Baseline serum testosterone and differential efficacy of bipolar androgen therapy and enzalutamide in the randomized TRANSFORMER trial.

Bipolar androgen therapy (BAT) is effective in a subset of metastatic castration-resistant prostate cancer (mCRPC) patients. Treatment selection biomarkers are needed due to other therapies that can be equally efficacious. We performed post-hoc analysis to determine whether baseline serum testosterone (T) is a treatment selection marker in the TRANSFORMER study, a randomized trial of abiraterone-pretreated mCRPC patients assigned to BAT (n = 94) or enzalutamide (n = 101). The findings suggest that patients with poor outcomes to abiraterone and serum T ≥ 20 ng/dL may benefit preferentially from BAT over enzalutamide. Baseline testosterone could be considered in the treatment selection process when BAT is an option.

Characterization of structural, biochemical, pharmacokinetic, and pharmacodynamic properties of the LSD1 inhibitor bomedemstat in preclinical models.

INTRODUCTION: Lysine-specific demethylase 1 (LSD1) is emerging as a critical mediator of tumor progression in metastatic castration-resistant prostate cancer (mCRPC). Neuroendocrine prostate cancer (NEPC) is increasingly recognized as an adaptive mechanism of resistance in mCRPC patients failing androgen receptor axis-targeted therapies. Safe and effective LSD1 inhibitors are necessary to determine antitumor response in prostate cancer models. For this reason, we characterize the LSD1 inhibitor bomedemstat to assess its clinical potential in NEPC as well as other mCRPC pathological subtypes. METHODS: Bomedemstat was characterized via crystallization, flavine adenine dinucleotide spectrophotometry, and enzyme kinetics. On-target effects were assessed in relevant prostate cancer cell models by measuring proliferation and H3K4 methylation using western blot analysis. In vivo, pharmacokinetic (PK) and pharmacodynamic (PD) profiles of bomedemstat are also described. RESULTS: Structural, biochemical, and PK/PD properties of bomedemstat, an irreversible, orally-bioavailable inhibitor of LSD1 are reported. Our data demonstrate bomedemstat has >2500-fold greater specificity for LSD1 over monoamine oxidase (MAO)-A and -B. Bomedemstat also demonstrates activity against several models of advanced CRPC, including NEPC patient-derived xenografts. Significant intra-tumoral accumulation of orally-administered bomedemstat is measured with micromolar levels achieved in vivo (1.2 ± 0.45 µM at the 7.5 mg/kg dose and 3.76 ± 0.43 µM at the 15 mg/kg dose). Daily oral dosing of bomedemstat at 40 mg/kg/day is well-tolerated, with on-target thrombocytopenia observed that is rapidly reversible following treatment cessation. CONCLUSIONS: Bomedemstat provides enhanced specificity against LSD1, as revealed by structural and biochemical data. PK/PD data display an overall safety profile with manageable side effects resulting from LSD1 inhibition using bomedemstat in preclinical models. Altogether, our results support clinical testing of bomedemstat in the setting of mCRPC.

Therapeutic Approaches to Targeting Androgen Receptor Splice Variants.

Therapeutic options for advanced prostate cancer have vastly expanded over the last decade and will continue to expand in the future. Drugs targeting the androgen receptor (AR) signaling pathway, i.e., androgen receptor targeting agents (ARTAs), remain the mainstream treatments that are increasingly transforming the disease into one that can be controlled for an extended period of time. Prostate cancer is inherently addicted to AR. Under the treatment pressure of ARTA, molecular alterations occur, leading to the clonal expansion of resistant cells in a disease state broadly categorized as castration-resistant prostate cancer (CRPC). One castration resistance mechanism involves AR splice variants (AR-Vs) lacking the ligand-binding domain. Some AR-Vs have been identified as constitutively active, capable of activating AR signaling pathways without androgenic ligands. Among these variants, AR-V7 is the most extensively studied and may be measured non-invasively using validated circulating tumor cell (CTC) tests. In the context of the evolving prostate cancer treatment landscape, novel agents are developed and evaluated for their efficacy in targeting AR-V7. In patients with metastatic CRPC (mCRPC), the availability of the AR-V7 tests will make it possible to determine whether the treatments are effective for CTC AR-V7-positive disease, even though the treatments may not be specifically designed to target AR-V7. In this review, we will first outline the current prostate cancer treatment landscape, followed by an in-depth review of relatively newer prostate cancer therapeutics, focusing on AR-targeting agents under clinical development. These drugs are categorized from the standpoint of their activities against AR-V7 through direct or indirect mechanisms.

