RNA-loaded CD40-activated B cells stimulate antigen-specific T-cell responses in dogs with spontaneous lymphoma

Mason NJ, Coughlin CM, Overley B, Cohen JN, Mitchell EL, Colligon TA, Clifford CA, Zurbriggen A, Sorenmo KU, Vonderheide RH
Source: Gene Ther
Publication Date: (2008)
Issue: 15(13): 955-65
Research Area:
Cancer Research/Cell Biology
Immunotherapy / Hematology
Cells used in publication:
Species: human
Tissue Origin: blood
B cell, human
Species: human
Tissue Origin: blood
Nucleofector® I/II/2b
For RNA electroporation of CD40-B cells, CD40-B cells were harvested after 2 weeks of culture, washed twice in PBS, resuspended at 2–2.5106 per 100 ml of Nucleofector B solution (Amaxa, Cologne, Germany) and electroporated with 2 mg mRNA/sample (pulse program U08). Expression of electroporated mRNA in CD40-B cells was determined by GFP expression 24 h after electroporation.
Cell-based vaccination strategies to induce functional tumor-specific T cells in cancer patients have focused on using autologous dendritic cells. An alternative approach is to use RNA-loaded CD40 activated B cells (CD40-B) that are highly efficient antigen-presenting cells capable of priming naive T cells, boosting memory T-cell responses and breaking tolerance to tumor antigens. The use of tumor RNA as the antigenic payload allows for gene transfer without viruses or vectors and permits major histocompatibility complex (MHC)-independent, multiple-antigen targeting. Here, we use CD40L transfected K562 cells to generate functional CD40-B cells from the peripheral blood of humans and dogs. Testing of RNA-loaded CD40-B cells in dogs allows not only for its development in veterinary medicine but also for determination of its safety and efficacy in a large animal model of spontaneous cancer prior to initiation of human clinical trials. We found that CD40-B cells from healthy humans, healthy dogs and tumor-bearing dogs express increased levels of immune molecules such as MHC and CCR7. Moreover, RNA-loaded CD40-B cells induce functional, antigen-specific T cells from healthy dogs and dogs with lymphoma. These findings pave the way for immunotherapy trials using tumor RNA-loaded CD40-B cells to stimulate antitumor immunity in a large animal model of spontaneous neoplasia.