Measurement of ctDNA Tumor Fraction Identifies Informative Negative Liquid Biopsy Results and Informs Value of Tissue Confirmation.

PURPOSE: Liquid biopsy (LBx) for tumor profiling is increasingly used, but concerns remain regarding negative results. A lack of results may truly reflect tumor genomics, or it may be a false negative that would be clarified by tissue testing. A method of distinguishing between these scenarios could help clarify when follow-on tissue testing is valuable. EXPERIMENTAL DESIGN: Here we evaluate circulating tumor DNA (ctDNA) tumor fraction (TF), a quantification of ctDNA in LBx samples, for utility in identifying true negative results.We assessed concordance between LBx and tissue-based results, stratified by ctDNA TF, in a real-world genomic data set of paired samples across multiple disease types. We also evaluated the frequency of tissue results identifying driver alterations in lung cancer patients after negative LBx in a real-world clinicogenomic database. RESULTS: The positive percent agreement and negative predictive value between liquid and tissue samples for driver alterations increased from 63% and 66% for all samples to 98% and 97% in samples with ctDNA TF ≥1%. Among 505 lung cancer patients with no targetable driver alterations found by LBx who had subsequent tissue-based profiling, 37% had a driver, all of which had ctDNA TF <1%. CONCLUSIONS: Lung cancer patients with negative LBx and ctDNA TF ≥1% are unlikely to have a driver detected on confirmatory tissue testing; such informative negative results may benefit instead from prompt treatment initiation. Conversely, negative LBx with ctDNA TF <1% will commonly have a driver identified by follow-on tissue testing and should be prioritized for reflex testing.

Association of Timely Comprehensive Genomic Profiling With Precision Oncology Treatment Use and Patient Outcomes in Advanced Non-Small-Cell Lung Cancer.

PURPOSE: Timely biomarker testing remains out of reach for many patients with advanced non-small-cell lung cancer (aNSCLC). Here, we studied the quality-of-care implications of closing the gap in timely receipt of comprehensive genomic profiling (CGP) to inform first-line (1L) decisions. METHODS: Using a real-world clinicogenomic database, we studied testing and 1L treatment patterns in aNSCLC after the approval of pembrolizumab in combination with pemetrexed and carboplatin (May 10, 2017). To estimate the association of timely CGP results with therapy selection and patient outcomes, we identified patients with no previous genomic testing beyond PD-L1 immunohistochemistry and dichotomized patients by whether CGP results were available before or after 1L therapy initiation. RESULTS: In total, 2,694 patients were included in the 1L therapy decision impact assessment. Timely CGP increased matched targeted therapy use by 14 percentage points (17% with CGP v 2.8% without) and precision immune checkpoint inhibitor (ICPI) use by 14 percentage points (18% with CGP v 3.9% without). Receipt of timely CGP resulted in an estimated 31 percentage point decrease in ICPI use among ALK/EGFR/RET/ROS1-positive patients at an expected per-patient reduction in ineffective ICPI therapy cost of $13,659.37 with timely CGP to inform 1L treatment selection. Patient benefit of CGP extended to real-world time to therapy discontinuation (median time to therapy discontinuation: 3.9 v 10 months [hazard ratio, HR, 0.54 [95% CI, 0.42 to 0.70]; P = 1.9E-06; adjusted hazard ratio [aHR], 0.50 [95% CI, 0.38 to 0.67]; P = 2.0E-06) in 1L driver-positive patients. This effect was not significant for real-world overall survival (median overall survival: 32 v 29 months [HR, 1.2 [95% CI, 0.84 to 1.67]; P = .33; aHR, 1.4 [95% CI, 0.92 to 1.99]; P = .12). CONCLUSION: Timely CGP is associated with the quality of patient care as measured by 1L matched targeted therapy use, time to therapy discontinuation, and avoidance of ineffective, costly ICPIs.

Liquid biopsy-based circulating tumour (ct)DNA analysis of a spectrum of myeloid and lymphoid malignancies yields clinically actionable results.