Clinical and Functional Analyses of an African-ancestry Gain-of-function HOXB13 Variant Implicated in Aggressive Prostate Cancer.

BACKGROUND: Recent reports have uncovered a HOXB13 variant (X285K) predisposing to prostate cancer in men of West African ancestry. The clinical relevance and protein function associated with this inherited variant are unknown. OBJECTIVE: To determine the clinical relevance of HOXB13 (X285K) in comparison with HOXB13 (G84E) and BRCA2 pathogenic/likely pathogenic (P/LP) variants, and to elucidate the oncogenic mechanisms of the X285K protein. DESIGN, SETTING, AND PARTICIPANTS: Real-world data were collected from 21,393 men with prostate cancer undergoing genetic testing from 2019 to 2022, and in vitro cell-line models were established for the evaluation of oncogenic functions associated with the X285K protein. OUTCOME MEASUREMENTS AND STATISTICAL ANALYSIS: Genetic testing results were compared among patient groups according to self-reported race/ethnicity, Gleason scores, and American Joint Committee on Cancer stages using the exact test. Oncogenic functions of X285K were evaluated by RNA sequencing, chromatin immunoprecipitation sequencing, and Western blot analyses. RESULTS AND LIMITATIONS: HOXB13 (X285K) was significantly enriched in self-reported Black (1.01%) versus White (0.01%) patients. We observed a trend of more aggressive disease in the HOXB13 (X285K) and BRCA2 P/LP carriers than in the HOXB13 (G84E) carriers. Replacement of the wild-type HOXB13 protein with the X285K protein resulted in a gain of an E2F/MYC signature, validated by the elevated expression of cyclin B1 and c-Myc, without affecting the androgen response signature. Elevated expression of cyclin B1 and c-Myc was explained by enhanced binding of the X285K protein to the promoters and enhancers of these genes. The limitations of the study are the lack of complete clinical outcome data for all patients studied and the use of a single cell line in the functional analysis. CONCLUSIONS: HOXB13 (X285K) is significantly enriched in self-reported Black patients, and X285K carriers detected in the real-world clinical setting have aggressive prostate cancer features similar to the BRCA2 carriers. Functional studies revealed a unique gain-of-function oncogenic mechanism of X285K protein in regulating E2F/MYC signatures. PATIENT SUMMARY: The HOXB13 (X285K) variant is clinically and functionally linked to aggressive prostate cancer, supporting genetic testing for X285K in Black men and early disease screening of carriers of this variant.

CD38-Induced Apoptosis and Mitochondrial Damage is Restored by Nicotinamide in Prostate Cancer.

Nicotinamide adenine dinucleotide (NAD+) is an essential molecule for living organisms. CD38 is a key NAD+-dependent enzyme which breaks down NAD+ to cyclic ADP-ribose (ADPR) and nicotinamide (NAM, vitamin B3), and NAM can be recycled to synthesize NAD+. CD38 expression is consistently silenced by methylation in prostate cancer and progressively downregulated in advanced castration-resistant prostate cancer, suggesting a connection between NAD+ and prostate carcinogenesis as well as prostate cancer progression. However, the functional interplay between NAD+, CD38, and NAM remains largely uncharacterized in prostate cancer cells. In this study, we generated stable LNCaP95 cell clones expressing varying levels of CD38 upon induction by doxycycline. We demonstrate that CD38 overexpression resulted in growth suppression and apoptosis accompanied by cleavage of poly (ADP-ribose) polymerase 1 (PARP1). CD38 overexpression also dramatically reduced intracellular NAD+ levels and decreased mitochondrial respiration as measured by oxygen consumption rate. We further show that some but not all of these CD38-induced phenotypes could be rescued by exogenous NAM. Treatment of cells with NAM rescued CD38-induced apoptosis and mitochondrial stress but did not restore intracellular NAD+ levels. We also found that NAM demonstrated biphasic effect on mitochondria function, a finding that can be explained by the dual role of NAM as both a precursor of NAD+ and also as a suppressor of a number of NAD+-dependent enzymes. Collectively, these findings provide additional insight supporting the functional relevance of CD38 loss in prostate cancer by linking cell-autonomous regulation of mitochondrial function and prostate cancer.

