Oncogenic Role of THOR, a Conserved Cancer/Testis Long Non-coding RNA.

Large-scale transcriptome sequencing efforts have vastly expanded the catalog of long non-coding RNAs (lncRNAs) with varying evolutionary conservation, lineage expression, and cancer specificity. Here, we functionally characterize a novel ultraconserved lncRNA, THOR (ENSG00000226856), which exhibits expression exclusively in testis and a broad range of human cancers. THOR knockdown and overexpression in multiple cell lines and animal models alters cell or tumor growth supporting an oncogenic role. We discovered a conserved interaction of THOR with IGF2BP1 and show that THOR contributes to the mRNA stabilization activities of IGF2BP1. Notably, transgenic THOR knockout produced fertilization defects in zebrafish and also conferred a resistance to melanoma onset. Likewise, ectopic expression of human THOR in zebrafish accelerated the onset of melanoma. THOR represents a novel class of functionally important cancer/testis lncRNAs whose structure and function have undergone positive evolutionary selection.

A CXCR4 inhibitor (balixafortide) enhances docetaxel-mediated antitumor activity in a murine model of prostate cancer bone metastasis.

BACKGROUND: Prostate cancer (PCa) bone metastases have been shown to be more resistant to docetaxel than soft tissue metastases. The proinflammatory chemokine receptor CXCR4 has been shown to confer resistance to docetaxel (DOC) in PCa cells. Balixafortide (BLX) is a protein epitope mimetic inhibitor of CXCR4. Accordingly, we hypothesized that BLX would enhance DOC-mediated antitumor activity in PCa bone metastases. METHODS: PC-3 luciferase-labeled cells were injected into the tibia of mice to model bone metastases. Four treatment groups were created: vehicle, DOC (5 mg/kg), BLX (20 mg/kg), and combo (receiving both DOC and BLX). Mice were injected twice daily subcutaneously with either vehicle or BLX starting on Day 1 and weekly intraperitoneally with DOC starting on Day 1. Tumor burden was measured weekly via bioluminescent imaging. At end of study (29 days), radiographs were taken of the tibiae and blood was collected. Serum levels of TRAcP, IL-2, and IFNγ levels were measured using ELISA. Harvested tibiae were decalcified and stained for Ki67, cleaved caspase-3, and CD34 positive cells or microvessels were quantified. RESULTS: Tumor burden was lower in the combo group compared to the DOC alone group. Treatment with the combination had no impact on the number of mice with osteolytic lesions, however the area of osteolytic lesions was lower in the combo group compared to the vehicle and BLX groups, but not the DOC group. Serum TRAcP levels were lower in the combo compared to vehicle group, but not the other groups. No significant difference in Ki67 staining was found among the groups; whereas, cleaved caspase-3 staining was lowest in the Combo group and highest in the BLX group. The DOC and combo groups had more CD34+ microvessels than the control and BLX groups. There was no difference between the treatment groups for IL-2, but the combo group had increased levels of IFNγ compared to the DOC group. CONCLUSIONS: Our data demonstrate that a combination of BAL and DOC has greater antitumor activity in a model of PCa bone metastases than either drug alone. These data support further evaluation of this combination in metastatic PCa.

Transcription factor network analysis based on single cell RNA-seq identifies that Trichostatin-a reverses docetaxel resistance in prostate Cancer.

BACKGROUND: Overcoming drug resistance is critical for increasing the survival rate of prostate cancer (PCa). Docetaxel is the first cytotoxic chemotherapeutical approved for treatment of PCa. However, 99% of PCa patients will develop resistance to docetaxel within 3 years. Understanding how resistance arises is important to increasing PCa survival. METHODS: In this study, we modeled docetaxel resistance using two PCa cell lines: DU145 and PC3. Using the Passing Attributes between Networks for Data Assimilation (PANDA) method to model transcription factor (TF) activity networks in both sensitive and resistant variants of the two cell lines. We identified edges and nodes shared by both PCa cell lines that composed a shared TF network that modeled changes which occur during acquisition of docetaxel resistance in PCa. We subjected the shared TF network to connectivity map analysis (CMAP) to identify potential drugs that could disrupt the resistant networks. We validated the candidate drug in combination with docetaxel to treat docetaxel-resistant PCa in both in vitro and in vivo models. RESULTS: In the final shared TF network, 10 TF nodes were identified as the main nodes for the development of docetaxel resistance. CMAP analysis of the shared TF network identified trichostatin A (TSA) as a candidate adjuvant to reverse docetaxel resistance. In cell lines, the addition of TSA to docetaxel enhanced cytotoxicity of docetaxel resistant PCa cells with an associated reduction of the IC50 of docetaxel on the resistant cells. In the PCa mouse model, combination of TSA and docetaxel reduced tumor growth and final weight greater than either drug alone or vehicle. CONCLUSIONS: We identified a shared TF activity network that drives docetaxel resistance in PCa. We also demonstrated a novel combination therapy to overcome this resistance. This study highlights the usage of novel application of single cell RNA-sequencing and subsequent network analyses that can reveal novel insights which have the potential to improve clinical outcomes.

