Targeting SphK2 Reverses Acquired Resistance of Regorafenib in Hepatocellular Carcinoma

Abstract

Background: Regorafenib is a second-line therapy drug used for advanced hepatocellular carcinoma (HCC). Unfortunately, the survival benefit of the patients receiving this treatment is modest, which may be attributed to drug resistance. In the present study, sphingosine kinase 2 (SphK2) was targeted to reverse regorafenib resistance in HCC. Methods: The functions of SphK2 and sphingosine-1-phosphate (S1P), the catalytic product of SphK2 in regorafenib resistance of HCC cells, were evaluated by cell counting kit-8 assay, colony formation, cell cycle evaluation, and annexin V-fluorescein isothiocyanate/propidium iodide double-staining assay. The antitumor activity of combined treatment of regorafenib and the SphK2-specific inhibitor ABC294640 was examined in HCC cells in vitro and xenograft model in vivo. The molecular mechanisms of SphK2/S1P-mediating regorafenib resistance were investigated using cell line establishment and Western blot analysis. Results: Well-developed regorafenib-resistant HCC cells indicated high expression levels of SphK2. The sensitivity to regorafenib of regorafenib-resistant HCC cells was restored following SphK2 knockdown or pharmacological inhibition by ABC294640. In addition, ectopic expression of SphK2 and exogenous addition of S1P decreased the sensitivity of HCC cells to regorafenib. Furthermore, the combination treatment with ABC294640 sensitized resistant tumor to regorafenib in xenograft model of HCC. The phosphorylation levels of nuclear factor κB (NF-κB), as well as those of signal transducer and activator of transcription 3 (STAT3), were positively associated with SphK2 and S1P. Conclusions: SphK2/S1P mediates regorafenib resistance of HCC through NF-κB and STAT3 activation. Targeting SphK2 by ABC294640 potently reduces regorafenib resistance of HCC cells both in vitro and in vivo. The combination of ABC294640 and regorafenib could be developed as a novel potential treatment strategy for advanced HCC.

S1P induces proliferation of pulmonary artery smooth muscle cells by promoting YAP-induced Notch3 expression and activation

“…Studies have shown that S1P promotes the activation of STAT3 in non-PASMCs ( 20 , 21 ). To examine the effect of S1P on STAT3 in PASMCs, cells were stimulated with 1000-nM S1P at different time points, and Western blotting was performed to detect the phosphorylation level of STAT3 and total STAT3 level.…”

“…MicroRNAs are small noncoding RNAs that bind to the 3' untranslated regions of the target genes, thereby inhibiting translation and/or inducing degradation of target mRNAs. It has been shown that signal transducers and activators of transcription 3 (STAT3), an important downstream of S1P ( 20 , 21 ), mediates the expression of miR-135b in lymphoma cells ( 22 , 23 ). Meanwhile, miR-135b can downregulate the expression of the β-transduction repeat-containing protein (β-TrCP) in lung cancer cells ( 24 ) and hepatocellular carcinoma cells ( 25 ).…”

Targeting Sphingolipids for Cancer Therapy

“…The decrease in proliferation is mediated by inhibition of SPHK2 activity ( 82 , 85 , 97 ), S1P depletion ( 76 , 79 , 84 , 85 , 95 , 97 , 99 ), accumulation of ceramide ( 79 , 84 , 85 , 89 , 99 ), induction of apoptosis (increased caspase cleavage, decreased Bcl-2 levels, and decreased NOXA transcription) ( 76 , 78 , 79 , 84 – 91 , 94 – 96 , 98 , 99 , 102 , 103 ), induction of autophagy (increased LC3-II and beclin-1 levels) ( 77 , 83 , 98 ), estrogen/androgen receptor signaling (decrease in progesterone or androgen receptor levels) ( 75 , 81 , 82 ), cell cycle arrest (increased Myt1, p-cdc2, p53, and p21 levels and decreased pRb, cyclin B1, and cyclin D1 levels) ( 81 , 85 – 87 , 100 , 101 ), and modulation of cell survival pathways (decrease or inhibition of NF-κB, pERK1/2, pJNK, pAKT, c-Myc, and survivin expression, as well as p21-activated kinase 1 (PAK1)/p-Lin-11/Isl-1/Mec-3 kinase 1 (LIMK1)/Cofilin1 signaling) ( 77 , 78 , 81 , 82 , 84 , 87 , 90 – 92 , 100 – 103 ) ( Figure 3 ). The combination of ABC294640 with other drugs, such as regorafenib, sorafenib, PDMP, and ABT-199, induces synergistic potentiation of the treatment effect, reducing chemoresistance in various cancer types ( 98 , 99 , 103 , 104 ). For example, SPHK2/SPP1 arbitrates regorafenib resistance by activating signal transducer and activator of transcription 3 (STAT3) and nuclear factor kappa light chain enhancer of activated B cells (NF-κB).…”

The Role of IGF/IGF-1R Signaling in Hepatocellular Carcinomas: Stemness-Related Properties and Drug Resistance

“…Meanwhile, the Hippo and IGF/IGF-1R signaling pathways were demonstrated to be linked in conferring sorafenib resistance in HCC cells [ 163 ]. Additional investigations into mechanisms behind regorafenib resistance in HCC found that resistance is mediated via proteins controlling the cell cycle and proliferation—NF-κB, STAT3, Akt, cMyc [ 252 , 253 , 254 , 255 ]. Thus, the IGF/IGF-1R signaling may be a potential target for combination treatment with regorafenib.…”