Integrated multi-omics assessment of lineage plasticity in a prostate cancer patient with brain and dural metastases.

Metastases to the brain are rare in prostate cancer. Here, we describe a patient with two treatment-emergent metastatic lesions, one to the brain with neuroendocrine prostate cancer (NEPC) histology and one to the dural membrane of adenocarcinoma histology. We performed genomic, transcriptomic, and proteomic characterization of these lesions and the primary tumor to investigate molecular features promoting these metastases. The two metastatic lesions had high genomic similarity, including TP53 mutation and PTEN deletion, with the most striking difference being the additional loss of RB1 in the NEPC lesion. Interestingly, the dural lesion expressed both androgen receptor and neuroendocrine markers, suggesting amphicrine carcinoma (AMPC). When analyzing pioneer transcription factors, the AMPC lesion exhibited elevated FOXA1 activity while the brain NEPC lesion showed elevated HOXC10, NFYB, and OTX2 expression suggesting novel roles in NEPC formation or brain tropism. Our results highlight the utility of performing multi-omic characterization, especially in rare cancer subtypes.

Current uses and resistance mechanisms of enzalutamide in prostate cancer treatment.

INTRODUCTION: Prostate cancer continues to be a major cause of morbidity and mortality for men worldwide. Enzalutamide, a second-generation non-steroidal antiandrogen that blocks androgen receptor (AR) transcriptional activity, is a treatment for biochemically recurrent, metastatic, castration-sensitive, and castration-resistant tumors. Unfortunately, most patients ultimately develop resistance to enzalutamide, making long-term treatment with this agent challenging. AREAS COVERED: We performed a literature search of PubMed without date restrictions to investigate the literature surrounding enzalutamide and discuss the current uses of enzalutamide, proposed mechanisms driving resistance, and summarize current efforts to mitigate this resistance. EXPERT OPINION: Enzalutamide is an effective prostate cancer therapy that is currently used in biochemically recurrent and metastatic disease and for both castration-sensitive and castration-resistant tumors. Unfortunately, resistance to enzalutamide occurs in each of these scenarios. In the clinical setting, enzalutamide-resistant tumors are either AR-driven or AR-indifferent. AR-dependent resistance mechanisms include genomic or epigenomic events that result in enhanced AR signaling. Tumors that do not require AR signaling instead may depend on alternative oncogenic pathways. There are numerous strategies to mitigate enzalutamide resistance, including concurrent use of PARP inhibitors or immune therapies. Additional work is required to uncover novel approaches to treat patients in the enzalutamide-resistant setting.

MYBL2 Drives Prostate Cancer Plasticity: Inhibiting Its Transcriptional Target CDK2 for RB1-Deficient Neuroendocrine Prostate Cancer.

Phenotypic plasticity is a recognized mechanism driving therapeutic resistance in patients with prostate cancer. Although underlying molecular causations driving phenotypic plasticity have been identified, therapeutic success is yet to be achieved. To identify putative master regulator transcription factors (MR-TF) driving phenotypic plasticity in prostate cancer, this work utilized a multiomic approach using genetically engineered mouse models of prostate cancer combined with patient data to identify MYB proto-oncogene like 2 (MYBL2) as a significantly enriched transcription factor in prostate cancer exhibiting phenotypic plasticity. Genetic inhibition of Mybl2 using independent murine prostate cancer cell lines representing phenotypic plasticity demonstrated Mybl2 loss significantly decreased in vivo growth as well as cell fitness and repressed gene expression signatures involved in pluripotency and stemness. Because MYBL2 is currently not druggable, a MYBL2 gene signature was employed to identify cyclin-dependent kinase-2 (CDK2) as a potential therapeutic target. CDK2 inhibition phenocopied genetic loss of Mybl2 and significantly decreased in vivo tumor growth associated with enrichment of DNA damage. Together, this work demonstrates MYBL2 as an important MR-TF driving phenotypic plasticity in prostate cancer. Furthermore, high MYBL2 activity identifies prostate cancer that would be responsive to CDK2 inhibition. SIGNIFICANCE: Prostate cancers that escape therapy targeting the androgen receptor signaling pathways via phenotypic plasticity are currently untreatable. Our study identifies MYBL2 as a MR-TF in phenotypic plastic prostate cancer and implicates CDK2 inhibition as a novel therapeutic target for this most lethal subtype of prostate cancer.

