Identification of a Biomarker Panel from Genome-Wide Methylation to Detect Early HPV-Associated Oropharyngeal Cancer.

As oropharyngeal cancer (OPC) associated with human papillomavirus (HPV) increases in men, the need for a screening test to diagnose OPC early is crucial. This study agnostically identified differentially methylated CpG sites to identify additional biomarkers to improve screening for early OPC.DNA was extracted from oral gargles of 89 early cases and 108 frequency matched healthy controls, and processed for genome-wide methylation using the Illumina Infinium MethylationEPIC BeadChip. Selected sites were combined with our prior methylation data in the EPB41L3 gene (CpG sites 438, 427, and 425) and oral HPV16 and HPV18 status were considered as binary variables (positive/negative). Lasso regression identified CpG sites strongly associated with early OPC. ROC curves with AUC were generated. The panel was validated utilizing bootstrap resampling.Machine learning analyses identified 14 markers that are significantly associated with early OPC, including one EPB41L3 CpG site (438) and oral HPV16 status. A final model was trained on all available samples using the discovered panel and was able to predict early OPC compared with controls with an AUC of 0.970 on the training set. In the bootstrap validation sets, the average AUC was 0.935, indicating adequate internal validity.Our data suggest that this panel can detect OPC early, however external validation of this panel is needed. Further refinement of a panel of biomarkers to diagnose OPC earlier is urgently needed to prevent complex treatment of OPC and associated comorbidities, while reducing risk of recurrence. PREVENTION RELEVANCE: This study identified biomarkers using genome-wide methylation to create a panel capable of discerning early oropharyngeal cancer (OPC) from those without OPC. Such a biomarker panel would be an effective tool to detect OPC early and prevent complications of treatment associated with later diagnosis.

Increased spatial coupling of integrin and collagen IV in the immunoresistant clear cell renal cell carcinoma tumor microenvironment.

Background: Immunotherapy (IO) has improved survival for patients with advanced clear cell renal cell carcinoma (ccRCC), but resistance to therapy develops in most patients. We use cellular-resolution spatial transcriptomics in patients with IO naïve and IO exposed primary ccRCC tumors to better understand IO resistance. Spatial molecular imaging (SMI) was obtained for tumor and adjacent stroma samples. Spatial gene set enrichment analysis (GSEA) and autocorrelation (coupling with high expression) of ligand-receptor transcript pairs were assessed. Multiplex immunofluorescence (mIF) validation was used for significant autocorrelative findings and the cancer genome atlas (TCGA) and the clinical proteomic tumor analysis consortium (CPTAC) databases were queried to assess bulk RNA expression and proteomic correlates. Results: 21 patient samples underwent SMI. Viable tumors following IO harbored more stromal CD8+ T cells and neutrophils than IO naïve tumors. YES1 was significantly upregulated in IO exposed tumor cells. The epithelial-mesenchymal transition pathway was enriched on spatial GSEA and the associated transcript pair COL4A1-ITGAV had significantly higher autocorrelation in the stroma. Fibroblasts, tumor cells, and endothelium had the relative highest expression. More integrin αV+ cells were seen in IO exposed stroma on mIF validation. Compared to other cancers in TCGA, ccRCC tumors have the highest expression of both COL4A1 and ITGAV. In CPTAC, collagen IV protein was more abundant in advanced stages of disease. Conclusions: On spatial transcriptomics, COL4A1 and ITGAV were more autocorrelated in IO-exposed stroma compared to IO-naïve tumors, with high expression amongst fibroblasts, tumor cells, and endothelium. Integrin represents a potential therapeutic target in IO treated ccRCC.

Epigenetic Activation of TUSC3 Sensitizes Glioblastoma to Temozolomide Independent of MGMT Promoter Methylation Status.

