Assessing selectivity and whole-transcriptomic effects of algae extracts on C2 canine mastocytoma cell line: Insights into molecular mechanisms and pathways

Algal extracts have attracted attention in cancer research thanks to their richness in bioactive molecules known for their antiproliferative properties. However, a comprehensive understanding of the underlying cytotoxic mechanisms remains limited, especially concerning specific canine malignancies such as mastocytoma. This study utilizes a genome-wide transcriptome analysis to elucidate the antitumor mechanisms of an algal extract obtained from the red alga Gracilaria cervicornis (GC). Furthermore, this master's project conducts a selectivity assessment by testing the cytotoxic effects of the algal extract on canine tumors and normal cells. The cancer cells include canine mastocytoma C2 cells and canine CLBL-1 lymphoma cells. The normal cells consist of Cf2Th cells derived from normal fetal canine thymus and MDCK cells, isolated from the dog’s kidney. The cell viability was assessed in-vitro using the Alamar Blue test. Our investigation reveals that GC extract exerts dose-dependent cytotoxicity against canine mastocytoma C2 cells, with a median inhibitory concentration (IC50) of 26.41 μg/mL. We employed RNA-seq to unravel the molecular mechanism of GC extract's cytotoxic effects on C2 cells. A total of 1837 differentially expressed genes (DEGs) were detected in response to the highest GC extract dose (GC17), with 674 downregulated and 1163 upregulated. Additionally, 22 DEGs were identified in response to the lower doses (GC8). 21 DEGs were shared between GC17 and GC8, indicating common molecular responses to different extract concentrations. Gene Ontology (GO) analysis revealed that downregulated genes were associated with cell cycle regulation and chromosome segregation, suggesting a disruption of cell division and potential aneuploidy in treated cells. In contrast, upregulated genes were linked to immune responses, cytokine production, inflammatory regulation, and angiogenesis inhibition, collectively creating an unfavorable environment for cancer cells. These results suggest that GC extract modulates C2 cell biology by inhibiting cell cycle progression and activating immunomodulatory pathways. Assessing the selectivity of GC extract, we found that it exhibited higher cytotoxicity against cancer cells compared to normal cells. Indeed, the lowest IC50 values were found with the cancer cells C2 and CLBL-1, reaching 26.41 and 12.63 μg/mL, respectively. On the other side, the highest IC50 values were exhibited by the normal cells. They reached 40.87 μg/mL with Cf2Th cells and 34.25 μg/mL with MDCK cells. Our results suggest that the algal extracts possess a mild selective cytotoxicity with a more negative impact on cancer cell lines. Overall, our study advances the understanding of GC extract's anticancer properties and its selective cytotoxicity. These findings offer hope for the development of canine cancer treatments with reduced adverse effects.