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Purpose: Glioblastoma (GBM) is an incurable primary brain tumor that has not benefited from immunotherapy to date. More than 90% of GBM expresses the tryptophan (Trp) metabolic enzyme, indoleamine 2,3-dioxygenase 1 (IDO). This observation supported the historical hypothesis that IDO suppresses the antitumor immune response solely through a mechanism that requires intratumoral Trp depletion. However, recent findings led us to investigate the alternative hypothesis that IDO suppresses the anti-GBM immune response independent of its association with Trp metabolism. Experimental Design: IDO-deficient GBM cell lines reconstituted with IDO wild-type or IDO enzyme–null cDNA were created and validated in vitro and in vivo. Microarray analysis was conducted to search for genes that IDO regulates, followed by the analysis of human GBM cell lines, patient GBM and plasma, and The Cancer Genome Atlas (TCGA) database. Ex vivo cell coculture assays, syngeneic and humanized mouse GBM models, were used to test the alternative hypothesis. Results: Nonenzymic tumor cell IDO activity decreased the survival of experimental animals and increased the expression of complement factor H (CFH) and its isoform, factor H like protein 1 (FHL-1) in human GBM. Tumor cell IDO increased CFH and FHL-1 expression independent of Trp metabolism. Increased intratumoral CFH and FHL-1 levels were associated with poorer survival among patients with glioma. Similar to IDO effects, GBM cell FHL-1 expression increased intratumoral regulatory T cells (Treg) and myeloid-derived suppressor cells while it decreased overall survival in mice with GBM. Conclusions: Our study reveals a nonmetabolic IDO-mediated enhancement of CFH expression and provides a new therapeutic target for patients with GBM.

Harnessing natural killer (NK) cell activity, either endogenous NK or NK-cell-based therapy, is in the forefront development for many tumor types. There are compelling clinical evidence demonstrating the significant relevance of loss of NK cell activity with prostate cancer (PCa) disease progression to the lethal phenotype, metastatic castration-resistant PCa (mCRPC). Given that mCRPC is generally regarded as a “cold” tumor and has only elicited marginal response to immune checkpoint blockade therapy to revive CD8 T cell activity, NK cell targeted therapy may present a viable opportunity not only to directly target mCRPC and also to “fire up” the cold mCRPC tumor. However, current efforts on harnessing NK cell activity for treating mCRPC is very limited with only one open NK-cell-based clinical trial. This minireview will present the clinical significance of NK cell activity in controlling metastatic PCa and provide perspectives on harnessing NK cell activity for treating mCRPC.

Background Melanoma patients who have detectable serum soluble NKG2D ligands either at the baseline or post-treatment of PD1/PDL1 blockade exhibit poor overall survival. Among families of soluble human NKG2D ligands, the soluble human MHC I chain-related molecule (sMIC) was found to be elevated in melanoma patients and mostly associated with poor response to PD1/PDL1 blockade therapy. Methods In this study, we aim to investigate whether co-targeting tumor-released sMIC enhances the therapeutic outcome of PD1/PDL1 blockade therapy for melanoma. We implanted sMIC-expressing B16F10 melanoma tumors into syngeneic host and evaluated therapeutic efficacy of anti-sMIC antibody and anti-PDL1 antibody combination therapy in comparison with monotherapy. We analyzed associated effector mechanism. We also assessed sMIC/MIC prevalence in metastatic human melanoma tumors. Results We found that the combination therapy of the anti-PDL1 antibody with an antibody targeting sMIC significantly improved animal survival as compared to monotherapies and that the effect of combination therapy depends significantly on NK cells. We show that combination therapy significantly increased IL-2Rα (CD25) on NK cells which sensitizes NK cells to low dose IL-2 for survival. We demonstrate that sMIC negatively reprograms gene expression related to NK cell homeostatic survival and proliferation and that antibody clearing sMIC reverses the effect of sMIC and reprograms NK cell for survival. We further show that sMIC/MIC is abundantly present in metastatic human melanoma tumors. Conclusions Our findings provide a pre-clinical proof-of-concept and a new mechanistic understanding to underscore the significance of antibody targeting sMIC to improve therapeutic efficacy of anti-PD1/PDL1 antibody for MIC/sMIC+ metastatic melanoma patients.

Expressed prostatic secretions (EPS), also called post digital rectal exam urines, are proximal fluids of the prostate that are widely used for diagnostic and prognostic assays for prostate cancer. These fluids contain an abundant number of glycoproteins and extracellular vesicles secreted by the prostate gland, and the ability to detect changes in their N-glycans composition as a reflection of disease state represents potential new biomarker candidates. Methods to characterize these N-glycan constituents directly from clinical samples in a timely manner and with minimal sample processing requirements are not currently available. In this report, an approach is described to directly profile the N-glycan constituents of EPS urine samples, prostatic fluids and urine using imaging mass spectrometry for detection. An amine reactive slide is used to immobilize glycoproteins from a few microliters of spotted samples, followed by peptide N-glycosidase digestion. Over 100 N-glycan compositions can be detected with this method, and it works with urine, urine EPS, prostatic fluids, and urine EPS-derived extracellular vesicles. A comparison of the N-glycans detected from the fluids with tissue N-glycans from prostate cancer tissues was done, indicating a subset of N-glycans present in fluids derived from the gland lumens. The developed N-glycan profiling is amenable to analysis of larger clinical cohorts and adaptable to other biofluids.

BackgroundAfrican Americans (AAs) have higher colorectal cancer (CRC) incidence and mortality rate than Caucasian Americans (CAs). Recent studies suggest that immune responses within CRCs contribute to the disparities. If racially distinct immune signatures are present in the early phases of carcinogenesis, they could be used to develop interventions to prevent or slow disease.MethodsWe selected a convenience sample of 95 patients (48 CAs, 47 AAs) with preinvasive colorectal adenomas from the surgical pathology laboratory at the Medical University of South Carolina. Using immunofluorescent-conjugated antibodies on tissue slides from the lesions, we quantified specific immune cell populations: mast cells (CD117+), Th17 cells (CD4+RORC+), and NK cell ligand (MICA/B) and inflammatory cytokines, including IL-6, IL-17A, and IFN-γ. We compared the mean density counts (MDCs) and density rate ratios (RR) and 95% CI of immune markers between AAs to CAs using negative binomial regression analysis. We adjusted our models for age, sex, clinicopathologic characteristics (histology, location, dysplasia), and batch.ResultsWe observed no racial differences in age or sex at the baseline endoscopic exam. AAs compared to CAs had a higher prevalence of proximal adenomas (66% vs. 40%) and a lower prevalence of rectal adenomas (11% vs. 23%) (p =0.04) but no other differences in pathologic characteristics. In age, sex, and batch adjusted models, AAs vs. CAs had lower RRs for cells labeled with IFNγ (RR 0.50 (95% CI 0.32-0.81); p=0.004) and NK cell ligand (RR 0.67 (0.43-1.04); p=0.07). In models adjusted for age, sex, and clinicopathologic variables, AAs had reduced RRs relative to CAs for CD4 (p=0.02), NK cell ligands (p=0.01), Th17 (p=0.005), mast cells (p=0.04) and IFN-γ (p< 0.0001).ConclusionsOverall, the lower RRs in AAs vs. CAs suggests reduced effector response capacity and an immunosuppressive (‘cold’) tumor environment. Our results also highlight the importance of colonic location of adenoma in influencing these differences; the reduced immune responses in AAs relative to CAs may indicate impaired immune surveillance in early carcinogenesis. Future studies are needed to understand the role of risk factors (such as obesity) in influencing differences in immune responses by race.

Black men die more often of prostate cancer yet, interestingly, may derive greater survival benefits from immune-based treatment with sipuleucel-T. Since no signatures of immune-responsiveness exist for prostate cancer, we explored race-based immune-profiles to identify vulnerabilities. Here we show in multiple independent cohorts comprised of over 1,300 patient samples annotated with either self-identified race or genetic ancestry, prostate tumors from Black men or men of African ancestry have increases in plasma cell infiltrate and augmented markers of NK cell activity and IgG expression. These findings are associated with improved recurrence-free survival following surgery and nominate plasma cells as drivers of prostate cancer immune-responsiveness., A recent report suggested Black men with prostate cancer were more responsive to immunotherapy. Here, the authors analysed prostate cancer gene expression profiles and show tumours from Black men and men with African ancestry have an increased proportion of plasma cells compared to those of White men and this correlates with improved outcome following treatment.