AIMS: Liquid biopsy (LBx)-based next-generation sequencing (NGS) of circulating tumour DNA (ctDNA) can facilitate molecular profiling of haematopoietic neoplasms (HNs), particularly when tissue-based NGS is infeasible. METHODS AND RESULTS: We studied HN LBx samples tested with FoundationOne Liquid CDx, FoundationOne Liquid, or FoundationACT between July 2016 and March 2022. We identified 271 samples: 89 non-Hodgkin lymphoma (NHL), 43 plasma-cell neoplasm (PCN), 41 histiocytoses, 27 myelodysplastic syndrome (MDS), 25 diffuse large B-cell lymphoma (DLBCL), 22 myeloproliferative neoplasm (MPN), 14 Hodgkin lymphoma (HL), and 10 acute myeloid leukaemia (AML). Among 73.4% with detectable pathogenic alterations, median maximum somatic allele frequency (MSAF) was 16.6%, with AML (36.2%), MDS (19.7%), and MPN (44.5%) having higher MSAFs than DLBCL (3.9%), NHL (8.4%), HL (1.5%), PCN (2.8%), and histiocytoses (1.8%) (P = 0.001). LBx detected characteristic alterations across HNs, including in TP53, KRAS, MYD88, and BTK in NHLs; TP53, KRAS, NRAS, and BRAF in PCNs; IGH in DLBCL; TP53, ATM, and PDCD1LG2 in HL; BRAF and MAP2K1 in histiocytoses; TP53, SF3B1, DNMT3A, TET2, and ASXL1 in MDS; JAK2 in MPNs; and FLT3, IDH2, and NPM1 in AML. Among 24 samples, the positive percent agreement by LBx was 75.7% for variants present in paired buffy coat, marrow, or tissues. Also, 75.0% of pairs exhibited alterations only present on LBx. These were predominantly subclonal (clonal fraction of 3.8%), reflecting the analytical sensitivity of LBx. CONCLUSION: These data demonstrate that LBx can detect relevant genomic alterations across HNs, including at low clonal fractions, suggesting a potential clinical utility for identifying residual or emerging therapy-resistant clones that may be undetectable in site-specific tissue biopsies.

Real-World Clinical Performance of a DNA-Based Comprehensive Genomic Profiling Assay for Detecting Targetable Fusions in Nonsquamous NSCLC.

BACKGROUND: Genomic fusions are potent oncogenic drivers across cancer types and many are targetable. We demonstrate the clinical performance of DNA-based comprehensive genomic profiling (CGP) for detecting targetable fusions. MATERIALS AND METHODS: We analyzed targetable fusion genes in >450 000 tissue specimens profiled using DNA CGP (FoundationOne CDx, FoundationOne). Using a de-identified nationwide (US-based) non-small cell lung cancer (NSCLC) clinico-genomic database, we assessed outcomes in patients with nonsquamous NSCLC (NonSqNSCLC) who received matched therapy based on a fusion identified using DNA CGP. Lastly, we modeled the added value of RNA CGP for fusion detection in NonSqNSCLC. RESULTS: We observed a broad diversity of fusion partners detected with DNA CGP in conjunction with targetable fusion genes (ALK, BRAF, FGFR2, FGFR3, NTRK1/2/3, RET, and ROS1). In NonSqNSCLC with oncogenic ALK, NTRK, RET, and ROS1 fusions detected by DNA CGP, patients treated with a matched tyrosine kinase inhibitor had better real-world progression-free survival than those receiving alternative treatment regimens and benefit was observed regardless of the results of orthogonal fusion testing. An estimated 1.3% of patients with NonSqNSCLC were predicted to have an oncogenic driver fusion identified by RNA, but not DNA CGP, according to a model that accounts for multiple real-world factors. CONCLUSION: A well-designed DNA CGP assay is capable of robust fusion detection and these fusion calls are reliable for informing clinical decision-making. While DNA CGP detects most driver fusions, the clinical impact of fusion detection is substantial for individual patients and exhaustive efforts, inclusive of additional RNA-based testing, should be considered when an oncogenic driver is not clearly identified.

Liquid Biopsy of Lung Cancer Before Pathological Diagnosis Is Associated With Shorter Time to Treatment.

PURPOSE: Studies have investigated the early use of liquid biopsy (LBx) during the diagnostic workup of patients presenting with clinical evidence of advanced lung cancer, but real-world adoption and impact has not been characterized. The aim of this study was to determine whether the use of LBx before diagnosis (Dx; LBx-Dx) enables timely comprehensive genomic profiling (CGP) and shortens time until treatment initiation for advanced non-small-cell lung cancer (aNSCLC). MATERIALS AND METHODS: This study used the Flatiron Health-Foundation Medicine electronic health record-derived deidentified clinicogenomic database of patients with aNSCLC from approximately 280 US cancer clinics. RESULTS: Of 1,076 patients with LBx CGP ordered within 30 days prediagnosis/postdiagnosis, we focused on 56 (5.2%) patients who ordered LBx before diagnosis date (median 8 days between order and diagnosis, range, 1-28). Compared with 1,020 patients who ordered LBx after diagnosis (Dx-LBx), LBx-Dx patients had similar stage and ctDNA tumor fraction (TF). LBx-Dx patients received CGP results a median of 1 day after Dx versus 25 days for Dx-LBx patients. Forty-three percent of LBx-Dx were positive for an National Comprehensive Cancer Network driver, and 32% had ctDNA TF >1% but were driver negative (presumed true negatives). In 748 patients with previously untreated aNSCLC, median time from Dx to therapy was shorter in the LBx-Dx versus Dx-LBx group (21 v 35 days; P < .001). CONCLUSION: Early LBx in anticipation of pathologic diagnosis of aNSCLC was uncommon in this real-world cohort, yet this emerging paradigm was associated with an abbreviated time to CGP results and faster therapy initiation. Forthcoming prospective studies will clarify the utility of LBx in parallel with biopsy for diagnostic confirmation for patients presenting with suspected advanced lung cancer.

