Role of proton-coupled folate transporter in pemetrexed resistance of mesothelioma: clinical evidence and new pharmacological tools

E Giovannetti, P A Zucali, Y G Assaraf, N Funel, M Gemelli, M Stark, E Thunnissen, Z Hou, I B Muller, E A Struys, M Perrino, G Jansen, L H Matherly, G J Peters

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Abstract

Background: Thymidylate synthase (TS) has a predictive role in pemetrexed treatment of mesothelioma; however, additional chemoresistance mechanisms are poorly understood. Here, we explored the role of the reduced-folate carrier (RFC/SLC19A1) and proton-coupled folate transporter (PCFT/SLC46A1) in antifolate resistance in mesothelioma.

Patients and methods: PCFT, RFC and TS RNA and PCFT protein levels were determined by quantitative RT-PCR of frozen tissues and immunohistochemistry of tissue-microarrays, respectively, in two cohorts of pemetrexed-treated patients. Data were analyzed by t-test, Fisher's/log-rank test and Cox proportional models. The contribution of PCFT expression and PCFT-promoter methylation to pemetrexed activity were evaluated in mesothelioma cells and spheroids, through 5-aza-2'-deoxycytidine-mediated demethylation and siRNA-knockdown.

Results: Pemetrexed-treated patients with low PCFT had significantly lower rates of disease control, and shorter overall survival (OS), in both the test (N = 73, 11.3 versus 20.1 months, P = 0.01) and validation (N = 51, 12.6 versus 30.3 months, P = 0.02) cohorts. Multivariate analysis confirmed PCFT-independent prognostic role. Low-PCFT protein levels were also associated with shorter OS. Patients with both low-PCFT and high-TS levels had the worst prognosis (OS, 5.5 months), whereas associations were neither found for RFC nor in pemetrexed-untreated patients. PCFT silencing reduced pemetrexed sensitivity, whereas 5-aza-2'-deoxycytidine overcame resistance.

Conclusions: These findings identify for the first time PCFT as a novel mesothelioma prognostic biomarker, prompting prospective trials for its validation. Moreover, preclinical data suggest that targeting PCFT-promoter methylation might eradicate pemetrexed-resistant cells characterized by low-PCFT expression.

Original languageEnglish
Pages (from-to)2725-2732
Number of pages8
JournalAnnals of oncology : official journal of the European Society for Medical Oncology
Volume28
Issue number11
DOIs
Publication statusPublished - Nov 1 2017

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Proton-Coupled Folate Transporter
Pemetrexed
Mesothelioma
Pharmacology
decitabine
Thymidylate Synthase
Methylation
Survival
Reduced Folate Carrier Protein
Folic Acid Antagonists
Proportional Hazards Models
Small Interfering RNA
Proteins
Multivariate Analysis
Biomarkers
Immunohistochemistry
RNA

Keywords

  • Journal Article

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Role of proton-coupled folate transporter in pemetrexed resistance of mesothelioma : clinical evidence and new pharmacological tools. / Giovannetti, E; Zucali, P A; Assaraf, Y G; Funel, N; Gemelli, M; Stark, M; Thunnissen, E; Hou, Z; Muller, I B; Struys, E A; Perrino, M; Jansen, G; Matherly, L H; Peters, G J.

In: Annals of oncology : official journal of the European Society for Medical Oncology, Vol. 28, No. 11, 01.11.2017, p. 2725-2732.

Research output: Contribution to journalArticle

Giovannetti, E, Zucali, PA, Assaraf, YG, Funel, N, Gemelli, M, Stark, M, Thunnissen, E, Hou, Z, Muller, IB, Struys, EA, Perrino, M, Jansen, G, Matherly, LH & Peters, GJ 2017, 'Role of proton-coupled folate transporter in pemetrexed resistance of mesothelioma: clinical evidence and new pharmacological tools', Annals of oncology : official journal of the European Society for Medical Oncology, vol. 28, no. 11, pp. 2725-2732. https://doi.org/10.1093/annonc/mdx499
Giovannetti, E ; Zucali, P A ; Assaraf, Y G ; Funel, N ; Gemelli, M ; Stark, M ; Thunnissen, E ; Hou, Z ; Muller, I B ; Struys, E A ; Perrino, M ; Jansen, G ; Matherly, L H ; Peters, G J. / Role of proton-coupled folate transporter in pemetrexed resistance of mesothelioma : clinical evidence and new pharmacological tools. In: Annals of oncology : official journal of the European Society for Medical Oncology. 2017 ; Vol. 28, No. 11. pp. 2725-2732.
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T1 - Role of proton-coupled folate transporter in pemetrexed resistance of mesothelioma

