Chemoresistant Prostate Cancer Cells Research Articles

Abstract C020: Nuclear encoded mitochondrial protein MAGMAS as a potential predictor of taxane sensitivity in advanced prostate cancer patients and differential diagnostic marker of tumor aggressiveness in men of African ancestry

Abstract Prostate cancer (PCa) is the most commonly diagnosed malignancy and the second leading cause of cancer-related death among men in the United States (U.S.). Strikingly, men of African ancestry (AA) are 1.7 times more likely to be diagnosed with PCa and more than twice as likely to die of PCa than men of European origin (EU). These disparities are driven by socioeconomic, lifestyle, environmental, and biological/genetic factors. Prostate-specific antigen screening is limited in identifying men with aggressive PCa and exposes men to increased harm with minimal reduction in mortality from PCa. Specifically, several studies report that men of AA are considered to have low-risk prostate cancer by clinical parameters, and those who enrolled in active surveillance programs progress to a higher Gleason-grade cancer and high-volume status than men of EA. Also, men of AA in low to moderate-risk groups appear to be at a higher risk for biochemical recurrence after curative-intent treatment. Therefore, there is a need to understand molecular mechanisms underlying PCa aggressiveness in men of AA that are translatable to the clinic as reliable biomarkers and for the development of novel therapeutic targets. Evidence supports that the mitochondria have a fundamental influence on all steps of oncogenesis, including malignant transformation, tumor progression, and treatment resistance. Regarding cancer health disparities, genes encoding mitochondrial complexes involved in oxidative phosphorylation are upregulated to a greater extent in tumor specimens from AA patients than those from EA patients, supporting the rationale for investigating the role of mitochondria in cancer health disparities. This study investigates Magmas, a nuclear-encoded mitochondrial protein, as a potential mitochondrial marker for PCa tumor aggressiveness and a therapeutic target for advanced PCa. Several observations support the rationale for this study: 1) Magmas overexpression in multiple aggressive cancer subtypes, including high-grade prostate, ovarian cancer, and malignant gliomas. 2) AA PCa cell line overexpresses Magmas similar to chemo-resistant PCa cells, whereas the EA cell line is shown to exhibit lower expression. 3) In human cells, overexpression of Magmas suppresses oxidative stress by regulating the reactive oxygen species (ROS) levels. 4) In chemo-resistant PCa cell lines, pharmacological inhibition of Magmas re-sensitized PCa cells to taxane treatment; and 5) non-cytotoxic pharmacological inhibition of Magmas in chemo-resistant PCa cells dramatically reduced clonogenicity potential. We will evaluate the hypothesis that AA PCa tissue will overexpress Magmas and that overexpression will predict tumor aggressiveness, and 2) that pharmacological inhibition of Magmas will overcome treatment resistance in advanced PCa models. The proposed exploratory study is highly relevant as it will reveal the biological basis for race-related differential expression of Magmas, providing critical insights into mitochondrial determinants that contribute to PCa mortality disparities. Citation Format: Alfonso Duran, Kristen Whitley, Krystal Santiago, Bhaskar Das, Carlos Casiano, Frankis Almaguel. Nuclear encoded mitochondrial protein MAGMAS as a potential predictor of taxane sensitivity in advanced prostate cancer patients and differential diagnostic marker of tumor aggressiveness in men of African ancestry [abstract]. In: Proceedings of the 17th AACR Conference on the Science of Cancer Health Disparities in Racial/Ethnic Minorities and the Medically Underserved; 2024 Sep 21-24; Los Angeles, CA. Philadelphia (PA): AACR; Cancer Epidemiol Biomarkers Prev 2024;33(9 Suppl):Abstract nr C020.

Docetaxel-Induced Cell Death Is Regulated by a Fatty Acid-Binding Protein 12-Slug-Survivin Pathway in Prostate Cancer Cells.

Chemotherapy is an important treatment option for advanced prostate cancer, especially for metastatic prostate cancer (PCa). Resistance to first-line chemotherapeutic drugs such as docetaxel often accompanies prostate cancer progression. Attempts to overcome resistance to docetaxel by combining docetaxel with other biological agents have been mostly unsuccessful. A better understanding of the mechanisms underlying docetaxel resistance may provide new avenues for the treatment of advanced PCa. We have previously found that the fatty acid-binding protein 12 (FABP12)-PPARγ pathway modulates lipid-related bioenergetics and PCa metastatic transformation through induction of Slug, a master driver of epithelial-to-mesenchymal transition (EMT). Here, we report that the FABP12-Slug axis also underlies chemoresistance in PCa cells. Cell sensitivity to docetaxel is markedly suppressed in FABP12-expressing cells, along with induction of Survivin, a typical apoptosis inhibitor, and inhibition of cleaved PARP, a hallmark of programmed cell death. Importantly, Slug depletion down-regulates Survivin and restores cell sensitivity to docetaxel in FABP12-expressing cells. Finally, we also show that high levels of Survivin are associated with poor prognosis in PCa patients, with FABP12 status determining its prognostic significance. Our research identifies a FABP12-Slug-Survivin pathway driving docetaxel resistance in PCa cells, suggesting that targeting FABP12 may be a precision approach to improve chemodrug efficacy and curb metastatic progression in PCa.

