Cisplatin-Induced Metabolic Responses Measured with Raman Spectroscopy in Cancer Cells, Spheroids, and Canine-Derived Organoids

<i>Ex vivo</i> assessment of drug response with conventional cell viability assays remains the standard practice for guiding initial therapeutic choices. However, such ensemble approaches fail to capture heterogeneities in treatment response and cannot identify early markers of response. Here, we leverage Raman spectroscopy (RS) as an accurate, low-cost, extraction-free, and label-free approach to track metabolic changes in cancer cells, spheroids, and organoids in response to cisplatin treatment. We identified 12 statistically significant metabolites in cells and 19 metabolites in spheroids and organoids as a function of depth. We show that the cisplatin treatment of 4T1 cells and spheroids results in a shift in metabolite levels; metabolites including nucleic acids such as DNA, 783 cm^-1 with <i>p</i> = 0.00021 for cells; <i>p</i> = 0.02173 for spheroids, major amino acids such as threonine, 1338 cm^-1 with <i>p</i> = 0.00045 for cells; <i>p</i> = 0.01022 for spheroids, proteins such as amide III, 1248 cm^-1 with <i>p</i> = 0.00606 for cells; <i>p</i> = 0.00511 for spheroids serve as early predictors of response. Our RS findings were also applicable to canine-derived organoids, showing spatial variations in metabolic changes as a function of organoid depth in response to cisplatin. Further, the metabolic pathways such as tricarboxylic acid (TCA)/citric acid cycle and glyoxylate and dicarboxylate metabolism that drive drug response showed significant differences based on organoid depth, replicating the heterogeneous treatment response seen in solid tumors where there is a difference from the periphery to the tumor core. Our study showcases the versatility of RS as a predictive tool for treatment response applicable from cells to organotypic cultures, that has the potential to decrease animal burden and readout time for preclinical drug efficacy.