Tumor Lactic Acidosis: Protecting Tumor by Inhibiting Cytotoxic Activity Through Motility Arrest and Bioenergetic Silencing

Abstract

Adoptive T cell therapy (ACT) is highly effective in the treatment of hematologic malignancies, but shows limited success in solid tumors. Inactivation of T cells in the tumor milieu is a major hurdle to a wider application of ACT. Cytotoxicity is the most relevant activity for tumor eradication. Here, we document that cytotoxic T cells (CTL) in lactic acidosis exhibited strongly reduced tumor cell killing, which could be compensated partly by increasing the CTL to tumor cell ratio. Lactic acid intervened at multiple steps of the killing process. Lactic acid repressed the number of CTL that performed lytic granule exocytosis (degranulation) in tumor cell co-culture, and, additionally impaired the quality of the response, as judged by the reduced intensity of degranulation and lower secretion of cytotoxins (perforin, granzyme B, granzyme A). CTL in lactic acid switched to a low bioenergetic profile with an inability to metabolize glucose efficiently. They responded to anti-CD3 stimulation poorly with less extracellular acidification rate (ECAR). This might explain their repressed granule exocytosis activity. Using live cell imaging, we show that CTL in lactic acid have reduced motility, resulting in lower field coverage. Many CTL in lactic acidosis did not make contact with tumor cells; however, those which made contact, adhered to the tumor cell much longer than a CTL in normal medium. Reduced motility together with prolonged contact duration hinders serial killing, a defining feature of killing potency, but also locally confines cytotoxic activity, which helps to reduce the risk of collateral organ damage. These activities define lactic acid as a major signaling molecule able to orchestrate the spatial distribution of CTL inside inflamed tissue, such as cancer, as well as moderating their functional response. Lactic acid intervention and strategies to improve T cell metabolic fitness hold promise to improve the clinical efficacy of T cell–based cancer immunotherapy.

Lactate Is a Metabolic Mediator That Shapes Immune Cell Fate and Function

“…In our studies, 12.5-mM lactic acid suppressed cytokine production and reduced the pH of the buffered media to 6.7 . It remained significantly lower than control media for 1 h. Fischbeck et al (2020) observed a similar drop in pH with 10-mM lactic acid and a drop to pH 6.3 with 20-mM lactic acid addition. Additionally, we observed that formic acid, with a similar pKa, also suppressed IL-6 secretion, and sodium lactate above 20 mM could mimic lactic acid.…”

“…Lactate and the associated H + ions have similar inhibitory effects on glycolysis in immune cells ( Figure 1 ). Lactic acid, which rapidly dissociates to lactate and H + , suppressed glycolysis in the myeloid lineage and T cells, as measured by Seahorse metabolic flux analysis ( Dietl et al, 2010 ; Errea et al, 2016 ; Ratter et al, 2018 ; Fischbeck et al, 2020 ; Quinn et al, 2020 ). Lactic acid suppressed glucose uptake, lactate export, glycolytic enzyme expression, and intracellular ATP levels in monocytes and mast cells ( Dietl et al, 2010 ; Caslin et al, 2019 ).…”

“…In NK cells, lactic acid (both lactate and acidity) inhibited cytolytic function ( Husain et al, 2013 ). In CD8 + T cells, lactic acid reduced proliferation, degranulation, motility, cytolytic activity, and inflammatory mediator secretion (interferon (IFN)γ, perforin, and granzyme; Fischer et al, 2007 ; Mendler et al, 2012 ; Haas et al, 2015 ; Fischbeck et al, 2020 ). In CD8 + T cells, these effects are likely due to altered receptor signaling, as lactic acid suppressed protein kinase activity ( Mendler et al, 2012 ), and suppression was dependent on MCT-1 ( Haas et al, 2015 ; for signaling, see Figure 1C ).…”

“…Lactate can not only provide a survival advantage for tumor cells by upregulating oncogenes and inducing angiogenesis; it promotes immune evasion and is considered an oncometabolite ( Choi et al, 2013 ; San-Millán and Brooks, 2017 ; San-Millán et al, 2019 ). Lactic acid from the tumor microenvironment suppressed CD8 + T cell activation and tumor killing ( Fischbeck et al, 2020 ). Additionally, lactic acid promoted both M2 differentiation and a mixed M1/M2 macrophage phenotype characteristic of tumor-associated macrophages that aid in immune escape ( Colegio et al, 2014 ; Helm et al, 2014 ; Penny et al, 2016 ; Mu et al, 2018 ; de-Brito et al, 2020 ; Paolini et al, 2020 ).…”

Conserved angio-immune subtypes of the cancer microenvironment predict response to immune checkpoint blockade therapy

“…Moreover, under sub-optimal perfusion, glycolytic co-option produces a highly acidic environment that suppresses T-cell effector functions 8 . Additionally, angiogenic tumor endothelial cells express death signals like FAS ligand (FASLG) that induce apoptosis in infiltrating cytotoxic T-cells 9 .…”