Background: Approximately 40% of malignant pleural mesothelioma (MPM) patients respond to the combination treatment with the antifolate pemetrexed (PMX) and platinum compounds. Several studies highlighted the predictive role of thymidylate synthase (TS), but additional resistance mechanisms are poorly understood. This study aimed to evaluate the role of the reduced folate carrier (RFC/SLC19A1) and proton-coupled folate transporter (PCFT/SLC46A1) as potential biological markers of intrinsic PMX-resistance in MPM.

Materials and methods: PCFT promoter methylation was evaluated by combined bisulfite restriction analysis as well as MethyLight PCR in 14 cell lines and 10 MPM tissues, while PCFT and RFC mRNA levels were studied by Real-Time-PCR in MPM specimens from a test and a validation cohort of 73 and 51 PMX-treated patients, respectively. PCFT protein expression was evaluated by immunohistochemistry in a tissue microarray with representative MPM core biopsies from 35 patients. Data were analyzed by t-test, Fisher/log-rank test and Cox proportional hazards models.

Results: PCFT expression inversely correlated with promoter methylation and treatment with 5-Aza-2′-deoxycytidine (5-Aza-dC) induced its marked restoration in MPM cells. Patients with low PCFT mRNA levels had significantly shorter overall survival (OS), in both the test (median OS, 11.3 vs. 17.6 months, P = 0.01) and in the validation cohort (OS, 12.6 vs. 30.3 months, P = 0.02). Multivariate analysis, including age, sex, EORTC prognostic score and histology, confirmed PCFT prognostic relevance (HR, 2.1 and 2.4 in the test and validation cohorts, respectively). Low PCFT protein expression was also associated with significantly shorter OS (11.8 vs. 30.3 months, P = 0.02).

Remarkably, the subgroup of patients with both low PCFT and high TS mRNA levels (30% of the patients in the test cohort) had the worse prognosis (median OS, 5.5 months, P<0.01), while no correlation was found in a series of 40 MPM patients not PMX-treated. No significant associations were found between RFC and clinical outcome.

Conclusions: These results define PCFT as a novel prognostic factor for MPM patients treated with PMX. Prospective trials for the validation of the prognostic/predictive role of PCFT in MPM patients treated with PMX-based regimens are warranted. Furthermore preclinical data suggest that exposure to 5-Aza-dC might restore PCFT expression and result in efficient eradication of antifolate-resistant cells.

Citation Format: Elisa Giovannetti, Paolo A. Zucali, Yehuda G. Assaraf, Niccola Funel, Maria Gemelli, Michal Stark, Leticia G. Leon, Zhanjun Hou, Matteo Perrino, Larry H. Matherly, Godefridus J. Peters. Role of proton-coupled folate transporter expression in resistance of mesothelioma patients treated with pemetrexed. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 4335. doi:10.1158/1538-7445.AM2015-4335