Periodic Reporting for period 2 - Immunometabolomics (CD8+ T cell metabolism in anti-tumor response)
Période du rapport: 2019-07-01 au 2020-06-30
We aimed to understand basic aspects of metabolism in CD8+ T cells, and how nutrient-deprived conditions in the tumor microenvironment shape CD8+ T cell metabolism and function, to see if it is possible to identify pharmacological or nutritional interventions that could synergize with existing checkpoint inhibitors for immunotherapy through the modulation of 1C and NADPH metabolism. To this end, we have combined nutrient and pharmacological manipulations in vitro and in vivo, using mass spectrometry and isotope tracers to read out pathway activity and cell surface markers and released/intracellular cytokines to read out immune cell functional status.
By using stable isotope tracers coupled to LC-MS-based metabolomics detection we have been able to deepen in the knowledge of 1C/folate and NADPH metabolism in CD8+ T cells and T cell derived hematological malignancies.
NADPH reducing power can come from various pathways. Using Deuterium-labeled substrates, we identified the oxidative pentose phosphate pathway (oxPPP, which catalyzes the oxidation of glucose to ribose) as the main source of NADPH in T cells. Inhibition of the pathway with an in house developed inhibitor (G6PDi-1) leads to NADPH depletion and loss of inframmatory cytokine production in effector T cells. The results were published in: “A small molecule G6PD inhibitor reveals immune dependence on pentose phosphate pathway” (Ghergurovich and Garcia-Canaveras et al, Nature Chemical Biology, 2020).
The enzyme serine hydroxymethyltransferase (SHMT) catalyzes the conversion of serine into glycine and folate-bound 1C units, that are essential for nucleotide biosynthesis and thus enable cell proliferation. We have developed SHIN2, the first SHMT inhibitor with favorable PK/PD profile for in vivo use and that effectively inhibits SHMT in vivo. SHIN2 inhibits proliferation of CD8+ T cells in vivo in a mouse model of effector CD8+ T cell response thus suggesting a key role for serine catabolism through SHMT1/2 in T cell proliferation. More importantly, SHIN2 increases survival in a NOTCH1-driven mouse primary T-ALL in vivo and in a human patient-derived xenograft in vivo, and synergizes with methotrexate in both settings. The results were published in: “SHMT inhibition is effective and synergizes with methotrexate in T-cell acute lymphoblastic leukemia” (Garcia-Canaveras and Lancho et al, Leukemia, 2020).
Antifolates (e.g. methotrexate and pemetrexed) are well-stablished treatments for cancer and autoimmune diseases. However, development of resistance is a common phenomena. The mitochondrial isoform of SHMT (SHMT2) is strongly upregulated in cancer and is the main source of 1C units for nucleotide biosynthesis in several cancers. SHMT 1/2 inhibitors effectively inhibit cancer cell proliferation in vitro. We have developed the first dual SHMT1/2 inhibitor, SHIN2, with favorable PK/PD profile for in vivo use and demonstrated in vivo target engagement by using a newly developed method combining isotope tracers and LC-MS-based analysis of serum metabolites. We showed that SHIN2 alone is effective and increases survival in a T-ALL mouse model in vivo and that synergizes with the antifolate drug methotrexate both in vitro and in vivo in mouse models of T-ALL. Moreover, cell rendered resistant to methotrexate remain sensitive to SHMT1/2 inhibition by SHIN2/. Thus, not only we have provided tools for evaluating SHMT 1/2 in vivo, but we have also identified that SHMT inhibition offers a complementary strategy in the treatment of T-ALL.