Synthesis and anti-cancer potential of potent peripheral MAOA inhibitors designed to limit blood:brain penetration.

Monoamine oxidases (MAOA/MAOB) are enzymes known for their role in neurotransmitter regulation in the central nervous system (CNS). Irreversible and non-selective MAO inhibitors (MAOi's) were the first class of antidepressants, thus subsequent work on drugs such as the selective MAOA inhibitor clorgyline has focussed on selectivity and increased CNS penetration. MAOA is highly expressed in high grade and metastatic prostate cancer with a proposed effect on prostate cancer growth, recurrence, and drug resistance. A Phase II Clinical Trial has demonstrated the therapeutic effects of the irreversible nonselective MAOi phenelzine for prostate cancer. However, neurologic adverse effects led to early withdrawal in 25% of the enrolled patient-population. In this work, we revised the clorgyline scaffold with the goal of decreasing CNS penetration to minimize CNS-related side effects while retaining or enhancing MAOA inhibition potency and selectivity. Using the known co-crystal structure of clorgyline bound with FAD co-factor in the hMAOA active site as a reference, we designed and synthesized a series of compounds predicted to have lower CNS penetration (logBB). All synthesized derivatives displayed favorable drug-like characteristics such as predicted Caco-2 permeability and human oral absorption, and exhibited highly selective hMAOA binding interactions. Introduction of an HBD group (NH2 or OH) at position 5 of the phenyl ring clorgyline resulted in 3x more potent hMAOA inhibition with equivalent or better hMAOB selectivity, and similar prostate cancer cell cytotoxicity. In contrast, introduction of larger substituents at this position or at the terminal amine significantly reduced the hMAOA inhibition potency, attributed in part to a steric clash within the binding pocket of the MAOA active site. Replacement of the N-methyl group by a more polar, but larger 2-hydroxyethyl group did not enhance potency. However, introduction of a polar 2-hydroxy in the propyl chain retained the highly selective MAOA inhibition and cancer cell cytotoxicity of clorgyline while reducing its CNS score from 2 to 0. We believe that these results identify a new class of peripherally directed MAOIs that may allow safer therapeutic targeting of MAOA for a variety of anti-cancer and anti-inflammatory indications.

Immune biomarkers associated with COVID-19 disease severity in an urban, hospitalized population.

Objectives: We sought to identify immune biomarkers associated with severe Coronavirus disease 2019 (COVID-19) in patients admitted to a large urban hospital during the early phase of the SARS-CoV-2 pandemic. Design: The study population consisted of SARS-CoV-2 positive subjects admitted for COVID-19 (n = 58) or controls (n = 14) at the Los Angeles County University of Southern California Medical Center between April 2020 through December 2020. Immunologic markers including chemokine/cytokines (IL-6, IL-8, IL-10, IP-10, MCP-1, TNF-α) and serologic markers against SARS-CoV-2 antigens (including spike subunits S1 and S2, receptor binding domain, and nucleocapsid) were assessed in serum collected on the day of admission using bead-based multiplex immunoassay panels. Results: We observed that body mass index (BMI) and SARS-CoV-2 antibodies were significantly elevated in patients with the highest COVID-19 disease severity. IP-10 was significantly elevated in COVID-19 patients and was associated with increased SARS-CoV-2 antibodies. Interactions among all available variables on COVID-19 disease severity were explored using a linear support vector machine model which supported the importance of BMI and SARS-CoV-2 antibodies. Conclusions: Our results confirm the known adverse association of BMI on COVID-19 severity and suggest that IP-10 and SARS-CoV-2 antibodies could be useful to identify patients most likely to experience the most severe forms of the disease.

Cell State and Cell Type: Deconvoluting Circulating Tumor Cell Populations in Liquid Biopsies by Multi-Omics.

