Douglas A. Mata

Background: Inactivating BRCA1/2 mutations confer sensitivity to poly(ADP‐ribose) polymerase inhibitors (PARPi) in prostate cancer (PCA). However, secondary BRCA1/2 reversion mutations ( BRCA rev) can restore BRCA function and mediate acquired PARPi resistance. While BRCA rev typically arise under PARPi selective pressure, their occurrence in PARPi‐naïve settings remains incompletely understood. We sought to characterize the frequency, clinical context, and therapeutic correlates of BRCA rev detection in men with advanced PCA, including those without prior PARPi exposure. Methods: We restrospectively analyzed clinical liquid biopsy results from men with PCA using FoundationOneLiquid CDx between January and December 2023. BRCA rev were defined as sequence alterations predicted to restore open reading frame of BRCA1 or BRCA2 harboring pathogenic inactivating variants. Clinical and treatment histories, including chemotherapy and PARPi exposure, were abstracted from medical records. Results: Over a one‐year period, we identified 10 PCA patients with inactivating BRCA1/2 alterations, BRCA rev and available treatment history. BRCA rev were detected in three of 10 (30%) patients who had not received prior PARPi therapy. All three PARP‐naïve patients had previously received chemotherapy (platinum or docetaxel) and radiation, and each exhibited multiple BRCA rev events. The remaining 7 patients (70%) had prior olaparib exposure. Among these, the duration of PARPi response was longer in chemo‐naïve patients compared with previously treated with chemotherapy (median: 22 vs. 7.5 months, respectively). Conclusions: BRCA rev can emerge in PCA in the absence of prior PARPi therapy, particularly following exposure to cytotoxic chemotherapy and radiation. These findings support that DNA‐damaging therapies may promote BRCA rev formation, potentially predisposing to primary PARPi resistance. Early integration of PARPi therapy before chemotherapy may enhance clinical benefit and circumvent emergence of primary resistance.

A mixed phenotype (MP) is a characteristic of de novo MP acute leukemia (MPAL), but it can also be found in other leukemias. It poses substantial classification and management dilemmas. Herein, we report a large cohort of acute leukemia with MP and define acute myeloid leukemia with MP (AML-MP) and MPAL as 2 distinct groups by characterizing clinical, genetic, and transcriptomic features. Clinically, patients with AML-MP and MPAL were both treated with either AML- or acute lymphoblastic leukemia (ALL)–directed induction regimens. AML-MP has inferior responses (hazard ratio, 12.5; 95% confidence interval, 2.72-57.8; P = .001), whereas MPAL has better responses to ALL-directed treatment. Genetically, AML-MP harbors more frequent RUNX1 (23/52 [44%]) and TP53 (12/52 [23.1%]) mutations. In contrast, RUNX1 mutations are less frequent in MPAL (8/35 [23%]; P = .01 vs AML-MP) and TP53 mutations as a driver are virtually absent in MPAL. Transcriptionally, AML-MP shows enrichment for stemness signatures and a relative deficit of transcription factors critical for myeloid and lymphoid differentiation. Furthermore, AML-MP rarely switches to a lymphoid immunophenotype after treatment, in contrast to MPAL (1/40 [2.5%] vs 10/28 [35.7%]; P = .0003). Last, a genomic classification framework is proposed for future studies. Together, these data support the designation of AML-MP as a diagnosis distinct from MPAL and provide novel insights into the pathogenesis and therapies of acute leukemia with MP.

Microsatellite instability high (MSI-H) and mismatch repair deficient (dMMR) tumor status have been demonstrated to predict patient response to immunotherapies. We developed and validated a next-generation sequencing (NGS)-based companion diagnostic (CDx) to detect MSI-H solid tumors via a comprehensive genomic profiling (CGP) assay, FoundationOne®CDx (F1CDx). To determine MSI status, F1CDx calculates the fraction of unstable microsatellite loci across >2000 loci using a fraction-based (FB) analysis. Across solid tumor types, F1CDx demonstrated a high analytical concordance with both PCR (n = 264) and IHC (n = 279) with an overall percent agreement (OPA) of 97.7% and 97.8%, respectively. As part of a retrospective bridging clinical study from KEYNOTE-158 Cohort K and KEYNOTE-164, patients with MSI-H tumors as determined by F1CDx demonstrated an objective response rate (ORR) of 43.0% to pembrolizumab. In real-world cancer patients from a deidentified clinicogenomic database, F1CDx was at least equivalent in assessing clinical outcome following immunotherapy compared with MMR IHC. Demonstrated analytical and clinical performance of F1CDx led to the pan-tumor FDA approval in 2022 of F1CDx to identify MSI-H solid tumor patients for treatment with pembrolizumab. F1CDx is an accurate, reliable, and FDA-approved method for the identification of MSI-H tumors for treatment with pembrolizumab.

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.

Gene fusions involving EWSR1 or FUS as the 5′ partner have been reported in a diverse array of sarcomas. Here, we characterize the histopathology and genomics of six tumors harboring a gene fusion between EWSR1 or FUS and POU2AF3, an understudied, putative colorectal cancer predisposition gene. Striking morphologic features reminiscent of synovial sarcoma were observed including a biphasic appearance with variable fusiform to epithelioid cytomorphology and staghorn‐type vasculature. RNA sequencing demonstrated variable breakpoints in EWSR1/FUS along with similar breakpoints in POU2AF3 that encompassed a 3′ portion of this gene. For cases in which additional information was available, the behavior of these neoplasms was aggressive with local spread and/or distant metastases. Although further studies are needed to confirm the functional significance of our findings, POU2AF3 fusions to EWSR1 or FUS may define a novel type of POU2AF3‐rearranged sarcomas with aggressive, malignant behavior.

Clear cell mesothelioma is uncommon and shows predominance of clear cells with resemblance to clear cell carcinomas. Clinicopathologic and molecular descriptions of clear cell mesothelioma remained limited. In this study, we identified an index patient with clear cell mesothelioma, confirmed by immunohistochemical and ultrastructural studies. Targeted next‐generation sequencing revealed the presence of an inactivating VHL mutation. We then systematically searched for VHL‐mutant mesotheliomas in a comprehensive genomic profiling database of 1532 mesotheliomas. Collectively, we identified a cohort of four VHL‐mutant clear cell mesotheliomas, including three peritoneal and one pleural tumors from three females and one male, with age range of 47–68 (median 63) years. Histologically, each tumor showed a microcystic to tubulopapillary architecture with prominent clear cells. By next‐generation DNA sequencing, each of the four clear cell mesotheliomas harbored inactivating VHL mutations, while lacking other alterations typical of mesotheliomas such as BAP1, NF2, SETD2, CDKN2A, CDKN2B, TP53, and PTEN. By using low‐pass whole genome sequencing on the index case and targeted next‐generation sequencing on the remaining three cases, we identified extensive loss of heterozygosity throughout the genome but consistently sparing chromosomes 5, 7, and 20, characteristic of genomic near‐haploidization. In summary, clear cell mesotheliomas were characterized by inactivating VHL mutations and genomic near‐haploidization and appeared to represent a distinct clinicopathologic and molecular category of mesotheliomas. Our findings implicate VHL in the pathogenesis of a subset of mesotheliomas, particularly those with clear cell morphology.