Therapeutic targeting of BET bromodomain proteins in castration-resistant prostate cancer.

Men who develop metastatic castration-resistant prostate cancer (CRPC) invariably succumb to the disease. Progression to CRPC after androgen ablation therapy is predominantly driven by deregulated androgen receptor (AR) signalling. Despite the success of recently approved therapies targeting AR signalling, such as abiraterone and second-generation anti-androgens including MDV3100 (also known as enzalutamide), durable responses are limited, presumably owing to acquired resistance. Recently, JQ1 and I-BET762 two selective small-molecule inhibitors that target the amino-terminal bromodomains of BRD4, have been shown to exhibit anti-proliferative effects in a range of malignancies. Here we show that AR-signalling-competent human CRPC cell lines are preferentially sensitive to bromodomain and extraterminal (BET) inhibition. BRD4 physically interacts with the N-terminal domain of AR and can be disrupted by JQ1 (refs 11, 13). Like the direct AR antagonist MDV3100, JQ1 disrupted AR recruitment to target gene loci. By contrast with MDV3100, JQ1 functions downstream of AR, and more potently abrogated BRD4 localization to AR target loci and AR-mediated gene transcription, including induction of the TMPRSS2-ERG gene fusion and its oncogenic activity. In vivo, BET bromodomain inhibition was more efficacious than direct AR antagonism in CRPC xenograft mouse models. Taken together, these studies provide a novel epigenetic approach for the concerted blockade of oncogenic drivers in advanced prostate cancer.

Targeted Notch1 inhibition with a Notch1 antibody, OMP-A2G1, decreases tumor growth in two murine models of prostate cancer in association with differing patterns of DNA damage response gene expression.

Notch plays a protumorigenic role in many cancers including prostate cancer (PCa). Global notch inhibition of multiple Notch family members using γ-secretase inhibitors has shown efficacy in suppressing PCa growth in murine models. However, global Notch inhibition is associated with marked toxicity due to the widespread function of many different Notch family members in normal cell physiology. Accordingly, in the current study, we explored if specific inhibition of Notch1 would effectively inhibit PCa growth in a murine model. The androgen-dependent VCaP and androgen-independent DU145 cell lines were injected subcutaneously into mice. The mice were treated with either control antibody 1B7.11, anti-Notch1 antibody (OMP-A2G1), docetaxel or the combination of OMP-A2G1 and docetaxel. Tumor growth was measured using calipers. At the end of the study, tumors were assessed for proliferative response, apoptotic response, Notch target gene expression, and DNA damage response (DDR) expression. OMP-A2G1 alone inhibited tumor growth of both PCa cell lines to a greater extent than docetaxel alone. There was no additive or synergistic effect of OMP-A2G1 and docetaxel. The primary toxicity was weight loss that was controlled with dietary supplementation. Proliferation and apoptosis were affected differentially in the two cell lines. OMP-A2G1 increased expression of the DDR gene GADD45α in VCaP cells but downregulated GADD45α in Du145 cells. Taken together, these data show that Notch1 inhibition decreases PCa xenograft growth but does so through different mechanisms in the androgen-dependent VCaP cell line vs the androgen-independent DU145 cell line. These results provide a rationale for further exploration of targeted Notch inhibition for therapy of PCa.

Raf kinase inhibitor protein (RKIP) deficiency decreases latency of tumorigenesis and increases metastasis in a murine genetic model of prostate cancer.