Bi-directional crosstalk between the tumor and the tumor microenvironment (TME) has been shown to increase the rate of tumor evolution and to play a key role in neoplastic progression, therapeutic resistance, and a patient's overall survival. Here, we set out to use a comprehensive liquid-biopsy analysis to study cancer and specific TME cells in circulation and their association with disease status. Cytokeratin+, CD45- circulating rare cells (CRCs) from nine breast and four prostate cancer patients were characterized through morphometrics, single-cell copy number analysis, and targeted multiplexed proteomics to delineate cancer cell lineage from other rare cells originating in the TME. We show that we can detect epithelial circulating tumor cells (EPI.CTC), CTCs undergoing epithelial-to-mesenchymal transition (EMT.CTC) and circulating endothelial cells (CECs) using a universal rare event detection platform (HDSCA). Longitudinal analysis of an index patient finds that CTCs are present at the time of disease progression, while CECs are predominately present at the time of stable disease. In a small cohort of prostate and breast cancer patients, we find high inter-patient and temporal intra-patient variability in the expression of tissue specific markers such as ER, HER2, AR, PSA and PSMA and EpCAM. Our study stresses the importance of the multi-omic characterization of circulating rare cells in patients with breast and prostate carcinomas, specifically highlighting overlapping and cell type defining proteo-genomic characteristics of CTCs and CECs.

Effect of Periodized Resistance Training on Skeletal Muscle During Androgen Deprivation Therapy for Prostate Cancer: A Pilot Randomized Trial.

PURPOSE: Prostate cancer survivors (PCS) receive androgen deprivation therapy (ADT) as treatment for recurrent cancer, yet ADT is associated with loss of skeletal muscle and physical function. Resistance training can counter both muscle and physical function loss; however, an understanding of the molecular responses of skeletal muscle to resistance training during ADT is still undefined. This sub-analysis of the original randomized, controlled pilot trial investigated effects of 12 weeks of periodized resistance training on mRNA expression of the anabolic genes IGF-1, myogenin, PGC-1α4 and the catabolic genes myostatin and MuRF-1 in skeletal muscle of PCS on ADT. Secondary aims investigated if changes in lean mass and physical function correlated with changes in mRNA expression. METHODS: PCS on ADT (n = 17) were randomized to 12 weeks of supervised resistance training (EXE, n = 9) or home-based stretching (STRETCH, n = 8) 3 days per week. Outcomes were assessed at baseline and post-intervention. Muscle biopsies were analyzed by RT-PCR for mRNA expression. Body composition was assessed through dual-energy X-ray absorptiometry, and physical function through muscular strength, timed up and go, stair climb, and 400 m walk. RESULTS: MuRF-1 mRNA expression was significantly greater in EXE compared to STRETCH post-intervention (P = .005). Change in MuRF-1 mRNA expression significantly correlated with improvements in strength and physical function (P < .05), while change in IGF-1 expression correlated with change in lean mass (P = .015). CONCLUSION: Twelve weeks of resistance training increased mRNA expression of MuRF-1 in skeletal muscle of PCS on ADT. Elevations in resting mRNA expression of IGF-1, myogenin and PGC-1α4, and reduction in mRNA expression of myostatin that are typically expected following resistance training were not observed.

Phase 2 trial of monoamine oxidase inhibitor phenelzine in biochemical recurrent prostate cancer.

PURPOSE: Monoamine oxidase A (MAOA) influences prostate cancer growth and metastasis in pre-clinical models. We examined effects of phenelzine (a monoamine oxidase inhibitor) in patients with biochemical recurrent castrate-sensitive prostate cancer. MATERIALS AND METHODS: An open-label single arm clinical trial enrolled subjects with biochemical recurrent prostate cancer defined by PSA ≥ 0.4 ng/ml (post prostatectomy) or PSA ≥ 2 ng/ml above nadir (post-radiation therapy); no evidence of metastasis on imaging; and normal androgen levels. Subjects received phenelzine 30 mg orally twice daily. Mood symptoms were assessed with the hospital anxiety depression score (HADS) questionnaire. The primary endpoint was the proportion of patients who achieved a PSA decline of ≥50% from baseline. RESULTS: Characteristics of the 20 eligible patients enrolled included: mean ± SD age 66.9 ± 4.8 years and PSA 4.7 ± 5.8 ng/dl. Maximal PSA declines ≥30% and ≥50% were observed in 25% (n = 5/20) and 10% (n = 2/20) of subjects, respectively. At 12 weeks, 17 subjects remained on treatment with PSA declines ≥30% and ≥50% of 24% (n = 4/17) and 6% (n = 1/17), respectively. Common toxicities observed included dizziness (grade 1 = 45%, grade 2 = 35%), hypertension (grade ≥ 2 = 30%), and edema (grade 1 = 25%, grade 2 = 10%). There was one episode of grade 4 hypertension (cycle 4) and two episodes of grade 3 syncope (cycle 12 and cycle 14) requiring treatment discontinuation. HADS questionnaires demonstrated a significant decrease in anxiety with no change in depressive symptoms on treatment. CONCLUSIONS: Phenelzine demonstrated efficacy in patients with biochemical recurrent castrate-sensitive prostate cancer. Most treatment-related toxicities were mild, but rare significant and reversible cardiovascular toxicities were observed. Therapies directed at MAOA may represent a new avenue for treatment in patients with recurrent prostate cancer.

