J., Cun Y., Ozreti? L., Kong G., Leenders F., Lu X., Fernndez-Cuesta L., Bosco G., Mller C., Dahmen I., Jahchan N. Collectively, we characterize a role of historically defined general oncogenes, c-Myc and L-Myc, for regulating lineage plasticity across molecular and histological subtypes. INTRODUCTION Small cell lung cancer (SCLC) represents about 15% of all lung cancers with a median survival time of approximately 10 months and 5-year overall survival at 6% (and expression, in addition to a cluster with low expression of both (and mouse (RP) harbored stochastic amplifications or overexpression associated with classic SCLC histopathology (amplification (is commonly amplified across all three major lung cancer subtypeslung adenocarcinomas, squamous cell lung carcinomas, and SCLC (and are uniquely amplified in SCLC, in a manner suggestive of their role as lineage-amplified genes. In this study, we investigated a previously undescribed of c-Myc and L-Myc as lineage-specific factors to associate SCLC molecular subtypes with histological classes. We investigated the potential of L-Myc and c-Myc to regulate lineage state and identified transcriptional programs unique to each Myc family member, wherein L-Myc regulates neuronal developmental pathways and c-Myc regulates epithelial-to-mesenchymal transition and Notch signaling, biological pathways that are associated with distinct molecular subsets. We showed that c-Myc expression is required to maintain lineage state marker NeuroD1 in NeuroD1-positive SCLC. In addition, c-Myc is incompatible with ASCL1-positive SCLC that ultimately leads to transdifferentiation to NeuroD1-SCLC, consistent with previous findings (and groups and examined mRNA expression and to select cell lines for c-Myc with high expression of and low expression of and vice versa (fig. S1B). We identified 457 differentially expressed genes (test, 0.01; fold change, 1.5), 147 and RTA-408 310 genes overexpressed in and SCLC cell lines, respectively, and defined them as their introductory gene signatures (fig. S1C and table S1). Open RTA-408 in a separate window Fig. 1 Bayesian network analysis reveals unique L-Myc and c-Myc networks associated with distinct biological processes.(A) Schematic of workflow to use SCLC Bayesian causal gene regulatory network to identify networks involving c-Myc and L-Myc. (B) L-Myc subnetwork showing directionality and association of genes when L-Myc gene signature (fig. S1C and table S1) is projected to SCLC Bayesian network. Circles colored in pink represent nodes from L-Myc gene signature. Size of pink circles is directly proportional to the number of outgoing nodes. Nodes indicated in larger text are key RTA-408 drivers of the subnetwork (table S2). (C) Gene ontology (GO) analysis for L-Myc neighbor subnetwork. Enriched functions for these genes are identified on the basis of hypergeometric test against GO terms. (D) Three c-Myc subnetworks showing directionality and association of genes when c-MycCassociated gene signature (fig. S1C and table S1) is projected to SCLC Bayesian network. Circles colored in blue represent nodes from c-Myc gene signature. Size of blue circles is directly proportional to the number of outgoing nodes. Nodes indicated in larger text are key drivers of the subnetwork (table S3). (E) GO analysis for corresponding c-Myc neighbor subnetwork. Enriched functions for these genes are identified on the basis of hypergeometric test against GO terms. To explore the subnetworks associated with RTA-408 L-Myc, we projected the genes up-regulated in the L-MycCexpressing subset onto the network and collected all nodes within two layers from them (see Methods). We identified one large closed subnetwork (L1; Fig. 1B) that comprises 959 gene nodes that included 120 of 310 genes from the L-Myc signature. To identify master regulators of the L-Myc subnetwork, we performed key driver analysis (see Methods) that revealed 13 statistically significant genes (table S2). Examining protein expression of Smad2, a node in the L-Myc subnetwork, revealed higher expression in L-MycCclassified cell lines compared to c-MycCclassified cell lines (fig. S1D). Gene ontology (GO) analysis of this L-Myc subnetwork revealed enrichments of two biological processes: cell cycle progression and neuronal development (Fig. 1C). These processes have been previously implicated as core descriptors of classic SCLC (and loci (pink, L-MycCclassified cell lines; blue, c-MycCclassified cell lines). (F) Heatmap showing 2808 differentially accessible regions [fold change, 5; false discovery rate (FDR), 0.05] between three L-Myc cell lines shown in pink and three c-Myc cell Nkx1-2 lines shown in blue. (G) Enriched ontology by GREAT (Genomic Regions Enrichment of Annotations Tool) analyses for regions differentially accessible.