BACKGROUND: Raf kinase inhibitor protein (RKIP) has been shown to act as a metastasis suppressor gene in multiple models of cancer. Loss of RKIP expression promotes invasion and metastasis in cell transplantation animal models. However, it is unknown if RKIP expression can impact the progression of cancer in an autochthonous model of cancer. The goal of this study was to determine if loss of RKIP expression in a genetic mouse model of prostate cancer (PCa) impacts metastasis. METHODS: Endogenous RKIP expression was measured in the primary tumors and metastases of transgenic adenocarcinoma of the mouse prostate (TRAMP(+) ) mice. RKIP knockout mice (RKIP(-/-) ) were crossbred with (TRAMP(+) ) mice to create RKIP(-/-) TRAMP(+) mice. Mice were euthanized at 10, 20, and 30 weeks for evaluation of primary and metastatic tumor development. To determine if loss of RKIP alone promotes metastasis, RKIP was knocked down in the low metastatic LNCaP prostate cancer cell line. RESULTS: Endogenous RKIP expression decreased in TRAMP(+) mice as tumors progressed. Primary tumors developed earlier in RKIP(-/-) TRAMP(+) compared to TRAMP(+) mice. At 30 weeks of age, distant metastases were identified only the RKIP(-/-) TRAMP(+) mice. While prostate epithelial cell proliferation rates were higher at 10 and 20 weeks in RKIP(-/-) TRAMP(+) compared to TRAMP(+) mice, by 30 weeks there was no difference. Apoptosis rates in both groups were similar at all timepoints. Decreased RKIP expression did not impact the metastatic rate of LNCaP in an orthotopic PCa model. CONCLUSIONS: These results demonstrate that loss of RKIP decreases latency of tumor development and promotes distant metastasis in the TRAMP mouse model in the context of a pro-metastatic background; but loss of RKIP alone is insufficient to promote metastasis. These findings suggest that in addition to its known metastasis suppressor activity, RKIP may promote tumor progression through enhancing tumor initiation. Prostate 75:292-302, 2015. © 2014 Wiley Periodicals, Inc.

Raf kinase inhibitor protein (RKIP) in cancer.

Raf kinase inhibitory protein (RKIP) was initially identified as phosphatidylethanolamine binding protein in bovine brain. It was later identified as a protein that inhibits Raf kinase activation of MEK. Further exploration has revealed that RKIP modulates several other signaling pathways including NF-κB and G-protein signaling. A gene array screen revealed that RKIP expression was low in a metastatic compared with non-metastatic prostate cancer cell line. Further experiments revealed that RKIP fits the criteria for a metastasis suppressor gene. RKIP expression has been shown to be downregulated in metastatic tissues, compared with non-metastatic tissue in multiple cancers, suggesting that loss of RKIP metastasis suppressor activity is a broad mechanism leading to metastasis. Additionally, loss of RKIP has been shown to impact therapy through conferring radioresistance and chemoresistance. Taken together, these data indicate understanding RKIP's contributions to cancer may lead to important therapeutic strategies to prevent metastasis and promote therapeutic efficacy.

Cilengitide (EMD 121974, NSC 707544) in asymptomatic metastatic castration resistant prostate cancer patients: a randomized phase II trial by the prostate cancer clinical trials consortium.

BACKGROUND: Integrins are involved in prostate cancer metastasis by regulating cell adhesion, migration, invasion, motility, angiogenesis and bone metabolism. We evaluated the efficacy of two dose levels of cilengitide in patients (pts) with castrate resistant prostate cancer (CRPC). METHODS: Chemotherapy-naïve, asymptomatic metastatic CRPC pts were randomized to cilengitide 500 mg or 2,000 mg IV twice weekly using parallel 2-stage design. The primary endpoint was rate of objective clinical progression at 6-months. Secondary endpoints included clinical and PSA response rates, safety and effects of cilengitide treatment on circulating tumor cells (CTCs) and bone remodeling markers. RESULTS: Forty-four pts were accrued to first stage (22/arm). Median number of cycles was three in both arms (500 mg arm: 1-8; 2,000 mg arm: 1-15). At 6 months, two pts (9%) on the 500 mg arm and five pts (23%) on the 2,000 mg arm had not progressed. Best objective response was stable disease (SD) in seven pts for 9.9[8.1,20.9] months. There were three grade 3 and no grade 4 toxicities. At 12 weeks, analysis of bone markers did not reveal significant trends. At progression, bone specific alkaline phosphatase and N-telopeptide increased in all pts, less so in pts on the 2,000 mg arm and in pts on both arms who obtained SD at 6 months. CTCs increased over time in both arms. CONCLUSION: Cilengitide was well tolerated with modest clinical effect in favor of the higher dose. The unique trial design including a shift from response rate to objective progression as the endpoint, and not acting on PSA increases was feasible.

