P18-08. Characterization of CD34+ derived dendritic cells generated in vitro and transfected with HIV gene as potential therapeutic vaccine in macaque

International audiencen.

Preclinical evaluation of therapeutic pDNA and mRNA vaccines for chronic hepatitis B

Abstract: Yearly, 1.5 million people get infected with the hepatitis B virus, although preventive vaccines are available. People who are not able to take control of the virus, especially infants, develop a chronic infection. As HBV targets the hepatocytes, a chronic infection can permanently damage the liver and can lead to the development of hepatocellular carcinoma. The hepatitis B virus is able to form circular covalently closed (ccc)DNA in infected hepatocytes and to integrate its DNA in the genome of the host. Both integrated DNA and cccDNA are a source for persistent high concentrations of viral antigens like HBsAg, and cause an immune system tolerant to it, with exhausted T-cells unable to kill infected hepatocytes. Current therapies can not eliminate the virus completely and there is need for a therapy that can reach functional cure, defined as the loss of systemic HBsAg levels and undetectable levels of HBV DNA after a finite treatment. It is believed that a successful treatment consists of multiple modalities including antivirals and immunomodulators that target the high viral and antigen burden as well as inadequate host immune responses. One such immunomodulator is a therapeutic vaccine that introduces HBV viral antigens to the body thereby eliciting a specific immune response. This thesis describes the development and preclinical evaluation of a pDNA vaccine encoding core and pol HBV antigen and a second vaccine in the form of mRNA encoding core, pol and envelope antigen. Robust immune responses were induced in several naive animals with the mRNA vaccine being superior in terms of magnitude of induced CD8 T-cell responses, but specifically the core-specific CD4 T-cell responses were negatively impacted compared to the pDNA vaccine. In AAV-HBV transduced mice, containing an HBV-tolerant, immune suppressed liver environment, the elicited immune responses after two immunizations with pDNA vaccine, were not powerful enough to reduce viral parameters while responses after three immunizations with the mRNA vaccine, reduced viral parameters for 50% of the tested mice. The used platforms allow multiple improvement such as coadministration with adjuvants, altering dosing scheme, finetuning the formulation, exploration of a heterologous regimens, ... The combination of a vaccine with modalities to reduce viral replication and reduce the suppressive HBV antigen levels, should strengthen, and enhance it effects. Such a multimodal therapy is believed to be essential to reduce changes of developing liver damage caused by the constant presence of hepatitis B virus

Role of dendritic cells in HIV-immunotherapy

HIV remains one of the most important deadly infections today, due to the lack of a preventive vaccine and limited access to medical care in developing countries. In developed countries, antiretroviral therapy is available, but it can not eliminate the virus, implying that life-long therapy is necessary. Therefore, it is important that other strategies such as therapeutic vaccination will be developed to control virus replication or even eliminate the virus. The major obstacles towards such a strategy are the huge variability of the virus and the profound HIV-induced immune dysfunction. In this review we focus on dendritic cell based immunotherapies against HIV. To develop an efficient immunotherapy, several elements should be taken into account such as which antigen and loading strategy to use, how to deliver the immunogen, how to optimize the interaction between antigenic peptide and T cells and avoid tolerance. Clearly, to develop an immunotherapy to complement the effect of HAART, it is not sufficient to enhance T cell responses against a consensus sequence or against the prevailing plasma virus. Broad and potent immune response are needed to suppress the entire quasispecies, including the latent reservoir, and to prevent any escape

Simultaneous activation of viral antigen-specific memory CD4+ and CD8+ T-cells using mRNA-electroporated CD40-activated autologous B-cells

Recently, it has become obvious that not only CD8 T-cells, but also CD4 T-helper cells are required for the induction of an effective, long-lasting cellular immune response. In view of the clinical importance of cytomegalovirus (CMV) and human immunodeficiency virus (HIV) infection, we developed 2 strategies to simultaneously reactivate viral antigen-specific memory CD4 and CD8 T-cells of CMV-seropositive and HIV-seropositive subjects using mRNA-electroporated autologous CD40-activated B cells. In the setting of HIV, we provide evidence that CD40-activated B cells can be cultured from HAART-naive HIV-1 seropositive patients. These cells not only express and secrete the HIV p24 antigen after electroporation with codon-optimized HIV-1 gag mRNA, but can also be used to in vitro reactivate Gag antigen-specific interferon-gamma-producing CD4 and CD8 autologous T-cells. For the CMV-specific approach, we applied mRNA coding for the pp65 protein coupled to the lysosomal-associated membrane protein-1 to transfect CD40-activated B cells to induce CMV antigen-specific CD4 and CD8 T-cells. More detailed analysis of the activated interferon-gamma-producing CMV pp65 tetramer positive CD8 T-cells revealed an effector memory phenotype with the capacity to produce interleukin-2. Our findings clearly show that the concomitant activation of both CD4 and CD8 (memory) T-cells using mRNA-electroporated CD40-B cells is feasible in CMV and HIV-1-seropositive persons, which indicates the potential value of this approach for application in cellular immunotherapy of infectious diseases