P28. Development of patient derived organotypic spheroids to study immune checkpoint inhibition in colorectal liver metastasis following preoperative injection with OX40-ligand expressing oncolytic virus.

Patients with colorectal liver metastasis (CRLM) have a 5-year overall survival of 15% with high risk of recurrence following curative surgery. Development of model systems to study alternative therapies is essential. We hypothesize that intra-tumoral injection of an oncolytic virus expressing OX-40 ligand (DNX-2440) would lead to innate and adaptive immunologic response against CRLM following immune check point inhibition (ICI). We present the development and characterization of patient derived organotypic spheroids (PDOTs) using CRLM injected with DNX-2440 followed by ex-vivo ICI treatment.

 


Methods:

Patients with 2 or more CRLM undergoing curative intent surgical resection were enrolled in a phase 1 trial testing pre-operative injection of DNX-2440 in a target lesion, followed by resection of both target and bystander (non-injected) lesions.  Designated patient tumor tissue was harvested, and PDOTs were created, incubated for 4-5 days and tested with single or dual ICI.  Culture media from the incubations experiments was used to perform cytokine profiling analysis (Bio-Plex Pro Human cytokine 10-plex assay). Flow cytometry using cell lineage markers, and Live/Dead staining was used to characterize PDOTs and ICI response.

 


Results:

Among 7 treated patients,13 tumors were harvested, and PDOTs successfully created from 6 paired target and bystander lesions and 1 unpaired target lesion. Flow cytometry analysis of PDOTs revealed presence of immune and non-immune cellular populations. Notably, we noted significant heterogeneity in cellular populations and tumor viability across all PDOTs. Cytokine analysis revealed consistent expression of IL-6 in 4 of 6 paired patient samples, while TNF-alpha, IL-10 and IL-1 cytokines were only detected in 2 of 6 paired patient’s samples.  We observed variability to single and dual ICI treatment, ranging from cell death induction to no response. Interestingly, similar treatment responses were seen in 3 of 6 patients paired lesions suggesting an abscopal effect.

 


Conclusions:

Colorectal liver metastasis PDOTs can be successfully generated as a model system to recapitulate the CRLM tumor microenvironment, and as such represents a good model to study ex-vivo personalized treatments.  PDOTs assays revealed tumor heterogeneity following DNX-2440 and ICI treatment highlighting the importance of personalized treatment for patients. Future studies will aim to understand the intra-patient tumoral heterogeneity observed in this study.