Abstract 1678: Docetaxel induces Rac1 and Cdc42 Rho GTPase activation in prostate cancer cells

Abstract Prostate cancer (PCa) is a major health burden that affects the quality of life of men worldwide. In the United States, PCa is the second leading cause of mortality among cancers that affect men. Treatment of advanced PCa aims to suppress androgen signaling and proliferation in cancer cells using anti-androgens and chemotherapy. However, the development of drug resistance is a major limitation to curing advanced prostate cancer and strategies to block mechanisms of therapy resistance would enhance treatment efficacy and potentially improve patient prognosis. Cdc42 and Rac1, members of the family of Rho GTPases, have been associated with the progression of different types of hormone-associated cancer, such as breast, ovary, and prostate, among others. The current study was designed to evaluate the role of Cdc42 and Rac1 in chemoresistant prostate cancer cells, including under castration-resistant and androgen-independent conditions. We did not observe significant differences in active (Rac-GTP) or total Rac1 levels in chemoresistant compared to parental PC3 or DU145 cells; similar findings were observed for Ccd42. Treatment of drug-naïve cells with a sublethal dose of docetaxel (10uM) induced gradual increases in Rac1-GTP up to 30 minutes in 22rv1, 60 minutes in LNCaP and DU145, and 24 hours in PC3 cells. Cdc42-GTP levels also increased with subacute docetaxel treatment up to 24 hours in 22rv1 and DU145. Chemoresistant 22rv1, PC3, and DU145 exhibited similar sensitivity to Rac1 and Ccd42 inhibitors including MB1-167, MBQ-168, EHop-097, with an IC50 of about 1uM for each compound and line. Ongoing studies are evaluating the role of these Rho-GTPases in 3-dimensional growth and survival. Citation Format: Andrea Paola López González, Jessica Colón González, María C. Santa María Fuentes, Surangani Dharmawardhane, Carlos J. Diaz Osterman. Docetaxel induces Rac1 and Cdc42 Rho GTPase activation in prostate cancer cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 1678.

Abstract 7200: Non-cytotoxic inhibition of nuclear encoded mitochondrial protein MAGMAS resensitize treatment resistant prostate cancer cells to taxane therapy

Abstract Metastatic prostate cancer (PCa) is the leading cause of PCa-associated deaths and one of the leading causes of cancer mortality in men. Novel therapeutics, such as Prostate Specific Membrane Antigen (PSMA) ligand therapy, are emerging as effective treatment strategies for metastatic prostate cancer. However, most men succumb to Metastatic Castration Resistant Prostate Cancer (mCRPC) due to treatment resistance. MAGMAS, a 13.8 kDa mitochondrial protein, imports nuclear-encoded mitochondrial proteins into the mitochondrial matrix. MAGMAS is overexpressed in several aggressive tumors, such as breast cancer, glioblastoma, and prostate cancer. When overexpressed, MAGMAS plays a role as a cytoprotective protein by acting as a reactive oxygen species (ROS) scavenger that maintains sufficient ROS levels to promote cell proliferation but below the threshold levels that could lead to apoptosis. This study was designed to investigate the pro-survival and therapy-resistance functions of MAGMAS in treatment-resistant PCa cells. Predictively, we observed that the docetaxel (DTX)-resistant PC3-DR and DU145-DR PCa cell lines expressed significantly higher levels of MAGMAS compared to their drug-sensitive parental cell lines. We hypothesized that inhibiting Magmas reduces the aggressive properties of highly proliferative DTX-resistant metastatic PCa cell lines and re-sensitizes them to DTX. MAGMAS knockdown in DTX-resistant PC3 and DU145 cells in the presence of increasing DTX treatments enhanced sensitivity to DTX. Consistent with these results, pharmacological inhibition of MAGMAS with the novel BT#9 inhibitor in combination with increasing DTX concentrations also enhanced sensitivity to DTX in these cell lines. Additional studies using cell migration/invasion assays and tumor-sphere formation assays are underway to further establish MAGMAS as a potential therapeutic target to attenuate the aggressive properties of chemo-resistant PCa cells. Lastly, we will investigate the diagnostic potential of MAGMAS as a biomarker for advanced PCa and BT#9 as an imaging/therapeutic-specific agent for treatment-resistant tumors. Citation Format: Alfonso Duran, Kristen Whitley, Krystal Santiago, Adil Mohammed, Carlos Casiano, Bhaskar Das, Frankis Almaguel. Non-cytotoxic inhibition of nuclear encoded mitochondrial protein MAGMAS resensitize treatment resistant prostate cancer cells to taxane therapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 7200.

Glucocorticoid Receptor Regulates and Interacts with LEDGF/p75 to Promote Docetaxel Resistance in Prostate Cancer Cells.