Delineating the Molecular Events Underlying Development of Prostate Cancer Variants with Neuroendocrine/Small Cell Carcinoma Characteristics.

The treatment landscape of prostate cancer has changed dramatically following the advent of novel systemic therapies, most of which target the androgen receptor (AR). Agents such as abiraterone, enzalutamide, apalutamide, darolutamide were designed to further suppress androgen receptor signaling following gonadal suppression achieved by first-line androgen deprivation therapies. These potent AR targeting agents are increasingly used in the earlier stages of the disease spectrum with the goal of delaying disease progression and extending survival. Although these therapies are effective in controlling prostate tumors dependent on or addicted to AR signaling, prostate tumors surviving the onslaught of potent treatments may evolve and develop drug resistance. A substantial proportion of treatment failures can be explained by the development of treatment-induced aggressive prostate cancer variants such as neuroendocrine/small cell carcinoma. These emerging disease entities demand detailed characterization and precise definitions. We postulate that these treatment-induced prostate cancer entities should be defined molecularly to overcome the drawbacks associated with the current clinical and pathological definitions. A precise molecular definition conforms with current knowledge on the molecular evolution of this disease entity and will enable early detection and early intervention.

AR Splicing Variants and Resistance to AR Targeting Agents.

Over the past decade, advances in prostate cancer research have led to discovery and development of novel biomarkers and effective treatments. As treatment options diversify, it is critical to further develop and use optimal biomarkers for the purpose of maximizing treatment benefit and minimizing unwanted adverse effects. Because most treatments for prostate cancer target androgen receptor (AR) signaling, aberrations affecting this drug target are likely to emerge following the development of castration-resistant prostate cancer (CRPC), and it is conceivable that such aberrations may play a role in drug resistance. Among the many AR aberrations, we and others have been studying androgen receptor splice variants (AR-Vs), especially AR-V7, and have conducted preclinical and clinical studies to develop and validate the clinical utility of AR-V7 as a prognostic and potential predictive biomarker. In this review, we first describe mechanisms of AR-V generation, regulation and their functions from a molecular perspective. We then discuss AR-Vs from a clinical perspective, focusing on the significance of AR-Vs detected in different types of human specimens and AR-Vs as potential therapeutic targets.

Abiraterone Rechallenge Based on Sequential Testing of Androgen Receptor Splice Variant 7 Expression in Circulating Tumor Cells: A Case Report.

Serial analysis of circulating tumor cells (CTCs) such as androgen receptor splice variant 7 is useful in selecting treatments for castration-resistant prostate cancer (CRPC). We report a case who had been positive for androgen receptor splice variant 7 in CTCs before docetaxel, and was subsequently treated with abiraterone rechallenge because of the negative conversion of androgen receptor splice variant 7 following docetaxel. Although, the rechallenge of anti-androgen agent based on CTCs analysis is expected to be an effective approach, it is yet to be reported. Thus, we chose the candidate for abiraterone rechallenge based on serial CTCs analyses by the AdnaTest. As a result, the patient responded to abiraterone that he once had developed resistance to. Our findings reinforce the utility of AR-V7 as a biomarker in the setting of post-chemo androgen-targeted-therapy rechallenge.

Prostate-specific membrane antigen in circulating tumor cells is a new poor prognostic marker for castration-resistant prostate cancer.

