Therapeutic Effects of Inhibition of Sphingosine-1-Phosphate Signaling in HIF-2α Inhibitor-Resistant Clear Cell Renal Cell Carcinoma

Abstract

Specific inhibitors of HIF-2α have recently been approved for the treatment of ccRCC in VHL disease patients and have shown encouraging results in clinical trials for metastatic sporadic ccRCC. However, not all patients respond to therapy and pre-clinical and clinical studies indicate that intrinsic as well as acquired resistance mechanisms to HIF-2α inhibitors are likely to represent upcoming clinical challenges. It would be desirable to have additional therapeutic options for the treatment of HIF-2α inhibitor resistant ccRCCs. Here we investigated the effects on tumor growth and on the tumor microenvironment of three different direct and indirect HIF-α inhibitors, namely the HIF-2α-specific inhibitor PT2399, the dual HIF-1α/HIF-2α inhibitor Acriflavine, and the S1P signaling pathway inhibitor FTY720, in the autochthonous Vhl/Trp53/Rb1 mutant ccRCC mouse model and validated these findings in human ccRCC cell culture models. We show that FTY720 and Acriflavine exhibit therapeutic activity in several different settings of HIF-2α inhibitor resistance. We also identify that HIF-2α inhibition strongly suppresses T cell activation in ccRCC. These findings suggest prioritization of sphingosine pathway inhibitors for clinical testing in ccRCC patients and also suggest that HIF-2α inhibitors may inhibit anti-tumor immunity and might therefore be contraindicated for combination therapies with immune checkpoint inhibitors.

Hypoxia-inducible factors: cancer progression and clinical translation

“…The development of belzutifan is a major advance in the treatment of RCC, but preclinical studies revealed that some RCC cell line–derived tumors and patient-derived xenografts showed resistance to HIF-2 inhibitors ( 162 , 164 ). One mouse model of RCC that was resistant to PT2385 responded to treatment with acriflavine ( 169 ), which binds to HIF-2α (and HIF-1α) at a site different from that of PT2385 ( 47 , 170 ). There are multiple registrations for clinical trials involving belzutifan at ClinicalTrials.gov ( Supplemental Table 4 ).…”

Lipids as Targets for Renal Cell Carcinoma Therapy

“…Due to the anti-apoptotic and pro-angiogenic roles of S1P, the SphK/S1P/S1PR axis attracts special interest in cancer treatment [ 23 ]. More specifically, this axis seems to be a master regulator of hypoxia by regulating HIF-1α and HIF-2α protein levels in human cancer cell lines including VHL-deficient ccRCC [ 24 ].…”

“…Fingolimod (FTY720) which is an approved immunomodulator for the treatment of multiple sclerosis, acts as an unspecific agonist of all S1P receptors, except S1P 2 , and as a functional antagonist of S1P 1 [ 38 ]. In several HIF-2α-resistant ccRCC cell lines, as well as in HIF-2α-resistant mouse ccRCC models, FTY720 showed anti-proliferative and anti-tumor effects [ 24 ]. Whether the modulation of all S1PRs or only one of them is needed for these actions is not well understood.…”

“…On the other hand, siponimod, which is an S1P 1+5 modulator, was devoid of anti-proliferative effects in RCC colony formation assays, which was attributed to the inability of siponimod to target the S1P 3 receptor [ 24 ]. Notably, RNA sequencing of five human ccRCC cell lines (769-P, A498, 786-O, SLR22, and RCC4) revealed that different cell lines expressed different levels of the five genes, with S1P 1 and S1P 3 being the most abundantly expressed subtypes [ 24 ].…”

“…On the other hand, siponimod, which is an S1P 1+5 modulator, was devoid of anti-proliferative effects in RCC colony formation assays, which was attributed to the inability of siponimod to target the S1P 3 receptor [ 24 ]. Notably, RNA sequencing of five human ccRCC cell lines (769-P, A498, 786-O, SLR22, and RCC4) revealed that different cell lines expressed different levels of the five genes, with S1P 1 and S1P 3 being the most abundantly expressed subtypes [ 24 ]. In RCC, the increased expression of insulin-like growth factor 2 mRNA-binding proteins enhances the stability of S1P 3 mRNA promoting cell proliferation and migration [ 40 ], while patients with RCC characterized by a high expression of S1P 3 have significantly worse overall survival [ 41 ].…”