c-Myc In Prostate Cancer Research Articles

USP11 promotes prostate cancer progression by up-regulating AR and c-Myc activity

Androgen receptor (AR) is a main driver for castration-resistant prostate cancer (CRPC). c-Myc is an oncogene underlying prostate tumorigenesis. Here, we find that the deubiquitinase USP11 targets both AR and c-Myc in prostate cancer (PCa). USP11 expression was up-regulated in metastatic PCa and CRPC. USP11 knockdown (KD) significantly inhibited PCa cell growth. Our RNA-seq studies revealed AR and c-Myc as the top transcription factors altered after USP11 KD. ChIP-seq analysis showed that either USP11 KD or replacement of endogenous USP11 with a catalytic-inactive USP11 mutant significantly decreased chromatin binding by AR and c-Myc. We find that USP11 employs two mechanisms to up-regulate AR and c-Myc levels: namely, deubiquitination of AR and c-Myc proteins to increase their stability and deubiquitination of H2A-K119Ub, a repressive histone mark, on promoters of AR and c-Myc genes to increase their transcription. AR and c-Myc reexpression in USP11-KD PCa cells partly rescued cell growth defects. Thus, our studies reveal a tumor-promoting role for USP11 in aggressive PCa through upregulation of AR and c-Myc activities and support USP11 as a potential target against PCa.

DDX52 knockdown inhibits the growth of prostate cancer cells by regulating c-Myc signaling

BackgroundDDX52 is a type of DEAD/H box RNA helicase that was identified as a novel prostate cancer (PCa) genetic locus and possible causal gene in a European large-scale transcriptome-wide association study. However, the functions of DDX52 in PCa remain undetermined. The c-Myc oncogene plays a crucial role in the development of PCa, but the factors that regulate the activity of c-Myc in PCa are still unknown.MethodsWe determined DDX52 protein levels in PCa tissues using immunohistochemistry (IHC). DDX52 expression and survival outcomes in other PCa cohorts were examined using bioinformatics analysis. The inhibition of DDX52 via RNA interference with shRNA was used to clarify the effects of DDX52 on PCa cell growth in vitro and in vivo. Gene set enrichment analysis and RNA sequencing were used to explore the signaling regulated by DDX52 in PCa. Western blotting and IHC were used to determine the possible DDX52 signaling mechanism in PCa.ResultsDDX52 expression was upregulated in PCa tissues. Bioinformatics analysis showed that the level of DDX52 further increased in advanced PCa, with a high DDX52 level indicating a poor outcome. In vitro and in vivo experiments showed that downregulating DDX52 impeded the growth of PCa cells. High DDX52 levels contributed to activating c-Myc signaling in PCa patients and PCa cells. Furthermore, DDX52 expression was regulated by c-Myc and positively correlated with c-Myc expression in PCa.ConclusionDDX52 was overexpressed in PCa tissues in contrast to normal prostate tissues. DDX52 knockdown repressed the growth of PCa cells in vitro and in vivo. Deleting c-Myc inhibited DDX52 expression, which affected the activation of c-Myc signaling.

Abstract 2450: Methyl CpG Binding Protein 2 suppresses Myc targeting miRNAs to promote context dependent tumor proliferation