Background Advanced age is a major risk factor for the development of many diseases including those affecting the central nervous system. Wild-type isocitrate dehydrogenase glioblastoma (IDHwt GBM) is the most common primary malignant brain cancer and accounts for ≥90% of all adult GBM diagnoses. Patients with IDHwt GBM have a median age of diagnosis at 68–70 years of age, and increasing age is associated with an increasingly worse prognosis for patients with this type of GBM. Methods The Surveillance, Epidemiology, and End Results, The Cancer Genome Atlas, and the Chinese Glioma Genome Atlas databases were analyzed for mortality indices. Meta-analysis of 80 clinical trials was evaluated for log hazard ratio for aging to tumor survivorship. Results Despite significant advances in the understanding of intratumoral genetic alterations, molecular characteristics of tumor microenvironments, and relationships between tumor molecular characteristics and the use of targeted therapeutics, life expectancy for older adults with GBM has yet to improve. Conclusions Based upon the results of our analysis, we propose that age-dependent factors that are yet to be fully elucidated, contribute to IDHwt GBM patient outcomes.

Background & aims Tumor-infiltrating neutrophils (polymorphonuclear neutrophils [PMNs]) are a prominent feature of colorectal cancer (CRC), where they can promote cytotoxicity or exacerbate disease outcomes. We recently showed that in acute colon injury, PMNs can increase DNA double-strand break (DSB) burden and promote genomic instability via microRNA-dependent inhibition of homologous recombination (HR) repair. In this study, we aimed to establish whether in inflamed colon, neutrophils shape the DSB-repair responses to impact CRC progression and sensitivity/resistance to DNA-repair targeted therapy. Methods Human sporadic CRC biopsies, The Cancer Genome Atlas gene expression analyses, tumor xenografts, and murine CRC models, as well as small-molecule inhibition of key DSB-repair factors were leveraged to investigate changes in the DSB-repair landscape and identify unique CRC responses with/without tumor infiltration by PMNs. Results We reveal that neutrophils exert a functional dualism in cancer cells, driving temporal modulation of the DNA damage landscape and resolution of DSBs. PMNs were found to promote HR deficiency in low-grade CRC by miR-155-dependent downregulation of RAD51, thus attenuating tumor growth. However, neutrophil-mediated genotoxicity due to accumulation of DSBs led to the induction of non-homologous end-joining (NHEJ), allowing for survival and growth of advanced CRC. Our findings identified a PMN-induced HR-deficient CRC phenotype, featuring low RAD51 and low Ku70 levels, rendering it susceptible to synthetic lethality induced by clinically approved PARP1 inhibitor Olaparib. We further identified a distinct PMN-induced HR-deficient CRC phenotype, featuring high Ku70 and heightened NHEJ, which can be therapeutically targeted by specific inhibition of NHEJ. Conclusions Our work delineates 2 mechanism-based translatable therapeutic interventions in sporadic CRC.

BackgroundInflammatory bowel disease (IBD) has been implicated as a risk factor for prostate cancer, however, the mechanism of how IBD leads to prostate tumorigenesis is not known. Here, we investigated whether chronic intestinal inflammation leads to pro-inflammatory changes associated with tumorigenesis in the prostate.MethodsUsing clinical samples of men with IBD who underwent prostatectomy, we analyzed whether prostate tumors had differences in lymphocyte infiltrate compared to non-IBD controls. In a mouse model of chemically-induced intestinal inflammation, we investigated whether chronic intestinal inflammation could be transferred to the wild-type mouse prostate. In addition, mouse prostates were evaluated for activation of pro-oncogenic signaling and genomic instability.ResultsA higher proportion of men with IBD had T and B lymphocyte infiltration within prostate tumors. Mice with chronic colitis showed significant increases in prostatic CD45 + leukocyte infiltration and elevation of three pro-inflammatory cytokines-TIMP-1, CCL5, and CXCL1 and activation of AKT and NF-kB signaling pathways. Lastly, mice with chronic colitis had greater prostatic oxidative stress/DNA damage, and prostate epithelial cells had undergone cell cycle arrest.ConclusionsThese data suggest chronic intestinal inflammation is associated with an inflammatory-rich, pro-tumorigenic prostatic phenotype which may explain how gut inflammation fosters prostate cancer development in men with IBD.

Immune responses vary in colorectal cancers, which strongly influence prognosis. However, little is known about the variance in immune response within preinvasive lesions. The study aims to investigate how the immune contexture differs by clinicopathologic features (location, histology, dysplasia) associated with progression and recurrence in early carcinogenesis. We performed a cross-sectional study using preinvasive lesions from the surgical pathology laboratory at the Medical University of South Carolina. We stained the tissues with immunofluorescence antibodies, then scanned and analyzed expression using automated image analysis software. We stained CD117 as a marker of mast cells, CD4/RORC to indicate Th17 cells, MICA/B as a marker of NK-cell ligands, and also used antibodies directed against cytokines IL6, IL17A, and IFNγ. We used negative binomial regression analysis to compare analyte density counts by location, histology, degree of dysplasia adjusted for age, sex, race, and batch. All immune markers studied (except IL17a) had significantly higher density counts in the proximal colon than distal colon and rectum. Increases in villous histology were associated with significant decreases in immune responses for IL6, IL17a, NK ligand, and mast cells. No differences were observed in lesions with low- and high-grade dysplasia, except in mast cells. The lesions of the proximal colon were rich in immune infiltrate, paralleling the responses observed in normal mucosa and invasive disease. The diminishing immune response with increasing villous histology suggests an immunologically suppressive tumor environment. Our findings highlight the heterogeneity of the immune responses in preinvasive lesions, which may have implications for prevention strategies. Prevention Relevance: Our study is focused on immune infiltrate expression in preinvasive colorectal lesions; our results suggest important differences by clinicopathologic features that have implications for immune prevention research.

BackgroundInflammatory bowel disease (IBD) is an established risk factor for colorectal cancer. Recent reports suggesting IBD is also a risk factor for prostate cancer (PC) require further investigation.ObjectiveTo test the association between IBD with incident PC.Design, setting, and participantsWe studied 218,084 men in the population-based UK Biobank cohort, aged 40-69 at study entry between 2006 and 2010, with follow-up through mid-2015.Outcome measurements and statistical analysisWe assessed the association between IBD and subsequent PC using multivariable Cox regression analyses, adjusting for age at assessment, ethnic group, UK region, smoking status, alcohol drinking frequency, body mass index, Townsend Deprivation Index, family history of prostate cancer, and previous prostate-specific antigen testing.Results and limitationsMean age at study entry was 56 years, 94% of the men were white, and 1.1% (n=2,311) had a diagnosis of IBD. After a median follow-up of 78 months, men with IBD had an increased risk of PC (adjusted hazards ratio [aHR] = 1.31, 95% Confidence interval [CI] = 1.03-1.67, p = 0.029). Separately analyzing the IBD subtypes of ulcerative colitis (UC) and Crohn’s disease (CD), the association with PC was only among men with the former (UC; aHR = 1.47, 95% CI = 1.11-1.95, p=0.0070), and not the latter (CD; aHR 1.06, 95% CI = 0.63-1.80, p = 0.82). Results are limited by lack of data on frequency of health care interactions.ConclusionsIn a large-scale, prospective cohort study, we detected an association between IBD, and UC specifically, with incident PC diagnosis.Patient summaryThis study of over 200,000 men in the UK suggests that men with inflammatory bowel disease may be at a higher risk of prostate cancer than the general population.

