Assessing drug uptake and response differences in 2D and 3D cellular environments using stimulated Raman scattering microscopy

Cells cultured in 3D environments have been reported to exhibit higher resistance than 2D monolayer cells during drug treatment. Techniques that provide experimental evidence of this phenomenon with spatial molecular profiling data are limited. In this work, we present a three-band stimulated Raman scattering (SRS) imaging system that integrates intracellular drug uptake and growth inhibition measurements of cells in both 2D and 3D environments. We demonstrate that despite having similar drug uptake levels in 2D and 3D cells, the growth of 3D spheroids was less impacted. We further investigate the influences of drug penetration and the extracellular matrix on the 3D cellular drug response. Our study provides valuable insights into the intricate mechanisms of increased drug resistance in 3D tumor models during cancer drug treatments.