Circulating Tumor DNA Enables Sensitive Detection of Actionable Gene Fusions and Rearrangements Across Cancer Types.

PURPOSE: Genomic rearrangements can generate potent oncogenic drivers or disrupt tumor suppressor genes. This study examines the landscape of fusions and rearrangements detected by liquid biopsy (LBx) of circulating tumor DNA (ctDNA) across different cancer types. EXPERIMENTAL DESIGN: LBx from 53,842 patients with 66 solid tumor types were profiled using FoundationOneLiquid CDx, a hybrid-capture sequencing platform that queries 324 cancer-related genes. Tissue biopsies (TBx) profiled using FoundationOneCDx were used as a comparator. RESULTS: Among all LBx, 7,377 (14%) had ≥1 pathogenic rearrangement detected. A total of 3,648 (6.8%) LBx had ≥1 gain-of-function (GOF) oncogene rearrangement, and 4,428 (8.2%) LBx had ≥1 loss-of-function rearrangement detected. Cancer types with higher prevalence of GOF rearrangements included those with canonical fusion drivers: prostate cancer (19%), cholangiocarcinoma (6.4%), bladder (5.5%), and non-small cell lung cancer (4.4%). Although the prevalence of driver rearrangements was lower in LBx than TBx overall, the frequency of detection was comparable in LBx with a tumor fraction (TF) ≥1%. Rearrangements in FGFR2, BRAF, RET, and ALK, were detected across cancer types, but tended to be clonal variants in some cancer types and potential acquired resistance variants in others. CONCLUSIONS: In contrast to some prior literature, this study reports detection of a wide variety of rearrangements in ctDNA. The prevalence of driver rearrangements in tissue and LBx was comparable when TF ≥1%. LBx presents a viable alternative when TBx is not available, and there may be less value in confirmatory testing when TF is sufficient.

Real-World Genomic Profile of EGFR Second-Site Mutations and Other Osimertinib Resistance Mechanisms and Clinical Landscape of NSCLC Post-Osimertinib.

INTRODUCTION: The emergence of osimertinib as standard of care for EGFR-mutant NSCLC has renewed the need to understand and overcome drug resistance. We sought to understand the genomics and real-world treatment landscape of NSCLC with EGFR C797S and other on- and off-target resistance mechanisms. METHODS: Comprehensive genomic profiling (CGP) results from tissue or blood samples from 93,065 patients with NSCLC were queried for osimertinib EGFR second-site resistance mutations (ssEGFRms; C797, L718, G724, G796, L792). A real-world electronic health record-derived deidentified clinicogenomic database of patients with NSCLC undergoing CGP from approximately 280 U.S. cancer clinics was queried to assess post-osimertinib resistance and clinical treatment outcomes. RESULTS: A ssEGFRm was identified in 239 of 8845 (2.7%) EGFR-driven (L858R or exon 19 deletion) NSCLCs, most frequently C797 (71%), L718 (15%), and G724 (9.5%). ssEGFRms were not equally distributed across drivers; C797 and G724 changes strongly favored exon 19 deletion and L718, G796 and L792 favored L858R. Post-osimertinib CGP detected ssEGFRm in 19% of the cases (39 of 205); in paired pre-/post-osimertinib samples, on- and off-target resistance was largely mutually exclusive and observed in 24% and 27% of the cases, respectively. Of 391 patients with post-osimertinib treatment data, 62% received a chemotherapy-based regimen, whereas 25% received a targeted therapy or clinical study drug. Median real-world overall survival was 11.4 months from osimertinib progression. CONCLUSIONS: The osimertinib resistance landscape is diverse with on-target ssEGFRm and off-target resistance detected in tissue and liquid biopsy. Post-osimertinib, patients are receiving primarily chemotherapy-based regimens with poor outcomes, and CGP at resistance may offer an opportunity to inform therapeutic development and improve treatment selection.

Real-World comprehensive genomic profiling data for diagnostic clarity in pulmonary Large-Cell neuroendocrine carcinoma.