Patients with advanced prostate cancer (PCa) invariably develop resistance to anti-androgen therapy and taxane-based chemotherapy. Glucocorticoid receptor (GR) has been implicated in PCa therapy resistance; however, the mechanisms underlying GR-mediated chemoresistance remain unclear. Lens epithelium-derived growth factor p75 (LEDGF/p75, also known as PSIP1 and DFS70) is a glucocorticoid-induced transcription co-activator implicated in cancer chemoresistance. We investigated the contribution of the GR-LEDGF/p75 axis to docetaxel (DTX)-resistance in PCa cells. GR silencing in DTX-sensitive and -resistant PCa cells decreased LEDGF/p75 expression, and GR upregulation in enzalutamide-resistant cells correlated with increased LEDGF/p75 expression. ChIP-sequencing revealed GR binding sites in the LEDGF/p75 promoter. STRING protein-protein interaction analysis indicated that GR and LEDGF/p75 belong to the same transcriptional network, and immunochemical studies demonstrated their co-immunoprecipitation and co-localization in DTX-resistant cells. The GR modulators exicorilant and relacorilant increased the sensitivity of chemoresistant PCa cells to DTX-induced cell death, and this effect was more pronounced upon LEDGF/p75 silencing. RNA-sequencing of DTX-resistant cells with GR or LEDGF/p75 knockdown revealed a transcriptomic overlap targeting signaling pathways associated with cell survival and proliferation, cancer, and therapy resistance. These studies implicate the GR-LEDGF/p75 axis in PCa therapy resistance and provide a pre-clinical rationale for developing novel therapeutic strategies for advanced PCa.

Nicardipine is a putative EED inhibitor and has high selectivity and potency against chemoresistant prostate cancer in preclinical models.

It is imperative to develop novel therapeutics to overcome chemoresistance, a significant obstacle in the clinical management of prostate cancer (PCa) and other cancers. A phenotypic screen was performed to identify novel inhibitors of chemoresistant PCa cells. The mechanism of action of potential candidate(s) was investigated using in silico docking, and molecular and cellular assays in chemoresistant PCa cells. The in vivo efficacy was evaluated in mouse xenograft models of chemoresistant PCa. Nicardipine exhibited high selectivity and potency against chemoresistant PCa cells via inducing apoptosis and cell cycle arrest. Computational, molecular, and cellular studies identified nicardipine as a putative inhibitor of embryonic ectoderm development (EED) protein, and the results are consistent with a proposed mechanism of action that nicardipine destabilised enhancer of zeste homologue 2 (EZH2) and inhibited key components of noncanonical EZH2 signalling, including transducer and activator of transcription 3, S-phase kinase-associated protein 2, ATP binding cassette B1, and survivin. As a monotherapy, nicardipine effectively inhibited the skeletal growth of chemoresistant C4-2B-TaxR tumours. As a combination regimen, nicardipine synergistically enhanced the in vivo efficacy of docetaxel against C4-2 xenografts. Our findings provided the first preclinical evidence supporting nicardipine as a novel EED inhibitor that has the potential to be promptly tested in PCa patients to overcome chemoresistance and improve clinical outcomes.

Bromocriptine monotherapy overcomes prostate cancer chemoresistance in preclinical models

Chemoresistance is a major obstacle in the clinical management of metastatic, castration-resistant prostate cancer (PCa). It is imperative to develop novel strategies to overcome chemoresistance and improve clinical outcomes in patients who have failed chemotherapy. Using a two-tier phenotypic screening platform, we identified bromocriptine mesylate as a potent and selective inhibitor of chemoresistant PCa cells. Bromocriptine effectively induced cell cycle arrest and activated apoptosis in chemoresistant PCa cells but not in chemoresponsive PCa cells. RNA-seq analyses revealed that bromocriptine affected a subset of genes implicated in the regulation of the cell cycle, DNA repair, and cell death. Interestingly, approximately one-third (50/157) of the differentially expressed genes affected by bromocriptine overlapped with known p53-p21- retinoblastoma protein (RB) target genes. At the protein level, bromocriptine increased the expression of dopamine D2 receptor (DRD2) and affected several classical and non-classical dopamine receptor signal pathways in chemoresistant PCa cells, including adenosine monophosphate-activated protein kinase (AMPK), p38 mitogen-activated protein kinase (p38 MAPK), nuclear factor kappa B (NF-κB), enhancer of zeste homolog 2 (EZH2), and survivin. As a monotherapy, bromocriptine treatment at 15 mg/kg, three times per week, via the intraperitoneal route significantly inhibited the skeletal growth of chemoresistant C4-2B-TaxR xenografts in athymic nude mice. In summary, these results provided the first preclinical evidence that bromocriptine is a selective and effective inhibitor of chemoresistant PCa. Due to its favorable clinical safety profiles, bromocriptine could be rapidly tested in PCa patients and repurposed as a novel subtype-specific treatment to overcome chemoresistance.

