Increased killing of SCCVII squamous cell carcinoma cells after the combination of Pc 4 photodynamic therapy and dasatinib is associated with enhanced caspase-3 activity and ceramide synthase 1 upregulation

Abstract

Photodynamic therapy (PDT) is not always effective as an anticancer treatment, therefore, PDT is combined with other anticancer agents for improved efficacy. The combination of dasatinib and PDT with the silicone phthalocyanine photosensitizer Pc 4 was assessed for increased killing of SCCVII mouse squamous cell carcinoma cells, a preclinical model of head and neck squamous cell carcinoma, using apoptotic markers and colony formation as experimental end-points. Because each of these treatments regulates the metabolism of the sphingolipid ceramide, their effects on mRNA levels of ceramide synthase, a ceramide-producing enzyme, and the sphingolipid profile were determined. PDT + dasatinib induced an additive loss of clonogenicity. Unlike PDT alone or PDT + dasatinib, dasatinib induced zVAD-fmk-dependent cell killing. PDT or dasatinib-induced caspase-3 activation was potentiated after the combination. PDT alone induced mitochondrial depolarization, and the effect was inhibited after the combination. Annexin V+ and propidium iodide+ cells remained at control levels after treatments. In contrast to PDT alone, dasatinib induced upregulation of ceramide synthase 1 mRNA, and the effect was enhanced after the combination. Dasatinib induced a modest increase in C20:1-and C22-ceramide but had no effect on total ceramide levels. PDT increased the levels of 12 individual ceramides and total ceramides, and the addition of dasatinib did not affect these increases. PDT alone decreased substantially sphingosine levels and inhibited the activity of acid ceramidase, an enzyme that converts ceramide to sphingosine. The data suggest that PDT-induced increases in ceramide levels do not correlate with ceramide synthase mRNA levels but rather with inhibition of ceramidase. Cell killing was zVAD-fmk-sensitive after dasatinib but not after either PDT or the combination and enhanced cell killing after the combination correlated with potentiated caspase-3 activation and upregulation of ceramide synthase 1 mRNA but not the production of ceramide. The data imply potential significance of the combination for cancer treatment.

Interaction of acid ceramidase inhibitor LCL521 with tumor response to photodynamic therapy and photodynamic therapy‐generated vaccine

“…). This finding of the interaction of a sphingolipid metabolism‐modulating drug and PDT at the cellular level is consistent with previously published in vitro studies documenting the enhancement of PDT‐mediated cell killing by short‐chain cationic ceramide LCL29, or drugs modifying the activity of ceramide‐generating enzymes such as dasatinib and fenretinide …”

C6-pyridinium ceramide sensitizes SCC17B human head and neck squamous cell carcinoma cells to photodynamic therapy

“…SCC17B cells were chosen for the study as a clinically-relevant HNSCC model because the cells were derived from a non-metastatic cancer of the larynx, a PDT-treatable HNSCC type [18]. We used Pc4 as the photosensitizer, because our in vitro data showed that combining Pc4PDT with dasatinib, a clinically-approved anticancer agent, enhances cell killing [19]. Also Pc4PDT has been shown to be a promising anticancer treatment in human patients [20, 21].…”

In Vitro Photodynamic Activity of N‐Methylated and N‐Oxidised Tripyridyl Porphyrins with Long Alkyl Chains and Their Inhibitory Activity in Sphingolipid Metabolism

“…Emphasis was placed on the enzymes regulating the metabolism of two sphingolipids with opposing functions, namely pro‐apoptotic ceramide and pro‐proliferative sphingosine 1‐phosphate. In several previous investigations, ceramide was shown to be de novo synthesised and accumulate in response to PDT, and in a recent study by Separovic et al., PDT decreased sphingosine levels and inhibited the activity of acid ceramidase (ASAH) …”

“…In severalp reviousi nvestigations,c eramide was shownt ob ed enovo synthesised and accumulate in response to PDT, [37] and in ar ecent study by Separovic et al,P DT decreasedsphingosine levels and inhibited the activity of acidceramidase(ASAH). [40] The tested compounds upon irradiationo fH CT116 cells did not induce significant changes in the expression levels of ASAH,c eramide synthase 6( LASS6) and phospho-sphingosine kinase 1( SphK1;F igure 10). This finding suggests that these enzymes do not mediate the antitumour effects of the tested compounds in HCT116 cells.…”

“…[41] Both sphingolipids have been implicated in drug resistanceo f cancers,t hus also inhibition of acidc eramidase was suggested as ap otential treatment for cancers with overexpressed acid ceramidase. [40] Furthermore,t he combination of ac eramide analoguea nd PDT in the treatment of mouse SCCVII tumours led to enhanced long-term cures relative to PDT alone. [42] Our results suggest that the porphyrins with inhibitory activity toward either SphK1 and/ora cid ceramidase could be considered as multifunctional PSs, and investigations to developf urther such specific PSs and evaluate their potentialina nticancer therapy are underway.…”