Androgen production, uptake, and conversion (APUC) genes define prostate cancer patients with distinct clinical outcomes.

BACKGROUNDProstate cancer (PC) is driven by aberrant signaling of the androgen receptor (AR) or its ligands, and androgen deprivation therapies (ADTs) are a cornerstone of treatment. ADT responsiveness may be associated with germline changes in genes that regulate androgen production, uptake, and conversion (APUC).METHODSWe analyzed whole-exome sequencing (WES) and whole-transcriptome sequencing (WTS) data from prostate tissues (SU2C/PCF, TCGA, GETx). We also interrogated the Caris Precision Oncology Alliance (POA) DNA (592-gene/whole exome) and RNA (whole transcriptome) next-generation sequencing databases. Algorithm for Linking Activity Networks (ALAN) was used to quantify all pairwise gene-to-gene associations. Real-world overall survival was determined from insurance claims data using Kaplan-Meier estimates.RESULTSSix APUC genes (HSD3B1, HSD3B2, CYP3A43, CYP11A1, CYP11B1, CYP17A1) exhibited coalescent gene behavior in a cohort of metastatic tumors (n = 208). In the Caris POA dataset, the 6 APUC genes (APUC-6) exhibited robust clustering in primary prostate (n = 4,490) and metastatic (n = 2,593) biopsies. Surprisingly, tumors with elevated APUC-6 expression had statically lower expression of AR, AR-V7, and AR signaling scores, suggesting ligand-driven disease biology. APUC-6 genes instead associated with the expression of alternative steroid hormone receptors, ESR1/2 and PGR. We used RNA expression of AR or APUC-6 genes to define 2 subgroups of tumors with differential association with hallmark pathways and cell surface targets.CONCLUSIONSThe APUC-6-high/AR-low tumors represented a subgroup of patients with good clinical outcomes, in contrast with the AR-high or neuroendocrine PCs. Altogether, measuring the aggregate expression of APUC-6 genes in current genomic tests identifies PCs that are ligand (rather than AR) driven and require distinct therapeutic strategies.FUNDINGNCI/NIH 1R37CA288972-01, NCI Cancer Center Support P30 CA077598, DOD W81XWH-22-2-0025, R01 CA249279.

Targeting the multifaceted BRAF in cancer: New directions.

Activating mutations in the mitogen-activated protein kinase (MAPK) pathway represent driver alterations governing tumorigenesis, metastasis, and therapy resistance. MAPK activation predominantly occurs through genomic alterations in RAS and BRAF. BRAF is an effector kinase that functions downstream of RAS and propagates this oncogenic activity through MEK and ERK. Across cancers, BRAF alterations include gain-of-function mutations, copy-number alterations, and structural rearrangements. In cancer patients, BRAF-targeting precision therapeutics are effective against Class I BRAF alterations (p.V600 hotspot mutations) in tumors such as melanomas, thyroid cancers, and colorectal cancers. However, numerous non-Class I BRAF inhibitors are also in development and have been explored in some cancers. Here we discuss the diverse forms of BRAF alterations found in human cancers and the strategies to inhibit them in patients harboring cancers of distinct origins.

Epigenomic signatures of sarcomatoid differentiation to guide the treatment of renal cell carcinoma.