CCR2-targeted micelles for anti-cancer peptide delivery and immune stimulation.

Signaling between the CC chemokine receptor 2 (CCR2) with its ligand, monocyte chemoattractant protein-1 (MCP-1) promotes cancer progression by directly stimulating tumor cell proliferation and downregulating the expression of apoptotic proteins. Additionally, the MCP-1/CCR2 signaling axis drives the migration of circulating monocytes into the tumor microenvironment, where they mature into tumor-associated macrophages (TAMs) that promote disease progression through induction of angiogenesis, tissue remodeling, and suppression of the cytotoxic T lymphocyte (CTL) response. In order to simultaneously disrupt MCP-1/CCR2 signaling and target CCR2-expressing cancer cells for drug delivery, KLAK-MCP-1 micelles consisting of a CCR2-targeting peptide sequence (MCP-1 peptide) and the apoptotic KLAKLAK peptide were synthesized. In vitro, KLAK-MCP-1 micelles were observed to bind and induce cytotoxicity to cancer cells through interaction with CCR2. In vivo, KLAK-MCP-1 micelles inhibited tumor growth (34 ± 11%) in a subcutaneous B16F10 murine melanoma model despite minimal tumor accumulation upon intravenous injection. Tumors treated with KLAK-MCP1 demonstrated reduced intratumor CCR2 expression and altered infiltration of TAMs and CTLs as evidenced by immunohistochemical and flow cytometric analysis. These studies highlight the potential application of CCR2-targeted nanotherapeutic micelles in cancer treatment.

Randomized Phase II Trial of Abiraterone Alone or With Dasatinib in Men With Metastatic Castration-resistant Prostate Cancer (mCRPC).

BACKGROUND: Signaling via the Src pathway is thought to be a mediator of resistance to androgen targeted therapy in prostate cancer. We studied whether adding the Src inhibitor dasatinib to abiraterone would delay progression. PATIENTS AND METHODS: Eligible patients had metastatic castration-resistant prostate cancer (mCRPC), without prior chemotherapy. Abiraterone was prescribed at 1000 mg daily with prednisone 5 mg twice daily in both arms, and dasatinib 100 mg daily was added for Arm B. The primary endpoint was progression-free survival (PFS). The interim analysis was planned after 48 subjects, but the study was terminated early. PFS was evaluated using a 1-sided log rank test. The Fisher exact test was used for other categorical data analyses. Circulating tumor cells (CTCs) were identified with the Epic platform. RESULTS: With 26 men randomized and a median follow up of 41.8 months, the median PFS was 15.7 months (95% confidence interval, 8.2-49.0+ months) for Arm B and 9.0 months (95% confidence interval, 4.4-30.7 months) for Arm A (P = .15). Response Evaluation Criteria in Solid Tumors responses were seen in 5 (36%) of 14 patients, including 2 complete responses (CRs) on Arm B, and 2 (17%) of 12 responses without CR on Arm A (P = .39). Grade ≥ 3 toxicities more common in Arm B included hypertension, pleural effusion/dyspnea, and gastrointestinal effects. CTCs were detected at baseline in 10 of 19 evaluable patients (median, 2.7/mL blood [range 0.41-59.7]). At week 4, CTCs increased in 1 (10%) of 10 patients on Arm A and 4 (44%) of 9 patients on Arm B. CONCLUSION: Dasatinib did not significantly prolong PFS in combination with abiraterone, although power was limited owing to the incomplete study cohort. Treatment with the combination was associated with robust objective responses, including Response Evaluation Criteria in Solid Tumors CRs.

Monitoring dynamic cytotoxic chemotherapy response in castration-resistant prostate cancer using plasma cell-free DNA (cfDNA).