RRNA-Derived Small RNA rsRNA-28S Regulates the Chemoresistance of Prostate Cancer Cells by Targeting PTGIS.

rRNA-derived small RNAs (rsRNAs) represent a novel class of small non-coding RNAs (sncRNAs), produced by the specific cleavage of rRNAs; however, their roles in tumor development are unclear. In the present study, we explored the effect of a kind of rsRNA-28S, which originates from 28S rRNA, on the chemoresistance of prostate cancer cells and the mechanisms underlying its effect. Quantitative reverse transcription PCR (RT-PCR) was performed to quantify rsRNA-28S levels in serum samples taken from prostate cancer patients. DU-145R cells, which are resistant to both paclitaxel and docetaxel, were generated from parental DU-145 cells. Northern blot was conducted to detect cellular rsRNA-28S levels following drug treatments. To verify the effect of rsRNAs-28S on chemoresistance, antisense oligonucleotides were utilized to block rsRNA-28S functions, and a series of assays were further performed, such as cell viability, cell proliferation, colony formation and tumor sphere formation. The target gene of rsRNA-28S was explored using dual-luciferase reporter gene assay. The rsRNA-28S level was reduced in the serum samples of patients who received chemotherapy compared to that of patients who did not. Furthermore, the rsRNA-28S level was remarkably declined in DU-145R cells, and drug treatments decreased the levels of rsRNA-28S in DU-145 and DU-145R cells. Moreover, rsRNA-28S inhibition enhanced the chemoresistance of prostate cancer cells as well as their cancer stem cell characteristics. Mechanistically, the prostaglandin I2 synthase (PTGIS) gene transcript was verified as a target of rsRNA-28S, as rsRNA-28S inhibited the translation of PTGIS mRNA by directly binding the 3' untranslated region of PTGIS mRNA. rsRNA-28S inhibition was also found to increase PTGIS abundance, and PTGIS overexpression significantly enhanced prostate cancer cell chemoresistance. Our findings indicate that rsRNA-28S attenuates prostate cancer cell chemoresistance by downregulating its target gene PTGIS. This study not only greatly contributes to systematic identification and functional elucidation of chemoresistance relevant rsRNAs, but also promotes rsRNA-included combinatorial therapeutic regimens for cancer.

The prognostic implications of macrophage polarization in advanced and therapy-resistant prostate cancer

Abstract Tumor-associated macrophages (TAMs) are components of the spatial microenvironment of many solid neoplasms. In prostate cancer (PCa), TAMs are linked to a poor prognosis. PCa is characterized by enhanced tumor lipid production and TAM lipid accumulation. However, the interplay between lipid-producing tumor cells and TAM lipid uptake has yet to be determined. Human prostate tumor tissue microarrays were evaluated for TAM (CD68+) and M2/M1 (CD206+/CD86+) levels correlated with tumor aggressiveness by multiplex immunohistochemistry. Standard PCa cells 22rv1, PC3 and DU145 were used to develop chemoresistant sublines which were validated using dose-response viability assays and immunoblotting for resistance-associated markers and lipid-producing enzymes. Cancer cell-derived extracellular vesicles (EVs) were isolated from cell culture supernatants by size exclusion chromatography and were incubated with U937-derived macrophages, followed by flow cytometric evaluation of M1 and M2 markers. Proteomic evaluation of human prostate tumors indicates that CD68+ TAMs correlate with aggressive, metastatic PCa. Furthermore, a higher M2-to-M1 ratio also correlates with metastatic disease compared to normal or localized tumors. Chemoresistant PCa cells exhibited an enhanced lipogenic phenotype compared to naive cells and the over-production of lipids was linked to EV-mediated reprogramming of macrophages toward an M2-like phenotype. These findings indicate that alternative activation of macrophages is linked to aggressive, therapy resistant PCa. Moreover, our data suggest that tumor lipid metabolism may influence tumor-associated macrophage phenotype and function through EV-mediated intercellular signaling. Puerto Rico Science Research and Technology Trust

Correction to "A Novel Semisynthetic Inhibitor of the FRB Domain of Mammalian Target of Rapamycin Blocks Proliferation and Triggers Apoptosis in Chemoresistant Prostate Cancer Cells".

Correction to "A Novel Semisynthetic Inhibitor of the FRB Domain of Mammalian Target of Rapamycin Blocks Proliferation and Triggers Apoptosis in Chemoresistant Prostate Cancer Cells".