BACKGROUND: Pulmonary large-cell neuroendocrine carcinoma (LCNEC) is an uncommon subtype of lung cancer believed to represent a spectrum of tumors sharing characteristics of both small cell lung cancer (SCLC) and non-small cell lung cancer (NSCLC). Other groups have proposed genomic LCNEC subtypes, including small cell-like, non-small cell-like, and carcinoid-like subtypes. The primary goal of this study was to better define the NSCLC-like subtype with comprehensive genomic profiling (CGP). METHODS: An institutional database was queried to identify tissue specimens (TBx, N = 1,426) and liquid biopsies (LBx, N = 39) submitted for CGP during routine clinical care (8/2014 - 7/2023) with a disease ontology of LCNEC. TBx were profiled with FoundationOne® (F1) or F1CDx, using hybrid-capture technology to detect genomic alterations (GAs). RESULTS: 1,426 LCNEC samples were genomically profiled. The presence of RB1 and TP53 genomic alterations (GAs) were used to define a SCLC-like subtype (n = 557). A carcinoid-like group was defined by the presence of MEN1 mutation in the absence of TP53 GAs (n = 25). The remaining 844 samples were compared to the SCLC-like group and GAs enriched relative to the SCLC-like samples with a false discovery rate (FDR) < 0.0001 were used to define a NSCLC-like group. These NSCLC-like subtype-defining GAs included SMARCA4, KRAS, FGF3/4/19, STK11, CDKN2A/B, MTAP, and CCND1. Under this schema, 530 samples were classified as NSCLC-like and 314 remained unclassified. CONCLUSIONS: Large-scale CGP can better characterize biologically distinct molecular subtypes in LCNEC. Further studies to define how these molecular subtypes may help inform treatment decisions in this complex and challenging malignancy are warranted.

Evaluation of tissue- and plasma-derived tumor mutational burden (TMB) and genomic alterations of interest in CheckMate 848, a study of nivolumab combined with ipilimumab and nivolumab alone in patients with advanced or metastatic solid tumors with high TMB.

BACKGROUND: An accumulation of somatic mutations in tumors leads to increased neoantigen levels and antitumor immune response. Tumor mutational burden (TMB) reflects the rate of somatic mutations in the tumor genome, as determined from tumor tissue (tTMB) or blood (bTMB). While high tTMB is a biomarker of immune checkpoint inhibitor (ICI) treatment efficacy, few studies have explored the clinical utility of bTMB, a less invasive alternative for TMB assessment. Establishing the correlation between tTMB and bTMB would provide insight into whether bTMB is a potential substitute for tTMB. We explored the tumor genomes of patients enrolled in CheckMate 848 with measurable TMB. The correlation between tTMB and bTMB, and the factors affecting it, were evaluated. METHODS: In the phase 2 CheckMate 848 (NCT03668119) study, immuno-oncology-naïve patients with advanced, metastatic, or unresectable solid tumors and tTMB-high or bTMB-high (≥10 mut/Mb) were prospectively randomized 2:1 to receive nivolumab plus ipilimumab or nivolumab monotherapy. Tissue and plasma DNA sequencing was performed using the Foundation Medicine FoundationOne CDx and bTMB Clinical Trial Assays, respectively. tTMB was quantified from coding variants, insertions, and deletions, and bTMB from somatic base substitutions. Correlations between tTMB and bTMB were determined across samples and with respect to maximum somatic allele frequency (MSAF). Assay agreement and variant composition were also evaluated. RESULTS: A total of 1,438 and 1,720 unique tissue and blood samples, respectively, were obtained from 1,954 patients and included >100 screened disease ontologies, with 1,017 unique pairs of tTMB and bTMB measurements available for assessment. Median tTMB and bTMB were 3.8 and 3.5 mut/Mb, respectively. A significant correlation between tTMB and bTMB (r=0.48, p<0.0001) was observed across all sample pairs, which increased to r=0.54 (p<0.0001) for samples with MSAF≥1%. Assay concordance was highest for samples with MSAF≥10% across multiple disease ontologies and observed for both responders and non-responders to ICI therapy. The variants contributing to tTMB and bTMB were similar. CONCLUSIONS: We observed that tTMB and bTMB had a statistically significant correlation, particularly for samples with high MSAF, and that this correlation applied across disease ontologies. Further investigation into the clinical utility of bTMB is warranted.

Pan-Cancer Analysis of Copy-Number Features Identifies Recurrent Signatures and a Homologous Recombination Deficiency Biomarker to Predict Poly (ADP-Ribose) Polymerase Inhibitor Response.