The aim of this study is to elucidate the clinical significance of prostate-specific membrane antigen (PSMA) expression in circulating tumor cells (CTCs) from castration-resistant prostate cancer (CRPC) patients. We analyzed a total of 203 CTC samples from 79 CRPC patients to investigate the proportion of positive mRNA expressions at different treatment phases. Among them, we elected to focus on specimens from 56 CRPC patients who progressed on therapy and were subsequently provided a new treatment (treatment-switch cohort). In this cohort, we investigated the association between PSMA expression in CTCs and treatment response. CTCs were detected in 55/79 patients and median serum PSA in CTC-positive patients was 67.0 ng/ml. In the treatment-switch cohort of 56 patients, 20 patients were positive for PSMA in CTCs. PSMA expression was inversely associated with percentage of change in prostate-specific antigen (PSA). The median PSA progression-free survival and overall survival were significantly shorter in the PSMA-positive cohort. Furthermore, PSMA expression was predictive of poorer treatment response, shorter PSA progression-free survival and overall survival. PSMA expression in circulating tumor cells may be a novel poor prognostic marker for CRPC.

Movement of target organs and anatomical landmarks caused by body position change during urological laparoscopic surgery.

INTRODUCTION: The aim of our study was to elucidate the impact of patients' physical characteristics on the movement of target organs and anatomical landmarks by comparing supine and lateral CT images. METHODS: This study consisted of 55 patients who underwent laparoscopic surgery in the lateral position. CT images of the area between the abdomen and pelvis were taken preoperatively with patients in both supine and lateral positions. We measured the moving distance of target organs and anatomical landmarks on the body surface used for access port settings. We investigated which covariates from patients' body composition most affected moving distance in correlation analysis. Then, using multiple linear regression analysis, we examined whether we could predict the movement of target organs and anatomical landmarks solely based on information obtained from supine CT images. RESULTS: The moving distance of both the hilum of the kidney and the outer edge of the rectus abdominis muscle were significantly associated with some physical characteristics. Multiple regression analysis showed that a larger visceral fat area could be a useful index for predicting the movement of the kidney toward the counter side. Lower CT density of back muscles and higher BMI could be useful indexes for predicting the movement of the rectus abdominis muscle. CONCLUSION: Our results suggested that body composition characteristics obtained from preoperative CT images can help predict the movement of target organs and anatomical landmarks used to determine proper port-site placement for laparoscopic surgery performed with the patient in the lateral position.

Significant association between urethral length measured by magnetic resonance imaging and urinary continence recovery after robot-assisted radical prostatectomy.

INTRODUCTION: To determine the clinical predictive factors affecting the recovery from postoperative urinary incontinence after robot-assisted radical prostatectomy (RARP). MATERIALS AND METHODS: We consecutively analyzed 320 patients who underwent RARP between January 2012 and March 2015. The restoration of urinary continence was defined as follows: the use of no pads/no leakage of urine or the use of a safety pad. Preoperative covariates were statistically assessed by multivariate logistic regression analysis to investigate their predict factor to recovery of urinary incontinence. Therefore, in this study, we sought to identify predictors of early urinary continence status in a single-center retrospective study of consecutive patients who underwent RARP. RESULTS: Continence rates at 1, 3, 6, and 12 months after the catheter was removed were 44%, 71%, 83%, and 93%, respectively. Age, body mass index, and prostate volume had no significant association with urinary continence recovery. In contrast to this, longer preoperative membranous urethral length (MUL) was significantly associated with earlier postoperative continence recovery. Multivariate analysis demonstrated that longer preoperative MUL is significantly associated with continence recovery at 1 month (P = 0.0235). CONCLUSION: Approximately 70% of patients achieved urinary continence within 3 months after RARP. Multivariate analysis showed that age, body mass index, and prostate volume had no significant association with urinary continence recovery. Preoperative MUL assessed by magnetic resonance imaging was an independent predictor of early recovery from urinary incontinence after RARP.

Sarcopenia is a poor prognostic factor of castration-resistant prostate cancer treated with docetaxel therapy.

