumor metabolic burden (P = 0.0006) regardless of cfDNA quantity (P = 0.2362)., Conclusion: This study supports the utility of using plasma-based genomic assays in cancer patients with high plasma MSAF level or high tumor metabolic burden.

Genomic profiling of circulating tumor DNA derived from cell-free DNA (cfDNA) in blood can provide a noninvasive method for detecting genomic biomarkers to guide clinical decision making for cancer patients. We developed a hybrid capture-based next-generation sequencing assay for genomic profiling of circulating tumor DNA from blood (FoundationACT). High-sequencing coverage and molecular barcode-based error detection enabled accurate detection of genomic alterations, including short variants (base substitutions, short insertions/deletions) and genomic re-arrangements at low allele frequencies (AFs), and copy number amplifications. Analytical validation was performed on 2666 reference alterations. The assay achieved >99% overall sensitivity (95% CI, 99.1%-99.4%) for short variants at AF >0.5%, >95% sensitivity (95% CI, 94.2%-95.7%) for AF 0.25% to 0.5%, and 70% sensitivity (95% CI, 68.2%-71.5%) for AF 0.125% to 0.25%. No false positives were detected in 62 samples from healthy volunteers. Genomic alterations detected by FoundationACT demonstrated high concordance with orthogonal assays run on the same clinical cfDNA samples. In 860 routine clinical FoundationACT cases, genomic alterations were detected in cfDNA at comparable frequencies to tissue; for the subset of cases with temporally matched tissue and blood samples, 75% of genomic alterations and 83% of short variant mutations detected in tissue were also detected in cfDNA. On the basis of analytical validation results, FoundationACT has been approved for use in our Clinical Laboratory Improvement Amendments-certified/College of American Pathologists-accredited/New York State-approved laboratory., (Copyright © 2018 American Society for Investigative Pathology and the Association for Molecular Pathology. Published by Elsevier Inc. All rights reserved.)

Rearrangements involving the neurotrophic receptor kinase genes (NTRK1, NTRK2, and NTRK3; hereafter referred to as TRK) produce oncogenic fusions in a wide variety of cancers in adults and children. Although TRK fusions occur in fewer than 1% of all solid tumors, inhibition of TRK results in profound therapeutic responses, resulting in Breakthrough Therapy FDA approval of the TRK inhibitor larotrectinib for adult and pediatric patients with solid tumors, regardless of histology. In contrast to solid tumors, the frequency of TRK fusions and the clinical effects of targeting TRK in hematologic malignancies are unknown. Here, through an evaluation for TRK fusions across more than 7,000 patients with hematologic malignancies, we identified TRK fusions in acute lymphoblastic leukemia (ALL), acute myeloid leukemia (AML), histiocytosis, multiple myeloma, and dendritic cell neoplasms. Although TRK fusions occurred in only 0.1% of patients (8 of 7,311 patients), they conferred responsiveness to TRK inhibition in vitro and in vivo in a patient-derived xenograft and a corresponding AML patient with ETV6-NTRK2 fusion. These data identify that despite their individual rarity, collectively, TRK fusions are present in a wide variety of hematologic malignancies and predict clinically significant therapeutic responses to TRK inhibition.