Indoleamine 2, 3-dioxygenase 1 (IDO; IDO1; INDO) is a rate-limiting enzyme that metabolizes the essential amino acid, tryptophan, into downstream kynurenines. Canonically, the metabolic depletion of tryptophan and/or the accumulation of kynurenine is the mechanism that defines how immunosuppressive IDO inhibits immune cell effector functions and/or facilitates T cell death. Non-canonically, IDO also suppresses immunity through non-enzymic effects. Since IDO targeting compounds predominantly aim to inhibit metabolic activity as evidenced across the numerous clinical trials currently evaluating safety/efficacy in patients with cancer, in addition to the recent disappointment of IDO enzyme inhibitor therapy during the phase III ECHO-301 trial, the issue of IDO non-enzyme effects have come to the forefront of mechanistic and therapeutic consideration(s). Here, we review enzyme-dependent and -independent IDO-mediated immunosuppression as it primarily relates to glioblastoma (GBM); the most common and aggressive primary brain tumor in adults. Our group's recent discovery that IDO levels increase in the brain parenchyma during advanced age and regardless of whether GBM is present, highlights an immunosuppressive synergy between aging-increased IDO activity in cells of the central nervous system that reside outside of the brain tumor but collaborate with GBM cell IDO activity inside of the tumor. Because of their potential value for the in vivo study of IDO, we also review current transgenic animal modeling systems while highlighting three new constructs recently created by our group. This work converges on the central premise that maximal immunotherapeutic efficacy in subjects with advanced cancer requires both IDO enzyme- and non-enzyme-neutralization, which is not adequately addressed by available IDO-targeting pharmacologic approaches at this time.

INTRODUCTION AND OBJECTIVE:Inflammatory bowel disease (IBD) increases one’s risk of colorectal cancer (CRC). Recent data implicating IBD as a risk factor for prostate cancer (PC) requires external ...

INTRODUCTION AND OBJECTIVE:Recent epidemiological data has implicated inflammatory bowel disease (IBD) as a risk factor for clinically significant prostate cancer (PC). One key driver of malignant ...

Inflammatory bowel disease (IBD) is an established risk factor for colorectal cancer. Recent reports suggesting IBD is also a risk factor for prostate cancer (PC) require further investigation. We studied 218 084 men in the population-based UK Biobank cohort, aged 40 to 69 at study entry between 2006 and 2010, with follow-up through mid-2015. We assessed the association between IBD and subsequent PC using multivariable Cox regression analyses, adjusting for age at assessment, ethnic group, UK region, smoking status, alcohol drinking frequency, body mass index, Townsend Deprivation Index, family history of PC and previous prostate-specific antigen testing. Mean age at study entry was 56 years, 94% of the men were white, and 1.1% (n = 2311) had a diagnosis of IBD. After a median follow-up of 78 months, men with IBD had an increased risk of PC (adjusted hazard ratio [aHR] = 1.31, 95% confidence interval [CI] = 1.03-1.67, P = .029). The association with PC was only among men with the ulcerative colitis (UC; aHR = 1.47, 95% CI = 1.11-1.95, P = .0070), and not Crohn's disease (aHR 1.06, 95% CI = 0.63-1.80, P = .82). Results are limited by lack of data on frequency of health care interactions. In a large-scale, prospective cohort study, we detected an association between IBD, and UC specifically, with incident PC diagnosis.

Purpose: Wild-type isocitrate dehydrogenase–expressing glioblastoma (GBM) is the most common and aggressive primary brain tumor with a median age at diagnosis of ≥65 years. It accounts for approximately 90% of all GBMs and has a median overall survival (OS) of Experimental Design: (i) Clinical data: GBM patient datasets from The Cancer Genome Atlas, Northwestern Medicine Enterprise Data Warehouse, and clinical studies evaluating ICB were stratified by age and compared for OS. (ii) Animal models: young, middle-aged, and older adult wild-type and indoleamine 2,3 dioxygenase (IDO)-knockout syngeneic mice were intracranially engrafted with CT-2A or GL261 glioma cell lines and treated with or without CTLA-4/PD-L1 mAbs, or radiation, anti–PD-1 mAb, and/or a pharmacologic IDO enzyme inhibitor. Results: Advanced age was associated with decreased GBM patient survival regardless of treatment with ICB. The advanced age–associated increase of brain IDO expression was linked to the suppression of immunotherapeutic efficacy and was not reversed by IDO enzyme inhibitor treatment. Conclusions: Immunosuppression increases in the brain during advanced age and inhibits antiglioma immunity in older adults. Going forward, it will be important to fully understand the factors and mechanisms in the elderly brain that contribute to the decreased survival of older patients with GBM during treatment with ICB.

Background: Compared with Caucasian Americans (CAs), African Americans (AAs) with colorectal cancer have poorer survival, especially younger-age patients. A robust lymphocytic reaction within colorectal cancers is strongly associated with better survival, but whether immune response impacts the disparity in colorectal cancer survival is unknown. Methods: The study population was comprised of 211 histologically confirmed colorectal cancers at the Medical University of South Carolina (Charleston, SC; 159 CAs and 52 AAs) diagnosed between Jan 01, 2000, and June 30, 2013. We constructed a lymphocyte score based on blinded pathologic assessment of the four different types of lymphocytic reactions. Cox proportional hazards regression was used to evaluate the association between the lymphocyte score and risk of death by race. Results: Colorectal cancers in AAs (vs. CAs) had a stronger lymphocytic reaction at diagnosis. A high lymphocyte score (vs. the lowest) was associated with better survival in AAs [HR 0.19; 95% confidence interval (CI), 0.04–0.99] and CAs (HR 0.47; 95% CI, 0.15–1.45). AAs with no lymphocytic reaction (vs. other categories) had poor survival HR 4.48 (1.58–12.7) whereas no difference was observed in CAs. The risk of death in AAs (vs. CA) was more pronounced in younger patients (HR 2.92; 95% CI, 1.18–7.22) compared with older (HR 1.20; 95% CI, 0.54–2.67), especially those with lymphocytic poor colorectal cancers. Conclusions: The lymphocytic reaction in tumor impacted the racial disparity in survival. Impact: Our results confirm the importance of the lymphocytic score on survival and highlight the need to fully characterize the immune environment of colorectal cancers by race. Cancer Epidemiol Biomarkers Prev; 27(7); 755–61. ©2018 AACR.

NKG2D is an activating immune receptor expressed by NK and effector T cells. Induced expression of NKG2D ligand on tumor cell surface during oncogenic insults renders cancer cells susceptible to immune destruction. In advanced human cancers, tumor cells shed NKG2D ligand to produce an immune soluble form as a means of immune evasion. Soluble NKG2D ligands have been associated with poor clinical prognosis in cancer patients. Harnessing NKG2D pathway is considered a viable avenue in cancer immunotherapy over recent years. In this review, we will discuss the progress and perspectives.

OBJECTIVE Indoleamine 2,3-dioxygenase 1 (IDO; IDO1) is an immune checkpoint that’s characterized as a potent immunosuppressive mediator through its ability to metabolize tryptophan and wild-type IDH patient-resected glioblastoma (GBM) expresses IDO in ≥ 95% of cases. Recent findings from our group led us to investigate the alternative hypothesis that IDO possesses immunosuppressive effects that are independent of its associated metabolic activity. METHODS Murine GBM cell lines that overexpress either wild-type or enzyme-null IDO were created for in vivo characterization of IDO enzyme-independent immunosuppressive function. Microarray was conducted to identify human IDO expression-correlated genes, which were further investigated in human GBM cell lines, patient GBM tissues and plasma, as well as the TCGA database. Ex vivo cell co-culture assays and syngeneic mouse orthotopic GBM models were employed to study immunosuppressive mechanisms. RESULTS Here, we demonstrate that non-enzymic IDO activity decreases survival in experimental animals and increases the expression of immunosuppressive complement factor H (CFH) in human GBM. CFH mRNA levels positively correlate with those of IDO and many other immunosuppressive genes in patient resected GBM and can be applied as a prognostic marker in both lower grade gliomas and GBM. Similar to IDO, the increased expression of CFH in patient-resected glioma was positively correlated with an increased signature for regulatory T cells (Tregs) and myeloid-derived suppressive cells (MDSCs). High expression of CFH in tumor cells increases intratumoral Tregs levels and decreases overall survival in mice with GBM, while inducing tumor associated macrophage cell differentiation. CONCLUSIONS Here, we demonstrated that glioblastoma (GBM) cell IDO promotes the accumulation of intratumoral FoxP3+ regulatory T cells (Tregs) and tumor progression while decreasing overall survival - independent of IDO enzyme activity. Our study reveals a targetable non-metabolic IDO-dependent mechanism for future therapeutic intervention in patients with GBM.