Abstract 2844: BMP inhibition overcomes chemoresistance of prostate cancer

Abstract Chemoresistance is a major therapeutic challenge to prostate cancer and its underlying molecular mechanism is poorly understood. Previously, it has been suggested that bone morphogenetic protein (BMP) signaling is down-regulated during the prostate cancer progression from the early androgen-sensitive stage to the metastatic castration-resistant stage. However, no literature reports are available for BMP signaling in the more advanced-chemoresistant prostate cancer. In this study, we found the expression levels of the BMP type I receptor members, Activin-like kinase-2 (ALK2) and Activin-like kinase-3 (ALK3), were significantly higher in the chemoresistant prostate cancer cells than those in the chemosensitive prostate cancer cells. In addition, the phospho-Smad1/5/9 proteins, the pivotal intracellular effectors of the BMP signaling, were notably elevated in the chemoresistant prostate cancer cells over the chemosensitive prostate cancer cells, indicating that BMP signaling is highly activated in the chemoresistant prostate cancer cells. We also found that BMP signaling inhibition with either DMH1 or the knockdown of ALK2/ALK3 sensitized chemoresistant prostate cancer cells to the chemotherapy drug docetaxel in a dose dependent manner. Our further study indicates that DMH1 suppressed the migration and invasion of chemoresistant prostate cancer cells in vitro, and attenuated chemoresistant prostate tumor growth in the mouse xenograft model in vivo. In addition, we showed that DMH1 disrupted the sphere formation in DU145-TxR and PC3-TxR cells, suppressed the expression of marker genes of the cancer stem cells (CSCs), and inhibited the efflux activity of multidrug resistance P-glycoprotein (P-gp1). In conclusion, our study demonstrates that BMP signaling is associated with prostate cancer chemoresistance and BMP signaling inhibition effectively overcomes the cancer chemoresistance potentially through the disruption of CSCs’ stemness and P-gp1 efflux activity. Citation Format: Jijun Hao, Chen Xie, Zhijun Wang, Yong Bar, Jose Aguilar, Austin Kyan, Li Zhing. BMP inhibition overcomes chemoresistance of prostate cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2844.

Abstract 1754: Targeting the mitochondrial protein MAGMAS increases sensitivity to docetaxel in neuroendocrine prostate cancer cells

Abstract Metastatic prostate cancer (PCa) is the leading cause of PCa-associated deaths and one of the leading causes of cancer mortality in men. Novel therapeutics, such as Prostate Specific Membrane Antigen (PSMA) ligand therapy, are emerging as effective treatment strategies for metastatic prostate cancer. However, neuroendocrine differentiated (NED) metastatic PCa, which lacks the PSMA antigen, is deprived of effective biomarkers for disease treatment. Magmas, a 13.8 kDa mitochondrial protein, imports nuclear-encoded mitochondrial proteins into the mitochondrial matrix. Magmas is overexpressed in several aggressive tumors, such as breast cancer, glioblastoma, and prostate cancer. When overexpressed, Magmas plays a role as a cytoprotective protein by acting as a reactive oxygen species (ROS) scavenger that maintains sufficient ROS levels to promote cell proliferation, but below the threshold levels that could lead to apoptosis. This study was designed to investigate the pro-survival and therapy-resistance functions of Magmas in PCa cells expressing NED markers. Predictively, we observed that the docetaxel (DTX)-resistant PC3-DR and DU145-DR PCa cell lines expressed significantly higher levels of Magmas compared to their drug-sensitive parental cell lines. We hypothesized that inhibiting Magmas reduces the aggressive properties of highly proliferative DTX- resistant metastatic PCa cell lines and re-sensitize them to DTX. Magmas knockdown in DTX-sensitive and DTX-resistant PC3 and DU145 cells in the presence of increasing DTX treatments enhanced sensitivity to DTX. Consistent with these results, pharmacological inhibition of Magmas with the novel BT#9 inhibitor in combination with increasing DTX concentrations also enhanced sensitivity to DTX in these cell lines. Additional studies using clonogenic assays, cell migration/invasion assays, and tumorsphere formation assays are currently underway to further establish Magmas as a potential therapeutic target to attenuate the aggressive properties of chemoresistant PCa cells. Lastly, we will investigate the diagnostic potential of Magmas as a biomarker for NED and BT#9 as an imaging/therapeutic-specific agent for NED. Citation Format: Alfonso M. Durán, Christian H. Yoo, Jennifer D. Tran, Krystal Santiago, Adil S. Mohammed, Carlos A. Casiano, Frankis A. Almaguel. Targeting the mitochondrial protein MAGMAS increases sensitivity to docetaxel in neuroendocrine prostate cancer cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 1754.

Necroptosis Induced by Delta-Tocotrienol Overcomes Docetaxel Chemoresistance in Prostate Cancer Cells

Prostate cancer (PCa) represents the fifth cause of cancer death in men. Currently, chemotherapeutic agents for the treatment of cancers, including PCa, mainly inhibit tumor growth by apoptosis induction. However, defects in apoptotic cellular responses frequently lead to drug resistance, which is the main cause of chemotherapy failure. For this reason, trigger non-apoptotic cell death might represent an alternative approach to prevent drug resistance in cancer. Several agents, including natural compounds, have been shown to induce necroptosis in human cancer cells. In this study we evaluated the involvement of necroptosis in anticancer activity of delta-tocotrienol (δ-TT) in PCa cells (DU145 and PC3). Combination therapy is one tool used to overcome therapeutic resistance and drug toxicity. Evaluating the combined effect of δ-TT and docetaxel (DTX), we found that δ-TT potentiates DTX cytotoxicity in DU145 cells. Moreover, δ-TT induces cell death in DU145 cells that have developed DTX resistance (DU-DXR) activating necroptosis. Taken together, obtained data indicate the ability of δ-TT to induce necroptosis in both DU145, PC3 and DU-DXR cell lines. Furthermore, the ability of δ-TT to induce necroptotic cell death may represent a promising therapeutical approach to overcome DTX chemoresistance in PCa.

