Targeted
therapies have yet to have significant impact on the survival of patients with
bladder cancer. In this study, we focused on the
urea cycle
enzyme argininosuccinate synthetase 1 (ASS1) as a therapeutic target in
bladder cancer, based on our discovery of the prognostic and functional import of ASS1 in this setting. ASS1 expression status in
bladder tumors from 183 Caucasian and 295 Asian patients was analyzed, along with its hypothesized prognostic impact and association with clinicopathologic features, including
tumor size and invasion. Furthermore, the genetics, biology, and therapeutic implications of ASS1 loss were investigated in urothelial
cancer cells. We detected ASS1 negativity in 40% of
bladder cancers, in which multivariate analysis indicated worse disease-specific and
metastasis-free survival. ASS1 loss secondary to epigenetic silencing was accompanied by increased
tumor cell proliferation and invasion, consistent with a
tumor-suppressor role for ASS1. In developing a treatment approach, we identified a novel targeted
antimetabolite strategy to exploit
arginine deprivation with
pegylated arginine deiminase (ADI-PEG20) as a therapeutic.
ADI-PEG20 was synthetically lethal in ASS1-methylated bladder cells and its exposure was associated with a marked reduction in intracellular levels of
thymidine, due to suppression of both uptake and de novo synthesis. We found that
thymidine uptake correlated with
thymidine kinase-1
protein levels and that
thymidine levels were imageable with [(18)F]-fluoro-L-
thymidine (FLT)-positron emission tomography (PET). In contrast, inhibition of de novo synthesis was linked to decreased expression of
thymidylate synthase and
dihydrofolate reductase. Notably, inhibition of de novo synthesis was associated with potentiation of
ADI-PEG20 activity by the
antifolate drug pemetrexed. Taken together, our findings argue that
arginine deprivation combined with
antifolates warrants clinical investigation in ASS1-negative urothelial and related
cancers, using FLT-PET as an early
surrogate marker of response.