African Americans (AAs) compared with European Americans (EAs) have poorer stage-specific survival from colorectal cancer (CRC). Recent reports have indicated that the racial difference in survival has worsened over time, especially among younger patients. To better characterize this association, we used population-based Surveillance, Epidemiology, and End Results registry data to evaluate the effect of race on stage IV CRC survival in patients aged 50 and ≥ 50 years.The population included 16,782 patients diagnosed with stage IV colon and rectal adenocarcinoma from January 1, 2004 and December 31, 2011. Cox proportional hazards regression was used to evaluate the association between race and other prognostic factors and the risk of death in each age group.Younger AAs compared with EAs had a greater prevalence of proximal CRC at diagnosis, a factor associated with a significantly greater risk of death in both races. Among patients 50 years old, AAs had a greater risk of death compared with EAs (hazard ratio, 1.35; 95% confidence interval, 1.20-1.51), which was attenuated in patients ≥ 50 years of age (hazard ratio, 1.10; 95% confidence interval, 1.04-1.16); P for interaction = .01.The results revealed poor overall survival for AAs compared with EAs, especially for those 50 years of age. The greater prevalence of proximal CRC at diagnosis among younger AAs (vs. EAs) might contribute to the racial difference in survival. Future studies are needed to understand how the colonic location affects the efficacy of treatment regimens.

GARP encoded by the Lrrc32 gene is the cell surface docking receptor for latent TGFβ, which is expressed naturally by platelets and regulatory T cells (Treg). Although Lrrc32 is amplified frequently in breast cancer, the expression and relevant functions of GARP in cancer have not been explored. Here, we report that GARP exerts oncogenic effects, promoting immune tolerance by enriching and activating latent TGFβ in the tumor microenvironment. We found that human breast, lung, and colon cancers expressed GARP aberrantly. In genetic studies in normal mammary gland epithelial and carcinoma cells, GARP expression increased TGFβ bioactivity and promoted malignant transformation in immunodeficient mice. In breast carcinoma–bearing mice that were immunocompetent, GARP overexpression promoted Foxp3+ Treg activity, which in turn contributed to enhancing cancer progression and metastasis. Notably, administration of a GARP-specific mAb limited metastasis in an orthotopic model of human breast cancer. Overall, these results define the oncogenic effects of the GARP–TGFβ axis in the tumor microenvironment and suggest mechanisms that might be exploited for diagnostic and therapeutic purposes. Cancer Res; 76(24); 7106–17. ©2016 AACR.

The need continues for new prostate cancer (PCa) biomarkers for use in early detection, recurrence and metastatic disease diagnostics, therapeutic monitoring, and prediction of treatment benefit. As the urology, radiology and oncology disciplines continue to evaluate how to minimize the overdiagnosis and overtreatment associated with current prostate specific antigen (PSA) serum tests [1], there has been significant emphasis placed on multiparametric magnetic resonance imaging (mpMRI) based detection of clinically significant PCa. It is used for multiple indications including stratification of patients into low, intermediate, and high risk groups that guide treatment regimens from active surveillance to radical prostatectomy. Although an important step forward, inherent limitations with the use of MR in the detection of clinically significant cancer are still present and contribute to both false positives and false negatives [2]. Therefore optimizing tissue biomarker discovery that can be used as “ground truth” for further optimization of clinical imaging protocols represents a significant unmet need in the field. As a potential alternative, genomic-based liquid biopsy targets of cell free DNA (cfDNA) and circulating tumor cells (CTC) have emerged [3]. A recent consensus report of clinical experts in circulating prostate cancer biomarkers highlights these needs, especially in regards to developing biomarkers for treatment decisions, efficacy and metastasis [4]. These opinions were all based on developing genomic-based cfDNA and CTC assays. Genomics-based classification and risk stratification have made great strides, but many limitations still exist due to the continued interpretation of complex genomic data, its relation to impact on clinical outcomes, and not every mutation may be clinically actionable. Therefore, employing complementary research strategies to better clarify the functional readout of the cancer genome, i.e, how pathways downstream of genome regulation enhance tumorigenesis, may lead to new approaches for diagnosis and treatment for advanced PCa, as well as new biomarker targets. One well-known functional readout of the cancer genome is metabolism. Carbohydrate, or sugar, metabolism, specifically glucose, has been documented as a hallmark of tumorigenesis that enhances cell energetics (e.g. ATP synthesis) and biosynthetic processes (e.g. nucleotide and amino acid synthesis) [5]. This provides the basis of fluorodeoxyglucose positron emission tomographic (FDG-PET) imaging where tumor FDG uptake inversely correlates with survival in numerous cancers, including prostate cancer [6]. However, carbohydrates also can be incorporated into multiple types of complex carbohydrate structures that comprise the cell surface glycocalyx. These glycans are represented by N-linked and O-linked glycoproteins, glycosphingolipids, and glycosaminoglycans attached to proteoglycans and can interact with multiple extracellular matrix (ECM) proteins in the stroma, many of which are also glycosylated, as well various cell types including the immune system [7]. The role of the stroma in promoting and supporting cancer progression is a major area of cancer research, including PCa. Malignant transformation of the ECM disrupts the homeostatic tissue microenvironment, altering tissue processes of adhesion, cell death, migration, and proliferation to promote tumor growth [8]. Overall, targeting carbohydrate utilization as a functional end-product of metabolism, combined with targeted analysis of ECM constituents, could lead to novel diagnostic and therapeutic biomarkers of PCa across the clinical spectrum of the disease. Herein, we propose a strategy for PCa biomarker discovery in tissues, that spatially registers tissue glycosylation and protein expression of tumor, ECM, and immune components by merging matrix assisted laser desorption ionization (MALDI) imaging mass spectrometry and clinical prostate multiparametric magnetic resonance imaging (mpMRI). Specifically, MRI scans that are performed prior to prostatectomy are used to generate histologic 3D printed molds that are used to section the prostate in the same orientation as the MRI scan [9]. This allows for the coregistration of MRI imaging biomarkers and histology. Once the tumor tissue is sectioned by the pathologist using these molds, it is formalin fixed and paraffin embedded. Molecular MALDI imaging mass spectrometry of glycans and ECM proteins from the same patient tissues are then performed on unstained slides that directly correspond to the clinical imaging slices (see Figure 1). Open in a separate window Figure 1. An integrated three dimensional multimodality work flow to evaluate new glycomic and extracellular matrix biomarker candidates for prostate cancer. A. Representative pre-prostatectomy MRI images for T2W (T2 Weighted Image) and DWI (Diffusion-weighted image). B. A representative 3D-mold from mpMRI used for preparing prostatectomy tissue co-registered with mpMRI coordinates. C. Gross pathology and standard H&E stained tissue section, with two representative MALDI-IMS images of tumor and stroma localized N-glycans and collagenase derived ECM peptides in the same tissue slice.

Prostate cancer (PCa) is the most common cancer in men, and the second leading cause of cancer related death in men in Western countries. The standard therapy for metastatic PCa is androgen suppression therapy (AST). Men undergoing AST eventually develop metastatic castration-resistant prostate cancer (mCRPC), of which there are limited treatment options available. Immunotherapy has presented substantial benefits for many types of cancer, but only a marginal benefit for mCRPC, at least in part, due to the immunosuppressive tumor microenvironment (TME). Current clinical trials are investigating monotherapies or combination therapies involving adoptive cellular therapy, viral, DNA vaccines, oncolytic viruses, and immune checkpoint inhibitors (ICI). Immunotherapies are also being combined with chemotherapy, radiation, and AST. Additionally, preclinical investigations show promise with the recent description of alternative ways to circumvent the immunosuppressive nature of the prostate tumor microenvironment, including harnessing the immune stimulatory NKG2D pathway, inhibiting myeloid derived suppressor cells, and utilizing immunomodulatory oncolytic viruses. Herein we provide an overview of recent preclinical and clinical developments in cancer immunotherapies and discuss the perspectives for future immunotherapies in PCa.

CD73 is a novel immune checkpoint associated with adenosine metabolism that promotes tumor progression by suppressing antitumor immune response and promoting angiogenesis. The inhibition of CD73, in combination with immune checkpoint blockade, targeted therapy or conventional therapy, improves antitumor effects in numerous preclinical mouse models of cancer. Emerging evidence suggests that the combination of anti-CD73 and immune checkpoint blockade has promising clinical activity in patients with advanced solid tumors. In this review, we will discuss the specific role of CD73 on both tumor cells and nontumor cells in regulating tumor immunity and tumorigenesis and provide an update on the current view of the antitumor activity of targeting CD73 by mAb or small molecule selective inhibitors in preclinical and clinical settings.