T2 - clinical evidence and new pharmacological tools

AU - Giovannetti, E

AU - Zucali, P A

AU - Assaraf, Y G

AU - Funel, N

AU - Gemelli, M

AU - Stark, M

AU - Thunnissen, E

AU - Hou, Z

AU - Muller, I B

AU - Struys, E A

AU - Perrino, M

AU - Jansen, G

AU - Matherly, L H

AU - Peters, G J

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N2 - Background: Thymidylate synthase (TS) has a predictive role in pemetrexed treatment of mesothelioma; however, additional chemoresistance mechanisms are poorly understood. Here, we explored the role of the reduced-folate carrier (RFC/SLC19A1) and proton-coupled folate transporter (PCFT/SLC46A1) in antifolate resistance in mesothelioma.Patients and methods: PCFT, RFC and TS RNA and PCFT protein levels were determined by quantitative RT-PCR of frozen tissues and immunohistochemistry of tissue-microarrays, respectively, in two cohorts of pemetrexed-treated patients. Data were analyzed by t-test, Fisher's/log-rank test and Cox proportional models. The contribution of PCFT expression and PCFT-promoter methylation to pemetrexed activity were evaluated in mesothelioma cells and spheroids, through 5-aza-2'-deoxycytidine-mediated demethylation and siRNA-knockdown.Results: Pemetrexed-treated patients with low PCFT had significantly lower rates of disease control, and shorter overall survival (OS), in both the test (N = 73, 11.3 versus 20.1 months, P = 0.01) and validation (N = 51, 12.6 versus 30.3 months, P = 0.02) cohorts. Multivariate analysis confirmed PCFT-independent prognostic role. Low-PCFT protein levels were also associated with shorter OS. Patients with both low-PCFT and high-TS levels had the worst prognosis (OS, 5.5 months), whereas associations were neither found for RFC nor in pemetrexed-untreated patients. PCFT silencing reduced pemetrexed sensitivity, whereas 5-aza-2'-deoxycytidine overcame resistance.Conclusions: These findings identify for the first time PCFT as a novel mesothelioma prognostic biomarker, prompting prospective trials for its validation. Moreover, preclinical data suggest that targeting PCFT-promoter methylation might eradicate pemetrexed-resistant cells characterized by low-PCFT expression.

AB - Background: Thymidylate synthase (TS) has a predictive role in pemetrexed treatment of mesothelioma; however, additional chemoresistance mechanisms are poorly understood. Here, we explored the role of the reduced-folate carrier (RFC/SLC19A1) and proton-coupled folate transporter (PCFT/SLC46A1) in antifolate resistance in mesothelioma.Patients and methods: PCFT, RFC and TS RNA and PCFT protein levels were determined by quantitative RT-PCR of frozen tissues and immunohistochemistry of tissue-microarrays, respectively, in two cohorts of pemetrexed-treated patients. Data were analyzed by t-test, Fisher's/log-rank test and Cox proportional models. The contribution of PCFT expression and PCFT-promoter methylation to pemetrexed activity were evaluated in mesothelioma cells and spheroids, through 5-aza-2'-deoxycytidine-mediated demethylation and siRNA-knockdown.Results: Pemetrexed-treated patients with low PCFT had significantly lower rates of disease control, and shorter overall survival (OS), in both the test (N = 73, 11.3 versus 20.1 months, P = 0.01) and validation (N = 51, 12.6 versus 30.3 months, P = 0.02) cohorts. Multivariate analysis confirmed PCFT-independent prognostic role. Low-PCFT protein levels were also associated with shorter OS. Patients with both low-PCFT and high-TS levels had the worst prognosis (OS, 5.5 months), whereas associations were neither found for RFC nor in pemetrexed-untreated patients. PCFT silencing reduced pemetrexed sensitivity, whereas 5-aza-2'-deoxycytidine overcame resistance.Conclusions: These findings identify for the first time PCFT as a novel mesothelioma prognostic biomarker, prompting prospective trials for its validation. Moreover, preclinical data suggest that targeting PCFT-promoter methylation might eradicate pemetrexed-resistant cells characterized by low-PCFT expression.

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