Renal cell carcinoma with sarcomatoid differentiation (sRCC) is associated with poor survival and a heightened response to immune checkpoint inhibitors (ICIs). Two major barriers to improving outcomes for sRCC are the limited understanding of its gene regulatory programs and the low diagnostic yield of tumor biopsies due to spatial heterogeneity. Herein, we characterized the epigenomic landscape of sRCC by profiling 107 epigenomic libraries from tissue and plasma samples from 50 patients with RCC and healthy volunteers. By profiling histone modifications and DNA methylation, we identified highly recurrent epigenomic reprogramming enriched in sRCC. Furthermore, CRISPRa experiments implicated the transcription factor FOSL1 in activating sRCC-associated gene regulatory programs, and FOSL1 expression was associated with the response to ICIs in RCC in two randomized clinical trials. Finally, we established a blood-based diagnostic approach using detectable sRCC epigenomic signatures in patient plasma, providing a framework for discovering epigenomic correlates of tumor histology via liquid biopsy.

Pan-Cancer Interrogation of B7-H3 (CD276) as an Actionable Therapeutic Target Across Human Malignancies.

B7-H3 (CD276) is a transmembrane glycoprotein of the B7 immune checkpoint superfamily that has emerged as a promising therapeutic target. To better understand the applicability of B7-H3-directed therapies, we analyzed 156,791 samples comprising 50 cancer types to interrogate the clinical, genomic, transcriptomic, and immunologic correlates of B7-H3 mRNA expression. DNA (592-gene/whole-exome) and RNA (whole-transcriptome) sequencing was performed from samples submitted to Caris Life Sciences. B7-H3 high versus low expression was based on top and bottom quartiles for each cancer type. Patients' overall survival was determined from insurance claims data. Pathway analysis was performed using gene set enrichment analyses. Immune cell fractions were inferred using quanTIseq. B7-H3 is expressed across several human malignancies including prostate, pancreatic, ovarian, and lung cancers. High B7-H3 expression is associated with differences in overall survival, possibly indicating a prognostic role of B7-H3 for some cancers. When examining molecular features across all cancer types, we did not identify recurrent associations between B7-H3 expression and genetic alterations in TP53, RB1, and KRAS. However, we find consistent enrichment of epithelial-to-mesenchymal transition, Wnt, TGFß, and Notch signaling pathways. In addition, tumors with high B7-H3 expression are associated with greater proportions of M1 macrophages, but lower fractions of CD8+ T cells. We have begun to define the genomic, transcriptomic, clinical, and immunologic features associated with B7-H3 expression in 50 cancer types. We report novel clinical and molecular features of B7-H3-high tumors which may inform how current B7-H3 therapeutics should be deployed and prioritized. SIGNIFICANCE: B7-H3-targeting therapeutics have shown promising results in initial clinical trials. In this pan-cancer analysis of B7-H3 mRNA expression, we found that B7-H3 exhibits robust expression in many common cancer types. These results may inform further development of B7-H3-targeting therapeutics and may guide clinical decisions for patients with limited treatment options.

Genomic Profiling of Rare Undifferentiated Sarcomatoid Subtypes of Pancreatic Carcinomas: In Search of Therapeutic Targets.

PURPOSE: The highly aggressive undifferentiated sarcomatoid carcinoma (USC) subtype of pancreatic ductal adenocarcinoma (PDAC) remains poorly characterized because of its rarity. Previous case reports suggest that immune checkpoint inhibitors could be a promising treatment strategy, but the prevalence of established predictive biomarkers of response is largely unknown. The objective of this study was to leverage comprehensive genomic profiling of USC PDAC tumors to determine the prevalence of biomarkers associated with potential response to targeted therapies. METHODS: USC tumors (n = 20) underwent central pathology review by a board-certified gastrointestinal pathologist to confirm the diagnosis. These samples were compared with non-USC PDAC tumors (N = 5,562). Retrospective analysis of DNA and RNA next-generation sequencing data was performed. RESULTS: USC PDACs were more frequently PD-L1+ by immunohistochemistry than non-USC PDAC (63% v 16%, respectively, P < .001). Furthermore, USC PDAC had an increase in neutrophils (8.99% v 5.55%, P = .005) and dendritic cells (1.08% v 0.00%, q = 0.022) and an increased expression of PDCD1LG2 (4.6% v 1.3%, q = 0.001), PDCD1 (2.0% v 0.8%, q = 0.060), and HAVCR2 (45.9% v 21.7%, q = 0.107) than non-USC PDAC. Similar to non-USC PDAC, KRAS was the most commonly mutated gene (86% v 90%, respectively, P = 1). CONCLUSION: To our knowledge, this work represents the largest molecular analysis of USC tumors to date and showed an increased expression of immune checkpoint genes in USC tumors. These findings provide evidence for further investigation into immune checkpoint inhibitors in USC tumors.