Purpose: Dabrafenib and trametinib are approved for the management of advanced non-small-cell lung cancers (NSCLCs) that harbor BRAF V600E mutations. Small series and pan-cancer analyses have identified non-V600 alterations as therapeutic targets. We sought to examine a large genomic data set to comprehensively characterize non-V600 B RAF alterations in lung cancer., Patients and Methods: A total of 23,396 patients with lung cancer provided data to assay with comprehensive genomic profiling. Data were reviewed for predicted pathogenic BRAF base substitutions, short insertions and deletions, copy number changes, and rearrangements., Results: Adenocarcinomas represented 65% of the occurrences; NSCLC not otherwise specified (NOS), 15%; squamous cell carcinoma, 12%; and small-cell lung carcinoma, 5%. BRAF was altered in 4.5% (1,048 of 23,396) of all tumors; 37.4% (n = 397) were BRAF V600E, 38% were BRAF non-V600E activating mutations, and 18% were BRAF inactivating. Rearrangements were observed at a frequency of 4.3% and consisted of N-terminal deletions (NTDs; 0.75%), kinase domain duplications (KDDs; 0.75%), and BRAF fusions (2.8%). The fusions involved three recurrent fusion partners: ARMC10 , DOCK4, and TRIM24 . BRAF V600E was associated with co-occurrence of SETD2 alterations, but other BRAF alterations were not and were instead associated with CDKN2A , TP53 , and STK11 alterations ( P < .05). Potential mechanisms of acquired resistance to BRAF V600E inhibition are demonstrated., Conclusion: This series characterized the frequent occurrence (4.4%) of BRAF alterations in lung cancers. Recurrent BRAF alterations in NSCLC adenocarcinoma are comparable to the frequency of other NSCLC oncogenic drivers, such as ALK , and exceed that of ROS1 or RET . This work supports a broad profiling approach in lung cancers and suggests that non-V600E BR AF alterations represent a subgroup of lung cancers in which targeted therapy should be considered., Competing Interests: The following represents disclosure information provided by authors of this manuscript. All relationships are considered compensated. Relationships are self-held unless noted. I = Immediate Family Member, Inst = My Institution. Relationships may not relate to the subject matter of this manuscript. For more information about ASCO's conflict of interest policy, please refer to www.asco.org/rwc or ascopubs.org/po/author-center. Yuri SheikineConsulting or Advisory Role: Foundation MedicineDean PavlickNo relationship to discloseSamuel KlempnerConsulting or Advisory Role: Lilly, Boston Biomedical, Celgene, Astellas Pharma Speakers' Bureau: Foundation Medicine Research Funding: Leap Therapeutics (Inst)Sally E. TrabuccoEmployment: Foundation Medicine Research Funding: Foundation Medicine Travel, Accommodations, Expenses: Foundation MedicineJon ChungEmployment: Foundation Medicine Stock and Other Ownership Interests: Foundation MedicineMark RosenzweigEmployment: Foundation Medicine Stock and Other Ownership Interests: Foundation MedicineKai WangNo relationship to discloseVamsidhar VelchetiHonoraria: Novartis, Foundation Medicine, Merck, Bristol-Myers Squibb, Genentech, Roche Consulting or Advisory Role: Clovis Oncology, Genentech, Bristol-Myers Squibb, Merck, Celgene, Foundation Medicine, AstraZeneca, MedImmune, Genoptix, Amgen, Takeda Research Funding: Genentech (Inst), Trovagene (Inst), Eisai (Inst), OncoPlex Diagnostics (Inst), ALkermes (Inst), NantOmics (Inst), GEnoptix (Inst), Altor BioScience (Inst), Merck (Inst), Bristol-Myers Squibb (Inst), Atreca (Inst), Heat Biologics (Inst), Leap Therapeutics (Inst) Travel, Accommodations, Expenses: AstraZeneca/MedImmune, Eisai, Foundation Medicine, MerckGarrett M. FramptonEmployment: Foundation Medicine Stock and Other Ownership Interests: Foundation MedicineNir PeledConsulting or Advisory Role: AstraZeneca, Boehringer Ingelheim, Bristol-Myers Squibb, Lilly, MSD, Novartis, Pfizer, Roche