Abstract Background: Epigenetic reader proteins maintain an imbalance between differentiation and self-renewal in cancer. Genetic alterations in the DNA methylation machinery (e.g., TET, DNMTs) and chromatin remodelers (e.g., KDMs, EZH1/2) are a hallmark of cancer. Methyl CpG binding domain protein 2 (MBD2) is an epigenetic reader protein which modulates regional gene transcription by recruiting co-repressor complexes to sites of CpG methylation. Our goal is to develop a novel class of anti-neoplastic epigenetic therapies targeting MBD2 to selectively reprogram tumor cells towards a terminally differentiated state and sensitize them to chemotherapy, radiation, and immunotherapy. Methods: To explore conserved mediators of MBD2 function in cancer, we used MBD2 targeting shRNA lentivirus to stably knockdown MBD2 in prostate cancer (PC3, DU145, LnCap), leukemia (SigM5, TET2WTK562 or TET2KOK562) and triple negative breast cancer (MDA-MB231-nanog-GFP) cell lines. Inhibition of MBD2 expression was confirmed by qRT-PCR and Western Blot. Proliferative potential was determined by cell counting and clonogenic potential was determined by methylcellulose-based colony forming assays. RNAseq analysis was performed on prostate cancer and leukemia cell lines. miRNA expression was analyzed using miScript miRNA PCR (Qiagen). In vivo tumor initiation capacity was analyzed using orthotropic and heterotopic xenografts in athymic NSG mice. Results: Our results show that MBD2 is required for the proliferation of triple negative breast cancer (TNBC), prostate cancer (PCa), and TET mutant leukemias (TML), in vitro. The functional mediators of MBD2's growth promoting effects were tissue/tumor context dependent. In leukemias, MBD2's growth promoting and tumor initiating effects were most pronounced in TET2 null cells (which accumulate 5mC). In TNBC, nanog-GFP reporter positive cells were more sensitive to MBD2 knockdown than reporter negative cells. In PCa and TNBC, RNAseq analysis revealed that knockdown of MBD2 led to downregulation of Myc pathway genes and increased the expression of Myc targeting microRNAs, miR33-5, miR34a, miR148a and miR363. Western blot analysis confirmed that MBD2 knockdown coordinately downregulated cMyc expression and activated p27 expression. We further demonstrated that inhibition of MBD2 diminished the tumor initiating capacity of TNBC and PCa in xenograft models and the in vivo engraftment rate of patient derived TET-/- AML. Conclusions: MBD2 knockdown diminished the proliferative capacity of PCa, TNBC and primary TET2 mutant leukemia cells in a genetic (TET2) and phenotypic (nanog+ stem/progenitor) context dependent manner. Delayed peak effect and altered differentiation markers after MBD2 inhibition suggest epigenetic reprogramming as the mechanism of growth suppression. MBD2 targets miRNA's upstream of cMyc in PCa and TNBC. MBD2 murine knockout models are developmentally normal, suggesting an acquired function in cancer with a favorable therapeutic window for targeting. Citation Format: Aysegul Balyimez, Yihong Guan, Emily Esakov, Shinjini Ganguly, Ofer Reizes, Daniel J. Lindner, Jaroslaw Maciejewski, Babal Jha, Omar Y. Mian. Methyl CpG Binding Protein 2 suppresses Myc targeting miRNAs to promote context dependent tumor proliferation [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 2450.

SOX4 is activated by C-MYC in prostate cancer.

Although MYC proto-oncogene (C-MYC) amplification has been consistently reported to be a potential marker for prostate cancer (PCa) progression and prognosis, the clinicopathological and prognostic significance of C-MYC protein expression remains controversial. Overexpression of SOX4 has been shown to play important roles in multiple cancers including PCa. However, the link between these two critical genetic aberrations is unclear. In the current study, we showed that C-MYC was overexpressed in 16.2% (17/105) of Chinese patients with localized PCa. Overexpression of C-MYC was significantly associated with high Gleason scores (P = 0.012) and high Ki67 labeling index (P = 0.005). C-MYC overexpression was correlated with cancer-related mortality and suggested to be an unfavorable prognostic factor in Chinese PCa patients (P = 0.018). Overexpression of C-MYC is associated with SOX4 overexpression in PCa tissues. Notably, SOX4 is a direct target gene of C-MYC; C-MYC activates SOX4 expression via binding to its promoter. In addition, Co-IP analysis demonstrated a physical interaction between C-MYC and SOX4 protein in PCa cells. Clinically, C-MYC+/SOX4+ characterized poor prognosis in a subset of PCa patients. In total, C-MYC overexpression may contribute to PCa progression by activating SOX4. Our findings highlight an important role of C-MYC/SOX4 in PCa progression in a subset of PCa patients.