PURPOSE: Copy-number (CN) features reveal the molecular state of cancers and may have predictive and prognostic value in the treatment of cancer. We sought to apply published CN analysis methods to a large pan-cancer data set and characterize ubiquitous CN signatures across tumor types, including potential utility for treatment selection. METHODS: We analyzed the landscape of CN features in 260,333 pan-cancer samples. We examined the association of 10 signatures with genomic alterations and clinical characteristics and trained a machine learning classifier using CN and insertion and deletion features to detect homologous recombination deficiency signature (HRDsig) positivity. Clinical outcomes were assessed using a real-world clinicogenomic database (CGDB) of comprehensive genomic profiling linked to deidentified, electronic health record-derived clinical data. RESULTS: CN signatures were prevalent across cancer types and associated with diverse processes including focal tandem duplications, seismic amplifications, genome-wide loss of heterozygosity (gLOH), and HRD. Our novel HRDsig outperformed gLOH in predicting BRCAness and effectively distinguished biallelic BRCA and homologous recombination-repair wild-type (HRRwt) samples pan-tumor, demonstrating high sensitivity to detect biallelic BRCA in ovarian (93%) and other HRD-associated cancers (80%-87%). Pan-tumor prevalence of HRDsig was 6.4%. HRRwt cases represented a significant fraction of the HRDsig-positive cohort, likely reflecting a population with nongenomic mechanisms of HRD. In ovarian and prostate CGDBs, HRDsig identified more patients than gLOH and had predictive value for poly (ADP-ribose) polymerase inhibitor (PARPi) benefit. CONCLUSION: Tumor CN profiles are informative, revealing diverse processes active in cancer. We describe the landscape of 10 CN signatures in a large pan-cancer cohort, including two associated with HRD. We trained a machine learning-based HRDsig that robustly identified BRCAness and associated with biallelic BRCA pan-tumor, and was predictive of PARPi benefit in real-world ovarian and prostate data sets.

Circulating Tumor DNA Monitoring on Chemo-immunotherapy for Risk Stratification in Advanced Non-Small Cell Lung Cancer.

PURPOSE: Chemoimmunotherapy (chemoIO) is a prevalent first-line treatment for advanced driver-negative non-small cell lung cancer (NSCLC), with maintenance therapy given after induction. However, there is significant clinical variability in the duration, dosing, and timing of maintenance therapy after induction chemoIO. We used circulating tumor DNA (ctDNA) monitoring to inform outcomes in patients with advanced NSCLC receiving chemoIO. EXPERIMENTAL DESIGN: This retrospective study included 221 patients from a phase III trial of atezolizumab+carboplatin+nab-paclitaxel versus carboplatin+nab-paclitaxel in squamous NSCLC (IMpower131). ctDNA monitoring used the FoundationOne Tracker involving comprehensive genomic profiling of pretreatment tumor tissue, variant selection using an algorithm to exclude nontumor variants, and multiplex PCR of up to 16 variants to detect and quantify ctDNA. RESULTS: ctDNA was detected (ctDNA+) in 96% of pretreatment samples (median, 93 mean tumor molecules/mL), and similar ctDNA dynamics were noted across treatment arms during chemoIO. ctDNA decrease from baseline to C4D1 was associated with improved outcomes across multiple cutoffs for patients treated with chemoIO. When including patients with missing plasma or ctDNA- at baseline, patients with ctDNA- at C4D1 (clearance), had more favorable progression-free survival (median 8.8 vs. 3.5 months; HR, 0.32;0.20-0.52) and OS (median not reached vs. 8.9 months; HR, 0.22; 0.12-0.39) from C4D1 than ctDNA+ patients. CONCLUSIONS: ctDNA monitoring during induction chemoIO can inform treatment outcomes in patients with advanced NSCLC. Importantly, monitoring remains feasible and informative for patients missing baseline ctDNA. ctDNA testing during induction chemoIO identifies patients at higher risk for disease progression and may inform patient selection for novel personalized maintenance or second-line treatment strategies.

Germline EGFR Mutations and Familial Lung Cancer.

PURPOSE: The genomic underpinnings of inherited lung cancer risk are poorly understood. This prospective study characterized the clinical phenotype of patients and families with germline EGFR pathogenic variants (PVs). METHODS: The Investigating Hereditary Risk from T790M study (ClinicalTrials.gov identifier: NCT01754025) enrolled patients with lung cancer whose tumor profiling harbored possible germline EGFR PVs and their relatives, either in person or remotely, providing germline testing and follow-up. RESULTS: A total of 141 participants were enrolled over a 5-year period, 100 (71%) remotely. Based upon previous genotyping, 116 participants from 59 kindreds were tested for EGFR T790M, demonstrating a pattern of Mendelian inheritance with variable lung cancer penetrance. In confirmed or obligate carriers of a germline EGFR PV from 39 different kindreds, 50/91 (55%) were affected with lung cancer with 34/65 (52%) diagnosed by age 60 years. Somatic testing of lung cancers in carriers revealed that 35 of 37 (95%) had an EGFR driver comutation. Among 36 germline carriers without a cancer diagnosis, 15 had computed tomography (CT) imaging and nine had lung nodules, including a 28-year-old with >10 lung nodules. Given geographic enrichment of germline EGFR T790M in the southeast United States, genome-wide haplotyping of 46 germline carriers was performed and identified a 4.1-Mb haplotype shared by 41 (89%), estimated to originate 223-279 years ago. CONCLUSION: To our knowledge, this is the first prospective description of familial EGFR-mutant lung cancer, identifying a recent founder germline EGFR T790M variant enriched in the Southeast United States. The high prevalence of EGFR-driver lung adenocarcinomas and lung nodules in germline carriers supports effort to identify affected patients and family members for investigation of CT-based screening for these high-risk individuals.