Speakers' Bureau: AstraZeneca, Boehringer Ingelheim, Bristol-Myers Squibb, Lilly, MSD, Novartis, Pfizer, RocheMolly MurrayEmployment: Foundation MedicineYoung Kwang ChaeConsulting or Advisory Role: Foundation Medicine, Boehringer Ingelheim, Biodesix, Counsyl, AstraZeneca, Guardant Health Speakers' Bureau: Merck, Genentech, Roche Travel, Accommodations, Expenses: HanmiLee A. AlbackerEmployment: Foundation Medicine Stock and Other Ownership Interests: Foundation MedicineLaurie GayEmployment: Foundation Medicine Stock and Other Ownership Interests: Gilead Sciences, Foundation MedicineHatim HusainConsulting or Advisory Role: AstraZeneca, Abbvie, Foundation Medicine Speakers' Bureau: AstraZeneca, Merck, Bristol-Myers Squibb Research Funding: Pfizer (Inst) Travel, Accommodations, Expenses: AstraZeneca, Merck, Bristol-Myers Squibb, Foundation Medicine, AbbvieJames SuhEmployment: Foundation Medicine Stock and Other Ownership Interests: Foundation MedicineSherri MillisEmployment: Foundation MedicineVenkataprasanth ReddyEmployment: Foundation Medicine Stock and Other Ownership Interests: Foundation Medicine Travel, Accommodations, Expenses: Foundation MedicineJulia ElvinEmployment: Foundation Medicine Stock and Other Ownership Interests: Foundation MedicineRyan J. HartmaierEmployment: Foundation Medicine Stock and Other Ownership Interests: Foundation Medicine Patents, Royalties, Other Intellectual Property: Nfe2L2 exon 2 and/or exon 3 loss from work conducted at Foundation Medicine (provisional patent filed) (Inst)Afshin DowlatiConsulting or Advisory Role: Abbvie, Stemcentrx, Ariad Research Funding: Lilly (Inst), ImClone (Inst), Amgen (Inst), Bristol-Myers Squibb (Inst), OncoMed (Inst), EMD Serono (Inst), MedImmune (Inst)Phil StephensEmployment: Foundation Medicine Leadership: Foundation Medicine Stock and Other Ownership Interests: Foundation MedicineJeffrey RossEmployment: Foundation Medicine Leadership: Foundation Medicine Stock and Other Ownership Interests: Foundation Medicine Honoraria: Pfizer, EMD Merck Serono Research Funding: Foundation MedicineTrever BivonaStock and Other Ownership Interests: Driver Honoraria: Novartis, AstraZeneca, Array BioPharma, Revolution Medicine Consulting or Advisory Role: Revolution Medicine, Array BioPharma, Novartis, AstraZeneca Speakers' Bureau: Novartis Research Funding: Ignyta, Revolution MedicineVincent MillerEmployment: Foundation Medicine Leadership: Foundation Medicine Stock and Other Ownership Interests: Foundation Medicine Patents, Royalties, Other Intellectual Property: Receive periodic royalties related to T790M patent awarded to Memorial Sloan Kettering Cancer CenterShridar GanesanEmployment: Merck (I) Stock and Other Ownership Interests: Ibris, Inspirata, Merck (I) Consulting or Advisory Role: Inspirata, Novartis Patents, Royalties, Other Intellectual Property: I hold two patents for digital imaging that may be licensed to Ibris and Inspirata Travel, Accommodations, Expenses: InspirataAlexa SchrockEmployment: Foundation Medicine Stock and Other Ownership Interests: Foundation MedicineSai-Hong OuHonoraria: Pfizer, Roche Pharma AG, Genentech, Roche, ARIAD, Takeda, Novartis, AstraZeneca, Foundation Medicine Consulting or Advisory Role: Pfizer, Roche, Genentech, Novartis, AstraZeneca, Takeda, ARIAD, Foundation Medicine Speakers' Bureau: Genentech, AstraZeneca, Takeda, ARIAD Research Funding: Pfizer (Inst), Roche Pharma AG (Inst), AstraZeneca (Inst), MedImmune (Inst), Clovis Oncology (Inst), ARIAD (Inst), Ignyta (Inst), Peregrine Pharmaceuticals (Inst), GlaxoSmithKline (Inst), Astellas Pharma (Inst), Chugai Pharma (Inst)Siraj AliEmployment: Foundation Medicine Stock and Other Ownership Interests: Exelixis, Blueprint Medicines, Agios Patents, Royalties, Other Intellectual Property: Patents via Foundation Medicine; patents via Seres Health on microbiomes in non-neoplastic disease (I), (© 2018 by American Society of Clinical Oncology.)