Cellular senescence is a tumor-suppressive mechanism that can paradoxically contribute to aging pathologies. Despite evidence of immune clearance in mouse models, it is not known how senescent cells (SnCs) persist and accumulate with age or in tumors in individuals. Here, we identify cooperative mechanisms that orchestrate the immunoevasion and persistence of normal and cancer human SnCs through extracellular targeting of natural killer receptor signaling. Damaged SnCs avoided immune recognition through MMP-dependent shedding of NKG2D ligands reinforced via paracrine suppression of NKG2D receptor–mediated immunosurveillance. These coordinated immunoediting processes were evident in residual, drug-resistant tumors from cohorts of more than 700 prostate and breast cancer patients treated with senescence-inducing genotoxic chemotherapies. Unlike in mice, these reversible senescence subversion mechanisms were independent of p53/p16 and exacerbated in oncogenic RAS-induced senescence. Critically, the p16(INK4A) tumor suppressor could disengage the senescence growth arrest from the damage-associated immune senescence program, which was manifest in benign nevus lesions, where indolent SnCs accumulated over time and preserved a non-proinflammatory tissue microenvironment maintaining NKG2D-mediated immunosurveillance. Our study shows how subpopulations of SnCs elude immunosurveillance and reveals potential secretome-targeted therapeutic strategies to selectively eliminate — and restore the clearance of — the detrimental SnCs that actively persist after chemotherapy and accumulate at sites of aging pathologies.

Persistent infection or chronic inflammation contributes significantly to tumourigenesis and tumour progression. C-X-C motif ligand 8 (CXCL8) is a chemokine that acts as an important multifunctional cytokine to modulate tumour proliferation, invasion and migration in an autocrine or paracrine manner. Studies have suggested that CXCL8 and its cognate receptors, C-X-C chemokine receptor 1 (CXCR1) and C-X-C chemokine receptor 2 (CXCR2), mediate the initiation and development of various cancers including breast cancer, prostate cancer, lung cancer, colorectal carcinoma and melanoma. CXCL8 also integrates with multiple intracellular signalling pathways to produce coordinated effects. Neovascularisation, which provides a basis for fostering tumour growth and metastasis, is now recognised as a critical function of CXCL8 in the tumour microenvironment. In this review, we summarize the biological functions and clinical significance of the CXCL8 signalling axis in cancer. We also propose that CXCL8 may be a potential therapeutic target for cancer treatment.

Cooperative therapeutic effect of anti-CTLA4 and anti-sMIC antibodies., Antibody therapy targeting cytotoxic T lymphocyte–associated antigen 4 (CTLA4) elicited survival benefits in cancer patients; however, the overall response rate is limited. In addition, anti-CTLA4 antibody therapy induces a high rate of immune-related adverse events. The underlying factors that may influence anti-CTLA4 antibody therapy are not well defined. We report the impact of a cancer-derived immune modulator, the human-soluble natural killer group 2D (NKG2D) ligand sMIC (soluble major histocompatibility complex I chain–related molecule), on the therapeutic outcome of anti-CTLA4 antibody using an MIC transgenic spontaneous TRAMP (transgenic adenocarcinoma of the mouse prostate)/MIC tumor model. Unexpectedly, animals with elevated serum sMIC (sMIChi) responded poorly to anti-CTLA4 antibody therapy, with significantly shortened survival due to increased lung metastasis. These sMIChi animals also developed colitis in response to anti-CTLA4 antibody therapy. Coadministration of an sMIC-neutralizing monoclonal antibody with the anti-CTLA4 antibody alleviated treatment-induced colitis in sMIChi animals and generated a cooperative antitumor therapeutic effect by synergistically augmenting innate and adoptive antitumor immune responses. Our findings imply that a new combination therapy could improve the clinical response to anti-CTLA4 antibody therapy. Our findings also suggest that prescreening cancer patients for serum sMIC may help in selecting candidates who will elicit a better response to anti-CTLA4 antibody therapy.

Background The causative role of the pro-inflammatory cytokine IL-6 in prostate cancer progression has been well established at molecular level. However, whether and how IL-6 may play a role in prostate cancer risk and development is not well defined. One limitation factor to acquiring this knowledge is the lack of appropriate animal models. Methods We generated a novel line of prostate-specific IL-6 transgenic mouse model. We compared the prostate pathology, tumorigenic signaling components, and prostate tumor microenvironment of the IL-6 transgenic mice with wild type littermates. Results With this model, we demonstrate that IL-6 induces prostate neoplasm autonomously. We further demonstrate that transgenic expression of IL-6 in the prostate activates oncogenic pathways, induces autocrine IL-6 secretion and steadily-state of STAT3 activation in the prostate tissue, upregulates paracrine insulin-like growth factor (IGF) signaling axis, reprograms prostate oncogenic gene expression, and more intriguingly, amplifies inflammation in the prostate and peri-prostatic adipose tissue. Conclusions The pro-inflammatory IL-6 is autonomous oncogene for the prostate. IL-6 induces prostate oncogenesis through amplifying local inflammation. We also presented a valuable animal model to study inflammation and prostate cancer development. Electronic supplementary material The online version of this article (doi:10.1186/s13045-016-0386-7) contains supplementary material, which is available to authorized users.

Background The SIX family homeobox genes have been demonstrated to be involved in the tumor initiation and progression, but their clinicopathological features and prognostic values in non-small cell lung cancer (NSCLC) have not been well defined. We analyzed relevant datasets and performed a systemic review and a meta-analysis to assess the profile of SIX family members in NSCLC and evaluate their importance as biomarkers for diagnosis and prediction of NSCLC. Methods This meta-analysis included 17 studies with 2358 patients. Hazard ratio (HR) and 95 % confidence interval (CI) were calculated to represent the prognosis of NSCLC with expression of the SIX family genes. Heterogeneity of the ORs and HRs was assessed and quantified using the Cochrane Q and I 2 test. Begg’s rank correlation method and Egger’s weighted regression method were used to screen for potential publication bias. Bar graphs of representative datasets were plotted to show the correlation between the SIX expression and clinicopathological features of NSCLC. Kaplan-Meier survival curves were used to validate our prognostic analysis by pooled HR. Results The systematic meta-analysis unveiled that the higher expressions of SIX1-5 were associated with the greater possibility of the tumorigenesis. SIX4 and SIX6 were linked to the lymph node metastasis (LNM). SIX2, SIX3, and SIX4 were correlated with higher TNM stages. Furthermore, the elevated expressions of SIX2, SIX4, and SIX6 predicted poor overall survival (OS) in NSCLC (SIX2: HR = 1.14, 95 % CI, 1.00–1.31; SIX4: HR = 1.39, 95 % CI, 1.16–1.66; SIX6: HR = 1.18, 95 % CI, 1.00–1.38) and poor relapse-free survival (RFS) in lung adenocarcinoma (ADC) (SIX2: HR = 1.42, 95 % CI, 1.14–1.77; SIX4: HR = 1.52, 95 % CI, 1.09–2.11; SIX6: HR = 1.25, 95 % CI, 1.01–1.56). Conclusions Our report demonstrated that the SIX family members play distinct roles in the tumorigenesis of NSCLC and can be potential biomarkers in predicting prognosis of NSCLC patients. Electronic supplementary material The online version of this article (doi:10.1186/s13045-016-0339-1) contains supplementary material, which is available to authorized users.

Glioblastoma (GBM) is resistant to most multimodal therapies. Clinical success of immune-checkpoint inhibitors (ICIs) has spurred interest in applying ICIs targeting CTLA4, PD1 or IDO1 against GBM. This amplifies the need to ascertain GBM's intrinsic susceptibility (or resistance) toward these ICIs, through clinical biomarkers that may also "guide and prioritize" preclinical testing. Here, we interrogated the TCGA and/or REMBRANDT human patient-cohorts to predict GBM's predisposition toward ICIs. We exploited various broad clinical biomarkers, including mutational or predicted-neoantigen burden, pre-existing or basal levels of tumor-infiltrating T lymphocytes (TILs), differential expression of immune-checkpoints within the tumor and their correlation with particular TILs/Treg-associated functional signature and prognostic impact of differential immune-checkpoint expression. Based on these analyses, we found that predictive biomarkers of ICI responsiveness exhibited inconsistent patterns in GBM patients, i.e., they either predicted ICI resistance (as compared with typical ICI-responsive cancer-types like melanoma, lung cancer or bladder cancer) or susceptibility to therapeutic targeting of CTLA4 or IDO1. On the other hand, our comprehensive literature meta-analysis and preclinical testing of ICIs using an orthotopic GL261-glioma mice model, indicated significant antitumor properties of anti-PD1 antibody, whereas blockade of IDO1 or CTLA4 either failed or provided very marginal advantage. These trends raise the need to better assess the applicability of ICIs and associated biomarkers for GBM.