Temozolomide (TMZ) is an important first-line treatment for glioblastoma (GBM), but there are limitations to TMZ response in terms of durability and dependence on the promoter methylation status of the DNA repair gene O6-methylguanine DNA methyltransferase (MGMT). MGMT-promoter-hypermethylated (MGMT-M) GBMs are more sensitive to TMZ than MGMT-promoter-hypomethylated (MGMT-UM) GBMs. Moreover, TMZ resistance is inevitable even in TMZ-sensitive MGMT-M GBMs. Hence, epigenetic reprogramming strategies are desperately needed in order to enhance TMZ response in both MGMT-M and MGMT-UM GBMs. In this study, we present novel evidence that the epigenetic reactivation of Tumor Suppressor Candidate 3 (TUSC3) can reprogram sensitivity of GBM stem cells (GSCs) to TMZ irrespective of MGMT promoter methylation status. Interrogation of TCGA patient GBM datasets confirmed TUSC3 promoter regulation of TUSC3 expression and also revealed a strong positive correlation between TUSC3 expression and GBM patient survival. Using a combination of loss-of-function, gain-of-function and rescue studies, we demonstrate that TUSC3 reactivation is associated with enhanced TMZ response in both MGMT-M and MGMT-UM GSCs. Further, we provide novel evidence that the demethylating agent 5-Azacitidine (5-Aza) reactivates TUSC3 expression in MGMT-M GSCs, whereas the combination of 5-Aza and MGMT inhibitor Lomeguatrib is necessary for TUSC3 reactivation in MGMT-UM GSCs. Lastly, we propose a pharmacological epigenetic reactivation strategy involving TUSC3 that leads to significantly prolonged survival in MGMT-M and MGMT-UM orthotopic GSCs models. Collectively, our findings provide a framework and rationale to further explore TUSC3-mediated epigenetic reprogramming strategies that could enhance TMZ sensitivity and outcomes in GBM. Mechanistic and translational evidence gained from such studies could contribute towards optimal design of impactful trials for MGMT-UM GBMs that currently do not have good treatment options.

CK1δ and CK1ε Signaling Sustains Mitochondrial Metabolism and Cell Survival in Multiple Myeloma.

Multiple myeloma remains an incurable malignancy due to acquisition of intrinsic programs that drive therapy resistance. Here we report that casein kinase-1δ (CK1δ) and CK1ε are therapeutic targets in multiple myeloma that are necessary to sustain mitochondrial metabolism. Specifically, the dual CK1δ/CK1ε inhibitor SR-3029 had potent in vivo and ex vivo anti-multiple myeloma activity, including against primary multiple myeloma patient specimens. RNA sequencing (RNA-seq) and metabolic analyses revealed inhibiting CK1δ/CK1ε disables multiple myeloma metabolism by suppressing genes involved in oxidative phosphorylation (OxPhos), reducing citric acid cycle intermediates, and suppressing complexes I and IV of the electron transport chain. Finally, sensitivity of multiple myeloma patient specimens to SR-3029 correlated with elevated expression of mitochondrial genes, and RNA-seq from 687 multiple myeloma patient samples revealed that increased CSNK1D, CSNK1E, and OxPhos genes correlate with disease progression and inferior outcomes. Thus, increases in mitochondrial metabolism are a hallmark of multiple myeloma progression that can be disabled by targeting CK1δ/CK1ε. SIGNIFICANCE: CK1δ and CK1ε are attractive therapeutic targets in multiple myeloma whose expression increases with disease progression and connote poor outcomes, and that are necessary to sustain expression of genes directing OxPhos.

Deregulated expression of the 14q32 miRNA cluster in clear cell renal cancer cells.

Clear cell renal cell carcinomas (ccRCC) are characterized by arm-wide chromosomal alterations. Loss at 14q is associated with disease aggressiveness in ccRCC, which responds poorly to chemotherapeutics. The 14q locus contains one of the largest miRNA clusters in the human genome; however, little is known about the contribution of these miRNAs to ccRCC pathogenesis. In this regard, we investigated the expression pattern of selected miRNAs at the 14q32 locus in TCGA kidney tumors and in ccRCC cell lines. We demonstrated that the miRNA cluster is downregulated in ccRCC (and cell lines) as well as in papillary kidney tumors relative to normal kidney tissues (and primary renal proximal tubule epithelial (RPTEC) cells). We demonstrated that agents modulating expression of DNMT1 (e.g., 5-Aza-deoxycytidine) could modulate 14q32 miRNA expression in ccRCC cell lines. Lysophosphatidic acid (LPA, a lysophospholipid mediator elevated in ccRCC) not only increased labile iron content but also modulated expression of a 14q32 miRNA. Through an overexpression approach targeting a subset of 14q32 miRNAs (specifically at subcluster A: miR-431-5p, miR-432-5p, miR-127-3p, and miR-433-3p) in 769-P cells, we uncovered changes in cellular viability and claudin-1, a tight junction marker. A global proteomic approach was implemented using these miRNA overexpressing cell lines which uncovered ATXN2 as a highly downregulated target. Collectively, these findings support a contribution of miRNAs at 14q32 in ccRCC pathogenesis.

