Pemetrexed: Preclinical Study With PD-1 Pathway Blockade

Introduction

Clinical data from cohort G of the KEYNOTE-021 trial and more recently from the phase 3 KEYNOTE-189 trial revealed compelling activity of pemetrexed/platinum chemotherapy in combination with pembrolizumab, a PD-1 antibody, in NSCLC patients. These results suggest a potential positive interaction between these therapeutics.^1,2

Pemetrexed inhibits enzymes in the folate pathway and disrupts cellular processes that are required for cell division. Rapidly dividing cancer cells are the favored targets of pemetrexed.³

The folate pathway is a component of the one-carbon metabolism that plays a critical role during T cell activation. Antifolate treatment like pemetrexed could thus interfere with the one-carbon metabolism and have a detrimental effect on effector T cells in the tumor microenvironment.^4,5 

Preclinical studies were completed to better understand the effects of pemetrexed on tumor immune microenvironment. The following key questions were evaluated using MC38 or Colon26 murine tumor models.

• Does pemetrexed have immune modulating properties?

• Does pemetrexed induce ICD leading to an immune response against tumors?

• Why and how does pemetrexed work with immunotherapy?³

In Vivo Data

In MC38 syngeneic mouse tumor model, pemetrexed monotherapy

• increased frequency of tumor-infiltrating lymphocytes, and

• increased expression of genes associated with the enrichment of 

  • macrophages 

  • dendritic cells/NK cells

  • T helper cells, and

  • interferon signaling.^3,6,7

Pemetrexed monotherapy showed more differentially expressed genes compared to paclitaxel, cisplatin or carboplatin monotherapy, and combination with carboplatin therapy.^3,7

The combination of pemetrexed and anti-PD-L1 improved the antitumor efficacy in MC38 and Colon26 syngeneic mouse tumor models.^6-8 Pathway analysis of gene expression in Colon26 tumors revealed that the combinatorial effect of pemetrexed and anti-PD-L1 was associated with a pronounced inflamed/immune activation phenotype characterized by

• improved antigen presentation

• enhanced T cell signaling and cytokine signaling, and

• engagement of CD4+ T cell-mediated immunity.^3,7,8

These changes correlated with an upregulation of MHC-I and MHC-II on monocytes, macrophages, and tumor cells, suggesting increased immune priming. Supporting this, blockade of T cell priming in lymph nodes after initiation of combination therapy resulted in a loss of combination efficacy.⁷

In Vitro Data

In both MC38 and Colon26 tumor cells, treatment with pemetrexed induced a robust release of HMGB1 indicative of immunogenic cell death.^3,7

Pemetrexed slightly reduced proliferation of primary human T cells, and lead to T cell-intrinsic effects as evidenced by

• enhanced mitochondrial respiratory capacity

• enhanced oxidative respiration, and

• increased expression of

  • cell surface molecules, and

  • immune-related genes.^3,6,7

Murine T cells primed in the presence of pemetrexed displayed improved antigen specific in vitro cytotoxic activity.⁷

References

1. Langer CJ, Gadgeel SM, Borghaei H, et al. Carboplatin and pemetrexed with or without pembrolizumab for advanced, non-squamous non-small-cell lung cancer: a randomised, phase 2 cohort of the open-label KEYNOTE-021 study. Lancet Oncol. 2016;17(11):1497-1508. https://doi.org/10.1016/S1470-2045(16)30498-3

2. Gandhi L, Rodríguez-Abreu D, Gadgeel S, et al. Pembrolizumab plus chemotherapy in metastatic non–small-cell lung cancer. N Engl J Med. 2018;378(22):2078-2092. http://dx.doi.org/10.1056/NEJMoa1801005

3. Schaer D, Geeganage S, Amaladas N, et al. Pemetrexed enhances anti-tumor efficacy of PD-L1 blockade by promoting intra-tumor immune response via tumor and T cell-intrinsic mechanisms. Poster presented at: International Association for the Study of Lung Cancer (IASLC) 19th World Conference on Lung Cancer (WCLC); September 23-26, 2018; Toronto, Canada. Abstract P1.04-07. https://library.iaslc.org/virtual-library-search?product_id=10&author=Schaer&category=

4. Ron-Harel N, Santos D, Ghergurovich JM, et al. Mitochondrial biogenesis and proteome remodeling promote one-carbon metabolism for T cell activation. Cell Metab. 2016;24(1):104-117. https://www.sciencedirect.com/science/article/pii/S1550413116302935?via%3Dihub

5. Tan H, Yang K, Li Y, et al. Integrative proteomics and phosphoproteomics profiling reveals dynamic signaling networks and bioenergetics pathways underlying T cell activation. Immunity. 2017;46(3):488-503. https://www.sciencedirect.com/science/article/pii/S1074761317300729?via%3Dihub

6. Novosiadly R, Schaer D, Lu Z, et al. P3.07-006 Pemetrexed exerts intratumor immunomodulatory effects and enhances efficacy of immune checkpoint blockade in MC38 syngeneic mouse tumor model. J Thorac Oncol. 2017;12(11, Supplement 2):S2300. http://dx.doi.org/10.1016/j.jtho.2017.09.1697

7. Schaer DA, Amaladas N, Lu ZH, et al. The folate pathway inhibitor pemetrexed pleiotropically enhances effects of cancer immunotherapy via immunogenic tumor cell death and T cell-intrinsic mechanisms. Poster presented at: 110th Annual Meeting of the American Association for Cancer Research (AACR); March 29-April 3, 2019; Atlanta, GA. https://www.abstractsonline.com/pp8/#!/6812/presentation/2706

8. Novosiadly RD, Schaer DA, Amaladas N, et al. Pemetrexed enhances anti-tumor efficacy of PD-1 pathway blockade by promoting intra-tumor immune response via immunogenic tumor cell death and T cell-intrinsic mechanisms. Presented as an abstract and poster at: 109th Annual Meeting of the American Association for Cancer Research (AACR); April 14-18, 2018; Chicago, IL. Abstract #4549.

Glossary

HMGB1 = high mobility group box 1 protein

ICD = immunogenic cell death

MHC = major histocompatibility complex

NK cells = Natural Killer cells

NSCLC = non-small cell lung cancer

PD-1 = programmed death-1

PD-L1 = programmed death-ligand 1