Obesity promotes immunotherapy efficacy by up-regulating the glycolytic-mediated histone lactacylation modification of CD8+ T cells

Abstract

The response rate of immune checkpoint blockade (ICB) therapy for non-small-cell lung cancer (NSCLC) remains limited. Recent evidence suggests that obese cancer patients are more likely to benefit from ICB therapy, however, the specific mechanism needs further research. In this study, we found that anti-PD-1 therapy was more effective in obese NSCLC patients compared to normal weight patients and this was verified in mouse NSCLC model. Further bioinformatics analysis indicated that the glycolytic metabolism was markedly elevated in obese NSCLC patients. In vitro co-culture experiment showed that both increased glycolysis of tumor cells and external addition of lactate promoted T cell PD-1 expression. And, PD-1 upregulation was related to monocarboxylate transporter 1 (MCT1)-mediated lactate transport and subsequent lysine lactylation of histones in T cells. Based on the aforementioned data, our study contributes to better application of anti-PD-1 therapy in NSCLC.

Keywords: MCT1; NSCLC; PD-1; histone lactacylation; immunotherapy.

FIGURE 1

Immunotherapy is more effective in obese NSCLC patients. (A) Comparison of clinical characteristics between obese (BMI ≥ 25.0) and normal weight (18.5 ≤ BMI < 25.0) NSCLC patients from our center; (B) PFS and (C) OS comparison between obese and normal weight NSCLC patients treated by Nivolumab; (D) PCA analysis of transcriptome sequencing data of 15 NSCLC patients’ tumor tissues from our center showed BMI contributed the primary difference; (E, F) CIBERSORT analysis of tumor infiltrating immune cells and comparison between the normal weight group and obese group.

FIGURE 2

In orthotopic CDX mouse model of LLC cell line, the obese group showed better response to anti-PD1 treatment and higher intracellular lactate and PD1 expression on CD8+ T cells. (A) Normal weight (CD10) and obese (HFD60) mice were incubated with LLC cell line to generate orthotopic CDX model, and treated by anti-PD1, isotype IgG, and Vehicle. The overall survival was analyzed and compared among different groups; (B) PD-1 expression on CD8+ T cells by flow cytometry; Intracellular lactate analysis of (C) CD45⁻tumor cells and (D) CD8+ T cells.

FIGURE 3

Obese NSCLC patients have higher glycolytic metabolism than normal weight NSCLC patients. (A) GO analysis of canonical glycolysis (GO 0061621) based on transcriptome data of 15 NSCLC patients’ tumor tissues from our center; (B) The heatmap of 14 main genes in canonical glycolytic pathway between the obese group and the normal weight group; (C) Volcano plot of DEGs between the obese group and the normal weight group; (D) Venn diagram of the intersection of DEGs and canonical glycolysis genes; (E) GO analysis of canonical glycolysis (GO 0061621) based on transcriptome data of colorectal adenocarcinoma patients in TCGA database.

FIGURE 4

NSCLC tumor cells elevate PD-1 expression of T cells in a co-culture system. (A) Lactate levels in Jurkat cells when co-cultured with NCI-H23 cells at different concentrations of glucose in 0 h, 6 h, 12 h and 24 h; (B) HK1, (C) PDK1, (D) LDHA mRNA expressions by qPCR in NCI-H23 cells at different concentrations of glucose in 0 h, 6 h, 12 h and 24 h; (E) The protein levels of PD-1 in Jurkat cells when co-cultured with NCI-H23 cells at different concentrations of glucose in 0 h, 6 h, 12 h and 24 h.

FIGURE 5

Lactate released from tumor cells elevates T cell PD-1 expression. (A) Lactate levels in Jurkat cells when co-cultured with NCI-H23 cells treated with glucose, glucose + oxamic acid, glucose + rotenone; (B) PD-1 protein expression in Jurkat cells when co-cultured with NCI-H23 cells treated with glucose, glucose + oxamic acid, glucose + rotenone by Western blot; Comparison of PD-1 expression on mouse spleen CD8+ T cells cultured with or without lactate by (C) flow cytometry, (D) Western blot, and (E) qPCR; (F) Correlation analysis of PD-1 and MCT1 in CD8+ T cells in tumor microenvironment by single-cell sequencing data from ArrayExpress database (data number. E-MTAB-6149); (G) The mRNA expressions of PD-1 and MCT1 in different clusters of tumor infiltrating immune cells by scRNA-seq.

FIGURE 6

Lactate released from tumor cells elevates PD-1 expression on T cell associated with the lysine lactylation of histones in T cells. (A) Diagram of the experimental design: lung tissue of CTRL group (CTRL-Lung), CDX group (CDX-lung) and tumor tissue (CDX-tumor) from CDX models (CD10 group, HFD40 group, HFD60) were harvested 2 weeks later and lysine lactylation of protein were investigated by Western blot; (B) Lysine lactylation of protein in CTRL-Lung, CDX-lung, and CDX-tumor tissues of CD10 group, HFD40 group, and HFD60 group by Western blot; (C) The protein levels of PD-1 expression in CTRL-Lung, CDX-lung, and CDX-tumor tissues of CD10 group, HFD40 group, and HFD60 group by Western blot; (D) The histone protein lysine lactylation of mouse spleen CD8+ T cells treated with or without lactate by Western blot.