Hepatocellular carcinoma (HCC) is a prevalent solid malignancy. Critically needed discovery of new therapeutics has been hindered by lack of an in vitro cell culture system that can effectively represent the in vivo tumor microenvironment. To address this need, a 3D in vitro HCC model was developed using a biocompatible, chitosan-alginate (CA) scaffold cultured with human HCC cell lines.The correlation between the cell function, such as secretion of growth factors and production of ECM in vitro, and the tumor growth and blood vessel recruitment in vivo was investigated.HCC cells grown on 3D CA scaffolds demonstrated morphological characteristics and increased expression of markers of highly malignant cells. Implantation of CA scaffolds cultured with human HCC cells in mice showed accelerated tumor growth. Histology revealed marked differences in morphology and organization of newly formed blood vessels between tumors produced by different pre-cultured conditions. Resistance to doxorubicin was significantly pronounced in CA scaffold-cultured HCC cells compared to 2D or Matrigel cultured HCC cells.This 3D model of HCC, with its ability to more closely mimic the in vivo tumor behavior, may serve as an invaluable model for study and application of novel anticancer therapeutics against HCC.

The insulin-like growth factor type I receptor (IGF-IR) has been suggested to play an important role in prostate cancer progression and possibly in the progression to androgen-independent (AI) disease. The term AI may not be entirely correct, in that recent data suggest that expression of androgen receptor (AR) and androgen-regulated genes is the primary association with prostate cancer progression after hormone ablation. Therefore, signaling through other growth factors has been thought to play a role in AR-mediated prostate cancer progression to AI disease in the absence of androgen ligand. However, existing data on how IGF-IR signaling interacts with AR activation in prostate cancer are conflicting. In this Prospect article, we review some of the published data on the mechanisms of IGF-IR/AR interaction and present new evidence that IGF-IR signaling may modulate AR compartmentation and thus alter AR activity in prostate cancer cells. Inhibition of IGF-IR signaling can result in cytoplasmic AR retention and a significant change in androgen-regulated gene expression. Translocation of AR from the cytoplasm to the nucleus may be associated with IGF-induced dephosphorylation. Since fully humanized antibodies targeting the IGF-IR are now in clinical trials, the current review is intended to reveal the mechanisms of potential therapeutic effects of these antibodies on AI prostate cancers.

The activating receptor NK cell group 2 member D (NKG2D) mediates antitumor immunity in experimental animal models. However, whether NKG2D ligands contribute to tumor suppression or progression clinically remains controversial. Here, we have described 2 novel lines of “humanized” bi-transgenic (bi-Tg) mice in which native human NKG2D ligand MHC class I polypeptide-related sequence B (MICB) or the engineered membrane-restricted MICB (MICB.A2) was expressed in the prostate of the transgenic adenocarcinoma of the mouse prostate (TRAMP) model of spontaneous carcinogenesis. Bi-Tg TRAMP/MICB mice exhibited a markedly increased incidence of progressed carcinomas and metastasis, whereas TRAMP/MICB.A2 mice enjoyed long-term tumor-free survival conferred by sustained NKG2D-mediated antitumor immunity. Mechanistically, we found that cancer progression in TRAMP/MICB mice was associated with loss of the peripheral NK cell pool owing to high serum levels of tumor-derived soluble MICB (sMICB). Prostate cancer patients also displayed reduction of peripheral NK cells and high sMIC levels. Our study has not only provided direct evidence in “humanized” mouse models that soluble and membrane-restricted NKG2D ligands pose opposite impacts on cancer progression, but also uncovered a mechanism of sMIC-induced impairment of NK cell antitumor immunity. Our findings suggest that the impact of soluble NKG2D ligands should be considered in NK cell–based cancer immunotherapy and that our unique mouse models should be valuable for therapy optimization.

The immune stimulatory cytokine interleukin-15 was recognized as one of the most promising cancer cure drug in an NIH guided review and is currently in clinical trial alone or as an adjuvant for certain types of metastatic solid tumors. IL-15 is an essential survival factor for natural killer (NK), natural killer-like T (NKT), and CD44hi memory CD8 T cells. The bioactivity of IL-15 in vivo is conferred mainly through a trans-presentation mechanism in which IL-15 is presented in complex with the α-subunit of soluble IL-15 receptor (IL-15R) to NK, NKT or T cells rather than directly interacts with membrane-bound IL-15R. With these understandings, recent studies have been focused on generating IL-15 agonists which consist of IL-15 and partial or whole of soluble IL-15R to improve its in vivo bioactivity. This minireview will summarize the key features of IL-15 as a potential cancer treatment cytokine and the most recent development of IL-15 agonists and preclinical studies. Critical milestones to translate the pre-clinical development to in-patients treatment are emphasized.

In response to oncogenic insult, human cells were induced to express a family of MHC I-chain related molecules A and B (MICA and MICB, generally termed MIC) on the surface which serve as the ligands for the activating immune receptor NKG2D expressed by all human NK, CD8 T, NKT, and subsets of γδ T cells. Theoretically, engagement of NKG2D by tumor cell surface MIC is deemed to signal and provoke the immune system to eliminate transformed cells. Clinically, almost all advanced tumors in cancer patients produce soluble MIC through proteolytic shedding mediated by metalloproteases, or by release in exosomes derived from the cell membrane. Tumor-derived sMIC is known to be highly immune suppressive and profoundly insults the immune system by downregulating receptor NKG2D expression on effector NK and T cells, driving the expansion of tumor-favoring myeloid suppression cells, skewing macrophages into alternatively activated phenotypes, and perturbing NK cell peripheral maintenance. High levels of serum sMIC significantly correlate with advanced diseases of many types of cancer. These observations clearly endorse sMIC to be a cancer immune therapeutic target. However, due to mice lack homologues to human MIC, this concept was not proven until our recent studies. Using a “humanized” MIC-transgenic spontaneous mouse model which recapitulates the NKG2D-mediated onco-immune dynamics of human cancer patients, we show that neutralizing circulating sMIC with a first-in-field non-blocking antibody B10 that does not block the interaction of MIC with NKG2D revamps endogenous innate and antigen-specific CD8 T cell responses and remodels tumor microenvironment to be more immune reactive. We show that therapy with the non-blocking sMIC-neutralizing antibody results in effectively debulk of primary tumor and elimination of metastasis, with no observed toxicity. Furthermore, we show that clearing sMIC with the first-in-class neutralizing antibody B10 also enhanced the efficacy of other cancer immunotherapeutic modalities, such as immune checkpoint blockade or adoptive cell-based therapy pre-clinically. Our study has launched a new avenue of cancer immunotherapy which can be readily translated into clinical trials. Citation Format: Jennifer D. Wu, Jinyu Zhang, Fahmin Basher, Mark Rubinstein. First-in-class antibody targeting soluble NKG2D ligand sMIC to enhance checkpoint cancer immunotherapy [abstract]. In: Proceedings of the Second CRI-CIMT-EATI-AACR International Cancer Immunotherapy Conference: Translating Science into Survival; 2016 Sept 25-28; New York, NY. Philadelphia (PA): AACR; Cancer Immunol Res 2016;4(11 Suppl):Abstract nr B024.

preoperative kidney volume in remaining kidney (p 0.022), diabetes mellitus (p 0.002), and preoperative GFR in remaining kidney (p 0.045) were significant predictors of lower GFR at 12 months postoperatively in radical nephrectomy group. Older age (p 0.001) and preoperative kidney volume in both kidney (p 0.037) were significant predictors of lower GFR at 12 months postoperatively in partial nephrectomy group. CONCLUSIONS: Kidney volume was correlated with renal function in radical or partial nephrectomy groups. Preoperative kidney volume is an independent predictor of lower GFR in patients who underwent radical or partial nephrectomy for renal cell carcinoma.