Interaction of pregnane X receptor with hypoxia-inducible factor-1 regulates chemoresistance of prostate cancer cells.

Aim: The nuclear pregnane X receptor (PXR) is a pivotal regulator of steroid and xenobiotics metabolism and plays an important role in shaping tumor cell responses to chemotherapy. Hypoxia within tumor tissue has multifaceted effects, including multiple drug resistance. The goal of this study was to determine whether PXR contributes to hypoxia-induced drug resistance. Methods: Metastatic prostate cancer cells were used to study the interaction of PXR and hypoxia-inducible factor-1 (HIF-1 in drug resistance associated with hypoxia. The activities of PXR and HIF-1 were determined by assays for its reporter gene or target gene expression. Co-immunoprecipitation (Co-IP) was used to determine the interaction of PXR and HIF-1. Ablation or inhibition of PXR or HIF-1 was used to determine their roles in hypoxia-induced chemoresistance. Results: PXR was activated by hypoxia, leading to increased expression of multidrug resistance protein 1 (MDR1). Inhibition of PXR by pharmacological compounds or depletion by shRNAs reduced the hypoxic induction of MDR1 and sensitized prostate cancer cells to chemotherapy under hypoxia. HIF-1 was required for PXR activation under hypoxia. Co-immunoprecipitation results showed that HIF-1 and PXR could physically interact with each other, leading to crosstalk between these two transcription factors. Conclusion: PXR contributes to hypoxia-induced drug resistance in prostate cancer cells through its interaction with HIF-1.

Abstract 5365: Repositioning nicardipine as a novel targeted therapy for chemoresistant prostate cancer

Abstract Background: It is imperative to develop novel therapeutics to overcome chemoresistance, a major obstacle in the clinical management of prostate cancer (PCa) and other cancers. Drug repositioning is being pursued as an attractive approach due to its potentially lower overall development costs and shorter timelines. The purpose of this study is to investigate the preclinical efficacy and mechanism of action of nicardipine, an FDA-approved hypertension drug, as a new treatment against chemoresistant prostate cancer. Methods: A panel of FDA-approved non-oncology drugs was screened for their selectivity and potency to inhibit chemoresistant PCa cells using a two-tier phenotypic screening platform (Theranostics, 2021, 11(14): 6873-6890). The mechanism of action of potential candidate(s) was evaluated using in silico docking, cellular and molecular assays in ARCaPE-shEPLIN and C4-2B-TaxR, two independent chemoresistant PCa cell lines. The in vivo efficacy was evaluated in clinically-relevant xenograft models of PCa. Results: Nicardipine exhibited high selectivity and potency against chemoresistant PCa cells via effectively inducing apoptosis and cell cycle arrest. The IC50 of nicardipine was determined as 0.5 μM and 2.1 μM in ARCaPE-shEPLIN and C4-2B-TaxR cells, respectively. Mechanically, nicardipine targeted a novel non-canonical enhancer of zeste homolog 2 (EZH2) signal pathway in chemoresistant PCa cells. Specifically, nicardipine might bind embryonic ectoderm development (EED), interrupt the integrity and stability of the polycomb repressive complex 2 (PRC2), reduce the phosphorylation and expression of EZH2, and subsequently inhibit downstream effectors including signal transducer and activator of transcription 3 (Stat3), S-phase kinase-associated protein 2 (SKP2), ATP binding cassette B1 (ABCB1) and survivin. In animal models, as a monotherapy, nicardipine significantly suppressed the intratibial growth of chemoresistant C4-2B-TaxR xenografts, with an average PSA level of 43.47 ng/ml, 36.63 ng/ml, and 28.78 ng/ml for the vehicle control, docetaxel, and nicardipine groups at the endpoint, respectively (p < 0.05 between the nicardipine group and the vehicle control or docetaxel groups). As a combination regime, nicardipine synergistically enhanced the efficacy of docetaxel against the intratibial growth of C4-2 xenografts, with an average PSA level of 79.17 ng/ml, 49.04 ng/ml, 42.80 ng/ml, and 23.23 ng/ml for the vehicle control, docetaxel, nicardipine, and combination groups at the endpoint, respectively (p < 0.0001 or 0.01 between the combination group and the vehicle control or docetaxel groups, respectively). Conclusion: These results demonstrated that, for the first time, nicardipine could be a novel EED inhibitor that has the potential to be promptly tested in PCa patients to overcome chemoresistance and improve clinical outcomes. Citation Format: Xin Li, Alira Danaher, Jason M. Wu, Yifei Wu, Zhong-Ru Xie, Nicholas Cook, Nathan Bowen, Daqing Wu. Repositioning nicardipine as a novel targeted therapy for chemoresistant prostate cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 5365.