Effects of Analgesics on Tumor Growth in Mouse Models of Prostate Cancer Bone Metastasis.

Murine models of tumor development often require invasive procedures for tumor implantation, potentially causing pain or distress. However, analgesics are often withheld during implantation because of concerns that they may adversely affect tumor development. Previous studies examining the effects of analgesics on the development and metastasis of various tumor lines show that the effect of analgesics depends on the tumor line and analgesic used. A blanket statement that analgesics affect the general growth of tumors is not adequate scientific justification for withholding pain relief, and pilot studies or references are recommended for each specific tumor cell line and treatment combination. In this study, we evaluated the effects of 2 commonly used analgesics on tumor growth in 2 models of prostate cancer (PCa) bone metastasis. We hypothesized that a one-time injection of analgesics at the time of intratibial injection of tumor cells would not significantly impact tumor growth. Either C57BL/6 or SCID mice were injected subcutaneously with an analgesic (carprofen [5 mg/kg], or buprenorphine [0.1 mg/kg]) or vehicle (0.1 mL of saline) at the time of intratibial injection with a PCa cell line (RM1 or PC3, n = 10 to 11 per group). Tumor growth (measured by determination of tumor burden and the extent of bone involvement) and welfare (measured by nociception, locomotion, and weight) were monitored for 2 to 4 wk. Neither carprofen or buprenorphine administration consistently affected tumor growth or indices of animal welfare as compared with the saline control for either cell line. This study adds to the growing body of literature demonstrating that analgesia can be compatible with scientific objectives, and that a decision to withhold analgesics must be scientifically justified and evaluated on a model-specific basis.

Androgen ablation augments human HLA2.1-restricted T cell responses to PSA self-antigen in transgenic mice.

BACKGROUND: In recent years, there has been an increasing interest in targeting human prostate tumor-associated antigens (TAAs) for prostate cancer immunotherapy as an alternative to other therapeutic modalities. However, immunologic tolerance to TAA poses a significant obstacle to effective, TAA-targeted immunotherapy. We sought to investigate whether androgen deprivation would result in circumventing immune tolerance to prostate TAA by impacting CD8 cell responses. METHODS: To this end, we generated a transgenic mouse that expresses the human prostate-specific antigen (PSA) specifically in the prostate, and crossed it to the HLA-A2.1 transgenic mouse to evaluate how androgen deprivation affects human HLA A2.1-resticted T cell responses following immunization of PSA-expressing mice by vaccinia-PSA (PROSTVAC). RESULTS: Our PSA transgenic mouse showed restricted expression of PSA in the prostate and detectable circulating PSA levels. Additionally, PSA expression was androgen-dependent with reduced PSA expression in the prostate within 1 week of castration, and undetectable PSA by day 42 after castration as evaluated by ELISA. Castration of the PSA/A2.1 hybrid mouse prior to immunization with a PSA-expressing recombinant vaccinia virus resulted in a significant augmentation of PSA-specific cytotoxic lymphocytes. CONCLUSIONS: This humanized hybrid mouse model provides a well-defined system to gain additional insight into the mechanisms of immune tolerance to PSA and to test novel strategies aiming at circumventing immune tolerance to PSA and other TAA for targeted prostate cancer immunotherapy.

Efficacy and Effect of Cabozantinib on Bone Metastases in Treatment-naive Castration-resistant Prostate Cancer.

BACKGROUND: Cabozantinib is active in advanced prostate cancer with improvement on bone scans in men on phase II trials. This trial evaluated the efficacy and changes in bone lesions in men with metastatic castration-resistant prostate cancer (mCRPC) treated with cabozantinib. PATIENTS AND METHODS: Eligible patients with mCRPC involving bone underwent biopsy of a bone lesion followed by cabozantinib starting at 60 mg daily and continuing until progression or intolerable toxicity. The primary study endpoint was progression-free survival at 12 weeks. The bone lesion was rebiopsied at 6 weeks. Expression of CMET, phospho-CMET, and VEGFR2 was assayed by immunohistochemistry. Serum was obtained at baseline, and at 3, 6, and 12 weeks and assayed for bone remodeling markers. RESULTS: A total of 25 patients were enrolled: 22 were evaluable, and 3 were excluded before receiving cabozantinib. At 12 weeks, 17 (77%) of 22 patients had stable disease or better. The median time on treatment was 24 weeks (range, 3-112 weeks). The overall median progression-free survival was 43.7 weeks (95% confidence interval, 23.7-97.0 weeks). Eight (36%) of 22 patients had markedly reduced uptake on bone scan. Patients with significant response on bone scan had higher bone morphogenic protein-2 levels at baseline, stable N-telopeptides levels, increased vascular endothelial growth factor receptor 2 expression, and a trend towards increased phospho-CMET while on cabozantinib compared with patients with stable disease. CONCLUSIONS: Cabozantinib is active in men with mCRPC, inducing significant changes on bone scan in one-third of patients with changes in markers of bone formation and the tumor microenvironment.

