Fig. 3

In vitro and in vivo validation of drug combinations in HNSCC cell lines and xenografts. a Dose-response curves of cell viability after palbociclib monotherapy and palbociclib combined therapy with 500 nM alpelisib and 50 nM GDC-0032 for 72 h in palbociclib sensitive and resistant HNSCC cell lines (red, PIK3CA mut/amp; blue, PIK3CA WT). b Dose-response curves of cell viability at gradient concentrations of palbociclib (blue), PI3K inhibitors (red), or combined Palbociclib-PI3K (purple) therapies for 72 h in representative PIK3CA mut/amp and PIK3CA WT HNSCC cell lines. See the remaining cell lines in each combination in Additional file 1: Fig. S3b-e. c Fa-CI dot plots show the CI values used to identify drug synergies between palbociclib and four PI3K inhibitors in all 8 experimentally examined HNSCC cell lines from each combination (red, PIK3CA mut/amp; blue, PIK3CA WT). Horizontal dashed line indicates CI = 1. Values below this threshold indicate drug synergy, while those above it represent drug antagonism. Fa (fraction affected), CI (combination index). d Cell proliferation measured by EdU analysis after treatment with DMSO vehicle (control; gray), palbociclib (blue), alpelisib (red), and palbociclib-alpelisib combination (purple) for 48 h (left). Population of cells at the G0/G1 phase of cell cycle estimated by flow cytometry analysis after the treatments specified above (right). The data are shown as the mean ± SD. * P < 0.05, ** P < 0.01, *** P < 0.001, and **** P < 0.0001, as estimated by two-way ANOVA. e Tumor growth of FADU cell xenograft tumors after treatment with vehicle, palbociclib, alpelisib, and palbociclib-alpelisib combination (n = 5). The data are shown as the mean ± SEM. ** P < 0.01, **** P < 0.0001, as estimated by two-way ANOVA. mut/amp, mutation or amplification; WT, wild type