The Polyamine-Hypusine Circuit Controls an Oncogenic Translational Program Essential for Malignant Conversion in MYC-Driven Lymphoma.

The MYC oncoprotein is activated in a broad spectrum of human malignancies and transcriptionally reprograms the genome to drive cancer cell growth. Given this, it is unclear if targeting a single effector of MYC will have therapeutic benefit. MYC activates the polyamine-hypusine circuit, which posttranslationally modifies the eukaryotic translation factor eIF5A. The roles of this circuit in cancer are unclear. Here we report essential intrinsic roles for hypusinated eIF5A in the development and maintenance of MYC-driven lymphoma, where the loss of eIF5A hypusination abolishes malignant transformation of MYC-overexpressing B cells. Mechanistically, integrating RNA sequencing, ribosome sequencing, and proteomic analyses revealed that efficient translation of select targets is dependent upon eIF5A hypusination, including regulators of G1-S phase cell-cycle progression and DNA replication. This circuit thus controls MYC's proliferative response, and it is also activated across multiple malignancies. These findings suggest the hypusine circuit as a therapeutic target for several human tumor types. SIGNIFICANCE: Elevated EIF5A and the polyamine-hypusine circuit are manifest in many malignancies, including MYC-driven tumors, and eIF5A hypusination is necessary for MYC proliferative signaling. Not-ably, this circuit controls an oncogenic translational program essential for the development and maintenance of MYC-driven lymphoma, supporting this axis as a target for cancer prevention and treatment. See related commentary by Wilson and Klein, p. 248. This article is highlighted in the In This Issue feature, p. 247.

TCR-Independent Metabolic Reprogramming Precedes Lymphoma-Driven Changes in T-cell Fate.

Chronic T-cell receptor (TCR) signaling in the tumor microenvironment is known to promote T-cell dysfunction. However, we reasoned that poorly immunogenic tumors may also compromise T cells by impairing their metabolism. To address this, we assessed temporal changes in T-cell metabolism, fate, and function in models of B-cell lymphoma driven by Myc, a promoter of energetics and repressor of immunogenicity. Increases in lymphoma burden most significantly impaired CD4+ T-cell function and promoted regulatory T cell (Treg) and Th1-cell differentiation. Metabolomic analyses revealed early reprogramming of CD4+ T-cell metabolism, reduced glucose uptake, and impaired mitochondrial function, which preceded changes in T-cell fate. In contrast, B-cell lymphoma metabolism remained robust during tumor progression. Finally, mitochondrial functions were impaired in CD4+ and CD8+ T cells in lymphoma-transplanted OT-II and OT-I transgenic mice, respectively. These findings support a model, whereby early, TCR-independent, metabolic interactions with developing lymphomas limits T cell-mediated immune surveillance.

Proteogenomic, Epigenetic, and Clinical Implications of Recurrent Aberrant Splice Variants in Clear Cell Renal Cell Carcinoma.

