Summary Cancer is a complex disease that relies on both oncogenic mutations and non-mutated genes for survival, and therefore coined as oncogene and non-oncogene addictions . The need for more effective combination therapies to overcome drug resistance in oncology has been increasingly recognized, but the identification of potentially synergistic drugs at scale remains challenging. Here we propose a gene-expression-based approach, which uses the recurrent perturbation-transcript regulatory relationships inferred from a large compendium of chemical and genetic perturbation experiments across multiple cell lines, to engender a testable hypothesis for combination therapies. These transcript-level recurrences were distinct from known compound-protein target counterparts, were reproducible in external datasets, and correlated with small-molecule sensitivity. We applied these recurrent relationships to predict synergistic drug pairs for cancer and experimentally confirmed two unexpected drug combinations in vitro . Our results corroborate a gene-expression-based strategy for combinatorial drug screening as a way to target non-mutated genes in complex diseases.

MYCN-amplified (MNA) neuroblastoma is an aggressive neural crest-derived pediatric cancer. However, MYCN is indispensable for development and transcriptionally regulates extensive network of genes. Integrating anti-MYCN ChIP-seq and gene expression profiles of neuroblastoma patients revealed the metabolic enzymes, MTHFD2 and PAICS, required for one-carbon metabolism and purine biosynthesis were concomitantly upregulated, which were more susceptible to metastatic neuroblastoma. Moreover, we found that MYCN mediated the folate cycle via MTHFD2, which contributed one-carbon unit to enhance purine synthesis, and further regulated nucleotide production by PAICS in response to cancer progression. Dual knockdown of the MYCN-targeted gene pair, MTHFD2 and PAICS, in MNA neuroblastoma cells synergically reduced cell proliferation, colony formation, migration ability, and DNA synthesis. By systematically screening the compound perturbagens, the gene expression levels of MTHFD2 and PAICS were specifically suppressed by anisomycin and apicidin across cell lines, and our co-treatment results also displayed synergistic inhibition of MNA neuroblastoma cell proliferation. Collectively, targeting a combination of MYCN-targeted genes that interrupts the interconnection of metabolic pathways may overcome drug toxicity and improve the efficacy of current therapeutic agents in MNA neuroblastoma.