The development of in vivo lung cancer models that faithfully mimic the human disease are crucial research tools for understanding the molecular mechanisms driving tumourigenesis. Subcutaneous transplantation assays are the most common, most likely due to their amenability to easily monitor tumour growth, However, this route of administration does not mirror tumours grown within the organ of interest. Importantly, subcutaneous tumours grow in an immune microenvironment distinct from that resident within the lung. This is important, and pertinent for in vivo CRISPR screening focused on identifying genes that influence anti-tumour immunity. To overcome this limitation, we optimised an orthotopic transplantation model, termed iPUL, based on previously published methods.
The iPUL transplantation method allows for the establishment of focal tumour development within the left lobe of recipient mice. This is achieved through the direct injection of either lung cancer cell lines from genetically engineered mouse models (GEMMs); or focused delivery of Cre-recombinase in GEMMs. Utilising mT/mG fluorescent reporter mice, we identified restricted reporter switching of lung epithelial cells in the left lung lobes following iPUL injection of adeno-Cre and lenti-Cre viruses. Consistent with these findings, iPUL delivery of Ad5-CMV-Cre in the KrasG12D/+ GEMM induced focal lung tumour formation in the left lobe of mice. Tumour formation has also been replicated with murine small cell lung cancer cell lines. The iPUL method is advantageous over traditional intranasal or intratracheal delivery methods, as lesions are restricted to one lobe, in contrast to spontaneous generation of multiple tumours. This enables the monitoring of tumour growth more readily over time using PET/CT imaging. iPUL orthotopic transplantation allows lung tumours to be grown in their relevant environment, creating an effective pre-clinical tool for drug studies and CRISPR screening in vivo.