Intrinsic and/or acquired resistance represents one of the great challenges in targeted cancer therapy. A deeper understanding of the molecular biology of cancer has resulted in more efficient strategies, where one or multiple drugs are adopted in novel therapies to tackle resistance. This beneficial effect of using combination treatments has also been observed in colorectal cancer patients harboring the BRAF(V600E) mutation, whereby dual inhibition of BRAF(V600E) and EGFR increases antitumor activity. Notwithstanding this success, it is not clear whether this combination treatment is the only or most effective treatment to block intrinsic resistance to BRAF inhibitors. Here, we investigate molecular responses upon single and multi-target treatments, over time, using BRAF(V600E) mutant colorectal cancer cells as a model system. Through integration of transcriptomic, proteomic and phosphoproteomics data we obtain a comprehensive overview, revealing both known and novel responses. We primarily observe widespread up-regulation of receptor tyrosine kinases and metabolic pathways upon BRAF inhibition. These findings point to mechanisms by which the drug-treated cells switch energy sources and enter a quiescent-like state as a defensive response, while additionally compensating for the MAPK pathway inhibition.

Breast cancer (BC) comprises multiple distinct subtypes that differ genetically, pathologically, and clinically. Here, we describe a robust protocol for long-term culturing of human mammary epithelial organoids. Using this protocol, >100 primary and metastatic BC organoid lines were generated, broadly recapitulating the diversity of the disease. BC organoid morphologies typically matched the histopathology, hormone receptor status, and HER2 status of the original tumor. DNA copy number variations as well as sequence changes were consistent within tumor-organoid pairs and largely retained even after extended passaging. BC organoids furthermore populated all major gene-expression-based classification groups and allowed in vitro drug screens that were consistent with in vivo xeno-transplantations and patient response. This study describes a representative collection of well-characterized BC organoids available for cancer research and drug development, as well as a strategy to assess in vitro drug response in a personalized fashion. The heterogeneity of breast cancer subtypes can be captured using organoid cultures that can facilitate drug screens that corroborate with patient responses.