Evaluating dendritic cell reprogramming in patient-derived cancer cells and cancer organoids

Abstract

Immunotherapy utilizes the patient’s own immune system for the treatment of cancer. Due to exceptional antigen-presentation capacity Dendritic Cells (DCs) have great potential for cancer immunotherapy. Using direct cellular reprogramming our group has identified a combination of transcription factors, PU.1, IRF8, and BATF3 that reprograms mouse and human fibroblasts and a range of cancer cell lines into antigen-presenting cells resembling type 1 conventional DCs (cDC1) in terms of morphology, transcriptional, epigenetic profiles, and functional features. To support clinical translation of this approach based on induced antigen presentation of cancer antigens, we addressed cDC1 reprogramming in primary cancer tissues obtained from patients with head and neck, urothelial, lung carcinoma, and melanoma. we showed that all primary samples tested were permissive to cDC1 reprogramming with varying efficiencies according to the cancer cell type of origin. Phenotypic analysis demonstrated that reprogrammed primary cancer cells upregulated expression CD45 and HLA-DR which represent well the reprogramming trajectory. Furthermore, reprogrammed cells expressed cDC1-specific marker CD226 as well as co-stimulatory molecules CD40 and CD80, suggesting that reprogrammed primary cells became competent for antigen presentation. In addition, reprogrammed tumor cells secreted pro-inflammatory cytokines- TNF-α and IL-12p70 which may further enhance anti-tumor immunity. To model cDC1 reprogramming within the tumor microenvironment (TME), we generated cancer derived organoids in the presence or absence of fibroblasts. We showed that reprogramming was feasible in organoid 3D models, despite an overall decrease in transduction efficiency, that could be improved by combining transduction protocols with dissociation methods. Interestingly, the efficiency of cDC1 reprogramming was not hampered in cancer organoids generated with fibroblasts, suggesting that the immunosuppressive TME does not negatively impact the reprogramming process. This study brings valuable information for clinical translation of cDC1 cancer cell reprogramming and contributes to the development of novel immunotherapies based on direct reprogramming.

Popular Abstract

Trojan DC in organoids (3D)

Even today, cancer remains one of the major reasons for death worldwide, regardless of the recent advances in the field of cancer therapeutics. Tumors elicit properties with which it can evade the immune system that follow due to rapid progression and mutation in cancer cells. This reinstates the need for better and novel approaches for the treatment of cancer. Immunotherapy, one of the recent advances to fight cancer, utilizes the patient’s own immune system to eradicate tumors. Although immunotherapy can deliver long-term remission even in patients with advanced disease, a vast majority of the patients do not respond to therapy.

The Pereira group from the Stem Cell Center at Lund University identified a combination of transcription factors- PIB (PU.1, IRF8, and BATF3), which is sufficient to convert mouse fibroblasts into antigen-presenting cells resembling type 1 conventional DCs (cDC1) by morphology and functions. These cells are responsible for capturing, processing and presenting antigens to the T-lymphocytes which thereby exhibits anti-tumor responses. The combination was then used to convert tumor cells into tumor-antigen presenting cells (APCs) making the cancer cells traitor to their own kind, hence the name Trojan DC! It is now understood from the data that this reprogramming can be applied to both mouse and human cancer cells. Generation of tumor-APCs holds potential for the development of novel reprogramming-based immunotherapy, surpassing current limitations like tumor cell heterogeneity, immune evasion, and neoantigen identification.

Primary cancer cells are the tumor cells in a patient which reflect better tumor heterogeneity and other properties of the parental tissue. Using the same reprogramming approach, primary samples from different origins were reprogrammed into tumor-APCs. An organoid system allows us to faithfully re-create tumor architecture and microenvironment, so we validated the reprogramming in cancer cells into tumor-APCs also in the 3D system first using cell lines. Based on the functional properties of the tumor-APCs derived from the different primary cells, we selected a set of primary cells to proceed with the study in organoids. The tumor organoids showed similar reprogramming patterns as the cancer cells in 2D, but the overall reprogramming efficiency was lower. However, it was clear that the reprogramming remained independent to the amount of PIB input to them, making lower amounts sufficient for this process. Since tumor microenvironment has a huge immunosuppressive effect on the tumor growth, we formed cancer organoids together with fibroblasts that are a major part of tumor microenvironment contributing to immunosuppression. Interestingly, we observed that with increase in the fibroblast numbers, there was decrease in the levels of transduction, but the levels of reprogramming remained consistent throughout.
The reprogramming of primary cancer cells and organoid study was successful and helps recapitulate the real tumor environment making it easy to understand the system which ultimately aids in development of a new cancer immunotherapy based on direct cellular reprogramming.

Master’s Degree in Molecular Biology, 60 credits, 2022
Department of Biology, Lund University
Supervisor: Filipe Pereira. Division of Molecular Medicine and Gene Therapy, Lund University

• author: Nair, Malavika Sreekumar
• supervisor: Filipe Pereira
• organization: Degree Projects in Molecular Biology
• course: MOBN03 20212
• year: 2022
• type: H2 - Master's Degree (Two Years)
• subject: Biology and Life Sciences
• language: English
• id: 9103308
• date added to LUP: 2022-11-17 15:39:13
• date last changed: 2022-11-17 15:39:13