Potential pathogenic germline variant reporting from tumor comprehensive genomic profiling complements classic approaches to germline testing.

Existing guidance regarding clinically informed germline testing for patients with cancer is effective for evaluation of classic hereditary cancer syndromes and established gene/cancer type associations. However, current screening methods may miss patients with rare, reduced penetrance, or otherwise occult hereditary risk. Secondary finding of suspected germline variants that may confer inherited cancer risk via tumor comprehensive genomic profiling (CGP) has the potential to help address these limitations. However, reporting practices for secondary finding of germline variants are inconsistent, necessitating solutions for transparent and coherent communication of these potentially important findings. A workflow for improved confidence detection and clear reporting of potential pathogenic germline variants (PPGV) in select cancer susceptibility genes (CSG) was applied to a research dataset from real-world clinical tumor CGP of > 125,000 patients with advanced cancer. The presence and patterns of PPGVs identified across tumor types was assessed with a focus on scenarios in which traditional clinical germline evaluation may have been insufficient to capture genetic risk. PPGVs were identified in 9.7% of tumor CGP cases using tissue- and liquid-based assays across a broad range of cancer types, including in a number of "off-tumor" contexts. Overall, PPGVs were identified in a similar proportion of cancers with National Comprehensive Cancer Network (NCCN) recommendations for germline testing regardless of family history (11%) as in all other cancer types (9%). These findings suggest that tumor CGP can serve as a tool that is complementary to traditional germline genetic evaluation in helping to ascertain inherited susceptibility in patients with advanced cancer.

Unique Spectrum of Activating BRAF Alterations in Prostate Cancer.

PURPOSE: Alterations in BRAF have been reported in 3% to 5% of prostate cancer, although further characterization is lacking. Here, we describe the nature of BRAF alterations in prostate cancer using a large cohort from commercially available tissue and liquid biopsies subjected to comprehensive genomic profiling (CGP). EXPERIMENTAL DESIGN: Tissue and liquid biopsies from patients with prostate cancer were profiled using FoundationOne CDx and FoundationOne Liquid CDx CGP assays, respectively. Tissue biopsies from non-prostate cancer types were used for comparison (n = 275,151). Genetic ancestry was predicted using a single-nucleotide polymorphism (SNP) based approach. RESULTS: Among 15,864 tissue biopsies, BRAF-activating alterations were detected in 520 cases (3.3%). The majority (463 samples, 2.9%) harbored class II alterations, including BRAF rearrangements (243 samples, 1.5%), K601E (101 samples, 0.6%), and G469A (58 samples, 0.4%). BRAF-altered prostate cancers were enriched for CDK12 mutations (OR, 1.87; 9.2% vs. 5.2%; P = 0.018), but depleted in TMPRSS2 fusions (OR, 0.25; 11% vs. 32%; P < 0.0001), PTEN alterations (OR, 0.47; 17% vs. 31%; P < 0.0001), and APC alterations (OR, 0.48; 4.4% vs. 8.9%; P = 0.018) relative to BRAF wild-type (WT) disease. Compared with patients of European ancestry, BRAF alterations were more common in tumors from patients of African ancestry (5.1% vs. 2.9%, P < 0.0001) and Asian ancestry (6.0% vs. 2.9%, P < 0.001). CONCLUSIONS: Activating BRAF alterations were detected in approximately 3% of prostate cancers, and most were class II mutations and rearrangements; BRAF V600 mutations were exceedingly rare. These findings suggest that BRAF activation in prostate cancer is unique from other cancers and supports further clinical investigation of therapeutics targeting the mitogen-activated protein kinase (MAPK) pathway.

Tumor Mutational Burden in Real-World Patients With Pancreatic Cancer: Genomic Alterations and Predictive Value for Immune Checkpoint Inhibitor Effectiveness.

