Browsing by Author "Pili, Roberto"

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    Androgen Receptor Modulation Optimized for Response (ARMOR) Phase I and II Studies: Galeterone for the Treatment of Castration-Resistant Prostate Cancer
    (AACR, 2016-03) Montgomery, Bruce; Eisenberger, Mario A.; Rettig, Matthew B.; Chu, Franklin; Pili, Roberto; Stephenson, Joseph J.; Vogelzang, Nicholas J.; Koletsky, Alan J.; Nordquist, Luke T.; Edenfield, William J.; Mamlouk, Khalid; Ferrante, Karen J.; Taplin, Mary-Ellen; Department of Medicine, IU School of Medicine
    Purpose: Galeterone is a selective, multitargeted agent that inhibits CYP17, antagonizes the androgen receptor (AR), and reduces AR expression in prostate cancer cells by causing an increase in AR protein degradation. These open-label phase I and II studies [Androgen Receptor Modulation Optimized for Response-1 (ARMOR1) and ARMOR2 part 1] evaluated the efficacy and safety of galeterone in patients with treatment-naive nonmetastatic or metastatic castration-resistant prostate cancer (CRPC) and established a dose for further study. Experimental Design: In ARMOR1, 49 patients received increasing doses (650–2,600 mg) of galeterone in capsule formulation; 28 patients in ARMOR2 part 1 received increasing doses (1,700–3,400 mg) of galeterone in tablet formulation for 12 weeks. Patients were evaluated biweekly for safety and efficacy, and pharmacokinetic parameters were assessed. Results: In ARMOR1, across all doses, 49.0% (24/49) achieved a ≥30% decline in prostate-specific antigen (PSA; PSA30) and 22.4% (11/49) demonstrated a ≥50% PSA decline (PSA50). In ARMOR2 part 1, across all doses, PSA30 was 64.0% (16/25) and PSA50 was 48.0% (12/25). In the 2,550-mg dose cohort, PSA30 was 72.7% (8/11) and PSA50 was 54.5% (6/11). Galeterone was well tolerated; the most common adverse events were fatigue, increased liver enzymes, gastrointestinal events, and pruritus. Most were mild or moderate in severity and required no action and there were no apparent mineralocorticoid excess (AME) events. Conclusions: The efficacy and safety from ARMOR1 and ARMOR2 part 1 and the pharmacokinetic results support the galeterone tablet dose of 2,550 mg/d for further study. Galeterone was well tolerated and demonstrated pharmacodynamic changes consistent with its selective, multifunctional AR signaling inhibition.
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    Anti-tumor activity and mechanistic characterization of APE1/Ref-1 inhibitors in bladder cancer
    (American Association for Cancer Research, 2019-08-14) Fishel, Melissa L.; Xia, Hanyu; McGeown, Jack; McIlwain, David W.; Elbanna, May; Craft, Ariel A.; Kaimakliotis, Hristos Z.; Sandusky, George E.; Zhang, Chi; Pili, Roberto; Kelley, Mark R.; Jerde, Travis J.; Pharmacology and Toxicology, School of Medicine
    Bladder cancer is the ninth most common cause of cancer-related deaths worldwide. Although cisplatin is used routinely in treating bladder cancer, refractory disease remains lethal for many patients. The recent addition of immunotherapy has improved patient outcomes; however, a large cohort of patients does not respond to these treatments. Therefore, identification of innovative molecular targets for bladder cancer is crucial. Apurinic/apyrimidinic endonuclease 1/redox factor-1 (APE1/Ref-1) is a multifunctional protein involved in both DNA repair and activation of transcription factors through reduction-oxidation (redox) regulation. High APE1/Ref-1 expression is associated with shorter patient survival time in many cancer types. In this study, we found high APE1/Ref-1 expression in human bladder cancer tissue relative to benign urothelium. Inhibition of APE1/Ref-1 redox signaling using APE1/Ref-1-specific inhibitors attenuates bladder cancer cell proliferation in monolayer, in three-dimensional cultures, and in vivo. This inhibition corresponds with an increase in apoptosis and decreased transcriptional activity of NF-κB and STAT3, transcription factors known to be regulated by APE1/Ref-1, resulting in decreased expression of downstream effectors survivin and Cyclin D1 in vitro and in vivo. We also demonstrate that in vitro treatment of bladder cancer cells with APE1/Ref-1 redox inhibitors in combination with standard-of-care chemotherapy cisplatin is more effective than cisplatin alone at inhibiting cell proliferation. Collectively, our data demonstrate that APE1/Ref-1 is a viable drug target for the treatment of bladder cancer, provide a mechanism of APE1/Ref-1 action in bladder cancer cells, and support the use of novel redox-selective APE1/Ref-1 inhibitors in clinical studies. SIGNIFICANCE: This work identifies a critical mechanism for APE1/Ref-1 in bladder cancer growth and provides compelling preclinical data using selective redox activity inhibitors of APE1/Ref-1 in vitro and in vivo.