Natural Killer Cell Infiltration in Prostate Cancers Predict Improved Patient Outcomes.

BACKGROUND: Natural killer (NK) cells are non-antigen specific innate immune cells that can be redirected to targets of interest using multiple strategies, although none are currently FDA-approved. We sought to evaluate NK cell infiltration into tumors to develop an improved understanding of which histologies may be most amenable to NK cell-based therapies currently in the developmental pipeline. METHODS: DNA (targeted/whole-exome) and RNA (whole-transcriptome) sequencing was performed from tumors from 45 cancer types (N = 90,916 for all cancers and N = 3365 for prostate cancer) submitted to Caris Life Sciences. NK cell fractions and immune deconvolution were inferred from RNA-seq data using quanTIseq. Real-world overall survival (OS) and treatment status was determined and Kaplan-Meier estimates were calculated. Statistical significance was determined using X2 and Mann-Whitney U tests, with corrections for multiple comparisons where appropriate. RESULTS: In both a pan-tumor and prostate cancer (PCa) -specific setting, we demonstrated that NK cells represent a substantial proportion of the total cellular infiltrate (median range 2-9% for all tumors). Higher NK cell infiltration was associated with improved OS in 28 of 45 cancer types, including (PCa). NK cell infiltration was negatively correlated with common driver mutations and androgen receptor variants (AR-V7) in primary prostate biopsies, while positively correlated with negative immune regulators. Higher levels of NK cell infiltration were associated with patterns consistent with a compensatory anti-inflammatory response. CONCLUSIONS: Using the largest available dataset to date, we demonstrated that NK cells infiltrate a broad range of tumors, including both primary and metastatic PCa. NK cell infiltration is associated with improved PCa patient outcomes. This study demonstrates that NK cells are capable of trafficking to both primary and metastatic PCa and are a viable option for immunotherapy approaches moving forward. Future development of strategies to enhance tumor-infiltrating NK cell-mediated cytolytic activity and activation while limiting inhibitory pathways will be key.

Unraveling the Global Proteome and Phosphoproteome of Prostate Cancer Patient-Derived Xenografts.

Resistance to androgen-deprivation therapies leads to metastatic castration-resistant prostate cancer (mCRPC) of adenocarcinoma (AdCa) origin that can transform into emergent aggressive variant prostate cancer (AVPC), which has neuroendocrine (NE)-like features. In this work, we used LuCaP patient-derived xenograft (PDX) tumors, clinically relevant models that reflect and retain key features of the tumor from advanced prostate cancer patients. Here we performed proteome and phosphoproteome characterization of 48 LuCaP PDX tumors and identified over 94,000 peptides and 9,700 phosphopeptides corresponding to 7,738 proteins. We compared 15 NE versus 33 AdCa samples, which included six different PDX tumors for each group in biological replicates, and identified 309 unique proteins and 476 unique phosphopeptides that were significantly altered and corresponded to proteins that are known to distinguish these two phenotypes. Assessment of concordance from PDX tumor-matched protein and mRNA revealed increased dissonance in transcriptionally regulated proteins in NE and metabolite interconversion enzymes in AdCa. IMPLICATIONS: Overall, our study highlights the importance of protein-based identification when compared with RNA and provides a rich resource of new and feasible targets for clinical assay development and in understanding the underlying biology of these tumors.