Expression and significance of C-MYC and thioredoxin-interacting protein in prostate cancer

Objective To study the expression and significance of C-MYC and thioredoxin-interacting protein (TXNIP) in prostate cancer and benign prostatic hyperplasia. Methods 50 cases of prostate cancer tissues and 49 cases of benign prostatic hyperplasia tissues were selected. The immunohistochemistry method was used to detect the expression level of C-MYC and TXNIP, and the correlation with clinical data. Results The positive rate of C-MYC in prostate cancer tissues was 64.0% (32/50) while in benign prostatic hyperplasia tissues was 10.2% (5/49, P 7 group, the positive rate of C-MYC was statistically significant (P 7 group. In the Gleason score 7 group, 7 group. Overexpression of C-MYC was associated with downregulation of TXNIP (Spearman’s r=-0.306, P<0.05). Conclusion Overexpression of C-MYC and downregulation of TXNIP might be involved in the pathogenesis, development and degree of malignancy of prostate cancer. TXNIP is expected to be a potential therapeutic target for prostate cancer. Key words: Prostate cancer; C-MYC; Thioredoxin-interacting protein

Abstract B61: c-Myc is regulated and therapeutically targetable via the miR-1207-3p/FNDC1/FN1/AR pathway in aggressive prostate cancer in men of African ancestry

Abstract Prostate cancer is the 2nd most common cancer in the world for men. For reasons still unclear, aggressive PCa disproportionately affects males of African ancestry (MoAA). Incidence and mortality rates are highest in MoAA as they have consistently shown a 2.3-3.0-fold higher risk of mortality compared to Caucasian men (CM). This aggressiveness of PCa may be due to specific biologic factors. Located downstream of c-Myc at chromosome 8q24 is PVT1, which encodes miR-1207-3p. Studies have shown that PVT1/MYC cooperation is a fundamental feature in all cancers with 8q24 amplification, and 98% of the 8q24 amplicons contained concurrent amplification of the MYC and PVT1 loci. Moreover, MYC has been linked to PCa aggressiveness and has been reported to be downstream of AR in some PCa. However, the mechanisms regulating c-MYC have never been studied in MoAA. We recently demonstrated that miR-1207-3p directly binds to FNDC1 to regulate a novel FNDC1/FN1/AR pathway upregulated in metastatic prostate cancer (PCa). However, the mechanisms regulating c-Myc in PCa remain unclear, and the relevance of our novel and clinically significant miR-1207-3p molecular pathway in PCa in MoAA is unknown. The aim of this study was to determine if c-Myc is regulated and therapeutically targetable via the miR-1207-3p/FNDC1/FN1/AR pathway in aggressive PCa in MoAA. We used qPCR, immunoblotting, RNA pulldown, proliferation, migration, and apoptosis assays to evaluate miR-1207-3p regulation of c-Myc in aggressive PCa in MoAA. Also, miR-1207-3p, FNDC1, FN1, AR, and c-Myc expression was analyzed in prostate tissues (normal = 21; benign = 41; tumor = 26) of patients who received prostatectomy or transrectal ultrasound-guided biopsies at the University College Hospital, Ibadan, Nigeria, a sub-Saharan Black African population. Seventeen patients had tumor tissues with Gleason score ≥ 8. Tissues were collected in compliance with Institutional Review Board-approved protocols. ANOVA, student's t-test, and Tukey post-hoc tests were used for statistical analysis. Prostate tissue analysis revealed that underexpression of miR-1207-3p and the overexpression of FNDC1, FN1, AR, and c-Myc is significantly associated with aggressive PCa in MoAA. Also, miR-1207-3p was underexpressed while FNDC1 and c-MYC were overexpressed in tumors with Gleason score ≥8 in comparison to those with Gleason score &amp;lt;8. Thus, miR-1207-3p and FNDC1 may be useful for risk stratification in PCa in MoAA. Next, we observed that our novel synthetic biotinylated miR-1207-3p duplex (NB1207, patent pending) significantly inhibited c-Myc protein expression in 7 PCa cell lines when compared with a biotinylated scramble duplex (NB-1, patent pending). RNA pulldown assay determined that c-Myc is not a direct molecular target of miR-1207-3p. However, siRNAs against FNDC1, FN1, and AR revealed significant suppression of c-Myc expression by &amp;gt;75% in the MoAA-derived indolent E006AA PCa cell line and the MoAA-derived aggressive/castration-resistant E006AA-hT PCa cell line, indicating that c-Myc is downstream of AR. c-Myc expression is higher in the E006AA-hT PCa cell line when compared to the E006AA PCa cell line, suggesting that c-Myc is associated with aggressive PCa. Moreover, NB1207 significantly inhibited migration and induced apoptosis in E006AA and E006AA-hT PCa cell lines. Next, we compared the efficacy of NB1207 in inhibiting proliferation to the commercially available drugs for treatment of CPRC (enzalutamide and abiraterone). NB1207 inhibited proliferation in the CRPC cell line E006AA-hT by nearly 50% while enzalutamide and abiraterone had no effect. In conclusion, miR-1207-3p regulates c-Myc expression via the miR-1207-3p/FNDC1/FN1/AR pathway in aggressive PCa in MoAA. miR-1207-3p may be a biomarker for risk stratification in PCa in MoAA. NB1207 has potential for therapeutic targeting of c-Myc for treatment of aggressive PCa in MoAA. Citation Format: Dibash K. Das, Akintunde T. Orunmuyi, Gabriel Olabiyi Ogun, S. Adekola Adebayo, A. Ayo Salako, Adeodat Ilboudo, Cuong Bach, E. O. Olapade-Olaopa, Olorunseun O. Ogunwobi. c-Myc is regulated and therapeutically targetable via the miR-1207-3p/FNDC1/FN1/AR pathway in aggressive prostate cancer in men of African ancestry [abstract]. In: Proceedings of the Tenth AACR Conference on the Science of Cancer Health Disparities in Racial/Ethnic Minorities and the Medically Underserved; 2017 Sep 25-28; Atlanta, GA. Philadelphia (PA): AACR; Cancer Epidemiol Biomarkers Prev 2018;27(7 Suppl):Abstract nr B61.

