The role of tumor-derived lactic acid in immunomodulation

Tumors show alterations in their metabolism to fulfill the requirements of excessive proliferation. The most prominent metabolic change is the switch from OXPHOS to aerobic glycolysis, termed Warburg effect. The increased uptake of glucose, subsequent conversion of pyruvate to lactate and the export of lactate in co-transport with protons lead to the extracellular accumulation of lactic acid and occur in the majority of malignant neoplasia. Extracellular lactic acid has been shown to exert a range of direct tumor-promoting effects and there is evidence that various immune cells are impaired by lactic acid. The aim of this dissertation was to clarify the immunomodulatory role of tumor-derived lactic acid. Ldha expression, coding for the lactate-generating enzyme LDH-5, was downregulated by means of shRNA technology in B16.SIY E12 mouse melanoma cells, resulting in low-lactate producing clones (Ldhalow). Cells transfected with scrambled shRNA were used as a control. Ldhalow clones showed higher OXPHOS than control cells due to increased mitochondrial content. Despite the metabolic differences in Ldhalow clones and control cells, proliferation and cell cycle properties were similar in vitro. In immunocompetent C57BL/6 mice, however, tumor growth of Ldhalow clones was significantly decreased compared to that of control cells. Bioluminescence analyses revealed 50% lower intratumoral lactate concentrations, along with reduced Ldha mRNA expression in Ldhalow tumors compared to control tumors. As other enzymes involved in tumorigenesis and immunosuppression, including IDO, ARG, COX and iNOS, did not show any differences in mRNA expression in Ldhalow tumors and control tumors, reduced tumor growth of Ldhalow clones could be attributed to their diminished lactate production. Ldhalow tumors contained more leukocytes and among them lower numbers of myeloid cells and B cells, which are said to be pro-tumorigenic, whereas the amount of infiltrating anti-tumoral cytotoxic T cells and NK cells was increased compared to control tumors. Moreover, cytotoxic T cells and NK cells from Ldhalow tumors produced significantly greater amounts of IFN-γ, thereby showing a higher activity than T cells and NK cells from control tumors. Consistently, IFN-γ production of T cells and NK cells was inhibited by lactic acid in vitro. This suggests that tumor immunosurveillance by activated T and NK cells is possible in a low-lactate tumor microenvironment but compromised in the presence of high amounts of lactate. Accordingly, in the absence of T cells and/or NK cells, tumor growth of Ldhalow clones was accelerated and became similar to that of control cells. Growth control of Ldhalow tumors was also lost in the absence of IFN-γ. This was accompanied by a diminished infiltration of NK cells and activated cytotoxic T cells, indicating a lactate-dependent regulation of IFN-γ production. In vitro analyses of cytotoxic T cells revealed a lactic-acid induced drop of intracellular pH, and subsequent compromised NFAT expression resulting in a diminished IFN-γ production. In summary, this dissertation highlights tumor-derived lactic acid as a potent immunomodulatory metabolite which limits effector cell recruitment into the tumor and impairs IFN-γ production of cytotoxic T and NK cells in highly glycolytic tumors. Thereby, tumor immunosurveillance is lost and tumor progression is facilitated. Interestingly, analyses of cutaneous metastases from melanoma patients displayed high lactate levels along with a reduced infiltration of activated cytotoxic T cells. These findings suggest that lactic acid has immunomodulatory relevance, not only in the mouse model but also for human malignancies

The predictive power of CD3+ T cell infiltration of oral squamous cell tumors is limited to non-diabetic patients