Recently, combined serological and molecular studies of autoantibodies have revealed that these antibodies play an important role in the normal function of the immune system and in the development of the B cell repertoire. Accordingly, we hypothesized that a homozygous deletion of a critical autoantibody-associated Ig variable (V) gene may alter the immune system and thus predispose the host to autoimmune disorders. Initial experiments revealed several restriction fragment length polymorphisms (RFLP) of the Humhv3005 gene, that is likely to encode heavy chains of rheumatoid factors, and the closely related 1.9III gene. By probing EcoR1-digested DNA with the Humhv3005/P1 probe, we found that one of the four major hybridizing bands was missing in approximately 20% of patients with either rheumatoid arthritis or systemic lupus erythematosus, but only 2% of normal subjects. To delineate the genetic basis of this polymorphism, we have now employed the PCR to amplify and analyze hv3005, 1.9III, and homologous genes in individuals with characteristic RFLP genotypes. Our results indicate that the human Vh gene repertoire contains several hv3005- and 1.9III-like genes, and that a complete deletion of the hv3005-like genes is relatively restricted to a subset of autoimmune patients. These findings provide initial evidence for deletion of developmentally regulated autoreactive V genes in autoimmune diseases.

DNA damage has been associated with prostate cancer risk. Men who were referred for initial prostate biopsy for elevated prostate-specific antigen or abnormal digital rectal examination are often found with no cancer but have a higher risk of developing prostate cancer than the general population of men in their lifetime. In this study, we investigated whether DNA damage is one of the factors that predispose these men referred for prostate biopsies to a higher risk of prostate cancer. We found significantly elevated levels of 8-oxo-2-deoxyguanosine immunoreactivity in the prostates of the referred men (n = 50) in comparison to the control prostates of men (n = 32) with no indication for referral for prostate biopsy. Twelve of these control men were healthy middle-aged men and 20 of them were older men whose conditions were diagnosed with bladder cancer but with normal serum prostate-specific antigen and digital rectal examination and no evidence of prostate disease. In all the 8-oxo-2-deoxyguanosine-positive prostates, we detected phosphorylation of the ataxia telangiectasia mutated kinase and expression of the immune-stimulatory molecule MIC in the prostate epithelium. These data suggest that: 1) oxidative DNA damage has occurred in the “referred” but pathologically normal prostates, indicating that these prostates may be subjected to genomic instability and eventually neoplastic transformation; 2) in response to DNA damage, two surveillance pathways, represented by ataxia telangiectasia mutated phosphorylation and induction of the NKG2D ligand MIC, were activated to prevent tumorigenesis.

Purpose: To determine the effect of inhibition of insulin-like growth factor-IR (IGF-IR) signaling with an antibody to the IGF-IR, A12, in conjunction with androgen withdrawal on prostate cancer progression in a human prostate xenograft model, LuCaP 35. Experimental Design: LuCaP 35 was implanted s.c. in severe combined immunodeficient mice. At the time of castration, mice were randomized to one of three groups. Group 1 was castrate only; group 2 received A12 40 mg/kg i.p. for 2 weeks beginning 1 week after castration; and group 3 received A12 40 mg/kg i.p. for 2 weeks beginning 2 weeks after castration. Results: In group 1, tumor volume decreased to 60% of the starting volume 4 weeks post-castration. In groups 2 and 3, tumor volumes nadired 6 weeks after castration at Conclusions: This study shows that the inhibition of IGF-IR enhances the effects of castration in prostate cancer. These effects are associated with a decrease in AR signaling and nuclear AR localization, and recurrence is associated with an increase in AR-regulated gene expression.

Human tumor-derived soluble NKG2D sMIC paralyzes the immune system through multiple pathways. Targeting soluble MIC with a nonblocking sMIC-neutralizing anti-MIC antibody effectuated and revamped endogenous innate and adoptive antitumor responses. Therapy induced regression of primary tumors and eliminated metastasis in preclinical models.

Expression of the MHC I-chain related molecules A and B (MICA/B) on epithelial cell surface in response to transformation or DNA damage can signal the immune system of the abnormality and thus initiate active immune surveillance by Natural Killer (NK) cells and cytotoxicity T cells. We and others have shown that malignant tumor cells can shed MICA/B to down regulate NKG2D expression and negatively impact NK and CD8 T cell function in cancer patients. Increased tumor-specific shedding of cell surface NKG2D ligand, MHC I chain related molecule (MIC), is associated with advanced stage and metastasis in many types of epithelial cancer. High serum levels of soluble MIC (sMIC) insults the immune system not only by down-regulating NKG2D expression on natural killer (NK) cells and effector T cells but also perturbing NK cell peripheral maintenance. Whether sMIC is an effective cancer therapeutic target has not been addressed due to the lack of specific sMIC-blocking reagent and clinically relevant pre-clinical animal models. Using a “humanized” clinically relevant spontaneous prostate carcinoma TRAMP/MIC bi-transgenic mouse model and multiple engineered syngeneic transplantable tumor models, we unprecedentedly show that therapy with a sMIC-specific monoclonal antibody induced rapid regression of primary tumors and metastasis without systemic toxicity. The therapy revamped a myriad of anti-tumor immune responses including NK cell homeostatic renewal and function, CD4 T cells to Th1 and Th17 responses. Using the well-defined melanoma model, we further show that anti-sMIC therapy augments Pmel-1 antigen-specific CD8 T cell responses. Notably, depletion of NK cells mitigated the therapeutic effect of anti-sMIC antibody and the effector function of both CD4 and CD8 T cells. We conclude that sMIC is an effective therapeutic target to potentiate innate and adoptive immune responses against MIC+ malignancies. Citation Format: Jennifer D. Wu, Fahmin Basher, Mark Rubinstein. Antibody targeting soluble NKG2D ligand sMIC induces regression of primary tumors and eliminates metastasis in multiple pre-clinical cancer models. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 2468. doi:10.1158/1538-7445.AM2015-2468

Purpose: A human type 1 insulin-like growth factor receptor antibody (A12) has been shown to effectively inhibit human xenograft tumor growth, including androgen-dependent and androgen-independent prostate tumors. Docetaxel, either as a single agent or combined with others, has shown a survival benefit in prostate cancer patients. Based on these data, we investigated the combined in vivo effect of A12 and docetaxel on human androgen-independent and osseous prostate tumor growth. Experimental Design: To study human androgen-independent prostate cancer model, LuCaP35V tumors were implanted s.c. into castrated severe combined immunodeficient mice. When tumors reached about 100 mm3, animals were treated with vehicle control docetaxel (10 or 20 mg/kg) and docetaxel in combination with A12 (40 μg/kg) for 4 weeks. To study human osseous prostate cancer model, LuCaP 23.1 tumors were implanted intratibiae. When serum prostate-specific antigen reached 5 to 10 ng/mL, treatments were initiated. Results: A12 markedly augmented the inhibition of docetaxel on tumor growth. When docetaxel is combined with A12, the inhibition of tumor growth continued after treatment cessation, which was associated with continued apoptosis and decreased proliferation of tumor cells. Gene expression profiles indicated that the posttreatment suppression of tumor growth may be due to enhanced negative regulation of cell cycle progression– and/or cell survival–associated genes, some of which have been shown to induce resistance to docetaxel. Conclusions: Our findings suggest that targeting type 1 insulin-like growth factor receptor can enhance the therapeutic effect of docetaxel on advanced prostate cancer. Our findings also suggest a potential mechanism to improve the treatment efficacy of docetaxel in prostate cancer.

Purpose: The type I insulin-like growth factor receptor (IGF-IR) and its ligands have been shown to play a critical role in prostate carcinoma development, growth, and metastasis. Targeting the IGF-IR may be a potential treatment for prostate cancer. A fully human monoclonal antibody, A12, specific to IGF-IR, has shown potent antitumor effects in breast, colon, and pancreatic cancers in vitro and in vivo. In this study, we tested the in vivo effects of A12 on androgen-dependent and androgen-independent prostate tumor growth. Experimental Design: Androgen-dependent LuCaP 35 and androgen-independent LuCaP 35V prostate tumors were implanted s.c. into intact and castrated severe combined immunodeficient mice, respectively. When tumor volume reached about 150 to 200 mm3, A12 was injected at 40 mg/kg body weight thrice a week for up to 5 weeks. Results: We find that A12 significantly inhibits growth of androgen-dependent LuCaP 35 and androgen-independent LuCaP 35V prostate xenografts, however, by different mechanisms. In LuCaP 35 xenografts, A12 treatment induces tumor cell apoptosis or G1 cycle arrest. In LuCaP 35V xenografts, A12 treatment induces tumor cell G2-M cycle arrest. Moreover, we find that blocking the function of IGF-IR down-regulates androgen-regulated gene expression in androgen-independent LuCaP 35V tumor cells. Conclusions: Our findings suggest that A12 is a therapeutic candidate for both androgen-dependent and androgen-independent prostate cancer. Our findings also suggest an IGF-IR–dependent activity of the androgen receptor in androgen-independent prostate cancer cells.