BACKGROUND: Alternative mRNA splicing can be dysregulated in cancer, resulting in the generation of aberrant splice variants (SVs). Given the paucity of actionable genomic mutations in clear cell renal cell carcinoma (ccRCC), aberrant SVs may be an avenue to novel mechanisms of pathogenesis. OBJECTIVE: To identify and characterize aberrant SVs enriched in ccRCC. DESIGN, SETTING, AND PARTICIPANTS: Using RNA-seq data from the Cancer Cell Line Encyclopedia, we identified neojunctions uniquely expressed in ccRCC. Candidate SVs were then checked for expression across normal tissue in the Genotype-Tissue Expression Project and primary tumor tissue from The Cancer Genome Atlas (TCGA), Clinical Proteomic Tumor Analysis Consortium (CPTAC), and our institutional Total Cancer Care database. OUTCOME MEASUREMENTS AND STATISTICAL ANALYSIS: Clinicopathologic, genomic, and survival data were available for all cohorts. Epigenetic data were available for the TCGA and CPTAC cohorts. Proteomic data were available for the CPTAC cohort. The association of aberrant SV expression with these variables was examined using the Kruskal-Wallis test, pairwise t test, Spearman correlation test, and Cox regression analysis. RESULTS AND LIMITATIONS: Our pipeline identified 16 ccRCC-enriched SVs. EGFR, HPCAL1-SV and RNASET2-SV expression was negatively correlated with gene-specific CpG methylation. We derived a survival risk score based primarily on the expression of five SVs (RNASET2, FGD1, PDZD2, COBLL1, and PTPN14), which was consistent and applicable across multiple cohorts on multivariate analysis. The splicing factor RBM4, which modulates splicing of HIF-1α, exhibited significantly lower expression at the protein level in the high-risk group, as defined by our SV-based score. CONCLUSIONS: We describe 16 aberrant SVs enriched in ccRCC, many of which are associated with disease biology and/or clinical outcomes. This study provides a novel strategy for identifying and characterizing disease-specific aberrant SVs. PATIENT SUMMARY: We describe a method to identify disease targets and biomarkers using transcriptomic analysis beyond somatic mutations or gene expression. Kidney tumors express unique splice variants that may provide additional prognostic information following surgery.

A Targetable Myeloid Inflammatory State Governs Disease Recurrence in Clear-Cell Renal Cell Carcinoma.

It is poorly understood how the tumor immune microenvironment influences disease recurrence in localized clear-cell renal cell carcinoma (ccRCC). Here we performed whole-transcriptomic profiling of 236 tumors from patients assigned to the placebo-only arm of a randomized, adjuvant clinical trial for high-risk localized ccRCC. Unbiased pathway analysis identified myeloid-derived IL6 as a key mediator. Furthermore, a novel myeloid gene signature strongly correlated with disease recurrence and overall survival on uni- and multivariate analyses and is linked to TP53 inactivation across multiple data sets. Strikingly, effector T-cell gene signatures, infiltration patterns, and exhaustion markers were not associated with disease recurrence. Targeting immunosuppressive myeloid inflammation with an adenosine A2A receptor antagonist in a novel, immunocompetent, Tp53-inactivated mouse model significantly reduced metastatic development. Our findings suggest that myeloid inflammation promotes disease recurrence in ccRCC and is targetable as well as provide a potential biomarker-based framework for the design of future immuno-oncology trials in ccRCC. SIGNIFICANCE: Improved understanding of factors that influence metastatic development in localized ccRCC is greatly needed to aid accurate prediction of disease recurrence, clinical decision-making, and future adjuvant clinical trial design. Our analysis implicates intratumoral myeloid inflammation as a key driver of metastasis in patients and a novel immunocompetent mouse model. This article is highlighted in the In This Issue feature, p. 2221.

Disrupting the MYC-TFEB Circuit Impairs Amino Acid Homeostasis and Provokes Metabolic Anergy.

MYC family oncoproteins are regulators of metabolic reprogramming that sustains cancer cell anabolism. Normal cells adapt to nutrient-limiting conditions by activating autophagy, which is required for amino acid (AA) homeostasis. Here we report that the autophagy pathway is suppressed by Myc in normal B cells, in premalignant and neoplastic B cells of Eµ-Myc transgenic mice, and in human MYC-driven Burkitt lymphoma. Myc suppresses autophagy by antagonizing the expression and function of transcription factor EB (TFEB), a master regulator of autophagy. Mechanisms that sustained AA pools in MYC-expressing B cells include coordinated induction of the proteasome and increases in AA transport. Reactivation of the autophagy-lysosomal pathway by TFEB disabled the malignant state by disrupting mitochondrial functions, proteasome activity, AA transport, and AA and nucleotide metabolism, leading to metabolic anergy, growth arrest, and apoptosis. This phenotype provides therapeutic opportunities to disable MYC-driven malignancies, including AA restriction and treatment with proteasome inhibitors. SIGNIFICANCE: MYC suppresses TFEB and autophagy and controls amino acid homeostasis by upregulating amino acid transport and the proteasome, and reactivation of TFEB disables the metabolism of MYC-driven tumors.