Abstract 399: The glucocorticoid receptor and β-catenin interact in prostate cancer cells and their co-inhibition attenuates stemness and docetaxel resistance

Abstract The efficacy of anti-androgen receptor (AR) therapies and taxane-based chemotherapy for the treatment of advanced prostate cancer (PCa) is hindered by the development of therapy resistance. Our previous studies demonstrated that docetaxel (DTX)-resistant PCa cells activates a transcriptomic program associated with stemness and display cancer stem cell (CSC) properties, consistent with the notion that CSCs within prostate tumors contribute to chemoresistance. Glucocorticoid receptor (GR) signaling is a critical mediator of resistance to AR-targeted therapy, and has been recently implicated in the development of PCa cell tumorsphere formation and chemoresistance. Like GR, β-catenin is overexpressed in metastatic and therapy-resistant tumors, and is considered a key regulator of cancer stemness and androgen-targeted therapy resistance. Given the structural and functional overlap between GR and AR, a known interacting partner of β-catenin, we hypothesized that GR also interacts with β-catenin to support stemness and chemoresistance in PCa cells. We observed that treatment with the glucocorticoid dexamethasone induces enhanced nuclear accumulation of both GR and β-catenin in DTX-resistant PCa cells compared to their DTX-sensitive counterparts. Knockdown studies revealed that GR and β-catenin do no regulate each other at the protein level. However, using whole cell and nuclear co-immunoprecipitation, we demonstrated the interaction between GR and β-catenin in both DTX-resistant and DTX-sensitive PC3, DU145, and 22RV1 cells. Pharmacological inhibition and RNA interference-mediated silencing of GR with concomitant inhibition of β-catenin enhanced DTX cytotoxicity in resistant PCa cells grown in both adherent and spheroid cultures, and decreased CD44+/CD24- cell populations in tumorspheres. These results indicate that both GR and β-catenin interact and influence stemness, and may be promising therapeutic targets to overcome PCa chemoresistance. Citation Format: Shannalee R. Martinez, Catherine C. Elix, Pedro T. Ochoa, Evelyn S. Sanchez-Hernandez, Hossam R. Alkashgari, Greisha L. Ortiz-Hernandez, Lubo Zhang, Carlos A. Casiano. The glucocorticoid receptor and β-catenin interact in prostate cancer cells and their co-inhibition attenuates stemness and docetaxel resistance [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 399.

Abstract 301: Repurposing engineered thymoquinone nanoparticles to inhibit drug-resistant prostate cancer

Abstract Despite advanced treatment strategies and therapeutics for prostate cancer (PCa), the metastatic castration-resistant prostate cancer (mCRPC) mechanism remains obscure. Although docetaxel (DTX) and other anti-mitotic chemotherapeutic drug are considered the first line drugs for PCa patients; the resulting side effects and drug resistance increase the demand for an alternative such as natural compounds for effective treatment. However, the natural compounds that achieved remarkable height to overcome drug resistance, their bioavailability and targeted delivery during treatment has become a new challenge. As prostate-specific membrane antigen (PSMA) receptors are expressed on the surface of most PCa cells, we anticipated that these receptors may provide the potential target for effective therapy. Considering these, we employed the natural compound, thymoquinone (TQ)-encapsulated planetary ball-milled nanoparticles (PBM-NPs) formulated with starch (FDA approved, natural polysaccharides) inner core and biocompatible and biodegradable co-polymers poly (ε-caprolactone)/poly (ethylene glycol), functionalized with a PSMA aptamer (A10), which can selectively recognize and bind to PSMA membrane. The functionalities of this strategy were analyzed by targeting A10-TQ-PBM-NP in DTX-resistant cells (C4-2B-R and LNCaP-R), which overexpresses PSMA protein and multidrug resistance (MDR) gene ABCB1, and validated through flow cytometry, immunofluorescence, qRT-PCR, and western blot techniques. The results showed that the aptamer-based nano delivery of a TQ can modulate the cellular and molecular pathways responsible for DTX resistance in PCa cells through the EGFR and ZEB1 dependent mechanism. The A10-TQ-PBM-NPs counteract ATP-binding cassette (ABC)-transporter-mediated chemoresistance in PCa cells. In both cell lines, these NPs rapidly internalized and induced significant cell death at five times lower concentrations than non-conjugated PBM-TQ-NP. Further, A10-TQ-PBM-NPs have high selectivity to the PSMA receptor and downregulated the EGFR dependent pathway, Akt-1, ERK1/2, NF-kB, STAT3, and epithelial-mesenchymal transition (EMT) markers in both C4-2B-R and LNCaP-R cells, thus constraining cell survival, proliferation, and migration. The A10-TQ-PBM-NP bioconjugates also reveal remarkable efficacy and reduced toxicity as measured by body weight loss in the LNCaP-R cells xenograft mice. Altogether, our findings demonstrate that targeting PCa cells with PBM-NPs may reverse the MDR and could be used as potential therapeutic options for patients. Citation Format: Santosh Kumar Singh, Manoj Kumar Mishra, Rajesh Singh. Repurposing engineered thymoquinone nanoparticles to inhibit drug-resistant prostate cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 301.