MYBL2 drives prostate cancer plasticity and identifies CDK2 as a therapeutic vulnerability in RB1-loss and neuroendocrine prostate cancer.

Phenotypic plasticity is a recognized mechanism driving therapeutic resistance in prostate cancer (PCa) patients. While underlying molecular causations driving phenotypic plasticity have been identified, therapeutic success is yet to be achieved. To identify putative master regulator transcription factors (MR-TF) driving phenotypic plasticity in PCa, this work utilized a multiomic approach using genetically engineered mouse models of prostate cancer combined with patient data to identify MYBL2 as a significantly enriched transcription factor in PCa exhibiting phenotypic plasticity. Genetic inhibition of Mybl2 using independent murine PCa cell lines representing phenotypic plasticity demonstrated Mybl2 loss significantly decreased in vivo growth as well as cell fitness and repressed gene expression signatures involved in pluripotency and stemness. Because MYBL2 is currently not druggable, a MYBL2 gene signature was employed to identify cyclin-dependent kinase-2 (CDK2) as a potential therapeutic target. CDK2 inhibition phenocopied genetic loss of Mybl2 and significantly decreased in vivo tumor growth associated with enrichment of DNA damage. Together, this work demonstrates MYBL2 as an important MR-TF driving phenotypic plasticity in PCa. Further, high MYBL2 activity identifies PCa that would be responsive to CDK2 inhibition. Significance: PCa that escapes therapy targeting the androgen receptor signaling pathways via phenotypic plasticity are currently untreatable. Our study identifies MYBL2 as a MR-TF in phenotypic plastic PCa and implicates CDK2 inhibition as novel therapeutic target for this most lethal subtype of PCa.

Divergent immune microenvironments in two tumor nodules from a patient with mismatch repair-deficient prostate cancer.

Patients with prostate cancer (PC) generally do not respond favorably to immune checkpoint inhibitors, which may be due to a low abundance of tumor-infiltrating lymphocytes even when mutational load is high. Here, we identified a patient who presented with high-grade primary prostate cancer with two adjacent tumor nodules. While both nodules were mismatch repair-deficient (MMRd), exhibited pathogenic MSH2 and MSH6 alterations, had a high tumor mutational burden (TMB), and demonstrated high microsatellite instability (MSI), they had markedly distinct immune phenotypes. The first displayed a dense infiltrate of lymphocytes ("hot nodule"), while the second displayed significantly fewer infiltrating lymphocytes ("cold nodule"). Whole-exome DNA analysis found that both nodules shared many identical mutations, indicating that they were derived from a single clone. However, the cold nodule appeared to be sub-clonal relative to the hot nodule, suggesting divergent evolution of the cold nodule from the hot nodule. Whole-transcriptome RNA analysis found that the cold nodule demonstrated lower expression of genes related to antigen presentation (HLA) and, paradoxically, classical tumor immune tolerance markers such as PD-L1 (CD274) and CTLA-4. Immune cell deconvolution suggested that the hot nodule was enriched not only in CD8+ and CD4 + T lymphocytes, but also in M1 macrophages, activated NK cells, and γδ T cells compared to the cold nodule. This case highlights that MMRd/TMB-high PC can evolve to minimize an anti-tumor immune response, and nominates downregulation of antigen presentation machinery (HLA loss) as a potential mechanism of adaptive immune evasion in PC.

Co-occurring BRCA2/SPOP Mutations Predict Exceptional Poly (ADP-ribose) Polymerase Inhibitor Sensitivity in Metastatic Castration-Resistant Prostate Cancer.