PROSTATE AND BREAST CANCER

Background: CK2, a serine/threonine, protein kinase, targets over and above300 substrates including c-Myc. CK2 expression is elevated in human cancers includingbreast cancer and prostate cancer. c-Myc protooncogene expression is also up-regulated inthese cancers. Objectives: To evaluate the co expression and correlation of CK2 and c-Mycin prostate cancer as compared to their correlation in breast cancer. Study Design: Crosssectional analytical study. Setting: Army Medical College and AFIP, Duration: Two years.Methods: A retrospective study of immunohistochemical analysis, approved by Armed ForcesInstitute of Pathology Ethical Committee. Paraffin embedded tissues of diagnosed prostatecancer, 30 in number, 30 cases of Benign Prostatic Hypertrophy (BPH) and 30 cases of breastadenocarcinoma, were included in the study. We stained tissue sections for CK2 and c-Mycand measured staining intensity for each protein expression. Data analysis was done by SPSSversion 20. Pearson correlation coefficient was used for correlating the expression of bothproteins. P-value was calculated. Results: A strong correlation of CK2 with c-Myc was seen inprostate cancer tissue, in comparison to BPH. There was a very significant correlation presentbetween CK2 and c-Myc, especially in invasive cases of breast cancer. Conclusion: CK2 andc-Myc expressions are highly and significantly correlated in prostate cancer and breast cancerespecially in invasive cases. CK2 has influence over c-Myc and both can be used for forecastingthe cancer phenotype and aggression of disease.

A COMPARISON OF CK2 MODULATION OF C-MYC IN PROSTATE AND BREAST CANCER

Background: CK2, a serine/threonine, protein kinase, targets over and above300 substrates including c-Myc. CK2 expression is elevated in human cancers includingbreast cancer and prostate cancer. c-Myc protooncogene expression is also up-regulated inthese cancers. Objectives: To evaluate the co expression and correlation of CK2 and c-Mycin prostate cancer as compared to their correlation in breast cancer. Study Design: Crosssectional analytical study. Setting: Army Medical College and AFIP, Duration: Two years.Methods: A retrospective study of immunohistochemical analysis, approved by Armed ForcesInstitute of Pathology Ethical Committee. Paraffin embedded tissues of diagnosed prostatecancer, 30 in number, 30 cases of Benign Prostatic Hypertrophy (BPH) and 30 cases of breastadenocarcinoma, were included in the study. We stained tissue sections for CK2 and c-Mycand measured staining intensity for each protein expression. Data analysis was done by SPSSversion 20. Pearson correlation coefficient was used for correlating the expression of bothproteins. P-value was calculated. Results: A strong correlation of CK2 with c-Myc was seen inprostate cancer tissue, in comparison to BPH. There was a very significant correlation presentbetween CK2 and c-Myc, especially in invasive cases of breast cancer. Conclusion: CK2 andc-Myc expressions are highly and significantly correlated in prostate cancer and breast cancerespecially in invasive cases. CK2 has influence over c-Myc and both can be used for forecastingthe cancer phenotype and aggression of disease.