The Impact of Epigenetic Modifications on Adaptive Resistance Evolution in Glioblastoma.

Glioblastoma (GBM) is a highly lethal cancer that is universally refractory to the standard multimodal therapies of surgical resection, radiation, and chemotherapy treatment. Temozolomide (TMZ) is currently the best chemotherapy agent for GBM, but the durability of response is epigenetically dependent and often short-lived secondary to tumor resistance. Therapies that can provide synergy to chemoradiation are desperately needed in GBM. There is accumulating evidence that adaptive resistance evolution in GBM is facilitated through treatment-induced epigenetic modifications. Epigenetic alterations of DNA methylation, histone modifications, and chromatin remodeling have all been implicated as mechanisms that enhance accessibility for transcriptional activation of genes that play critical roles in GBM resistance and lethality. Hence, understanding and targeting epigenetic modifications associated with GBM resistance is of utmost priority. In this review, we summarize the latest updates on the impact of epigenetic modifications on adaptive resistance evolution in GBM to therapy.

TFEB links MYC signaling to epigenetic control of myeloid differentiation and acute myeloid leukemia.

MYC oncoproteins regulate transcription of genes directing cell proliferation, metabolism and tumorigenesis. A variety of alterations drive MYC expression in acute myeloid leukemia (AML) and enforced MYC expression in hematopoietic progenitors is sufficient to induce AML. Here we report that AML and myeloid progenitor cell growth and survival rely on MYC-directed suppression of Transcription Factor EB (TFEB), a master regulator of the autophagy-lysosome pathway. Notably, although originally identified as an oncogene, TFEB functions as a tumor suppressor in AML, where it provokes AML cell differentiation and death. These responses reflect TFEB control of myeloid epigenetic programs, by inducing expression of isocitrate dehydrogenase-1 (IDH1) and IDH2, resulting in global hydroxylation of 5-methycytosine. Finally, activating the TFEB-IDH1/IDH2-TET2 axis is revealed as a targetable vulnerability in AML. Thus, epigenetic control by a MYC-TFEB circuit dictates myeloid cell fate and is essential for maintenance of AML.

KLK3 SNP-SNP interactions for prediction of prostate cancer aggressiveness.

Risk classification for prostate cancer (PCa) aggressiveness and underlying mechanisms remain inadequate. Interactions between single nucleotide polymorphisms (SNPs) may provide a solution to fill these gaps. To identify SNP-SNP interactions in the four pathways (the angiogenesis-, mitochondria-, miRNA-, and androgen metabolism-related pathways) associated with PCa aggressiveness, we tested 8587 SNPs for 20,729 cases from the PCa consortium. We identified 3 KLK3 SNPs, and 1083 (P < 3.5 × 10-9) and 3145 (P < 1 × 10-5) SNP-SNP interaction pairs significantly associated with PCa aggressiveness. These SNP pairs associated with PCa aggressiveness were more significant than each of their constituent SNP individual effects. The majority (98.6%) of the 3145 pairs involved KLK3. The 3 most common gene-gene interactions were KLK3-COL4A1:COL4A2, KLK3-CDH13, and KLK3-TGFBR3. Predictions from the SNP interaction-based polygenic risk score based on 24 SNP pairs are promising. The prevalence of PCa aggressiveness was 49.8%, 21.9%, and 7.0% for the PCa cases from our cohort with the top 1%, middle 50%, and bottom 1% risk profiles. Potential biological functions of the identified KLK3 SNP-SNP interactions were supported by gene expression and protein-protein interaction results. Our findings suggest KLK3 SNP interactions may play an important role in PCa aggressiveness.

