My PhD thesis is focused on overcoming melanoma-derived immunosuppression with an oncolytic virus (ORCA-010) and macrophage-targeting therapies. In melanoma, suppression will result in inactivation of antigen-presenting cells, and recruitment of pro-tumour T cells, such as regulatory T cells. So even with checkpoint blockade, the tumour could still evade recognition by the immune system. Thus, the goal of my project is to determine how two immunotherapies can be combined to overcome suppression.
In the tumour microenvironment, p38 mitogen-activated protein kinase (p38 MAPK) is involved in controlling myeloid cell differentiation. Studies have shown that M2-like/myeloid cell differentiation was blocked in melanoma-derived single-cell suspensions by the addition of a small molecule protein kinase inhibitor (PKI), such as p38 MAPK inhibitor (p38 MAPKi).
Macrophages are antigen presenting cells which respond to cytokines in the tumour environment to become M2-like, or M1-like macrophages. The M2-like cells have scavenger functions, secrete immunosuppressive cytokines, and are poor antigen presenters. M1-like macrophages have anti-tumor functions, because they secrete pro-inflammatory cytokines and can present antigens to T cells.
Using an in vitro co-culture model with melanoma cell lines and monocytes, the immune effects of an oncolytic virus and macrophage-targeted therapies can be studied by flow cytometry and cytokine analysis techniques. The oncolytic virus will specifically kill tumour cells, and p38 MAPKi will block the development of tumour-associated M2-like macrophages (TAMs) in favour of an M1-like macrophage phenotype.
The results from my co-culture experiments showed that the melanoma skews monocytes to an M2-like macrophage phenotype. Upon infection with the oncolytic virus and addition of p38 MAPKi, the macrophages switch to pro-inflammatory M1-like macrophages. These cells express co-stimulatory molecules CD80 and CD86, which are necessary for T cell stimulation. Further, in an allogeneic mixed lymphocyte reaction, unlike TAMs, these activated macrophages are functionally capable of expanding both CD4 and CD8 T cells and secrete cytokines in favour of a T helper type 1 (Th1) anti-tumour immune response.
In a B16.OVA immunocompetent mouse tumour model, a triple therapy strategy of ORCA-010, p38 MAPKi, and anti-PD-1 checkpoint inhibitor showed higher recruitment of T cells to the tumour, and more activation of T cells in the spleen. These results demonstrate that an oncolytic virus can function together with small molecule inhibitors to reduce the impact of macrophages on suppression in the tumor environment.
The Netherlands Society of Gene and Cell Therapy (NVGCT) held their annual Spring Symposium on the 6-8 March 2019, and the programme covered all aspects of research, from fundamental to clinical, showing clinical results of gene therapies that are making a significant improvement in the lives of patients. My abstract was selected for a presentation in the immunotherapy session, which provided me with the opportunity to share my results with a wide scientific audience, and from the discussions afterwards, I walked away with a new idea about testing the oncolytic virus in my work.
Highlights of the Symposium for me were talks the following speakers, including Stefaan de Koker (eTheRNA immunotherapies), who presented preclinical data demonstrating how mRNA can be used in combination with a checkpoint inhibitor to activate dendritic cells (DCs) for T cell activation. This is a DC vaccination approach that will mature the patient’s own DCs. Len Seymour (University of Oxford) showcased the benefits of combining an oncolytic virus with bispecific T cell engagers (BiTEs), which are innovative antibodies that allow T cells to recognize antigens directly on tumour cells.
Brian Kaspar (Avexis) showed impressive work in children with a genetic disease called spinal muscular atrophy (SMA) Type 1. With early treatment of Avexis’ gene therapy, most of the children were able to sit and stand unaided. Matthew H. Porteus (Stanford University) discussed how to overcome issues in allogeneic stem cell transplantation between donor and recipient by using gene therapy.
I also have to acknowledge the ESGCT for supporting research, and am honored to have received the ESGCT Best Abstract Award.