Overcoming prostate cancer drug resistance with a novel organosilicon small molecule.

A major challenge to the treatment of advanced prostate cancer (PCa) is the development of resistance to androgen-deprivation therapy (ADT) and chemotherapy. It is imperative to discover effective therapies to overcome drug resistance and improve clinical outcomes. We have developed a novel class of silicon-containing compounds and evaluated the anticancer activities and mechanism of action using cellular and animal models of drug-resistant PCa. Five organosilicon compounds were evaluated for their anticancer activities in the NCI-60 panel and established drug-resistant PCa cell lines. GH1504 exhibited potent in vitro cytotoxicity in a broad spectrum of human cancer cells, including PCa cells refractory to ADT and chemotherapy. Molecular studies identified several potential targets of GH1504, most notably androgen receptor (AR), AR variant 7 (AR-v7) and survivin. Mechanistically, GH1504 may promote the protein turnover of AR, AR-v7 and survivin, thereby inducing apoptosis in ADT-resistant and chemoresistant PCa cells. Animal studies demonstrated that GH1504 effectively inhibited the in vivo growth of ADT-resistant CWR22Rv1 and chemoresistant C4-2B-TaxR xenografts in subcutaneous and intraosseous models. These preclinical results indicated that GH1504 is a promising lead that can be further developed as a novel therapy for drug-resistant PCa.

The lncRNA OGFRP1/miR-149-5p/IL-6 axis regulates prostate cancer chemoresistance

BackgroundThe long non-coding RNA (lncRNA) OGFRP1 has been found to promote malignancy in prostate cancer (PC) and other cancer types. How this lncRNA functions in the regulation of PC chemoresistance, however, is poorly defined. MethodsqRT-PCR was employed to measure OGFRP1, miR-149-5p, and IL-6 expression in PC tissues and cells. IC50 values for paclitaxel and docetaxel in PC cells were assessed via a CCK-8 assay approach. Putative miR-149-5p binding targets were identified and validated through bioinformatics assays and luciferase reporter assays, respectively. The impact of OGFRP1 on PC chemoresistance in vivo was validated using a xenograft model system. ResultsDocetaxel-resistant PC (PC/DR) cells and tissues exhibited reduced OGFRP1 expression and increased miR-149-5p expression. Knocking down OGFRP1 augmented the sensitivity of these PC cells to docetaxel and paclitaxel in vitro and in vivo. Mechanistically, OGFRP1 was found to bind and sequester miR-149-5p within PC/DR cells, thereby indirectly regulating IL-6 expression. Consistent with this model, the overexpression of IL-6 reversed the OGFRP1 knockdown-mediated reductions in docetaxel and paclitaxel IC50 values for these PC cells. ConclusionsOGFRP1 can sequester miR-149-5p, thereby indirectly promoting IL-6 upregulation and thereby promoting chemoresistance in PC cells. This OGFRP1/miR-149-5p/IL-6 axis may thus be a promising target for therapeutic efforts aimed at PC chemosensitization and treatment.

Role of ZEB Family Members in Proliferation, Metastasis, and Chemoresistance of Prostate Cancer Cells: Revealing Signaling Networks.

Prostate cancer (PCa) is one of the leading causes of death worldwide. A variety of strategies, including surgery, chemotherapy, radiotherapy, and immunotherapy, are applied for PCa treatment. PCa cells are responsive towards therapy at early stages, but they can obtain resistance in the advanced stage. Furthermore, their migratory ability is high in advanced stages. It seems that genetic and epigenetic factors play an important role in this case. Zinc finger E-box-binding homeobox (ZEB) is a family of transcription with two key members, including ZEB1 and ZEB2. ZEB family members are known due to their involvement in promoting cancer metastasis via EMT induction. Recent studies have shown their role in cancer proliferation and inducing therapy resistance. In the current review, we focus on revealing the role of ZEB1 and ZEB2 in PCa. ZEB family members are able to significantly promote the proliferation and viability of cancer cells. ZEB1 and ZEB2 enhance migration and invasion of PCa cells via EMT induction. Overexpression of ZEB1 and ZEB2 is associated with a poor prognosis of PCa. ZEB1 and ZEB2 upregulation occurs during PCa progression and can provide therapy resistance to cancer cells. PRMT1, Smad2, and non-coding RNAs can function as upstream mediators of the ZEB family. Besides, Bax, Bcl-2, MRP1, Ncadherin, and E-cadherin can be considered as downstream targets of the ZEB family in PCa.