OBJECTIVE: Cell-free DNA (cfDNA) is an attractive cancer biomarker, as it is thought to reflect a component of the underlying genetic makeup of the tumor and is readily accessible in serial fashion. Because chemotherapy regimens are expected to act rapidly on cancer and cfDNA is cleared from the blood within minutes, we hypothesized that cfDNA would reflect immediate effects of treatment. Here, we developed a method for monitoring long cfDNA fragments, and report dynamic changes in response to cytotoxic chemotherapy. RESULTS: Peripheral blood was obtained from 15 patients with metastatic castration-resistant prostate cancer (CRPC) immediately before and after cytotoxic chemotherapy infusion. cfDNA was extracted and quantified for long interspersed nuclear elements (LINE1; 297 bp) using qPCR. Targeted deep sequencing was performed to quantify the frequency of mutations in exon 8 of the androgen receptor (AR), a mutational hotspot region in CRPC. Single nucleotide mutations in AR exon 8 were found in 6 subjects (6/15 = 40%). Analytical variability was minimized by pooling independent PCR reactions for each library. In 5 patients, tumor-derived long cfDNA levels were found to change immediately after infusion. Detailed analysis of one subject suggests that cytotoxic chemotherapy can produce rapidly observable effects on cfDNA.

Effect of Monoamine oxidase A (MAOA) inhibitors on androgen-sensitive and castration-resistant prostate cancer cells.

BACKGROUND: Monoamine oxidase A (MAOA) is best known for its role in neuro-transmitter regulation. Monoamine oxidase inhibitors are used to treat atypical depression. MAOA is highly expressed in high grade prostate cancer and modulates tumorigenesis and progression in prostate cancer. Here, we investigated the potential role of MAOA inhibitors (MAOAIs) in relation to the androgen receptor (AR) pathway and resistance to antiandrogen treatment in prostate cancer. METHODS: We examined MAOA expression and the effect of MAOI treatment in relation to AR-targeted treatments using the LNCaP, C4-2B, and 22Rv1 human prostate cancer cell lines. MAOA, AR-full length (AR-FL), AR splice variant 7 (AR-V7), and PSA expression was evaluated in the presence of MAOAIs (clorgyline, phenelzine), androgenic ligand (R1881), and antiandrogen (enzalutamide) treatments. An enzalutamide resistance cell line was generated to test the effect of MAOAI treatment in this model. RESULTS: We observed that MAOAIs, particularly clorgyline and phenelzine, were effective at decreasing MAOA activity in human prostate cancer cells. MAOAIs significantly decreased growth of LNCaP, C4-2B, and 22Rv1 cells and produced additive growth inhibitory effects when combined with enzalutamide. Clorgyline decreased expression of AR-FL and AR-V7 in 22Rv1 cells and was effective at decreasing growth of an enzalutamide-resistant C4-2B cell line with increased AR-V7 expression. CONCLUSIONS: MAOAIs decrease growth and proliferation of androgen-sensitive and castration-resistant prostate cancer cells. Clorgyline, in particular, decreases expression of AR-FL and AR-V7 expression and decreases growth of an enzalutamide-resistant cell line. These findings provide preclinical validation of MAOA inhibitors either alone or in combination with antiandrogens for therapeutic intent in patients with advanced forms of prostate cancer.

Loss of ER retention motif of AGR2 can impact mTORC signaling and promote cancer metastasis.

Anterior gradient 2 (AGR2) is a member of the protein disulfide isomerase (PDI) family, which plays a role in the regulation of protein homeostasis and the unfolded protein response pathway (UPR). AGR2 has also been characterized as a proto-oncogene and a potential cancer biomarker. Cellular localization of AGR2 is emerging as a key component for understanding the role of AGR2 as a proto-oncogene. Here, we provide evidence that extracellular AGR2 (eAGR2) promotes tumor metastasis in various in vivo models. To further characterize the role of the intracellular-resident versus extracellular protein, we performed a comprehensive protein-protein interaction screen. Based on these results, we identify AGR2 as an interacting partner of the mTORC2 pathway. Importantly, our data indicates that eAGR2 promotes increased phosphorylation of RICTOR (T1135), while intracellular AGR2 (iAGR2) antagonizes its levels and phosphorylation. Localization of AGR2 also has opposing effects on the Hippo pathway, spheroid formation, and response to chemotherapy in vitro. Collectively, our results identify disparate phenotypes predicated on AGR2 localization. Our findings also provide credence for screening of eAGR2 to guide therapeutic decisions.

Identification, characterization and application of a new peptide against anterior gradient homolog 2 (AGR2).