Correction: Quantitative Computed Tomographic Descriptors Associate Tumor Shape Complexity and Intratumor Heterogeneity with Prognosis in Lung Adenocarcinoma.

[This corrects the article DOI: 10.1371/journal.pone.0118261.].

Alcohol Intake and Alcohol-SNP Interactions Associated with Prostate Cancer Aggressiveness.

Excessive alcohol intake is a well-known modifiable risk factor for many cancers. It is still unclear whether genetic variants or single nucleotide polymorphisms (SNPs) can modify alcohol intake's impact on prostate cancer (PCa) aggressiveness. The objective is to test the alcohol-SNP interactions of the 7501 SNPs in the four pathways (angiogenesis, mitochondria, miRNA, and androgen metabolism-related pathways) associated with PCa aggressiveness. We evaluated the impacts of three excessive alcohol intake behaviors in 3306 PCa patients with European ancestry from the PCa Consortium. We tested the alcohol-SNP interactions using logistic models with the discovery-validation study design. All three excessive alcohol intake behaviors were not significantly associated with PCa aggressiveness. However, the interactions of excessive alcohol intake and three SNPs (rs13107662 [CAMK2D, p = 6.2 × 10-6], rs9907521 [PRKCA, p = 7.1 × 10-5], and rs11925452 [ROBO1, p = 8.2 × 10-4]) were significantly associated with PCa aggressiveness. These alcohol-SNP interactions revealed contrasting effects of excessive alcohol intake on PCa aggressiveness according to the genotypes in the identified SNPs. We identified PCa patients with the rs13107662 (CAMK2D) AA genotype, the rs11925452 (ROBO1) AA genotype, and the rs9907521 (PRKCA) AG genotype were more vulnerable to excessive alcohol intake for developing aggressive PCa. Our findings support that the impact of excessive alcohol intake on PCa aggressiveness was varied by the selected genetic profiles.

A Novel Role of BIRC3 in Stemness Reprogramming of Glioblastoma.

Stemness reprogramming remains a largely unaddressed principal cause of lethality in glioblastoma (GBM). It is therefore of utmost importance to identify and target mechanisms that are essential for GBM stemness and self-renewal. Previously, we implicated BIRC3 as an essential mediator of therapeutic resistance and survival adaptation in GBM. In this study, we present novel evidence that BIRC3 has an essential noncanonical role in GBM self-renewal and stemness reprogramming. We demonstrate that BIRC3 drives stemness reprogramming of human GBM cell lines, mouse GBM cell lines and patient-derived GBM stem cells (GSCs) through regulation of BMP4 signaling axis. Specifically, BIRC3 induces stemness reprogramming in GBM through downstream inactivation of BMP4 signaling. RNA-Seq interrogation of the stemness reprogramming hypoxic (pseudopalisading necrosis and perinecrosis) niche in GBM patient tissues further validated the high BIRC3/low BMP4 expression correlation. BIRC3 knockout upregulated BMP4 expression and prevented stemness reprogramming of GBM models. Furthermore, siRNA silencing of BMP4 restored stemness reprogramming of BIRC3 knockout in GBM models. In vivo silencing of BIRC3 suppressed tumor initiation and progression in GBM orthotopic intracranial xenografts. The stemness reprograming of both GSCs and non-GSCs populations highlights the impact of BIRC3 on intra-tumoral cellular heterogeneity GBM. Our study has identified a novel function of BIRC3 that can be targeted to reverse stemness programming of GBM.

Accessible Pipeline for Translational Research Using TCGA: Examples of Relating Gene Mechanism to Disease-Specific Outcomes.

Bioinformatic scientists are often asked to do widespread analyses of publicly available datasets in order to identify genetic alterations in cancer for genes of interest; therefore, we sought to create a set of tools to conduct common statistical analyses of The Cancer Genome Atlas (TCGA) data. These tools have been developed in response to requests from our collaborators to ask questions, validate findings, and better understand the function of their gene of interest. We describe here what data we have used, how to obtain it, and what figures we have found useful.

TMPRSS2-ERG fusion impacts anterior tumor location in men with prostate cancer.