BACKGROUND: Sarcopenia is a geriatric syndrome that is characterized by the gradual muscle loss and frailty in the elderly. Meanwhile, the prevalence of prostate cancer is on the rise worldwide. Mainstay treatments for metastatic prostate cancer are androgen-deprivation therapy and taxane-based chemotherapy. Owing to the indolent nature of prostate cancer, these treatments tend to be long-lasting, giving rise to the problem of tolerance to the treatments. Especially given the fact that long-term chemotherapy is closely associated with muscle loss, we aimed to elucidate the correlation between chemotherapy and sarcopenia in the clinical setting. MATERIALS AND METHODS: This study was a retrospective study. Participants with castration-resistant prostate cancer were recruited from November 2009 to September 2015.Participants were recruited at two hospitals, Juntendo and Teikyo University Hospital, Tokyo, Japan.Participants were 77 Japanese males with castration-resistant prostate cancer who underwent docetaxel chemotherapy.Sarcopenia was defined as L3-psoas muscle index < 5.7 cm2/m2. We statistically investigated whether the existence of sarcopenia has an impact on the survival time, and identified potential covariates that affect it. RESULTS: Out of 77 patients, 26 patients (34%) were diagnosed as sarcopenia. Analysis showed that sarcopenia is independently associated with mortality risk (hazards ratio = 2.74, P = 0.0055). Sarcopenic patients showed significant decrease in body mass index, pretreatment hemoglobin, C-related protein, and L3-psoas muscle index as compared with nonsarcopenic patients. The median observation period was 499 days (330-790). Thirty-five patients (45%) died of prostate cancer during that period. Sarcopenic patients showed significantly shorter survival time after the initiation of docetaxel treatments (P = 0.0055). CONCLUSION: Sarcopenia is an independent predictive factor for a poor tolerance to docetaxel treatment. Given that cessation of the treatment leads to death from the disease, our study identified sarcopenia as an independent factor that raises mortality risk.

Generation of transgenic mouse line with prostate-specific expression of codon-improved Cre recombinase.

BACKGROUND: Genetically engineered mouse models are useful tools to decipher molecular mechanisms of diseases. As for prostates, a rat probasin promoter has been widely used to drive prostate-specific gene expression. To optimize its codon usage to that of mammals, we used codon-improved Cre recombinase (iCre) for prostate-specific Cre-loxP recombination. MATERIALS AND METHODS: We generated transgenic mice that express iCre driven by conventional probasin promoter in a prostate-specific manner (PB-iCre). Linearized PB-iCre transgene deoxyribonucleic acids (DNAs) were microinjected into pronuclei of fertilized mouse embryos. The integration of the transgene was confirmed by Southern blot analysis. A line of transgenic mice expressing a sufficient amount of iCre mRNA in its prostate was selected. To test recombinase activity of PB-iCre in vivo, its offspring was crossbred with Ptenflox/flox mice in which murine prostate adenocarcinoma is reported to occur upon excision of loxP-flanked regions. RESULTS: Eight founder animals were obtained, all of which showed germ line integration of PB-iCre transgene by Southern blot analysis. Among them, the prostate from only one line (line 58) expressed a sufficient amount of iCre mRNA. This line was crossbred with Ptenflox/flox mice to generate PB-iCre58/Ptenflox/flox. As a result, 12-week-old PB-iCre58/Ptenflox/flox mice presented with prostate adenocarcinoma that was histologically similar to human cribriform prostate cancer of Gleason grade 4. CONCLUSIONS: We have successfully established a transgenic mouse line that expresses iCre in a prostate-specific manner.

The Surge in the Number of Circulating Tumor Cells Following Treatment with Sunitinib for Metastatic Renal Cell Carcinoma.

Circulating tumor cells (CTCs) are a promising biomarker for several cancers. We streamlined the experimental procedure of CTC immunofluorescent staining. We encountered a 72-year-old woman with metastatic right renal cell carcinoma (RCC) (clinical stage: T4N0M1), whose CTC number rapidly increased after the administration of sunitinib and then gradually decreased. The change in the CTC number appeared to coincide with laboratory data and hypertension, suggesting that a CTC analysis may be useful for promptly monitoring the treatment response. Our data provided the first evidence of an association between the CTC numbers and the treatment response in a metastatic RCC patient.

Role of Androgen Receptor Variants in Prostate Cancer: Report from the 2017 Mission Androgen Receptor Variants Meeting.