The cancer-associated protein Anterior Gradient 2 (AGR2) has been described, predominantly in adenocarcinomas. Increased levels of extracellular AGR2 (eAGR2) have been correlated with poor prognosis in cancer patients, making it a potential biomarker. Additionally, neutralizing AGR2 antibodies showed preclinical effectiveness in murine cancer models suggesting eAGR2 may be a therapeutic target. We set out to identify a peptide by mRNA display that would serve as a theranostic tool targeting AGR2. This method enables the selection of peptides from a complex (>1011) library and incorporates a protease incubation step that filters the selection for serum stable peptides. We performed six successive rounds of enrichment using a 10-amino acid mRNA display library and identified several AGR2 binding peptides. One of these peptides (H10), demonstrated high affinity binding to AGR2 with a binding constant (KD) of 6.4 nM. We developed an AGR2 ELISA with the H10 peptide as the capture reagent. Our H10-based ELISA detected eAGR2 from cancer cell spent media with a detection limit of (20-50 ng/ml). Furthermore, we investigated the therapeutic utility of H10 and discovered that it inhibited cell viability at IC50 (9-12 µmoles/L) in cancer cell lines. We also determined that 10 µg/ml of H10 was sufficient to inhibit cancer cell migration in breast and prostate cancer cell lines. A control peptide did not show any appreciable activity in these cells. The H10 peptide showed promise as both a novel diagnostic and a potential therapeutic peptide.

Loss of MAOA in epithelia inhibits adenocarcinoma development, cell proliferation and cancer stem cells in prostate.

Monoamine oxidase A (MAOA) is a mitochondrial enzyme, which degrades monoamine neurotransmitters and dietary amines and produces H2O2. Recent studies have shown increased MAOA expression in prostate cancer (PCa), glioma, and classical Hodgkin lymphoma. However, the biological function of MAOA in cancer development remains unknown. In this study, we investigated the role of MAOA in the development of prostate adenocarcinoma by creating a prostate-specific Pten/MAOA knockout (KO) mouse model, in which MAOA-floxP mouse was crossed with the conditional Pten KO PCa mouse that develops invasive PCa. In contrast to Pten KO mice, age-matched Pten/MAOA KO mice exhibited a significant decrease in both prostate size and the incidence of invasive cancer. We observed a significant decline in AKT phosphorylation and Ki67 expression in Pten/MAOA KO mice, which reduced epithelial cell growth and proliferation. As cancer stem cells (CSCs) are required for tumor initiation and growth, we investigated expression of OCT4 and NANOG in the setting of decreased MAOA expression. We found that both OCT4 and NANOG were significantly attenuated in the prostate epithelia of Pten/MAOA KO mice compared to Pten KO mice, which was confirmed with targeted knockdown of MAOA with a short-hairpin(sh) vector targeting MAOA compared to cells transfected with a control vector. Expression of other markers associated with the a stem cell phenotype, including CD44, α2ß1, and CD133 as well as HIF-1α+CD44+ stem cells were all decreased in shMAOA PCa cells compared with empty vector-transfected control cells. We also found spheroid formation ability in PCa cells was decreased when endogenous MAOA was suppressed by siRNA or MAOA inhibitor clorgyline in a colony formation assay. Using the TCGA database, elevated MAOA expression was associated with reduced Pten levels in high Gleason grade in patient samples. Further, we found that Pten-positive PCa cells were more resistant to clorgyline treatments than Pten-null cells in tumorigenicity and stemness. Taken together, these studies suggest that MAOA expression promotes PCa development by increasing cell proliferation and CSCs and highlights the potential use of MAOA inhibitors for the treatment of PCa.

Impact of resistance training on body composition and metabolic syndrome variables during androgen deprivation therapy for prostate cancer: a pilot randomized controlled trial.