BACKGROUND: In prostate cancer (PCa), lack of androgen receptor (AR) regulated TMPRSS2-ETS-related gene (ERG) gene fusion (ERGnegative ) status has been associated with African American race; however, the implications of ERG status for the location of dominant tumors within the prostate remains understudied. METHODS: An African American-enriched multiinstitutional cohort of 726 PCa patients consisting of both African American men (AAM; n = 254) and European American men (EAM; n = 472) was used in the analyses. Methods of categorical analysis were used. Messenger RNA (mRNA) expression differences between anterior and posterior tumor lesions were analyzed using Wilcoxon rank-sum tests with multiple comparison corrections. RESULTS: Anti-ERG immunohistochemistry staining showed that the association between ERG status and anterior tumors is independent of race and is consistently robust for both AAM (ERGnegative 81.4% vs. ERGpositive 18.6%; p = .005) and EAM (ERGnegative 60.4% vs. ERGpositive 39.6%; p < .001). In a multivariable model, anterior tumors were more likely to be IHC-ERGnegative (odds ratio [OR]: 3.20; 95% confidence interval [CI]: 2.14-4.78; p < .001). IHC-ERGnegative were also more likely to have high-grade tumors (OR: 1.73; 95% CI: 1.06-2.82; p = .02). In the exploratory genomic analysis, mRNA expression of location-dependent genes is highly influenced by ERG status and African American race. However, tumor location did not impact the expression of AR or the major canonical AR-target genes (KLK3, AMACR, and MYC). CONCLUSIONS: ERGnegative tumor status is the strongest predictor of anterior prostate tumors, regardless of race. Furthermore, AR expression and canonical AR signaling do not impact tumor location.

Sirt2 Inhibition Enhances Metabolic Fitness and Effector Functions of Tumor-Reactive T Cells.

Dysregulated metabolism is a key driver of maladaptive tumor-reactive T lymphocytes within the tumor microenvironment. Actionable targets that rescue the effector activity of antitumor T cells remain elusive. Here, we report that the Sirtuin-2 (Sirt2) NAD+-dependent deacetylase inhibits T cell metabolism and impairs T cell effector functions. Remarkably, upregulation of Sirt2 in human tumor-infiltrating lymphocytes (TILs) negatively correlates with response to TIL therapy in advanced non-small-cell lung cancer. Mechanistically, Sirt2 suppresses T cell metabolism by targeting key enzymes involved in glycolysis, tricarboxylic acid-cycle, fatty acid oxidation, and glutaminolysis. Accordingly, Sirt2-deficient murine T cells exhibit increased glycolysis and oxidative phosphorylation, resulting in enhanced proliferation and effector functions and subsequently exhibiting superior antitumor activity. Importantly, pharmacologic inhibition of Sirt2 endows human TILs with these superior metabolic fitness and effector functions. Our findings unveil Sirt2 as an unexpected actionable target for reprogramming T cell metabolism to augment a broad spectrum of cancer immunotherapies.

Anti-tumor efficacy of plasmid encoding emm55 in a murine melanoma model.

Emm55 is a bacterial gene derived from Streptococcus pyogenes (S. pyogenes) that was cloned into a plasmid DNA vaccine (pAc/emm55). In this study, we investigated the anti-tumor efficacy of pAc/emm55 in a B16 murine melanoma model. Intralesional (IL) injections of pAc/emm55 significantly delayed tumor growth compared to the pAc/Empty group. There was a significant increase in the CD8+ T cells infiltrating into the tumors after pAc/emm55 treatment compared to the control group. In addition, we observed that IL injection of pAc/emm55 increased antigen-specific T cell infiltration into tumors. Depletion of CD4+ or CD8+ T cells abrogated the anti-tumor effect of pAc/emm55. Combination treatment of IL injection of pAc/emm55 with anti-PD-1 antibody significantly delayed tumor growth compared to either monotherapy. pAc/emm55 treatment combined with PD-1 blockade enhanced anti-tumor immune response and improved systemic anti-tumor immunity. Together, these strategies may lead to improvements in the treatment of patients with melanoma.