MicroRNA-449a enhances radiosensitivity by downregulation of c-Myc in prostate cancer cells.

MicroRNAs (miRNAs) have been reported to be involved in DNA damage response induced by ionizing radiation (IR). c-Myc is reduced when cells treated with IR or other DNA damaging agents. It is unknown whether miRNAs participate in c-Myc downregulation in response to IR. In the present study, we found that miR-449a enhanced radiosensitivity in vitro and in vivo by targeting c-Myc in prostate cancer (LNCaP) cells. MiR-449a was upregulated and c-Myc was downregulated in response to IR in LNCaP cells. Overexpression of miR-449a or knockdown of c-Myc promoted the sensitivity of LNCaP cells to IR. By establishing c-Myc as a direct target of miR-449a, we revealed that miR-449a enhanced radiosensitivity by repressing c-Myc expression in LNCaP cells. Furthermore, we showed that miR-449a enhanced radiation-induced G2/M phase arrest by directly downregulating c-Myc, which controlled the Cdc2/CyclinB1 cell cycle signal by modulating Cdc25A. These results highlight an unrecognized mechanism of miR-449a-mediated c-Myc regulation in response to IR and may provide alternative therapeutic strategies for the treatment of prostate cancer.

SREBP-2 promotes stem cell-like properties and metastasis by transcriptional activation of c-Myc in prostate cancer.

Sterol regulatory element-binding protein-2 (SREBP-2) transcription factor mainly controls cholesterol biosynthesis and homeostasis in normal cells. The role of SREBP-2 in lethal prostate cancer (PCa) progression remains to be elucidated. Here, we showed that expression of SREBP-2 was elevated in advanced pathologic grade and metastatic PCa and significantly associated with poor clinical outcomes. Biofunctional analyses demonstrated that SREBP-2 induced PCa cell proliferation, invasion and migration. Furthermore, overexpression of SREBP-2 increased the PCa stem cell population, prostasphere-forming ability and tumor-initiating capability, whereas genetic silencing of SREBP-2 inhibited PCa cell growth, stemness, and xenograft tumor growth and metastasis. Clinical and mechanistic data showed that SREBP-2 was positively correlated with c-Myc and induced c-Myc activation by directly interacting with an SREBP-2-binding element in the 5′-flanking c-Myc promoter region to drive stemness and metastasis. Collectively, these clinical and experimental results reveal a novel role of SREBP-2 in the induction of a stem cell-like phenotype and PCa metastasis, which sheds light on translational potential by targeting SREBP-2 as a promising therapeutic approach in PCa.

The Long Non-Coding RNA PCAT-1 Promotes Prostate Cancer Cell Proliferation through cMyc

Long non-coding RNAs (lncRNAs) represent an emerging layer of cancer biology, contributing to tumor proliferation, invasion, and metastasis. Here, we describe a role for the oncogenic lncRNA PCAT-1 in prostate cancer proliferation through cMyc. We find that PCAT-1–mediated proliferation is dependent on cMyc protein stabilization, and using expression profiling, we observed that cMyc is required for a subset of PCAT-1–induced expression changes. The PCAT-1–cMyc relationship is mediated through the post-transcriptional activity of the MYC 3′ untranslated region, and we characterize a role for PCAT-1 in the disruption of MYC-targeting microRNAs. To further elucidate a role for post-transcriptional regulation, we demonstrate that targeting PCAT-1 with miR-3667-3p, which does not target MYC, is able to reverse the stabilization of cMyc by PCAT-1. This work establishes a basis for the oncogenic role of PCAT-1 in cancer cell proliferation and is the first study to implicate lncRNAs in the regulation of cMyc in prostate cancer.

