PROJECT SUMMARY/ABSTRACT Androgen indifferent prostate cancers account for a large proportion of the disease lethality and have limited therapy options, partly due to the lack of identifying biomarkers. To address the unmet need of developing effective therapies for this subset, we defined the aggressive variant prostate cancers (AVPC) criteria. Through a series of prospective trials and studies in mice, we showed that the AVPC criteria can enrich for prostate cancers that respond poorly to androgen inhibition and benefit from adding carboplatin to cabazitaxel. In recent trials we examined the contribution of PARP inhibitor maintenance, and of anti-PD1 inhibition to the chemotherapy backbone in men with AVPC. Early data show meaningful improvements in outcomes with these additions, but many men with AVPC are still progressing rapidly. Our overall goal is to arrive at rational, biologically-based combination therapies that effectively treat the AVPC. The analysis of samples from our trial participants, and preclinical studies, converge on altered arginine metabolism as a key metabolic pathway in androgen-indifferent prostate cancer biology. We also found evidence of argininosuccinate synthase 1 (ASS1) silencing with platinum chemotherapy in AVPC cell lines and patient samples. ASS1 deficiency renders cells dependent on extracellular arginine, and thus sensitive to arginine depletion with agents such as PEGylated arginine deiminase (ADI-PEG20). ADI-PEG20 depletes serum arginine levels, has activity in several malignancies (alone and in combination with chemotherapy), and has immunomodulatory effects. However, the effects of serum arginine depletion on intratumoral metabolite levels in patients are unknown, and its effects on the human immune tumor microenvironment (TME) remain poorly understood. We hypothesize that the addition of ADI-PEG20 will improve the efficacy of carboplatin+cabazitaxel by modifying intratumoral arginine metabolism and immune profiles in the AVPC TME. In AIM 1, we will conduct a phase I/II dose escalation trial to identify the optimal dose (in terms of safety and efficacy) of ADI-PEG20 to combine with carboplatin+cabazitaxel in men with AVPC. In AIM 2, peripheral blood and metastatic tumor biopsies obtained from trial participants at baseline, after 1 and after 6 cycles of treatment, and at parallel time-points in PDX and syngeneic mouse models, will be used to measure associations between: (a) serum levels of arginine and citrulline, (b) intratumoral levels of arginine, (c) ASS1 expression, and (d) the expression of other arginine metabolism enzymes, and (e) outcomes. In AIM 3 we will examine the effects of treatment on immune profiles and immune cell distribution within the TME. Our studies will provide a comprehensive evaluation of the effects of serum arginine depletion on the immune and non-immune AVPC TME, shed light on the mechanisms of synergy between ADI-PEG20 and cytotoxic chemotherapy, and ultimately, serve to ...