Primary prostate cancer educates bone stroma through exosomal pyruvate kinase M2 to promote bone metastasis.

Prostate cancer (PCa) metastasizes selectively to bone through unknown mechanisms. In the current study, we identified exosome-mediated transfer of pyruvate kinase M2 (PKM2) from PCa cells into bone marrow stromal cells (BMSCs) as a novel mechanism through which primary tumor-derived exosomes promote premetastatic niche formation. We found that PKM2 up-regulates BMSC CXCL12 production in a HIF-1α-dependent fashion, which subsequently enhances PCa seeding and growth in the bone marrow. Furthermore, serum-derived exosomes from patients with either primary PCa or PCa metastasis, as opposed to healthy men, reveal that increased exosome PKM2 expression is associated with metastasis, suggesting clinical relevance of exosome PKM2 in PCa. Targeting the exosome-induced CXCL12 axis diminished exosome-mediated bone metastasis. In summary, primary PCa cells educate the bone marrow to create a premetastatic niche through primary PCa exosome-mediated transfer of PKM2 into BMSCs and subsequent up-regulation of CXCL12. This novel mechanism indicates the potential for exosome PKM2 as a biomarker and suggests therapeutic targets for PCa bone metastasis.

Targeting cathepsin K diminishes prostate cancer establishment and growth in murine bone.

BACKGROUND: The processes of prostate cancer (PCa) invasion and metastasis are facilitated by proteolytic cascade involving multiple proteases, such as matrix metalloproteinases, serine proteases and cysteine proteases including cathepsin K (CatK). CatK is predominantly secreted by osteoclasts and specifically degrades collagen I leading to bone destruction. PCa and breast cancer preferentially metastasize to the bone. Importantly, CatK expression level is greater in PCa bone metastatic sites compared to primary tumor and normal prostate tissues. However, the underlying mechanism of CatK during PCa metastases into the bone remains to be elucidated. We investigated the functional role of CatK during the PCa establishment and growth process in the murine bone. METHODS: CatK mRNA expression was validated by RT-PCR, protein expression by immunoblotting in PCa LNCaP, C4-2B, and PC3 cells as well as in PCa tissues. Its protein production was measured using ELISA assay. The effect of both knockdowns via siRNA and CatK inhibitor was compared in regard to PCa cell invasion. We further studied the dose-dependent CatK inhibitor effect on conditioned media-induced bone resorption. In setting up an animal model, C4-2B cells were injected into the tibiae of SCID mice. The animals treated with either vehicle or CatK inhibitor for 8 weeks at the time of tumor cell injection (tumor establishment model; protocol I) or 4 weeks after tumor cell injection (tumor progression model; protocol II) were applied to histological and histomorphometric analyses. RESULTS: We confirmed CatK expression in PCa LNCaP, C4-2B, and PC3 cells as well as in PCa tissues. Furthermore, we observed the inhibitory effects of a selective CatK inhibitor on PCa cell invasion. The CatK inhibitor dose-dependently inhibited PCa-conditioned media-induced bone resorption. Upon injection of C4-2B cells into the tibiae of SCID mice, the selective CatK inhibitor significantly prevented the tumor establishment in protocol I, and reduced the tumor growth in bone in protocol II. It also decreased serum PSA levels in both animal models. The inhibitory effects of the CatK inhibitor were enhanced in combination with zoledronic acid (ZA). CONCLUSION: The selective CatK inhibitor may prevent the establishment and progression of PCa in bone, thus making it a novel therapeutic approach for advanced PCa.

Heat stress-induced heat shock protein 70 expression is dependent on ERK activation in zebrafish (Danio rerio) cells.

