Impact of hypoxia on chemoresistance of mesothelioma mediated by the proton-coupled folate transporter, and preclinical activity of new anti-LDH-A compounds

Our plain-English summary of Impact of hypoxia on chemoresistance of mesothelioma mediated by the proton-coupled folate transporter, and preclinical activity of new anti-LDH-A compounds, by G. Li Petri, B. El Hassouni, R. Sciarrillo, et al (British Journal of Cancer, 2020).

The study

• Authors: G. Li Petri, B. El Hassouni, R. Sciarrillo, N. Funel, G. Mantini, E.A. Zeeuw van der Laan, S. Cascioferro, A. Avan, P.A. Zucali, N. Zaffaroni, T. Lagerweij, B. Parrino, K. Smid, M. Deraco, C. Granchi, A. Braczko, R.T. Smolenski, L.H. Matherly, G. Jansen, Y.G. Assaraf, P. Diana, J. Cloos, G.J. Peters, F. Minutolo, E. Giovannetti
• Journal: British Journal of Cancer
• Published: 2020;123(4):644-656
• Cited by: 39 articles, as counted by NIH iCite
• Read it: PubMed · Publisher · Free full text

Abstract

Background: Expression of proton-coupled folate transporter (PCFT) is associated with survival of mesothelioma patients treated with pemetrexed, and is reduced by hypoxia, prompting studies to elucidate their correlation.

Methods: Modulation of glycolytic gene expression was evaluated by PCR arrays in tumour cells and primary cultures growing under hypoxia, in spheroids and after PCFT silencing. Inhibitors of lactate dehydrogenase (LDH-A) were tested in vitro and in vivo. LDH-A expression was determined in tissue microarrays of radically resected malignant pleural mesothelioma (MPM, N = 33) and diffuse peritoneal mesothelioma (DMPM, N = 56) patients.

Results: Overexpression of hypoxia marker CAIX was associated with low PCFT expression and decreased MPM cell growth inhibition by pemetrexed. Through integration of PCR arrays in hypoxic cells and spheroids and following PCFT silencing, we identified the upregulation of LDH-A, which correlated with shorter survival of MPM and DMPM patients. Novel LDH-A inhibitors enhanced spheroid disintegration and displayed synergistic effects with pemetrexed in MPM and gemcitabine in DMPM cells. Studies with bioluminescent hypoxic orthotopic and subcutaneous DMPM athymic-mice models revealed the marked antitumour activity of the LDH-A inhibitor NHI-Glc-2, alone or combined with gemcitabine.

Conclusions: This study provides novel insights into hypoxia/PCFT-dependent chemoresistance, unravelling the potential prognostic value of LDH-A, and demonstrating the preclinical activity of LDH-A inhibitors.

In plain English

This laboratory study, with an analysis of stored tumour tissue, examined why mesothelioma cells resist pemetrexed under low oxygen (hypoxia). Asbestos exposure was not studied. Of 33 malignant pleural mesothelioma (MPM) tumours, 28 were positive for the hypoxia marker CAIX; in these, high CAIX went with low PCFT, a pemetrexed transporter (P = 0.016). In three MPM cell lines, low oxygen reduced PCFT and increased pemetrexed resistance. Of 84 hypoxia-related genes tested, only the gene for the enzyme LDH-A rose significantly in all three conditions: 3D cell clumps, low oxygen, and PCFT reduced by gene silencing. High tumour LDH-A went with shorter overall survival: median 10.4 months (95% CI 7.1 to 13.7) versus 36.7 months (27.4 to 46.6) with low LDH-A in MPM, and 12.0 (5.3 to 18.7) versus 35.0 months (30.7 to 39.3) in 56 diffuse peritoneal mesothelioma patients. In mice, an experimental LDH-A blocker, NHI-Glc-2, slowed tumour growth, more so with gemcitabine. The authors concluded that high LDH-A suggests more aggressive mesothelioma that LDH-A inhibitors can target. They noted the patient groups were small and retrospective.

We wrote this plain-English summary from the full article with the help of AI, and a second AI model checked each sentence against the article. It is not the authors' wording; the abstract and the article are their own account.

