Modified 5-aminolevulinic acid photodynamic therapy suppresses cutaneous squamous cell carcinoma through blocking Akt/mTOR-mediated autophagic flux

Abstract

Background: We previously found that modified 5-aminolevulinic acid photodynamic therapy (M-PDT) is painless and effective in cutaneous squamous cell carcinoma (cSCC) treatment, however, the regulatory mechanism of M-PDT in cSCC is still unclear.Objective: To clarify the effect and relevant regulatory mechanism of M-PDT in cSCC.Methods: The cSCC apoptosis was examined by flow cytometry, TUNEL staining and Cleaved-caspase-3 immunofluorescence, respectively. The autophagy-related characterization was detected by monodansylcadaverine (MDC) staining, transmission electron microscopy (TEM), GFP-LC3B autophagic vacuoles localization and mRFP-EGFP tandem fluorescence-tagged LC3B construct, respectively. The expression of autophagy-related proteins and Akt/mTOR signaling molecules were examined by Western blot. ROS generation was measured by DCFH-DA probe.Results: We found that M-PDT induced cSCC apoptosis in a dose-dependent manner, and this result was related to autophagic flux blockage. The phenomenon is confirmed by the results that M-PDT could induce autophagosomes accumulation and upregulate LC3-II and p62 expression. M-PDT elevated co-localization of RFP and GFP tandem-tagged LC3B puncta in cSCC cell, reflecting autophagic flux blockage, and this was confirmed by transmission electron microscopy. Furthermore, we noticed that M-PDT induced accumulated autophagosomes-dependent apoptosis via targeting ROS-mediated Akt/mTOR signaling. Suppression of Akt potentiated M-PDT-induced upregulation of LC3-II and p62 levels, whereas Akt activation and ROS inhibition rendered resistance to these events. In addition, we observed that lysosomal dysfunction was involved in M-PDT-triggered accumulated autophagosomes-dependent cSCC apoptosis.Conclusion: Our data demonstrates that M-PDT inhibits cSCC through blocking Akt/mTOR-mediated autophagic flux.

Blue Light Aminolevulinic Acid Photodynamic Therapy Downregulates Cell Division and Proliferation Pathways in Cutaneous Squamous Cell Carcinoma

“…28 In laboratory studies, PDT treatment protocols decreased cSSC tumor proliferation and increased apoptosis and autophagy via regulation of MAPK protein, particularly AKT. [20][21][22] Compared to the present study, autophagy was one of upregulated KEGG 2021 pathways with a p < 0.05, but FDR > 0.05.…”

“…These findings using ALA PDT are similar to previously published studies using MAL PDT that indicate that residual cells post PDT undergo changes consistent with a less aggressive cancerous phenotype. 19,20 Additional laboratory and clinical studies need to be performed to confirm the efficacy of ALA PDT for cSCC.…”

“…Previous research has shown that PDT treatment of AKs with methyl-aminolevulinic acid (MAL) and red light may beneficially alter the expression of cancer genes. [19][20][21][22] MAL PDT has been shown to downregulate genes and pathways related to cellular proliferation while upregulating keratinocyte differentiation markers and autophagy. [19][20][21][22] The present study investigates whether 5-ALA blue light PDT similarly modulates gene expression and pathways related to proliferation and cancer pathogenesis in cSCC cells that survived PDT.…”

“…[19][20][21][22] MAL PDT has been shown to downregulate genes and pathways related to cellular proliferation while upregulating keratinocyte differentiation markers and autophagy. [19][20][21][22] The present study investigates whether 5-ALA blue light PDT similarly modulates gene expression and pathways related to proliferation and cancer pathogenesis in cSCC cells that survived PDT. Differential gene expression and pathway analysis of cSCC and human dermal fibroblasts (HDFs) were compared before and after 5-ALA blue light PDT using RNA-Sequencing (RNA-Seq).…”

Honokiol relieves hippocampal neuronal damage in Alzheimer's disease by activating the SIRT3‐mediated mitochondrial autophagy

“…The number of autophagosomes was determined under a fluorescence microscope (BX51, Olympus) by counting the number of LC3 puncta. 43 …”