BACKGROUND AND OBJECTIVE: BRCA2 mutations in metastatic castration-resistant prostate cancer (mCRPC) confer sensitivity to poly (ADP-ribose) polymerase (PARP) inhibitors. However, additional factors predicting PARP inhibitor efficacy in mCRPC are needed. Preclinical studies support a relationship between speckle-type POZ protein (SPOP) inactivation and PARP inhibitor sensitivity. We hypothesized that SPOP mutations may predict enhanced PARP inhibitor response in BRCA2-altered mCRPC. METHODS: We conducted a multicenter retrospective study involving 13 sites. We identified 131 patients with BRCA2-altered mCRPC treated with PARP inhibitors, 14 of which also carried concurrent SPOP mutations. The primary efficacy endpoint was prostate-specific antigen (PSA) response rate (≥50% PSA decline). The secondary endpoints were biochemical progression-free survival (PSA-PFS), clinical/radiographic progression-free survival (PFS), and overall survival (OS). These were compared by multivariable Cox proportional hazard models adjusting for age, tumor stage, baseline PSA level, Gleason sum, prior therapies, BRCA2 alteration types, and co-occurring mutations. KEY FINDINGS AND LIMITATIONS: Baseline characteristics were similar between groups. PSA responses were observed in 60% (70/117) of patients with BRCA2mut/SPOPwt disease and in 86% (12/14) of patients with BRCA2mut/SPOPmut disease (p = 0.06). The median time on PARP inhibitor treatment was 24.0 mo (95% confidence interval [CI] 19.2 mo to not reached) in this group versus 8.0 mo (95% CI 6.1-10.9 mo) in patients with BRCA2 mutation alone (p = 0.05). In an unadjusted analysis, patients with BRCA2mut/SPOPmut disease experienced longer PSA-PFS (hazard ratio [HR] 0.33 [95% CI 0.15-0.72], p = 0.005) and clinical/radiographic PFS (HR 0.4 [95% CI 0.18-0.86], p = 0.02), and numerically longer OS (HR 0.4 [95% CI 0.15-1.12], p = 0.08). In a multivariable analysis including histology, Gleason sum, prior taxane, prior androgen receptor pathway inhibitor, stage, PSA, BRCA2 alteration characteristics, and other co-mutations, patients with BRCA2mut/SPOPmut disease experienced longer PSA-PFS (HR 0.16 [95% CI 0.05-0.47], adjusted p = 0.001), clinical/radiographic PFS (HR 0.28 [95% CI 0.1-0.81], adjusted p = 0.019), and OS (HR 0.19 [95% CI 0.05-0.69], adjusted p = 0.012). In a separate cohort of patients not treated with a PARP inhibitor, there was no difference in OS between patients with BRCA2mut/SPOPmut versus BRCA2mut/SPOPwt disease (HR 0.97 [95% CI 0.40-2.4], p = 0.94). In a genomic signature analysis, Catalog of Somatic Mutations in Cancer (COSMIC) SBS3 scores predictive of homologous recombination repair (HRR) defects were higher for BRCA2mut/SPOPmut than for BRCA2mut/SPOPwt disease (p = 0.04). This was a retrospective study, and additional prospective validation cohorts are needed. CONCLUSIONS AND CLINICAL IMPLICATIONS: In this retrospective analysis, PARP inhibitors appeared more effective in patients with BRCA2mut/SPOPmut than in patients with BRCA2mut/SPOPwt mCRPC. This may be related to an increase in HRR defects in coaltered disease. PATIENT SUMMARY: In this study, we demonstrate that co-alteration of both BRCA2 and SPOP predicts superior clinical outcomes to treatment with poly (ADP-ribose) polymerase (PARP) inhibitors than BRCA2 alteration without SPOP mutation.

Unraveling the Global Proteome and Phosphoproteome of Prostate Cancer Patient-Derived Xenografts.