Abstract 3004: Functional analysis of ERG splice variants and their effect on c-myc expression in prostate cancer.

Abstract Introduction and Objectives: The oncogenic activation of ERG in prostate cancer has been well established in recent years. Towards understanding the functional role of ERG, our laboratory previously identified two major types of ERG splice variants in human prostate tumors: Type I, which encodes a full length prototypical ERG protein, and Type II, which encodes a truncated protein lacking the ETS DNA-binding domain. Analysis of clinico-pathologic data from patients and ratios of Type I/Type II ERG revealed that an increased ratio correlated with a higher Gleason sum and poorly differentiated phenotype. However, the functional significance of the ERG splice variants is not clearly understood. Studying the function and interaction of the individual splice variants can further enhance our understanding of ERG, and advance efforts towards its clinical utility in the treatment of prostate cancer. Methods: Exogenous Type II ERG was expressed in TMPRSS2-ERG fusion harboring VCaP cells and endogenous levels of Type I ERG and C-MYC proteins were evaluated. Type I and Type II ERG were also co-transfected in various cell lines to examine interaction among splice variants and regulatory efficiency with luciferase reporter constructs. In addition, quantitative RNA expression data of C-MYC and Type I/Type II ERG ratio were assessed by correlation analysis in patient samples. Results: Expression of exogenous Type II ERG resulted in a decrease in endogenous Type I ERG protein expression in VCaP cells in a dose dependent manner, with a concomitant decrease in C-MYC. This decrease was not observed in the other cell lines. Decreased regulatory efficiency of Type I ERG, by addition of Type II ERG, was also observed. Patient data analysis also revealed a positive correlation between C-MYC expression and increased Type I/Type II ratio. Conclusions: Decreased Type I/Type II ratios in VCaP cells showed decreases in C-MYC, suggesting that modulation of Type II ERG alters Type I ERG and thus its regulation of C-MYC. This interference was not observed in other cell lines, indicating the interaction is context dependent. While Type I ERG positively regulated downstream targets, addition of Type II ERG decreased their expression, implying an interaction between splice variants. Analysis of patient data affirmed in vitro results, thus we propose that modulation of Type I/Type II ratio may regulate downstream targets including oncogenic C-MYC in prostate cancer, providing a strong rationale for the biomarker and therapeutic utility of ERG splice variants. Funding source: DoD/CDMRP (PC073614) grant to S.S. Citation Format: Anshu Rastogi, Shyh-Han Tan, Gyorgy Petrovics, Yongmei Chen, Taduru Sreenath, David G. McLeod, Shiv Srivastava, Albert Dobi. Functional analysis of ERG splice variants and their effect on c-myc expression in prostate cancer. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3004. doi:10.1158/1538-7445.AM2013-3004 Note: This abstract was not presented at the AACR Annual Meeting 2013 because the presenter was unable to attend.

GLIPR1 Suppresses Prostate Cancer Development through Targeted Oncoprotein Destruction

Downregulation of the proapoptotic p53 target gene glioma pathogenesis-related protein 1 (GLIPR1) occurs frequently in prostate cancer, but the functional meaning of this event is obscure. Here, we report the discovery of functional relationship between GLIPR1 and c-Myc in prostate cancer where c-Myc is often upregulated. We found that the expression of GLIPR1 and c-Myc were inversely correlated in human prostate cancer. Restoration of GLIPR1 expression in prostate cancer cells downregulated c-myc levels, inhibiting cell-cycle progression. Downregulation was linked to a reduction in β-catenin/TCF4-mediated transcription of the c-myc gene, which was caused by GLIPR1-mediated redistribution of casein kinase 1α (CK1α) from the Golgi apparatus to the cytoplasm where CK1α could phosphorylate β-catenin and mediate its destruction. In parallel, GLIPR1 also promoted c-Myc protein ubiquitination and degradation by glycogen synthase kinase-3α- and/or CK1α-mediated c-Myc phosphorylation. Notably, genetic ablation of the mouse homolog of Glipr1 cooperated with c-myc overexpression to induce prostatic intraepithelial neoplasia and prostate cancer. Together, our findings provide evidence for CK1α-mediated destruction of c-Myc and identify c-Myc S252 as a crucial CK1α phosphorylation site for c-Myc degradation. Furthermore, they reveal parallel mechanisms of c-myc downregulation by GLIPR1 that when ablated in the prostate are sufficient to drive c-Myc expression and malignant development.