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    Clinical benefit of a precision medicine based approach for guiding treatment of refractory cancers
    (Impact Journals, 2016-08-30) Radovich, Milan; Kiel, Patrick J.; Nance, Stacy M.; Niland, Erin E.; Parsley, Megan E.; Ferguson, Meagan E.; Jiang, Guanglong; Ammakkanavar, Natraj R.; Einhorn, Lawrence H.; Cheng, Liang; Nassiri, Mehdi; Davidson, Darrell D.; Rushing, Daniel A.; Loehrer, Patrick J.; Pili, Roberto; Hanna, Nasser; Callaghan, J. Thomas; Skaar, Todd C.; Helft, Paul R.; Shahda, Safi; O’Neil, Bert H.; Schneider, Bryan P.; Medicine, School of Medicine
    Patients and methods: Patients with metastatic solid tumors who had progressed on at least one line of standard of care therapy were referred to the Indiana University Health Precision Genomics Program. Tumor samples were submitted for DNA & RNA next-generation sequencing, fluorescence in situ hybridization, and immunohistochemistry for actionable targets. A multi-disciplinary tumor board reviewed all results. For each patient, the ratio of progression-free survival (PFS) of the genomically guided line of therapy divided by the PFS of their prior line was calculated. Patients whose PFS ratio was ≥ 1.3 were deemed to have a meaningful improvement in PFS. Results: From April 2014-October 2015, 168 patients were evaluated and 101 patients achieved adequate clinical follow-up for analysis. 19 of 44 (43.2%) patients treated with genomically guided therapy attained a PFS ratio ≥ 1.3 vs. 3 of 57 (5.3%) treated with non-genomically guided therapy (p < 0.0001). Similarly, overall PFS ratios (irrespective of cutoff) were higher for patients with genomically guided therapy vs non-genomically guided therapy (p = 0.05). Further, patients treated with genomically guided therapy had a superior median PFS compared to those treated with non-genomically guided therapy (86 days vs. 49 days, p = 0.005, H.R. = 0.55, 95% C.I.:0.37-0.84). Conclusion: Patients with refractory metastatic cancer who receive genomically guided therapy have improved PFS ratios and longer median PFS compared to patients who do not receive genomically guided therapy.
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    The Combination of Low Skeletal Muscle Mass and High Tumor Interleukin-6 Associates with Decreased Survival in Clear Cell Renal Cell Carcinoma
    (MDPI, 2020-06-17) Kays, Joshua K.; Koniaris, Leonidas G.; Cooper, Caleb A.; Pili, Roberto; Jiang, Guanglong; Liu, Yunlong; Zimmers, Teresa A.; Medical and Molecular Genetics, School of Medicine
    Clear cell renal carcinoma (ccRCC) is frequently associated with cachexia which is itself associated with decreased survival and quality of life. We examined relationships among body phenotype, tumor gene expression, and survival. Demographic, clinical, computed tomography (CT) scans and tumor RNASeq for 217 ccRCC patients were acquired from the Cancer Imaging Archive and The Cancer Genome Atlas (TCGA). Skeletal muscle and fat masses measured from CT scans and tumor cytokine gene expression were compared with survival by univariate and multivariate analysis. Patients in the lowest skeletal muscle mass (SKM) quartile had significantly shorter overall survival versus the top three SKM quartiles. Patients who fell into the lowest quartiles for visceral adipose mass (VAT) and subcutaneous adipose mass (SCAT) also demonstrated significantly shorter overall survival. Multiple tumor cytokines correlated with mortality, most strongly interleukin-6 (IL-6); high IL-6 expression was associated with significantly decreased survival. The combination of low SKM/high IL-6 was associated with significantly lower overall survival compared to high SKM/low IL-6 expression (26.1 months vs. not reached; p < 0.001) and an increased risk of mortality (HR = 5.95; 95% CI = 2.86–12.38). In conclusion, tumor cytokine expression, body composition, and survival are closely related, with low SKM/high IL-6 expression portending worse prognosis in ccRCC.
