Functional precision oncology for follicular lymphoma with patient-derived xenograft in avian embryos

Topic: 20. Lymphoma Biology & Translational Research Background: Follicular lymphoma (FL) is an incurable type of B-cell malignancy that accounts for 25 percent of all lymphomas. Although most patients achieve remission with immuno-chemotherapy, almost all of them will experience multiple relapses after treatment. The prognosis is especially poor for around 20 percent of FL patients who relapse within two years after treatment initiation. The understanding of the heterogeneity of treatment responses is limited by the absence of in vitro or in vivo experimental models, mainly due to the strong dependence of tumor cells on their surrounding environment to survive. Aims: This project aimed to develop a primary FL cell xenograft model in chicken embryos to capture treatment response heterogeneity and analyze the mechanisms of response to immune-chemotherapy. Methods: The FL-AVI-PDXTM model was developed by transplanting primary FL cells into chicken embryos. We transplanted 20 biopsy samples, including good (n=11) and poor clinical responders (POD24, n=9), and treated each set of embryos with either RCHOP or vehicle intravenously. The effects of treatment were evaluated 24 hours later with tumor volume measurement by light-sheet microscopy and transcriptomic analysis at the single-cell level (scRNAseq). Results: All samples engrafted successfully in the aorto-gonado-mesonephros of chicken embryos. Tumor volume reduction achieved with RCHOP in the avian embryo was strongly correlated to patient’s clinical outcome (mean tumor volume decreased by 35% in the good responders (n=11) compared to an increase of 2% in the poor responders (n = 9), p < 0.05). Transcriptomic analysis of individual cells was conducted on 14 patients in three different conditions (before and after transplantation with RCHOP or vehicle treatment). We found a robust signature of 21 genes whose expression increased after exposure to RCHOP in all the patients. Among these genes was BAX, a key effector of the mitochondrial outer membrane permeabilization (MOMP), whose inactivation (by CRISPR-CAS9) conferred resistance to RCHOP in cell lines. Furthermore, Venetoclax, a BCL2 inhibitor that lowers the threshold of BAX level required for MOMP, had synergistic effects with RCHOP in cell lines and primary samples in vivo, validating the findings in the model. Summary/Conclusion: FL-AVI-PDXTM enables to study primary FL samples, and to capture the heterogeneity of clinical response to a complex therapeutic regimen. Moreover, it provides a unique opportunity to analyze the mechanisms of response to immune-chemotherapy and to identify therapeutic targets (such as BAX) that can be functionally assessed in the avian embryo.Keywords: Chemotherapy, Animal model, Follicular lymphoma, RNA-seq