Olfactory neuroblastoma mimics molecular subtypes and lineage trajectories of small cell lung cancer [Single cell RNA seq on RPM and RPMA GBC-derived allograft tumors, CellTagged]
The olfactory epithelium relies on active neuron regeneration from basal stem cells and is susceptible to olfactory neuroblastoma (ONB), a rare, aggressive tumor of unclear origins. Here, we establish a new, highly-penetrant, genetically-engineered mouse model of ONB with alterations in Rb1/Trp53/Myc that exhibit a NEUROD1+ immature neuronal state. ASCL1 loss leads to emergence of non-neuronal histopathologies, including a POU2F3+ microvillar-like state. We find ONB tumor heterogeneity to recapitulate developmental states of multipotent globose basal cells (GBCs), which our data demonstrate is a cell of origin for ONB. Similar to small cell lung cancer (SCLC), mouse and human ONB exhibit: mutually exclusive ASCL1, NEUROD1, and POU2F3- like states, an immune-cold tumor microenvironment, intratumoral subtype heterogeneity comprising neuronal and non-neuronal lineages, and subtype plasticity—as evidenced by barcode-based lineage tracing and single-cell transcriptomics. Collectively, our findings highlight conserved developmental trajectories between ONB and SCLC subtypes with significant implications for ONB classification and treatment. Overall design: We harvested normal globose basal cells (GBCs) via immunomagnetic selection for KIT from the pooled olfactory epithelium of n=3-4 Rb1 fl/fl; Trp53 fl/fl; Myc T58A (RPM) or Rb1 fl/fl; Trp53 fl/fl; Myc T58A; Ascl1 fl/fl (RPMA) mice, not exposed to Cre, 10 days following methimazole lesioning. RPM and RPMA GBCs were transformed by treatment with Cre and transduced with a lentiviral barcode-based lineage-tracing technology termed CellTag V1 (established by Samantha Morris's lab, Kong et al., Nature Protocols, 2020). Transformed, CellTagged  GBC's were  implanted with Matrigel into the flanks of Scid/Beige mice.  Upon outgrowth of the GBC-organoid-derived allografts, tumor cells were collected and  processed for single-cell RNA sequencing. Allograft tumors were processed into single cell suspensions, and subject to  10X Genomics single-cell gene expression library construction (Dual-Index 3' GEX) followed by downstream Illumina-based sequencing.