Determining argininosuccinate synthetase (ASS) expression in patients with melanoma treated with arginine depleting therapy.

10627 Background: In our phase II trial in advanced melanoma (ASCO 2010) with arginine depleting therapy using ADI-PEG20 (Polaris Pharmaceuticals), we found that response to therapy correlated with tumor expression of enzyme, ASS. Normal cells which possess ASS can recycle citrulline to arginine and hence evade cell death, whereas melanoma tumors lacking ASS do not. Thus, 11 of 17 patients (pts) with ASS(-) tumors (by immunohistochemistry or real-time PCR) had evidence of antitumor activity versus 1 of 10 pts with ASS (+) tumors, (P value = 0.01). To study why not all ASS(-) melanoma pts respond to ADI-PEG20, we have developed a sensitive method to detect small amounts of mRNA by PCR. Methods: Total RNA was prepared from primary culture or cell lines and used for cDNA synthesis. Real-time PCR of ASS was performed using specific primers. The reaction was based on SYBR Green and performed in a Bio-Rad iCycler PCR machine equipped with a MyiQ module. ΔΔCt method was used to calculate the relative ASS mRNA le...