Diabetes mellitus type II (DM) and immune cell infiltration determine patient outcome in many tumor entities. Here we studied a possible link between the metabolic and immune cell status of OSCC patients. Glucose transporter (GLUT) 1 mRNA expression was elevated in all tumor samples, whereas other glycolytic markers such as lactate dehydrogenase (LDH) A or monocarboxylate transporter (MCT) 1 were increased in tumor samples from patients with diabetes and these patients had a significantly worse prognosis compared to non-diabetic patients. Analyses of immune cell infiltration in tumors from diabetic and non-diabetic patients revealed an increased leukocyte (CD45(+)) infiltration compared to normal mucosa only in non-diabetic patients. In line, the amount of CD3(+) T cells per mm(2) tumor tissue, was elevated in patients without diabetes and crucial for patient outcome in OSCC patients without diabetes, as compared to healthy mucosa using fluorescence immunohistochemistry in tissue microarrays of 229 patients. Our results demonstrate that diabetes is a prognostic factor for OSCC patients and associates with decreased leukocyte and CD3(+) infiltration indicating that metabolic differences between diabetic and non-diabetic patients may alter tumor-infiltrating T cells and thereby determine patient outcome

Immunochip analysis identifies association of the <em>RAD50/IL13</em> region with human longevity

International audienceHuman longevity is characterized by a remarkable lack of confirmed genetic associations. Here, we report on the identification of a novel locus for longevity in the RAD50/IL13 region on chromosome 5q31.1 using a combined European sample of 3208 long-lived individuals (LLI) and 8919 younger controls. First, we performed a large-scale association study on 1458 German LLI (mean age 99.0years) and 6368 controls (mean age 57.2years) by targeting known immune-associated loci covered by the Immunochip. The analysis of 142136 autosomal single nucleotide polymorphisms (SNPs) revealed an Immunochip-wide significant signal (P-Immunochip=7.01x10(-9)) for the SNP rs2075650 in the TOMM40/APOE region, which has been previously described in the context of human longevity. To identify novel susceptibility loci, we selected 15 markers with P-Immunochip<5x10(-4) for replication in two samples from France (1257 LLI, mean age 102.4years; 1811 controls, mean age 49.1years) and Denmark (493 LLI, mean age 96.2years; 740 controls, mean age 63.1years). The association at SNP rs2706372 replicated in the French study collection and showed a similar trend in the Danish participants and was also significant in a meta-analysis of the combined French and Danish data after adjusting for multiple testing. In a meta-analysis of all three samples, rs2706372 reached a P-value of PImmunochip+Repl=5.42x10(-7) (OR=1.20; 95% CI=1.12-1.28). SNP rs2706372 is located in the extended RAD50/IL13 region. RAD50 seems a plausible longevity candidate due to its involvement in DNA repair and inflammation. Further studies are needed to identify the functional variant(s) that predispose(s) to a long and healthy life

Tumor immunoevasion via acidosis-dependent induction of regulatory tumor-associated macrophages

Many tumors evolve sophisticated strategies to evade the immune system, and these represent major obstacles for efficient antitumor immune responses. Here we explored a molecular mechanism of metabolic communication deployed by highly glycolytic tumors for immunoevasion. In contrast to colon adenocarcinomas, melanomas showed comparatively high glycolytic activity, which resulted in high acidification of the tumor microenvironment. This tumor acidosis induced Gprotein–coupled receptor–dependent expression of the transcriptional repressor ICER in tumor-associated macrophages that led to their functional polarization toward a non-inflammatory phenotype and promoted tumor growth. Collectively, our findings identify a molecular mechanism of metabolic communication between non-lymphoid tissue and the immune system that was exploited by high-glycolytic-rate tumors for evasion of the immune system

Tumor immunoevasion via acidosis-dependent induction of regulatory tumor-associated macrophages

Many tumors evolve sophisticated strategies to evade the immune system, and these represent major obstacles for efficient antitumor immune responses. Here we explored a molecular mechanism of metabolic communication deployed by highly glycolytic tumors for immunoevasion. In contrast to colon adenocarcinomas, melanomas showed comparatively high glycolytic activity, which resulted in high acidification of the tumor microenvironment. This tumor acidosis induced Gprotein-coupled receptor-dependent expression of the transcriptional repressor ICER in tumor-associated macrophages that led to their functional polarization toward a non-inflammatory phenotype and promoted tumor growth. Collectively, our findings identify a molecular mechanism of metabolic communication between non-lymphoid tissue and the immune system that was exploited by high-glycolytic-rate tumors for evasion of the immune system