BACKGROUND: Prostate cancer patients on androgen deprivation therapy (ADT) experience adverse effects such as lean mass loss, known as sarcopenia, fat gain, and changes in cardiometabolic factors that increase risk of metabolic syndrome (MetS). Resistance training can increase lean mass, reduce body fat, and improve physical function and quality of life, but no exercise interventions in prostate cancer patients on ADT have concomitantly improved body composition and MetS. This pilot trial investigated 12 weeks of resistance training on body composition and MetS changes in prostate cancer patients on ADT. An exploratory aim examined if a combined approach of training and protein supplementation would elicit greater changes in body composition. METHODS: Prostate cancer patients on ADT were randomized to resistance training and protein supplementation (TRAINPRO), resistance training (TRAIN), protein supplementation (PRO), or control stretching (STRETCH). Exercise groups (EXE = TRAINPRO, TRAIN) performed supervised exercise 3 days per week for 12 weeks, while non-exercise groups (NoEXE = PRO, STRETCH) performed a home-based stretching program. TRAINPRO and PRO received 50 g⋅day- 1 of whey protein. The primary outcome was change in lean mass assessed through dual energy x-ray absorptiometry. Secondary outcomes examined changes in sarcopenia, assessed through appendicular skeletal mass (ASM) index (kg/m2), body fat %, strength, physical function, quality of life, MetS score and the MetS components of waist circumference, blood pressure, glucose, high-density lipoprotein-cholesterol, and triglyceride levels. RESULTS: A total of 37 participants were randomized; 32 participated in the intervention (EXE n = 13; NoEXE n = 19). At baseline, 43.8% of participants were sarcopenic and 40.6% met the criteria for MetS. Post-intervention, EXE significantly improved lean mass (d = 0.9), sarcopenia prevalence (d = 0.8), body fat % (d = 1.1), strength (d = 0.8-3.0), and prostate cancer-specific quality of life (d = 0.9) compared to NoEXE (p < 0.05). No significant differences were observed between groups for physical function or MetS-related variables except waist circumference (d = 0.8). CONCLUSIONS: A 12-week resistance training intervention effectively improved sarcopenia, body fat %, strength and quality of life in hypogonadal prostate cancer patients, but did not change MetS or physical function. PRO did not offer additional benefit in improving body composition. TRIAL REGISTRATION: ClinicalTrials.gov: NCT01909440 . Registered 24 July 2013.

Safety and Efficacy of Docetaxel, Bevacizumab, and Everolimus for Castration-resistant Prostate Cancer (CRPC).

BACKGROUND: Previous data suggests that co-targeting mammalian target of rapamycin and angiogenic pathways may potentiate effects of cytotoxic chemotherapy. We studied combining mammalian target of rapamycin and vascular endothelial growth factor inhibition with docetaxel in castrate-resistant prostate cancer (CRPC). METHODS: Eligible patients had progressive, metastatic, chemotherapy-naive CRPC. Docetaxel and bevacizumab were given intravenously day 1 with everolimus orally daily on a 21-day cycle across 3 dose levels (75:15:2.5, 75:15:5, and 65:15:5; docetaxel mg/m2, mg/kg bevacizumab, and mg everolimus, respectively). Maintenance therapy with bevacizumab/everolimus without docetaxel was allowed after ≥ 6 cycles. RESULTS: Forty-three subjects were treated across all dose levels. Maximal tolerated doses for the combined therapies observed in the phase 1B portion of the trial were: docetaxel 75 mg/m2, bevacizumab 15 mg/kg, and everolimus 2.5 mg. Maximal prostate-specific antigen decline ≥ 30% and ≥ 50% was achieved in 33 (79%) and 31 (74%) of patients, respectively. Best response by modified Response Evaluation Criteria In Solid Tumors criteria in 25 subjects with measurable disease at baseline included complete or partial response in 20 (80%) patients. The median progression-free and overall survival were 8.9 months (95% confidence interval, 7.4-10.6 months) and 21.9 months (95% confidence interval, 18.4-30.3 months), respectively. Hematologic toxicities were the most common treatment-related grade ≥ 3 adverse events including: febrile neutropenia (12; 28%), lymphopenia (12; 28%), leukocytes (10; 23%), neutrophils (9; 21%), and hemoglobin (2; 5%). Nonhematologic grade ≥ 3 adverse events included: hypertension (8; 19%), fatigue (3; 7%), pneumonia (3; 7%), and mucositis (4; 5%). There was 1 treatment-related death owing to neutropenic fever and pneumonia in a patient treated at dose level 3 despite dose modifications and prophylactic growth factor support. CONCLUSIONS: Docetaxel, bevacizumab, and everolimus can be safely administered in CRPC and demonstrate a significant level of anticancer activity, meeting the predetermined response criteria. However, any potential benefit of combined therapy must be balanced against increased risk for toxicities. Our results do not support the hypothesis that this combination of agents improves upon the results obtained with docetaxel monotherapy in an unselected population of chemotherapy-naive patients with CRPC.