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    Combined inhibition of Ref‐1 and STAT3 leads to synergistic tumour inhibition in multiple cancers using 3D and in vivo tumour co‐culture models
    (Wiley, 2021-01) Caston, Rachel A.; Shah, Fenil; Starcher, Colton L.; Wireman, Randall; Babb, Olivia; Grimard, Michelle; McGeown, Jack; Armstrong, Lee; Tong, Yan; Pili, Roberto; Rupert, Joseph; Zimmers, Teresa A.; Elmi, Adily N.; Pollok, Karen E.; Motea, Edward A.; Kelley, Mark R.; Fishel, Melissa L.; Pediatrics, School of Medicine
    With a plethora of molecularly targeted agents under investigation in cancer, a clear need exists to understand which pathways can be targeted simultaneously with multiple agents to elicit a maximal killing effect on the tumour. Combination therapy provides the most promise in difficult to treat cancers such as pancreatic. Ref‐1 is a multifunctional protein with a role in redox signalling that activates transcription factors such as NF‐κB, AP‐1, HIF‐1α and STAT3. Formerly, we have demonstrated that dual targeting of Ref‐1 (redox factor‐1) and STAT3 is synergistic and decreases cell viability in pancreatic cancer cells. Data presented here extensively expands upon this work and provides further insights into the relationship of STAT3 and Ref‐1 in multiple cancer types. Using targeted small molecule inhibitors, Ref‐1 redox signalling was blocked along with STAT3 activation, and tumour growth evaluated in the presence and absence of the relevant tumour microenvironment. Our study utilized qPCR, cytotoxicity and in vivo analysis of tumour and cancer‐associated fibroblasts (CAF) response to determine the synergy of Ref‐1 and STAT3 inhibitors. Overall, pancreatic tumours grown in the presence of CAFs were sensitized to the combination of STAT3 and Ref‐1 inhibition in vivo. In vitro bladder and pancreatic cancer demonstrated the most synergistic responses. By disabling both of these important pathways, this combination therapy has the capacity to hinder crosstalk between the tumour and its microenvironment, leading to improved tumour response.
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    Data-Independent Acquisition Phosphoproteomics of Urinary Extracellular Vesicles Enables Renal Cell Carcinoma Grade Differentiation
    (Elsevier, 2023) Hadisurya, Marco; Lee, Zheng-Chi; Luo, Zhuojun; Zhang, Guiyuan; Ding, Yajie; Zhang, Hao; Iliuk, Anton B.; Pili, Roberto; Boris, Ronald S.; Tao, W. Andy; Urology, School of Medicine
    Translating the research capability and knowledge in cancer signaling into clinical settings has been slow and ineffective. Recently, extracellular vesicles (EVs) have emerged as a promising source for developing disease phosphoprotein markers to monitor disease status. This study focuses on the development of a robust data-independent acquisition (DIA) using mass spectrometry to profile urinary EV phosphoproteomics for renal cell cancer (RCC) grades differentiation. We examined gas-phase fractionated library, direct DIA (library-free), forbidden zones, and several different windowing schemes. After the development of a DIA mass spectrometry method for EV phosphoproteomics, we applied the strategy to identify and quantify urinary EV phosphoproteomes from 57 individuals representing low-grade clear cell RCC, high-grade clear cell RCC, chronic kidney disease, and healthy control individuals. Urinary EVs were efficiently isolated by functional magnetic beads, and EV phosphopeptides were subsequently enriched by PolyMAC. We quantified 2584 unique phosphosites and observed that multiple prominent cancer-related pathways, such as ErbB signaling, renal cell carcinoma, and regulation of actin cytoskeleton, were only upregulated in high-grade clear cell RCC. These results show that EV phosphoproteome analysis utilizing our optimized procedure of EV isolation, phosphopeptide enrichment, and DIA method provides a powerful tool for future clinical applications.
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    EZH2 modifies sunitinib resistance in renal cell carcinoma by kinome reprogramming
    (Cancer Research, 2017-12-01) Adelaiye-Ogala, Remi; Budka, Justin; Damayanti, Nur P.; Arrington, Justine; Ferris, Mary; Hsu, Chuan-Chih; Chintala, Sreenivasulu; Orillion, Ashley; Miles, Kiersten Marie; Shen, Li; Elbanna, May; Ciamporcero, Eric; Arisa, Sreevani; Pettazzoni, Piergiorgio; Draetta, Giulio F.; Seshadri, Mukund; Hancock, Bradley; Radovich, Milan; Kota, Janaiah; Buck, Michael; Keilhack, Heike; McCarthy, Brian P.; Persohn, Scott A.; Territo, Paul R.; Zang, Yong; Irudayaraj, Joseph; Tao, W. Andy; Hollenhorst, Peter; Pili, Roberto
    Acquired and intrinsic resistance to receptor tyrosine kinase inhibitors (RTKi) represent a major hurdle in improving the management of clear cell renal cell carcinoma (ccRCC). Recent reports suggest that drug resistance is driven by tumor adaptation via epigenetic mechanisms that activate alternative survival pathways. The histone methyl transferase EZH2 is frequently altered in many cancers including ccRCC. To evaluate its role in ccRCC resistance to RTKi, we established and characterized a spontaneously metastatic, patient-derived xenograft (PDX) model that is intrinsically resistant to the RTKI sunitinib but not to the VEGF therapeutic antibody bevacizumab. Sunitinib maintained its anti-angiogenic and anti-metastatic activity but lost its direct anti-tumor effects due to kinome reprogramming, which resulted in suppression of pro- apoptotic and cell cycle regulatory target genes. Modulating EZH2 expression or activity suppressed phosphorylation of certain RTK, restoring the anti-tumor effects of sunitnib in models of acquired or intrinsically resistant ccRCC. Overall, our results highlight EZH2 as a rational target for therapeutic intervention in sunitinib-resistant ccRCC as well as a predictive marker for RTKi response in this disease.