What the study did and found

• Design: Laboratory study (human mesothelioma cell lines, primary cell cultures, 3D spheroids, and experiments in mice) combined with a retrospective analysis of protein levels in stored patient tumour samples (tissue microarrays) linked to survival
• Who was studied: Tumour samples from 33 malignant pleural mesothelioma (MPM) patients enrolled at Humanitas Cancer Center, Milan, Italy, 2008 to 2013, and from 56 diffuse peritoneal mesothelioma (DMPM) patients enrolled at the National Cancer Institute, Milan, 1995 to 2013. The study also used three human MPM cell lines (H28, H2452, MSTO-211H), two primary DMPM cultures (MesoII, STO), and nu/nu athymic female mice.
• Exposure: Not asbestos. The study examined hypoxia (1% oxygen), silencing of the PCFT transporter, the chemotherapy drugs pemetrexed and gemcitabine, and the experimental LDH-A inhibitors NHI-2 and NHI-Glc-2. In mice, NHI-Glc-2 was given at 100 mg/kg (orthotopic model) or 50 mg/kg (subcutaneous model), and gemcitabine at 100 mg/kg.
• Main results:
  • In the MPM tissue samples, 84% (28 of 33) stained positive for the hypoxia marker CAIX.
  • Among the 28 CAIX-positive MPM samples, high CAIX was associated with low PCFT expression (P = 0.016).
  • In all three MPM cell lines grown under hypoxia, PCFT expression was reduced.
  • In H2452 cells tested with an 84-gene hypoxia PCR array, LDH-A was the only gene raised in all three conditions: spheroids, hypoxia and PCFT silencing.
  • High LDH-A expression was found in 20 of 33 MPM cases (60%) and in 36 of 56 DMPM cases (64%).
  • In MPM patients given pemetrexed-based chemotherapy, disease control was achieved in 92% of those with low LDH-A and in 43% of those with high LDH-A (P = 0.03).
  • In MPM patients, those with high LDH-A had a median overall survival of 10.4 months (95% CI 7.1 to 13.7) and a median progression-free survival of 6.9 months (95% CI 2.7 to 11.2). Those with low LDH-A had a median overall survival of 36.7 months (95% CI 27.4 to 46.6).
  • In DMPM patients, overall survival was 35.0 months (95% CI 30.7 to 39.3) for those with low LDH-A and 12.0 months (95% CI 5.3 to 18.7) for the remaining patients (P = 0.01).
  • In the multivariate analysis of DMPM patients, high LDH-A was associated with an increased risk of relapse (HR = 2.4, 95% CI 1.2 to 2.6, P = 0.01) and of death (HR = 2.7, 95% CI 1.7 to 3.6, P = 0.02).
  • In mice with subcutaneous DMPM tumours, NHI-Glc-2 at 50 mg/kg reduced tumour mass by 49% compared with controls at the end of week 7 (314 vs. 620 cubic millimetres, P < 0.001).
  • In the same mice, gemcitabine alone reduced tumour volume by 47% compared with controls (331 vs. 620 cubic millimetres, P < 0.001).
  • In the same mice, the combination of NHI-Glc-2 and gemcitabine gave smaller tumours than in controls (123 vs. 620 cubic millimetres, P < 0.001) and was also significantly better than either drug alone.
  • In both primary DMPM cell cultures, NHI-Glc-2 combined with gemcitabine showed synergistic or strongly synergistic effects in the laboratory.
• The authors' conclusion: The authors concluded that their clinical, laboratory and animal findings strongly suggest that mesothelioma is more aggressive when it has high LDH-A expression. They added that LDH-A can be targeted by specific LDH-A inhibitors, which they described as a promising avenue for prognosis and treatment.
• Limitations the authors state:
  • LDH-A was tested retrospectively in relatively small patient groups treated with different therapies, with and without pemetrexed, and in a small number of cell lines and primary cells.
  • The study did not follow the effects of treatment over the long term.
  • The imaging analysis in the orthotopic mouse model was done in only three animals, and the authors said its results should be treated with caution.
  • Bioluminescence imaging tends to underestimate hypoxic tumours.
  • No multivariate survival analysis was done for MPM patients because the sample was very small.

Topics

The National Library of Medicine indexes this study under subjects our site covers: Mesothelioma, Peritoneal mesothelioma.

Medical terms in this study, explained in our Lexicon: Hypoxia, Mesothelioma, Pemetrexed (Alimta), Lactate Dehydrogenase (LDH), Pleural Mesothelioma, Peritoneal Mesothelioma, Synergistic Effect, Gemcitabine.

Related studies

• Therapeutic targeting malignant mesothelioma with a novel 6-substituted pyrrolo[2,3-d]pyrimidine thienoyl antifolate via its selective uptake by the proton-coupled folate transporter — C. Cherian, S. Kugel Desmoulin, et al. (2013)
• The promise and challenges of exploiting the proton-coupled folate transporter for selective therapeutic targeting of cancer — L.H. Matherly, Z. Hou, et al. (2018)
• The human proton-coupled folate transporter: Biology and therapeutic applications to cancer — S.K. Desmoulin, Z. Hou, et al. (2012)
• A specific inhibitor of lactate dehydrogenase overcame the resistance toward gemcitabine in hypoxic mesothelioma cells, and modulated the expression of the human equilibrative transporter-1 — E. Giovannetti, L.G. Leon, et al. (2016)
• Carbonic anhydrase 9 confers resistance to ferroptosis/apoptosis in malignant mesothelioma under hypoxia — Z. Li, L. Jiang, et al. (2019)
• Splicing modulation as novel therapeutic strategy against diffuse malignant peritoneal mesothelioma — R. Sciarrillo, A. Wojtuszkiewicz, et al. (2019)

Browse more research in British Journal of Cancer, or return to medical research.