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12.02.2019 | Original Paper

Secretome profiling of heterotypic spheroids suggests a role of fibroblasts in HIF-1 pathway modulation and colorectal cancer photodynamic resistance

Zeitschrift:
Cellular Oncology
Autoren:
María Julia Lamberti, Mandy Rettel, Jeroen Krijgsveld, Viviana Alicia Rivarola, Natalia Belén Rumie Vittar
Wichtige Hinweise

Electronic supplementary material

The online version of this article (https://​doi.​org/​10.​1007/​s13402-018-00418-8) contains supplementary material, which is available to authorized users.

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Abstract

Purpose

Previous analyses of the tumor microenvironment (TME) have resulted in a concept that tumor progression may depend on interactions between cancer cells and its surrounding stroma. An important aspect of these interactions is the ability of cancer cells to modulate stroma behavior, and vice versa, through the action of a variety of soluble mediators. Here, we aimed to identify soluble factors present in the TME of colorectal cancer cells that may affect relevant pathways through secretome profiling.

Methods

To partially recapitulate the TME and its architecture, we co-cultured colorectal cancer cells (SW480, TC) with stromal fibroblasts (MRC-5, F) as 3D-spheroids. Subsequent characterization of both homotypic (TC) and heterotypic (TC + F) spheroid secretomes was performed using label-free liquid chromatography-mass spectrometry (LC-MS).

Results

Through bioinformatic analysis using the NCI-Pathway Interaction Database (NCI-PID) we found that the HIF-1 signaling pathway was most highly enriched among the proteins whose secretion was enhanced in the heterotypic spheroids. Previously, we found that HIF-1 may be associated with resistance of colorectal cancer cells to photodynamic therapy (PDT), an antitumor therapy that combines photosensitizing agents, O2 and light to create a harmful photochemical reaction. Here, we found that the presence of fibroblasts considerably diminished the sensitivity of colorectal cancer cells to photodynamic activity. Although the biological significance of the HIF-1 pathway of secretomes was decreased after photosensitization, this decrease was partially reversed in heterotypic 3D-spheroids. HIF-1 pathway modulation by both PDT and stromal fibroblasts was confirmed through expression assessment of the HIF-target VEGF, as well as through HIF transcriptional activity assessment.

Conclusion

Collectively, our results delineate a potential mechanism by which stromal fibroblasts may enhance colorectal cancer cell survival and photodynamic treatment resistance via HIF-1 pathway modulation.

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Zusatzmaterial
Table S1 Perseus correlation matrix. Correlation coefficients whose magnitude were greater than 0.7 were considered highly correlated (red numbers indicate correlation values <0.7) (DOCX 19 kb)
13402_2018_418_MOESM1_ESM.docx
Table S2 List of proteins identified in homotypic spheroids secretome. Available information: ID identified according to UniProt, name of the protein, corresponding gene, location of the signal peptide in those proteins secreted by the classical pathway (SignalP), NN-score of proteins secreted by non-classical pathways (SecretomeP, the NN-score should be greater than 0.5), LFQ (“label-free quantification”) value (mean and standard deviation). (DOCX 33 kb)
13402_2018_418_MOESM2_ESM.docx
Table S3 List of proteins identified in heterotypic spheroids secretome. Available information: ID identified according to UniProt, name of the protein, corresponding gene, location of the signal peptide in those proteins secreted by the classical pathway (SignalP), NN-score of proteins secreted by non-classical pathways (SecretomeP, the NN-score should be greater than 0.5), LFQ (“label-free quantification”) value (mean and standard deviation). (DOCX 36 kb)
13402_2018_418_MOESM3_ESM.docx
Table S4 List of proteins identified in PDT-treated homotypic spheroids secretome. Available information: ID identified according to UniProt, name of the protein, corresponding gene, location of the signal peptide in those proteins secreted by the classical pathway (SignalP), NN-score of proteins secreted by non-classical pathways (SecretomeP, the NN-score should be greater than 0.5), LFQ (“label-free quantification”) value (mean and standard deviation). (DOCX 45 kb)
13402_2018_418_MOESM4_ESM.docx
Table S5 List of proteins identified in PDT-treated heterotypic spheroids secretome. Available information: ID identified according to UniProt, name of the protein, corresponding gene, location of the signal peptide in those proteins secreted by the classical pathway (SignalP), NN-score of proteins secreted by non-classical pathways (SecretomeP, the NN-score should be greater than 0.5), LFQ (“label-free quantification”) value (mean and standard deviation). (DOCX 55 kb)
13402_2018_418_MOESM5_ESM.docx
Fig S1 Histograms. Histograms were performed using the logarithmic LFQ values (log10 (x)) to visualize the normal distribution of the data (Perseus software) (JPG 63 kb)
13402_2018_418_MOESM6_ESM.jpg
Fig S2 Multiscatter plot. The multiscatter plot was performed using the logarithmic LFQ values (log2 (x)) to visualize the correlation between the replicates (JPG 220 kb)
13402_2018_418_MOESM7_ESM.jpg
High Resolution (TIF 11850 kb)
13402_2018_418_MOESM8_ESM.tif
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