Collaborating Investigator: Michael Lim, MD 
Affiliation: Stanford University  

Funding Source: NIH
Grant Number: R01CA230285 
Project Period: 06/01/19-05/31/25  

Significance

A central challenge for brain cancer therapy is delivery to the “cold” tumor immune microenvironment (TIME). Dr. Lim at Stanford is investigating mechanisms by which glioblastoma (GBM) evades the immune system and how combination therapy can be used to enhance an effective immune response against tumors. mRNA delivery has the potential to reprogram the TIME by delivery to immune cells as well as delivery to tumor cells. For example, through delivery of mRNA to brain cancer cells expressing tumor antigens, one could force the expression of Signal 2 co-stimulatory molecules on the surface of tumor cells as well as driving the expression of immunostimulatory cytokines to be excreted into the microenvironment. In such a manner, the tumor cell could acquire similar signaling capacities to an antigen-presenting cell to activate CD8+ cytotoxic T cells against tumors. Immunological reprogramming approaches have great potential for novel immuno-oncology treatments.

Approach

Metabolically reprogramming CAR T cells by genetic disruption of TSC2
Aim 1: Evaluate NIM mRNA nanoparticles for gene delivery to glioma cells and evaluate the TIME reprogramming strategy in vitro.  GL261 glioma cells will be transfected with an array of LEAQ-based NIM particles encapsulating GFP mRNA to determine optimal nanoparticle formulation conditions for transfection. Leading NPs will then be used to deliver mRNA encoding Signal 2 (4-1BBL) and Signal 3 (IL-12) as a combination in the NPs. Splenocytes will be added to the transfected glioma cells in vitro and cells will be characterized for stimulation (such as secreted interferon IFN-γ as measured by ELISA as a surrogate for T cell activation).

Aim 2: Effective NIMs from Aim 1 will be evaluated for delivery of reprogramming mRNA to the TIME in a mouse glioma model. Immuno-competent C57Bl/6 mice will receive GL261 murine glioma tumors orthotopically that will be treated by local injection of the reprogramming NIM mRNA particles. Mice will be evaluated for long-term survival as well as immunological characterization of the TIME by immunohistochemistry, flow cytometry, and qPCR.

Push-Pull:

Push: The NCBIB will provide NIMs to modulate the TIME via delivery of mRNA to tumor cells and immune cells in the TIME and work in combination with local and systemic standards of care. The broadens the applicability of NIMs for immuno-oncology applications.
Pull: The NCBIB will benefit from Lim’s expertise in brain cancer and immunological analysis of the brain tumor microenvironment to better optimize and engineer the particles for TIME transfection and decide on the cargos to include for local tumor immunomodulation.