PURPOSE: Pancreatic ductal adenocarcinoma (PDAC) is largely considered a nonimmunogenic malignancy; however, approximately 1%, of patients may have tumors with deficient mismatch repair, high microsatellite instability, or high tumor mutational burden (TMB ≥10 mutations/Mb), which may be predictive of response to immune checkpoint inhibitor (ICI) therapy. We sought to analyze outcomes of patients with high-TMB and pathogenic genomic alterations observed in this population. METHODS: This study included patients with PDAC who underwent comprehensive genomic profiling (CGP) at Foundation Medicine (Cambridge, MA). Clinical data were obtained from a US-wide real-world clinicogenomic pancreatic database. We report genomic alterations in those with high and low TMB, and compare outcomes on the basis of receipt of single-agent ICI or therapy regimens not containing ICI. RESULTS: We evaluated 21,932 patients with PDAC who had tissue CGP data available, including 21,639 (98.7%) with low-TMB and 293 (1.3%) with high-TMB. Among patients with high-TMB, a greater number of alterations were observed in BRCA2, BRAF, PALB2, and genes of the mismatch repair pathway, whereas fewer alterations were observed in KRAS. Among patients who received an ICI (n = 51), those with high-TMB had more favorable median overall survival when compared with the low-TMB subset (25.7 v 5.2 months; hazard ratio, 0.32; 95% CI, 0.11 to 0.91; P = .034). CONCLUSION: Longer survival was observed in patients with high-TMB receiving ICI compared with those with low-TMB. This supports the role of high-TMB as a predictive biomarker for efficacy of ICI therapy in PDAC. Additionally, we report higher rates of BRAF and BRCA2 mutations and lower rates of KRAS mutation among patients with PDAC and high-TMB, which to our knowledge, is a novel finding.

Early Clearance of Plasma Epidermal Growth Factor Receptor Mutations as a Predictor of Outcome on Osimertinib in Advanced Non-Small Cell Lung Cancer; Exploratory Analysis from AURA3 and FLAURA.

PURPOSE: Plasma circulating tumor DNA (ctDNA) analysis is used for genotyping advanced non-small cell lung cancer (NSCLC); monitoring dynamic ctDNA changes may be used to predict outcomes. PATIENTS AND METHODS: This was a retrospective, exploratory analysis of two phase III trials [AURA3 (NCT02151981), FLAURA (NCT02296125)]. All patients had EGFR mutation-positive (EGFRm; ex19del or L858R) advanced NSCLC; AURA3 also included T790M-positive NSCLC. Osimertinib (FLAURA, AURA3), or comparator EGFR-tyrosine kinase inhibitor (EGFR-TKI; gefitinib/erlotinib; FLAURA), or platinum-based doublet chemotherapy (AURA3) was given. Plasma EGFRm was analyzed at baseline and Weeks 3/6 by droplet digital PCR. Outcomes were assessed by detectable/non-detectable baseline plasma EGFRm and plasma EGFRm clearance (non-detection) at Weeks 3/6. RESULTS: In AURA3 (n = 291), non-detectable versus detectable baseline plasma EGFRm had longer median progression-free survival [mPFS; HR, 0.48; 95% confidence interval (CI), 0.33-0.68; P < 0.0001]. In patients with Week 3 clearance versus non-clearance (n = 184), respectively, mPFS (months; 95% CI) was 10.9 (8.3-12.6) versus 5.7 (4.1-9.7) with osimertinib and 6.2 (4.0-9.7) versus 4.2 (4.0-5.1) with platinum-pemetrexed. In FLAURA (n = 499), mPFS was longer with non-detectable versus detectable baseline plasma EGFRm (HR, 0.54; 95% CI, 0.41-0.70; P < 0.0001). For Week 3 clearance versus non-clearance (n = 334), respectively, mPFS was 19.8 (15.1 to not calculable) versus 11.3 (9.5-16.5) with osimertinib and 10.8 (9.7-11.1) versus 7.0 (5.6-8.3) with comparator EGFR-TKI. Similar outcomes were observed by Week 6 clearance/non-clearance. CONCLUSIONS: Plasma EGFRm analysis as early as 3 weeks on-treatment has the potential to predict outcomes in EGFRm advanced NSCLC.

Not all TMB assays are the same: Clinical validity of robust algorithmic germline filtering.

Huang et al. propose that a tumor-only calculation of tumor mutational burden (TMB), which leverages algorithmic filtering of germline variants, maintains clinical validity across ancestries without necessitating germline sequencing.

A longitudinal circulating tumor DNA-based model associated with survival in metastatic non-small-cell lung cancer.