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    GCN2 eIF2 kinase promotes prostate cancer by maintaining amino acid homeostasis
    (eLife Sciences, 2022-09-15) Cordova, Ricardo A.; Misra, Jagannath; Amin, Parth H.; Klunk, Anglea J.; Damayanti, Nur P.; Carlson, Kenneth R.; Elmendorf, Andrew J.; Kim, Hyeong-Geug; Mirek, Emily T.; Elzey, Bennet D.; Miller, Marcus J.; Dong, X. Charlie; Cheng, Liang; Anthony, Tracy G.; Pili, Roberto; Wek, Ronald C.; Staschke, Kirk A.; Biochemistry and Molecular Biology, School of Medicine
    A stress adaptation pathway termed the integrated stress response has been suggested to be active in many cancers including prostate cancer (PCa). Here, we demonstrate that the eIF2 kinase GCN2 is required for sustained growth in androgen-sensitive and castration-resistant models of PCa both in vitro and in vivo, and is active in PCa patient samples. Using RNA-seq transcriptome analysis and a CRISPR-based phenotypic screen, GCN2 was shown to regulate expression of over 60 solute-carrier (SLC) genes, including those involved in amino acid transport and loss of GCN2 function reduces amino acid import and levels. Addition of essential amino acids or expression of 4F2 (SLC3A2) partially restored growth following loss of GCN2, suggesting that GCN2 targeting of SLC transporters is required for amino acid homeostasis needed to sustain tumor growth. A small molecule inhibitor of GCN2 showed robust in vivo efficacy in androgen-sensitive and castration-resistant mouse models of PCa, supporting its therapeutic potential for the treatment of PCa.
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    GCN2 eIF2 Kinase Promotes Prostate Cancer by Maintaining Amino Acid Homeostasis
    (2024-04) Cordova E., Ricardo A.; Wek, Ronald C.; Staschke, Kirk A.; Pili, Roberto; Mosley, Amber L.; Elmendorf, Jeffrey S.; Zhang, Ji
    Activation of the integrated stress response (ISR) contributes to the progression of many cancers, including prostate cancer (PCa). The ISR features a family of protein kinases that phosphorylate the eukaryotic translation initiation factor 2 (eIF2) during different stress conditions, repressing global protein synthesis. In parallel, eIF2 phosphorylation also enhances the translation of select gene transcripts, such as ATF4, which directs the transcription of ISR-target genes critical for stress adaptation. We reported that the eIF2 kinase GCN2 is a critical driver of the ISR in PCa and is crucial to maintaining amino acid (AA) homeostasis. GCN2 is activated in PCa due to AA limitation, resulting in increased expression of key AA transporters which providing nutrient import to fuel protein synthesis and metabolism that drive prostate tumor cell proliferation. Inhibition of GCN2 results in lowered expression of AA transporters, leading to severe depletion of intracellular AA and reduced proliferation in PCa. We identified purine biosynthesis as a key metabolic pathway dependent on GCN2. Inhibition of GCN2 and the accompanying depletion of AAs decreases purine levels in PCa cells, ultimately resulting in reduced ribosome biogenesis leading to the activation of a p53-dependent cell cycle checkpoint, termed the Impaired Ribosome Biogenesis Checkpoint (IRBC). Interestingly, induction of p53 promotes survival of PCa following GCN2 inhibition by halting cell cycle progression and reprogramming metabolism to restore metabolic homeostasis. We found that reductions in select AAs that impact nucleotide pools activate GCN2 and p53 in parallel, and that cooperation of these stress pathways is critical for maintaining AA and purine pools. Of importance, deletion of p53 sensitizes PCa cells to GCN2 inhibition suggesting that loss of p53 creates a dependency for GCN2. Of importance, we demonstrate that a small molecule inhibitor of GCN2 showed robust in vivo efficacy in androgen-sensitive and castrationresistant mouse models of PCa, supporting its therapeutic potential for the treatment of PCa.
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    Genomic profiling of collecting duct renal carcinoma
    (Impact Journals, LLC, 2016-10-26) Chintala, Sreenivasulu; Pili, Roberto; Department of Neurological Surgery, School of Medicine