Heat shock response is a common event that occurs in many species. Despite its evolutionary conservation, comparative studies of heat shock response have been largely unexplored. In mammals, heat shock response decreases with age through unclear mechanisms. Understanding how the age-related decline in heat shock response occurs may provide information to understanding the biology of aging. We have previously shown that heat shock response similarly declines with age in zebrafish. However, signaling pathways that regulate the heat shock response in zebrafish are unknown. In mammals there is evidence that mitogen-activated protein kinases (MAPKs) of the ERK family alter Hsp70 transcription, serving as a potential regulator of the heat shock response. We explored if heat stress-induced Hsp70 expression is altered by activation of ERK in the zebrafish Pac2 fibroblast cell line as occurs in mammalian cells. Heat stress induced both Hsp70 mRNA expression and phosphorylation of both ERK1 and ERK2 (ERK1/2) in Pac2 cells. ERK inhibitors PD98059 and U0126 blocked both heat stress-induced and plated-derived growth factor (PDGF)-induced ERK1/2 phosphorylation, and also diminished heat-induced Hsp70 expression. Pac2 cell viability was not affected by either the ERK inhibitors or heat stress. These results demonstrate that induction of Hsp70 in response to heat stress is dependent on ERK activation in Pac2 cells. This suggests that the heat shock response in zebrafish utilizes a similar signaling pathway to that of mammals and that zebrafish are a good model for comparative studies of heat shock response.

Inhibition of interleukin-6 with CNTO328, an anti-interleukin-6 monoclonal antibody, inhibits conversion of androgen-dependent prostate cancer to an androgen-independent phenotype in orchiectomized mice.

Initially, prostate cancer is androgen dependent. However, most cases progress to an androgen-independent state through unknown mechanisms. Interleukin-6 (IL-6) has been associated with prostate cancer progression including activation of the androgen receptor (AR). To determine if IL-6 plays a role in the conversion of prostate cancer from androgen dependent to androgen independent, we established androgen-dependent LuCaP 35 human prostate cancer xenografts in nude mice, castrated the mice, and blocked IL-6 activity using a neutralizing antibody (CNT0328) for a period of 18 weeks. IL-6 inhibition increased survival of mice and inhibited tumor growth, as reflected by decreased tumor volume and prostate-specific antigen levels, compared with that in mice receiving isotype control antibody. To test the effect of IL-6 inhibition on the conversion from androgen dependent to androgen independent, tumor cells from the treated mice were assessed for their androgen dependence both in vitro and by implanting them into sham-operated or orchiectomized mice. Tumor cells derived from the isotype-treated animals converted to androgen-independent state, whereas tumor cells from the anti-IL-6 antibody-treated mice were still androgen dependent in vitro and in vivo. Although there was no difference in AR levels between the androgen-independent and androgen-dependent tumors, IL-6 inhibition promoted both apoptosis and inhibited cell proliferation in tumors and blocked the orchiectomy-induced expression of histone acetylases, p300 and CBP, which are AR cofactors. These data show that IL-6 contributes to the development of androgen independence in prostate cancer and suggest that it mediates this effect, in part, through modulation of p300 and CBP.

Bone Microenvironment Changes in Latexin Expression Promote Chemoresistance.

Although docetaxel is the standard of care for advanced prostate cancer, most patients develop resistance to docetaxel. Therefore, elucidating the mechanism that underlies resistance to docetaxel is critical to enhance therapeutic intervention. Mining cDNA microarray from the PC-3 prostate cancer cell line and its docetaxel-resistant derivative (PC3-TxR) revealed decreased latexin (LXN) expression in the resistant cells. LXN expression was inversely correlated with taxane resistance in a panel of prostate cancer cell lines. LXN knockdown conferred docetaxel resistance to prostate cancer cells in vitro and in vivo, whereas LXN overexpression reduced docetaxel resistance in several prostate cancer cell lines. A mouse model of prostate cancer demonstrated that prostate cancer cells developed resistance to docetaxel in the bone microenvironment, but not the soft tissue microenvironment. This was associated with decreased LXN expression in prostate cancer cells in the bone microenvironment compared with the soft tissue microenvironment. It was identified that bone stromal cells decreased LXN expression through methylation and induced chemoresistance in prostate cancer in vitro These findings reveal that a subset of prostate cancer develops docetaxel resistance through loss of LXN expression associated with methylation and that the bone microenvironment promotes this drug resistance phenotype.Implications: This study suggests that the LXN pathway should be further explored as a viable target for preventing or reversing taxane resistance in prostate cancer. Mol Cancer Res; 15(4); 457-66. ©2017 AACR.