A pilot randomised controlled trial of a periodised resistance training and protein supplementation intervention in prostate cancer survivors on androgen deprivation therapy.

INTRODUCTION: Prostate cancer survivors (PCS) receiving androgen deprivation therapy (ADT) experience deleterious side effects such as unfavourable changes in cardiometabolic factors that lead to sarcopenic obesity and metabolic syndrome (MetS). While loss of lean body mass (LBM) compromises muscular strength and quality of life, MetS increases the risk of cardiovascular disease and may influence cancer recurrence. Exercise can improve LBM and strength, and may serve as an alternative to the pharmacological management of MetS in PCS on ADT. Prior exercise interventions in PCS on ADT have been effective at enhancing strength, but only marginally effective at enhancing body composition and ameliorating cardiometabolic risk factors. This pilot trial aims to improve on existing interventions by employing periodised resistance training (RT) to counter sarcopenic obesity in PCS on ADT. Secondary aims compare intervention effects on cardiometabolic, physical function, quality of life and molecular skeletal muscle changes. An exploratory aim examines if protein supplementation (PS) in combination with RT elicits greater changes in these outcomes. METHODS AND ANALYSIS: A 2×2 experimental design is used in 32 PCS on ADT across a 12-week intervention period. Participants are randomised to resistance training and protein supplementation (RTPS), RT, PS or control. RT and RTPS groups perform supervised RT three times per week for 12 weeks, while PS and RTPS groups receive 50 g whey protein per day. This pilot intervention applies a multilayered approach to ameliorate detrimental cardiometabolic effects of ADT while investigating molecular mechanisms underlying skeletal muscle changes in PCS. ETHICS AND DISSEMINATION: This trial was approved by the University of Southern California Institutional Review Board (HS-13-00315). Results from this trial will be communicated in peer-reviewed publications and scientific presentations. TRIAL REGISTRATION NUMBER: NCT01909440; Pre-results.

Automated analysis of co-localized protein expression in histologic sections of prostate cancer.

An automated approach based on routinely-processed, whole-slide immunohistochemistry (IHC) was implemented to study co-localized protein expression in tissue samples. Expression of two markers was chosen to represent stromal (CD31) and epithelial (Ki-67) compartments in prostate cancer. IHC was performed on whole-slide sections representing low-, intermediate-, and high-grade disease from 15 patients. The automated workflow was developed using a training set of regions-of-interest in sequential tissue sections. Protein expression was studied on digital representations of IHC images across entire slides representing formalin-fixed paraffin embedded blocks. Using the training-set, the known association between Ki-67 and Gleason grade was confirmed. CD31 expression was more heterogeneous across samples and remained invariant with grade in this cohort. Interestingly, the Ki-67/CD31 ratio was significantly increased in high (Gleason ≥ 8) versus low/intermediate (Gleason ≤7) samples when assessed in the training-set and the whole-tissue block images. Further, the feasibility of the automated approach to process Tissue Microarray (TMA) samples in high throughput was evaluated. This work establishes an initial framework for automated analysis of co-localized protein expression and distribution in high-resolution digital microscopy images based on standard IHC techniques. Applied to a larger sample population, the approach may help to elucidate the biologic basis for the Gleason grade, which is the strongest, single factor distinguishing clinically aggressive from indolent prostate cancer.

Androgen receptor in cancer-associated fibroblasts influences stemness in cancer cells.

Androgen receptor (AR) regulation pathways are essential for supporting the growth and survival of prostate cancer cells. Recently, sub-populations of prostate cancer cells have been identified with stem cell features and are associated with the emergence of treatment-resistant prostate cancer. Here, we explored the function of AR in prostate cancer-associated fibroblasts (CAFs) relative to growth and stem cell-associated characteristics. CAFs were isolated from the murine cPten-/-L prostate cancer model and cultured with human prostate cancer epithelial (hPCa) cells. A murine-specific AR antisense oligonucleotide (ASO) was used to suppress the expression of AR in the CAF cells. CAFs express low, but significant levels of AR relative to fibroblasts derived from non-malignant tissue. CAFs promoted growth and colony formation of hPCa cells, which was attenuated by the suppression of AR expression. Surprisingly, AR-depleted CAFs promoted increased stem cell marker expression in hPCa cells. Interferon gamma (IFN-γ) and macrophage colony-stimulating factor (M-CSF) were increased in AR-depleted CAF cells and exhibited similar effects on stem cell marker expression as seen in the CAF co-culture systems. Clinically, elevated IFN-γ expression was found to correlate with histologic grade in primary prostate cancer samples. In summary, AR and androgen-dependent signaling are active in CAFs and exert significant effects on prostate cancer cells. IFN-γ and M-CSF are AR-regulated factors secreted by CAF cells, which promote the expression of stem cell markers in prostate cancer epithelial cells. Understanding how CAFs and other constituents of stromal tissue react to anti-cancer therapies may provide insight into the development and progression of prostate cancer.

