Skip to main page content
Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
, 74 (6), 1133-1151

Multi-OMIC Profiling of Survival and Metabolic Signaling Networks in Cells Subjected to Photodynamic Therapy

Affiliations

Multi-OMIC Profiling of Survival and Metabolic Signaling Networks in Cells Subjected to Photodynamic Therapy

Ruud Weijer et al. Cell Mol Life Sci.

Abstract

Photodynamic therapy (PDT) is an established palliative treatment for perihilar cholangiocarcinoma that is clinically promising. However, tumors tend to regrow after PDT, which may result from the PDT-induced activation of survival pathways in sublethally afflicted tumor cells. In this study, tumor-comprising cells (i.e., vascular endothelial cells, macrophages, perihilar cholangiocarcinoma cells, and EGFR-overexpressing epidermoid cancer cells) were treated with the photosensitizer zinc phthalocyanine that was encapsulated in cationic liposomes (ZPCLs). The post-PDT survival pathways and metabolism were studied following sublethal (LC50) and supralethal (LC90) PDT. Sublethal PDT induced survival signaling in perihilar cholangiocarcinoma (SK-ChA-1) cells via mainly HIF-1-, NF-кB-, AP-1-, and heat shock factor (HSF)-mediated pathways. In contrast, supralethal PDT damage was associated with a dampened survival response. PDT-subjected SK-ChA-1 cells downregulated proteins associated with EGFR signaling, particularly at LC90. PDT also affected various components of glycolysis and the tricarboxylic acid cycle as well as metabolites involved in redox signaling. In conclusion, sublethal PDT activates multiple pathways in tumor-associated cell types that transcriptionally regulate cell survival, proliferation, energy metabolism, detoxification, inflammation/angiogenesis, and metastasis. Accordingly, tumor cells sublethally afflicted by PDT are a major therapeutic culprit. Our multi-omic analysis further unveiled multiple druggable targets for pharmacological co-intervention.

Keywords: Cancer therapy; Metallated phthalocyanines; Non-resectable perihilar cholangiocarcinoma; Reactive oxygen species; Therapeutic recalcitrance; Tumor targeting.

Conflict of interest statement

The authors declare that they have no competing interests.