CONTEXT: Although a number of studies have demonstrated the importance of constitutively active androgen receptor variants (AR-Vs) in prostate cancer, questions still remain about the precise role of AR-Vs in the progression of castration-resistant prostate cancer (CRPC). OBJECTIVE: Key stakeholders and opinion leaders in prostate cancer convened on May 11, 2017 in Boston to establish the current state of the field of AR-Vs. EVIDENCE ACQUISITION: The meeting "Mission Androgen Receptor Variants" was the second of its kind sponsored by the Prostate Cancer Foundation (PCF). This invitation-only event was attended by international leaders in the field and representatives from sponsoring organizations (PCF and industry sponsors). Eighteen faculty members gave short presentations, which were followed by in-depth discussions. Discussions focused on three thematic topics: (1) potential of AR-Vs as biomarkers of therapeutic resistance; (2) role of AR-Vs as functionally active CRPC progression drivers; and (3) utility of AR-Vs as therapeutic targets in CRPC. EVIDENCE SYNTHESIS: The three meeting organizers synthesized this meeting report, which is intended to summarize major data discussed at the meeting and identify key questions as well as strategies for addressing these questions. There was a critical consensus that further study of the AR-Vs is an important research focus in CRPC. Contrasting views and emphasis, each supported by data, were presented at the meeting, discussed among the participants, and synthesized in this report. CONCLUSIONS: This article highlights the state of knowledge and outlines the most pressing questions that need to be addressed to advance the AR-V field. PATIENT SUMMARY: Although further investigation is needed to delineate the role of androgen receptor (AR) variants in metastatic castration-resistant prostate cancer, advances in measurement science have enabled development of blood-based tests for treatment selection. Detection of AR variants (eg, AR-V7) identified a patient population with poor outcomes to existing AR-targeting therapies, highlighting the need for novel therapeutic agents currently under development.

Hyperactive mTOR induces neuroendocrine differentiation in prostate cancer cell with concurrent up-regulation of IRF1.

BACKGROUND: Neuroendocrine-differentiated prostate cancer (NEPCa) is refractory to androgen deprivation therapy and shows a poor prognosis. The underlying mechanisms responsible for neuroendocrine differentiation (NED) are yet to be clarified. In this study, we investigated the role of mammalian target of rapamycin (mTOR) in NEPCa. METHODS: We utilized a gain-of-function analysis by establishing a human PCa LNCaP stable line that expresses hyperactive mTOR (LNCaP-mTOR). Then, we employed a comprehensive mass spectrometric analysis to identify a key transcription factor in LNCaP-mTOR, followed by a loss-of-function analysis using CRISPR/Cas system. RESULTS: The activation of mTOR induced NED. We observed significant cell growth arrest in NED of LNCaP-mTOR, which accompanied increased expression of p21WAF1/CIP1 . A comprehensive mass spectrometric analysis identified interferon regulatory factor 1 (IRF1) as a key transcription factor in growth arrest of LNCaP-mTOR. The disruption of IRF1 gene in LNCaP-mTOR reversed cell growth arrest along with the suppression of its target p21WAF1/CIP1 . These results indicate that the growth arrest in NED is at least in part dependent on IRF1 through the induction of p21WAF1/CIP1 . CONCLUSIONS: We identified active mTOR as a novel inducer of NED, and elucidated a mechanism underlying the malignant transformation of NEPCa by recapitulating NED in vitro.

Aryl hydrocarbon receptor signaling involved in the invasiveness of LNCaP cells.

There is now mounting evidence that the aryl hydrocarbon receptor (AhR) plays an important role in physiologic responses such as development, cell cycle regulation, immune function and also malignant transformation in various tissues. The strong nuclear AhR expression is observed in the invasive phenotype, and an elevated nuclear AhR expression is associated with a poor prognosis of human prostate cancer. On the other hand, there are conflicting results that the AhR deficiency results in increased susceptibility to prostate tumors in mouse model. In the present study, we investigated AhR expression and its role in the growth and invasiveness of human prostate cancer cells. The AhR protein expression was detected in prostate cancer cell lines and human prostate cancer tissues. A small interfering RNA targeting AhR, constitutive active AhR expression vector, and AhR agonist and antagonist were used to moderate its expression and signaling. The induction of AhR signaling attenuated invasiveness of prostate cancer cells without affecting the cellular growth rate. These results suggest that AhR signaling in prostate cancer cells facilitates invasion of these cells, and modulation with this signaling can be a potential therapeutic target of invasive tumors.