Resistance to androgen deprivation therapies leads to metastatic castration-resistant prostate cancer (mCRPC) of adenocarcinoma (AdCa) origin that can transform to emergent aggressive variant prostate cancer (AVPC) which has neuroendocrine (NE)-like features. To this end, we used LuCaP patient-derived xenograft (PDX) tumors, clinically relevant models that reflects and retains key features of the tumor from advanced prostate cancer patients. Here we performed proteome and phosphoproteome characterization of 48 LuCaP PDX tumors and identified over 94,000 peptides and 9,700 phosphopeptides corresponding to 7,738 proteins. When we compared 15 NE versus 33 AdCa PDX samples, we identified 309 unique proteins and 476 unique phosphopeptides that were significantly altered and corresponded to proteins that are known to distinguish these two phenotypes. Assessment of protein and RNA concordance from these tumors revealed increased dissonance in transcriptionally regulated proteins in NE and metabolite interconversion enzymes in AdCa.

Saracatinib synergizes with enzalutamide to downregulate AR activity in CRPC.

Prostate cancer (PCa) remains the most diagnosed non-skin cancer amongst the American male population. Treatment for localized prostate cancer consists of androgen deprivation therapies (ADTs), which typically inhibit androgen production and the androgen receptor (AR). Though initially effective, a subset of patients will develop resistance to ADTs and the tumors will transition to castration-resistant prostate cancer (CRPC). Second generation hormonal therapies such as abiraterone acetate and enzalutamide are typically given to men with CRPC. However, these treatments are not curative and typically prolong survival only by a few months. Several resistance mechanisms contribute to this lack of efficacy such as the emergence of AR mutations, AR amplification, lineage plasticity, AR splice variants (AR-Vs) and increased kinase signaling. Having identified SRC kinase as a key tyrosine kinase enriched in CRPC patient tumors from our previous work, we evaluated whether inhibition of SRC kinase synergizes with enzalutamide or chemotherapy in several prostate cancer cell lines expressing variable AR isoforms. We observed robust synergy between the SRC kinase inhibitor, saracatinib, and enzalutamide, in the AR-FL+/AR-V+ CRPC cell lines, LNCaP95 and 22Rv1. We also observed that saracatinib significantly decreases AR Y534 phosphorylation, a key SRC kinase substrate residue, on AR-FL and AR-Vs, along with the AR regulome, supporting key mechanisms of synergy with enzalutamide. Lastly, we also found that the saracatinib-enzalutamide combination reduced DNA replication compared to the saracatinib-docetaxel combination, resulting in marked increased apoptosis. By elucidating this combination strategy, we provide pre-clinical data that suggests combining SRC kinase inhibitors with enzalutamide in select patients that express both AR-FL and AR-Vs.

Nicotinamide enhances natural killer cell function and yields remissions in patients with non-Hodgkin lymphoma.

Allogeneic natural killer (NK) cell adoptive transfer has shown the potential to induce remissions in relapsed or refractory leukemias and lymphomas, but strategies to enhance NK cell survival and function are needed to improve clinical efficacy. Here, we demonstrated that NK cells cultured ex vivo with interleukin-15 (IL-15) and nicotinamide (NAM) exhibited stable induction of l-selectin (CD62L), a lymphocyte adhesion molecule important for lymph node homing. High frequencies of CD62L were associated with elevated transcription factor forkhead box O1 (FOXO1), and NAM promoted the stability of FOXO1 by preventing proteasomal degradation. NK cells cultured with NAM exhibited metabolic changes associated with elevated glucose flux and protection against oxidative stress. NK cells incubated with NAM also displayed enhanced cytotoxicity and inflammatory cytokine production and preferentially persisted in xenogeneic adoptive transfer experiments. We also conducted a first-in-human phase 1 clinical trial testing adoptive transfer of NK cells expanded ex vivo with IL-15 and NAM (GDA-201) combined with monoclonal antibodies in patients with relapsed or refractory non-Hodgkin lymphoma (NHL) and multiple myeloma (MM) (NCT03019666). Cellular therapy with GDA-201 and rituximab was well tolerated and yielded an overall response rate of 74% in 19 patients with advanced NHL. Thirteen patients had a complete response, and 1 patient had a partial response. GDA-201 cells were detected for up to 14 days in blood, bone marrow, and tumor tissues and maintained a favorable metabolic profile. The safety and efficacy of GDA-201 in this study support further development as a cancer therapy.