Figures

Fig. 1
Fig. 1
Cell viability after ZPCL-PDT. HUVEC, RAW 264.7, SK-ChA-1, and A431 cells were incubated with ZPCLs (concentrations can be found in Table S1) and treated with PDT. Two hours (white bar), 6 h (light gray bar), and 24 h (dark gray bar) after PDT, cell viability was determined using the ad WST-1 and eh SRB assay (n = 8 per group). Readers are referred to the experimental section for the significance of the statistical symbols. Metab. act., metabolic activity
Fig. 2
Fig. 2
Gross transcriptional response 90 min after ZPCL-PDT. The Venn diagrams show the number of upregulated (red) and downregulated (green) genes compared to the control group (FDR < 0.05), as well as the overlapping genes between the vehicle (dark toxicity), LC50, and LC90 groups (n = 3 per group). The total number of upregulated and downregulated genes per PDT regimen (full circle) equals the sum of all values enveloped by the respective circle
Fig. 3
Fig. 3
Transcriptional response following ZPCL-PDT. Expression analysis of genes that are involved in NF-кB, UPR, HSF, NFE2L2, HIF-1, and AP-1 signaling as shown by the log2 fold-change (lower right corner). All comparisons were made between the PDT-treated groups versus the control group (n = 3 per group). A gene may correspond to multiple probes as indicated by horizontal splits. Each gene is divided in two halves corresponding to the LC50 (left) and LC90 (right) group. Gray boxes signify probes that exhibited poor quality or were not included in the gene expression analysis
Fig. 4
Fig. 4
Differentially expressed proteins observed after ZPCL-PDT of SK-ChA-1 cells in the LC90 group. Upregulated (in red) and downregulated (in green) proteins between the PDT-treated groups and control group (n = 4 per group) were analyzed using Reactome to assess functional interactions [47, 48]. Arrows indicate activating/catalyzing reactions, whereas straight and dashed lines indicate functional and predicted functional interactions, respectively. Proteins without functional interactions are not displayed in the figure
Fig. 5
Fig. 5
Phosphoproteomic analysis of SK-ChA-1 cells after ZPCL-PDT. The data (n = 4 per group) were analyzed with the Phosphopath plugin in Cytoscape [32]. Increased and decreased phosphorylation of proteins in the PDT-treated groups versus the control group are indicated in red and green, respectively. Straight lines and arrows indicate protein interactions (derived from the Biogrid database [78]) and kinase-substrate interactions (imported from PhosphoSitePlus [79]), respectively. Wikipathways was used for pathway analysis [80], where the dataset was queried against this database to identify pathways. For this figure, EGF, VEGF, insulin, FAK, and MAPK signaling pathways were selected
Fig. 6
Fig. 6
Metabolomic analysis of SK-ChA-1 cells after ZPCL-PDT. a Metabolites were classified into pathways and metabolite expression is depicted as the log2 fold-change (bottom left corner) between treated and control cells (n = 3 per group). Numerical values can be found in Table S6. b Log2 fold-change of metabolites in the category carbohydrate metabolism grouped per pathway. Changes in LC50- (left) and LC90-treated (right) SK-ChA-1 cells compared to control cells are depicted. Identical log2 fold-change values are plotted for 3PG and 2PG and for citrate and isocitrate, as these metabolites could not be resolved. Metabolites indicated in gray could not be quantified. TCA cycle, tricarboxylic acid cycle; PPP, pentose phosphate pathway; G6P, glucose-6-phosphate; F6P, fructose-6-phosphate; F1,6BP, fructose-1,6-bisphosphate; DHAP, dihydroxyacetone phosphate; G3P, glyceraldehyde-3-phosphate; 1,3 BPG, 1,3-bisphosphoglycerate; 3PG, 3-phosphoglycerate; 2PG, 2-phosphoglycerate; PEP, phosphoenolpyruvate; 6pG, 6-phosphogluconate; PenP, pentose-phosphate
Fig. 7
Fig. 7
Overview of the cellular response of SK-ChA-1 cells to supralethal (LC90) PDT. In response to PDT, SK-ChA-1 cells downregulate proteins involved in focal adhesion, tight and adherens junctions, and EGFR signaling. Metabolic processes that are dependent on mitochondria (TCA cycle, urea cycle) appear to be downmodulated, whereas the antioxidant response was activated. On the transcriptomic level, SK-ChA-1 cells exhibited upregulation of AP-1-, HSF-, and NF-кB-mediated signaling that may contribute to cell survival. Straight and dashed arrows indicate direct and indirect reactions, respectively. Phosphorylated proteins are indicated with (P)

Similar articles

See all similar articles

Cited by 3 articles

References

    1. Plaetzer K, Krammer B, Berlanda J, Berr F, Kiesslich T. Photophysics and photochemistry of photodynamic therapy: fundamental aspects. Lasers Med Sci. 2009;24(2):259–268. doi: 10.1007/s10103-008-0539-1. - DOI - PubMed
    1. Weijer R, Broekgaarden M, Kos M, van Vught R, Rauws EAJ, Breukink E, van Gulik TM, Storm G, Heger M. Enhancing photodynamic therapy of refractory solid cancers: Combining second-generation photosensitizers with multi-targeted liposomal delivery. J Photochem Photobiol C Photochem Rev. 2015;23:103–131. doi: 10.1016/j.jphotochemrev.2015.05.002. - DOI
    1. Sibille A, Lambert R, Souquet JC, Sabben G, Descos F. Long-term survival after photodynamic therapy for esophageal cancer. Gastroenterology. 1995;108(2):337–344. doi: 10.1016/0016-5085(95)90058-6. - DOI - PubMed
    1. Zeitouni NC, Shieh S, Oseroff AR. Laser and photodynamic therapy in the management of cutaneous malignancies. Clin Dermatol. 2001;19(3):328–338. doi: 10.1016/S0738-081X(01)00170-5. - DOI - PubMed
    1. Sun ZQ. Photodynamic therapy of nasopharyngeal carcinoma by argon or dye laser—an analysis of 137 cases. Zhonghua Zhong Liu Za Zhi. 1992;14(4):290–292. - PubMed

MeSH terms

Feedback