Abstract 3239: Synergy of PIM1 kinase with c-MYC in prostate cancer

Abstract The oncogenes PIM1 and c-MYC are frequently up-regulated in human prostate tumors. We found that PIM1 and c-MYC were co-expressed in human prostate cancer and were associated with higher Gleason scores. We next used a model of prostate regeneration coupled with lentiviral-mediated gene transfer to directly show that Pim1 dramatically enhanced c-MYC tumorigenicity in naïve adult mouse prostatic epithelium in a kinase-dependent manner. Microarray analysis revealed that many signaling pathways including MAPK, focal adhesion calcium signaling pathway were activated in MYC/Pim1 tumor. Knockdown of Pim1 in MYC/Pim1 tumor derived cell line reduced ERK and Akt phosphorylation. We further showed that targeting Pim1 by shRNA inhibited tumorigenicity in vitro and in vivo. Pim1 kinase inhibitor quercetagetin also inhibited cell growth in vitro. These results directly demonstrate that MYC and PIM1 cooperate in promoting tumor development. PIM1 can be considered as a drug target for human prostate cancer treatment. 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 3239.

Immortalization of Primary Human Prostate Epithelial Cells by c-Myc

A significant percentage of prostate tumors have amplifications of the c-Myc gene, but the precise role of c-Myc in prostate cancer is not fully understood. Immortalization of human epithelial cells involves both inactivation of the Rb/p16INK4a pathway and telomere maintenance, and it has been recapitulated in culture by expression of the catalytic subunit of telomerase, hTERT, in combination with viral oncoproteins. Here, we show the immortalization of human prostate epithelial cells (HPrEC) by a single genetic event, the expression of the c-Myc oncogene. Myc stabilizes telomere length in HPrEC through up-regulation of hTERT expression and overrides the accumulation of cell cycle inhibitory proteins, such as p16INK4a. Overall, HPrECs expressing c-Myc retain many characteristics of normal cells, such as the induction of a senescence-like growth arrest in response to oncogenic Ras, an intact p53 response, and an absence of gross karyotypic abnormalities. However, HPrECs expressing c-Myc lack a Rb/p16INK4a checkpoint and can be transformed without the need for additional genetic lesions in that pathway. These results give a partial explanation for the physiologic role of c-Myc overexpression in prostate cancer.

Mapping and Gene Expression Profile of the Minimally Overrepresented 8q24 Region in Prostate Cancer

We have recently reported that overrepresentation of 8q24 (c-myc) is associated with clinical progression in prostate cancer. In this study, we map the boundaries of the overrepresented region within 8q23-q24 using interphase fluorescent in situ hybridization analysis of paraffin-embedded prostate cancer specimens. One hundred primary prostate cancers and three prostate cancer cell lines were evaluated, and the minimally overrepresented region could be narrowed to the approximately 8.2-Mb region between D8S514 and H47317. This region includes c-myc and is wholly within 8q24. Eukaryotic translation initiation factor 3 subunit 3 does not seem to be overrepresented independent of c-myc in prostate cancer. The cell lines PC3 and DU145 have and do not have 8q24 overrepresentation, respectively. We then selected 39 expressed sequence tags (ESTs) within and surrounding the minimally overrepresented region and performed expression analysis using Northern blot hybridization. Five ESTs/genes including c-myc were overexpressed in both the PC3 cell line and DU145, but the PC3 to DU145 expression ratios were <2. Seven ESTs were overexpressed twofold or more in PC3 compared to DU145. This group included hyaluronan synthase 2, nephroblastoma-overexpressed gene, eukaryotic translation initiation factor 3 subunit 3, and an EST (R69368) encoding a hypothetical protein (BM009). These seven genes as well as c-myc are candidate target genes within the overrepresented 8q24 region and their overexpression may be associated with prostate cancer progression.