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.

Metastatic Prostate Cancers with BRCA2 versus ATM Mutations Exhibit Divergent Molecular Features and Clinical Outcomes.

PURPOSE: In patients with metastatic prostate cancer (mPC), ATM and BRCA2 mutations dictate differences in PARPi inhibitor response and other therapies. We interrogated the molecular features of ATM- and BRCA2-mutated mPC to explain the divergent clinical outcomes and inform future treatment decisions. EXPERIMENTAL DESIGN: We examined a novel set of 1,187 mPCs after excluding microsatellite-instable (MSI) tumors. We stratified these based on ATM (n = 88) or BRCA2 (n = 98) mutations. As control groups, mPCs with mutations in 12 other homologous recombination repair (HRR) genes were considered non-BRCA2/ATM HRR-deficient (HRDother, n = 193), whereas lack of any HRR mutations were considered HRR-proficient (HRP; n = 808). Gene expression analyses were performed using Limma. Real-world overall survival was determined from insurance claims data. RESULTS: In noncastrate mPCs, only BRCA2-mutated mPCs exhibited worse clinical outcomes to AR-targeted therapies. In castrate mPCs, both ATM and BRCA2 mutations exhibited worse clinical outcomes to AR-targeted therapies. ATM-mutated mPCs had reduced TP53 mutations and harbored coamplification of 11q13 genes, including CCND1 and genes in the FGF family. BRCA2-mutated tumors showed elevated genomic loss-of-heterozygosity scores and were often tumor mutational burden high. BRCA2-mutated mPCs had upregulation of cell-cycle genes and were enriched in cell-cycle signaling programs. This was distinct from ATM-mutated tumors. CONCLUSIONS: Tumoral ATM and BRCA2 mutations are associated with differential clinical outcomes when patients are stratified by treatments, including hormonal or taxane therapies. ATM- and BRCA2-mutated tumors exhibited differences in co-occurring molecular features. These unique molecular features may inform therapeutic decisions and development of novel therapies.

Saracatinib synergizes with enzalutamide to downregulate androgen receptor activity in castration resistant prostate cancer.

Prostate cancer (PCa) remains the most diagnosed non-skin cancer amongst the American male population. Treatment for localized prostate cancer consists of androgen deprivation therapies (ADTs), which typically inhibit androgen production and the androgen receptor (AR). Though initially effective, a subset of patients will develop resistance to ADTs and the tumors will transition to castration-resistant prostate cancer (CRPC). Second generation hormonal therapies such as abiraterone acetate and enzalutamide are typically given to men with CRPC. However, these treatments are not curative and typically prolong survival only by a few months. Several resistance mechanisms contribute to this lack of efficacy such as the emergence of AR mutations, AR amplification, lineage plasticity, AR splice variants (AR-Vs) and increased kinase signaling. Having identified SRC kinase as a key tyrosine kinase enriched in CRPC patient tumors from our previous work, we evaluated whether inhibition of SRC kinase synergizes with enzalutamide or chemotherapy in several prostate cancer cell lines expressing variable AR isoforms. We observed robust synergy between the SRC kinase inhibitor, saracatinib, and enzalutamide, in the AR-FL+/AR-V+ CRPC cell lines, LNCaP95 and 22Rv1. We also observed that saracatinib significantly decreases AR Y 534 phosphorylation, a key SRC kinase substrate residue, on AR-FL and AR-Vs, along with the AR regulome, supporting key mechanisms of synergy with enzalutamide. Lastly, we also found that the saracatinib-enzalutamide combination reduced DNA replication compared to the saracatinib-docetaxel combination, resulting in marked increased apoptosis. By elucidating this combination strategy, we provide pre-clinical data that suggests combining SRC kinase inhibitors with enzalutamide in select patients that express both AR-FL and AR-Vs.