The Retinal Determination Gene Network (RDGN), consisting of dachshund (dac), eyes absent (eya), and sin oculis (so), is best known for its role in eye specification. Recently, coordinate abnormal expression of the DACH1/Eya/Six complex has been observed in human tumors supporting a tumor promoting role of Six and Eya family members, and a tumor-suppressor function of DACH. Oncogenes sustain host-cell interactions, promoting invasiveness, recruiting inflammatory mediators and developing angiogenic networks. Ras and Myc promote tumor angiogenesis through up-regulation of VEGF and the inflammatory mediator, interleukin-8(IL-8) and interleukin-6 (IL-6). Altered expressions of the IL-6 and IL-8 genes are seen in many cancers including prostate cancer. Hormone-resistant prostate carcinoma cell lines produce large amounts of IL-6 and IL-8 compared to hormone responsive prostate cancer cell lines. The expression of IL-8 and IL-6 are linked to poor prognosis and metastatic phenotype in prostate cancer. Recent finding demonstrated that IL-6 regulates the dynamic equilibrium between cancer stem cells and non-stem cancer cells in human breast and prostate cancer. Moreover, IL-6 alone can transform non-tumorigenic benign prostate epithelial cells and further progress to invasive phenotype. We had previously reported that the expression of DACH1 was reduced in human prostate cancer and DACH1 inhibited androgen signaling transduction. To explore the role of DACH1 in androgen resistant prostate cancer, PC3 cells stably expressing DACH1 were established. DACH1 inhibited cellular proliferation evaluated by MTT assay and growth curve. Microarray analyses demonstrated DACH1 inhibited mRNA expression of GRO, IL-6, IL-8 by 75%, 80% and 90%, respectively. However, there was no change of mRNA expression for IL-6 and IL-8 receptor. Ectopic expression of DACH1 in PC-3 cells significantly decreased tumor growth compared with vector control group (p Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr LB-16. doi:1538-7445.AM2012-LB-16

In Drosophila, eye development is coordinately regulated by the retinal determination gene network (RDGN), mainly consisting of dachshund (dac), eyes absent (eya), and sin oculis (so). The mammalian homologues are Dach, Eya, and Six, respectively. RDGN, although best known for its role in eye specification, is essential for the development of many organs including retinal, kidney, brain, and limb in both flies and mammals. Recently, coordinate abnormal expression of the DACH1/Eya/Six complex has been observed in human tumors supporting a tumor promoting role of Six and Eya family members, and a tumor-suppressor function of DACH. Oncogenes sustain host-cell interactions, promoting invasiveness, recruiting inflammatory mediators and developing angiogenic networks. Ras and Myc promote tumor angiogenesis through up-regulation of VEGF and the inflammatory mediator, interleukin-8(IL-8) and interleukin-6 (IL-6). Our previous studies demonstrate DACH1 inhibits migration of MCF10A cells transformed with distinct oncogenes (Ha-Ras, c-Raf, c-Myc, and ErbB2) as well as the highly metastatic MDA-MB-231 human breast cancer cell line. An unbiased proteomic approach to identify candidate proteins regulated by DACH1 expression, using antibody array analysis, demonstrated DACH1 expression correlated with reduced secretion of IL-8 and the related chemokines IL-1, IL-6, GRO, and MIP1 (α, β, γ) from Ras or c-Myc transformed MCF-10A cells and repressed IL-8 promoter activity. Altered expressions of the IL-6 and IL-8 genes are seen in many cancers including prostate cancer. Hormone-resistant prostate carcinoma cell lines produce large amounts of IL-6 and IL-8 compared to hormone responsive prostate cancer cell lines. The expression of IL-8 and IL-6 are linked to poor prognosis and metastatic phenotype in prostate cancer. Recent finding demonstrated that IL-6 regulates the dynamic equilibrium between cancer stem cells and non-stem cancer cells in human breast and prostate cancer. IL-6 alone can transform non-tumorigenic benign prostate epithelial cells and further progress to invasive phenotype. To explore the role of DACH1 in androgen resistant prostate cancer, PC3 cells stably expressing DACH1 were established. DACH1 inhibited proliferation evaluated by MTT assay and growth curve. Microarray analyses demonstrated DACH1 inhibited mRNA expression of GRO, IL-6, IL-8 by 75%, 80% and 90%, respectively. However, there was no change of mRNA expression for IL-6 and IL-8 receptor. It has been reported that stable expression IL-6 in SV40T immortalized non-tumorigenic prostate epithelial cell line p69 induced EMT and acquired malignant phenotypes through autocrine loop IL6 and IGF-1R signaling. To further test the functional interaction of DACH1 with IL-6, a table cells expressing IL-6 alone or IL-6 with DACH1 were deployed. Ectopic expression of DACH1 attenuated activation of IGF-1 R by IL-6 as evaluated by phosphorylation of IGF-1R. Subcutaneous implantation study showed a 60% inhibition of in vivo tumor growth by DACH1. Together, our studies suggest that loss expression of DACH1 is an important cause of prostate tumorigenesis and restoration of DACH1 expression provides a potential therapeutic application. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the Second AACR International Conference on Frontiers in Basic Cancer Research; 2011 Sep 14-18; San Francisco, CA. Philadelphia (PA): AACR; Cancer Res 2011;71(18 Suppl):Abstract nr A21.

Background: Prostate cancer is the most common non-cutaneous malignant neoplasm in men in Western countries. Several studies have shown a significant correlation between the formation of a tumor and chronic prostatic inflammation, generally triggered by infectious agents or exposure to environmental factors (smoking, alcohol, burned meat, etc.), and/or various combination thereof. The incidence of chronic inflammation of the prostate is associated with the presence of inflammatory cells involved in the secretion of cytokines, which promote the migration and proliferation of cells implicated in the immune response. Our main objective of this project is to elucidate the mechanism involved in the promotion of prostate cancer mediated by IL-6 using different approaches such as Western Blots, qPCR, luciferase based reporter assays and immunofluorescence. Results: The effects of cytokines on prostate epithelial cells are not well understood, however some evidence showed that its presence may play a role in the initiation of prostate cancer of patients with chronic prostate inflammation. Preliminary data generated in our laboratory have shown that when the prostate cell lines p69 and BPH-1 (non- tumorigenic prostate cell lines) are treated with the cytokines IL-6 and TNF-α, their proliferation rate is higher when compared with non-treated cells. In addition we have observed that P69 cells cultured in presence of IL-6 suffer significant changes in their morphology as well as changes in the distribution of E-cadherin in the cell membrane. The activation of the MAPK pathway also was detected after the cytokine treatment but when we used the MAPK inhibitors (LY294002 and PD98059) the expression levels of E-cadherin and Vimentin were closed to those observed on non-treated cells. Previous reports have shown that the Snail transcription factor is involved in the regulation of the expression of E-cadherin and Vimentin. Interestingly, we have detected this transcription higher levels of this transcription factor (mRNA as well as protein) after stimulation of p69 and BPH-1 cells with IL-6. The presence of this transcription factor was not noticed in non-treated cells. Finally, we have performed a luciferase based assay to establish the occurrence of cell transformation and we have found a higher activation of the luciferase reporter in these cells when they were treated with IL-6. Interestingly, when we block the activity of the IGF-IR using the antibody A12, we have a reduction of the EMT phenotype observed in cells that were treated only with IL-6. Low effects of the antibody were detected in non-treated cells. It is possible that such modifications might help to facilitate epithelial cell invasion into the stromal and vasculature compartments. In summary, we are showing novel mechanisms by which IL-6 may play a key role in the initiation of prostate cancer in patients with chronic prostate inflammation. To have a better understanding of this event may help in the early treatment of this type of cancer. Further “in vivo” experiments will be performed to establish the importance of IL-6 in the development of tumors. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr LB-373.