BET Bromodomain Inhibitors Enhance Efficacy and Disrupt Resistance to AR Antagonists in the Treatment of Prostate Cancer.

UNLABELLED: Next-generation antiandrogen therapies, such as enzalutamide and abiraterone, have had a profound impact on the management of metastatic castration-resistant prostate cancer (mCRPC). However, mCRPC patients invariably develop resistance to these agents. Here, a series of clonal cell lines were developed from enzalutamide-resistant prostate tumor xenografts to study the molecular mechanism of resistance and test their oncogenic potential under various treatment conditions. Androgen receptor (AR) signaling was maintained in these cell lines, which acquired potential resistance mechanisms, including expression of AR-variant 7 (AR-v7) and glucocorticoid receptor. BET bromodomain inhibitors were shown previously to attenuate AR signaling in mCRPC; here, we demonstrate the efficacy of bromodomain and extraterminal (BET) inhibitors in enzalutamide-resistant prostate cancer models. AR antagonists, enzalutamide, and ARN509 exhibit enhanced prostate tumor growth inhibition when combined with BET inhibitors, JQ1 and OTX015, respectively. Taken together, these data provide a compelling preclinical rationale to combine BET inhibitors with AR antagonists to subvert resistance mechanisms. IMPLICATIONS: Therapeutic combinations of BET inhibitors and AR antagonists may enhance the clinical efficacy in the treatment of mCRPC. VISUAL OVERVIEW: http://mcr.aacrjournals.org/content/molcanres/14/4/324/F1.large.jpg

Targeting the MLL complex in castration-resistant prostate cancer.

Resistance to androgen deprivation therapies and increased androgen receptor (AR) activity are major drivers of castration-resistant prostate cancer (CRPC). Although prior work has focused on targeting AR directly, co-activators of AR signaling, which may represent new therapeutic targets, are relatively underexplored. Here we demonstrate that the mixed-lineage leukemia protein (MLL) complex, a well-known driver of MLL fusion-positive leukemia, acts as a co-activator of AR signaling. AR directly interacts with the MLL complex via the menin-MLL subunit. Menin expression is higher in CRPC than in both hormone-naive prostate cancer and benign prostate tissue, and high menin expression correlates with poor overall survival of individuals diagnosed with prostate cancer. Treatment with a small-molecule inhibitor of menin-MLL interaction blocks AR signaling and inhibits the growth of castration-resistant tumors in vivo in mice. Taken together, this work identifies the MLL complex as a crucial co-activator of AR and a potential therapeutic target in advanced prostate cancer.

ALDH activity indicates increased tumorigenic cells, but not cancer stem cells, in prostate cancer cell lines.

BACKGROUND: Cancer stem cells (CSCs) have been shown to be a small stem cell-like cell population which appears to drive tumorigenesis, tumor recurrence and metastasis. Thus, identification and characterization of CSCs may be critical to defining effective anticancer therapies. In prostate cancer (PCa), the CD44(+) cell population appears to have stem cell-like properties including being tumorigenic. The enzyme aldehyde dehydrogenase (ALDH) has been found to identify hematopoietic stem cells and our aim was to determine the utility of ALDH activity and CD44 in identifying PCa stem cell-like cells in PCa cell lines. MATERIALS AND METHODS: LNCaP cells and PC-3 cells were sorted based on their expression of CD44 and ALDH activity. The cell populations were investigated using colony-forming assays, invasion assays, sphere formation experiments in a non-adherent environment and 3-D Matrigel matrix culture to observe the in vitro stem-cell like properties. Different sorted cell populations were injected subcutaneously into NOD/SCID mice to determine the corresponding tumorigenic capacities. RESULTS: ALDH(hi) CD44(+) cells exhibit a higher proliferative, clonogenic and metastatic capacity in vitro and demonstrate higher tumorigenicity capacity in vivo than did ALDH(lo) CD44(-) cells. The tumors recapitulated the population of the original cell line. However, ALDHlo CD44(-) cells were able to develop tumors, albeit with longer latency periods. CONCLUSION: ALDH activity and CD44 do not appear to identify PCa stem cells; however, they do indicate increased tumorigenic and metastatic potential, indicating their potential importance for further exploration.