Protein Mimetic and Anticancer Properties of Monocyte-Targeting Peptide Amphiphile Micelles.

Monocyte chemoattractant protein-1 (MCP-1) stimulates the migration of monocytes to inflammatory sites, leading to the progression of many diseases. Recently, we described a monocyte-targeting peptide amphiphile micelle (MCP-1 PAM) incorporated with the chemokine receptor CCR2 binding motif of MCP-1, which has a high affinity for monocytes in atherosclerotic plaques. We further report here the biomimetic components of MCP-1 PAMs and the influence of the nanoparticle upon binding to monocytes. We report that MCP-1 PAMs have enhanced secondary structure compared to the MCP-1 peptide. As a result, MCP-1 PAMs displayed improved binding and chemoattractant properties to monocytes, which upregulated the inflammatory signaling pathways responsible for monocyte migration. Interestingly, when MCP-1 PAMs were incubated in the presence of prostate cancer cells in vitro, the particle displayed anticancer efficacy by reducing CCR2 expression. Given that monocytes play an important role in tumor cell migration and invasion, our results demonstrate that PAMs can improve the native biofunctional properties of the peptide and may be used as an effective inhibitor to prevent chemokine-receptor interactions that promote disease progression.

Blood-based gene expression profiling in castrate-resistant prostate cancer.

Castrate-resistant prostate cancer (CRPC), the most life-threatening form of prostate cancer, has recently been the focus of many successful new treatments. Contemporary trials highlight the heterogeneous prognosis of CRPC as overall survival times vary greatly across different patient sub-groups. As presented in BMC Medicine, Wang et al. identify a blood-based prognostic signature in CRPC. Their approach is notable for discovery and validation of a four-gene model based on a whole-blood expression signature sampled from three distinct clinical cohorts. Further, the marker selection process incorporates an understanding of biological pathways expressed in myeloid or lymphoid cells which may provide some insight into host-tumor interactions as reflected in the peripheral blood. While the study includes a multivariate analysis accounting for many important clinical variables, larger datasets with more complete clinical information and sufficient follow-up are needed to confirm the independent significance of the four-gene expression model in a way which may better inform the care of CRPC patients.Please see related article: http://www.biomedcentral.com/1741-7015/13/201 .

Salvage ablative therapy in prostate cancer: international multidisciplinary consensus on trial design.

INTRODUCTION: Salvage ablative therapy (SAT) has been developed as a form of localized treatment for localized recurrence of prostate cancers following radiation therapy. To better address the utility of SAT, prospective clinical trials must address the aspects of accepted standards in the initial evaluation, treatment, follow-up, and outcomes in the oncology community. We undertook this study to achieve consensus on uniform standardized trial design for SAT trials. METHODS: A literature search was performed and an international multidisciplinary group of experts was identified. A questionnaire was constructed and sent out to 71 participants in 3 consecutive rounds according to the Delphi method. The project was concluded with a face-to-face meeting in which the results were reviewed and conclusions were formulated. RESULTS: Patients with recurrent disease after radiation therapy were considered candidates for a SAT trial using any ablation scenario performed with cryotherapy or high-intensity focused ultrasound. It is feasible to compare different sources of energy or to compare with historical data on salvage radical prostatectomy outcomes. The primary objective should be to assess the efficacy of the treatment for negative biopsy rate at 12 months. Secondary objectives should include safety parameters and quality-of-life assessment. Exclusion criteria should include evidence of local or distant metastases. The optimal biopsy strategy is image-guided targeted biopsies. Follow-up includes multiparametric magnetic resonance imaging, prostate-specific antigen level, and quality of life for at least 5 years. CONCLUSIONS: A multidisciplinary board from international experts reached consensus on trial design for SAT in prostate cancer and provides a standard for designing a feasible SAT trial.