One of the great challenges in therapeutic oncology is determining who might achieve survival benefits from a particular therapy. Studies on longitudinal circulating tumor DNA (ctDNA) dynamics for the prediction of survival have generally been small or nonrandomized. We assessed ctDNA across 5 time points in 466 non-small-cell lung cancer (NSCLC) patients from the randomized phase 3 IMpower150 study comparing chemotherapy-immune checkpoint inhibitor (chemo-ICI) combinations and used machine learning to jointly model multiple ctDNA metrics to predict overall survival (OS). ctDNA assessments through cycle 3 day 1 of treatment enabled risk stratification of patients with stable disease (hazard ratio (HR) = 3.2 (2.0-5.3), P < 0.001; median 7.1 versus 22.3 months for high- versus low-intermediate risk) and with partial response (HR = 3.3 (1.7-6.4), P < 0.001; median 8.8 versus 28.6 months). The model also identified high-risk patients in an external validation cohort from the randomized phase 3 OAK study of ICI versus chemo in NSCLC (OS HR = 3.73 (1.83-7.60), P = 0.00012). Simulations of clinical trial scenarios employing our ctDNA model suggested that early ctDNA testing outperforms early radiographic imaging for predicting trial outcomes. Overall, measuring ctDNA dynamics during treatment can improve patient risk stratification and may allow early differentiation between competing therapies during clinical trials.

Osimertinib and Selpercatinib Efficacy, Safety, and Resistance in a Multicenter, Prospectively Treated Cohort of EGFR-Mutant and RET Fusion-Positive Lung Cancers.

PURPOSE: Acquired RET fusions have been reported at resistance to treatment with EGFR inhibitors in EGFR-mutant non-small cell lung cancer (NSCLC); however, a multicenter cohort of patients with EGFR-mutant lung cancers treated with osimertinib and selpercatinib for RET fusion-mediated osimertinib resistance has not previously been published. PATIENTS AND METHODS: Patients who received selpercatinib in combination with osimertinib on a prospective expanded access clinical trial (NCT03906331) and single-patient compassionate use programs across five countries were centrally analyzed. All patients had advanced EGFR-mutant NSCLC with a RET fusion detected from tissue or plasma following osimertinib therapy. Clinicopathologic and outcomes data were collected. RESULTS: Fourteen patients with EGFR-mutant and RET fusion-positive lung cancers who experienced prior progression on osimertinib received osimertinib and selpercatinib. EGFR exon 19 deletions (±T790M, 86%) and non-KIF5B fusions (CCDC6-RET 50%, NCOA4-RET 36%) predominated. Osimertinib 80 mg daily and selpercatinib 80 mg twice daily were the most commonly administered dosages. The response rate, disease control rate, and median treatment duration were 50% [95% confidence interval (CI), 25%-75%, n = 12], 83% (95% CI, 55%-95%), and 7.9 months (range, 0.8-25+), respectively. Resistance was complex, involving EGFR on-target (EGFR C797S), RET on-target (RET G810S), and off-target (EML4-ALK/STRN-ALK, KRAS G12S, BRAF V600E) mechanisms; RET fusion loss; or polyclonal mechanisms. CONCLUSIONS: For patients with EGFR-mutant NSCLC with an acquired RET fusion as a mechanism of EGFR inhibitor resistance, the addition of selpercatinib to osimertinib was feasible and safe and offered clinical benefit, supporting the prospective evaluation of this combination. See related commentary by Krebs and Popat, p. 2951.

BRAF V600E and RNF43 Co-mutations Predict Patient Outcomes With Targeted Therapies in Real-World Cases of Colorectal Cancer.

Anti-BRAF/EGFR therapy is approved for metastatic colorectal cancer (mCRC) with BRAFV600E mutations, although not all patients respond. Novel recent findings indicate the potential of RNF43 mutations to predict outcomes in patients with BRAF-mutated microsatellite stable (MSS) mCRC treated with anti-BRAF/EGFR therapy. This study aimed to independently and rapidly validate BRAFV600E/RNF43 co-mutations as predictive biomarkers of benefit to anti-EGFR/BRAF therapy. Clinical data were derived from electronic health record data from ~280 US cancer clinics between January 2011 and March 2022 from the Flatiron Health-Foundation Medicine real-world clinico-genomic mCRC database. Real-world cases of BRAFV600E-mutated mCRC, with patients receiving anti-BRAF/EGFR therapy (n = 49), were included. Patients who were MSS, with RNF43 mutations, had favorable progression-free survival (hazard ratio [HR] 0.29; 95% CI [CI], 0.13-0.65) and overall survival (HR 0.32, 95% CI, 0.12-0.84) compared with wild type. No difference in outcomes was observed between patient groups with RNF43-mutant versus wild-type receiving standard-of-care chemotherapy. BRAFV600E/RNF43 co-mutations predict mCRC anti-BRAF/EGFR outcomes in diverse clinical settings.