Creation of an immunodeficient HLA-transgenic mouse (HUMAMICE) and functional validation of human immunity after transfer of HLA-matched human cells.

Research on human immunology has been hindered by the lack of optimal small animal models, given that the protective immune responses of human and non-human species show significant differences. However, due to ethical constraints[1] and the high cost of clinical trials, it is urgent to improve the current animal models that can mimic faithfully human physiology, particularly the human immune system (HIS). HIS mice had been generated recently by engrafting human hematopoietic stem cells (hHSCs) or human peripheral mononuclear cells (hPBMCs) into highly immuno-deficient mice such as NSG, NOG or NRG mice. However, a major experimental drawback for studies using these models is the rapid onset of Graft-versus-Host Disease (GvHD). In the present study, we overcome this limitation by generating new immuno-deficient mice named "HUMAMICE" (HLA-A2+/+/DR1+/+/H-2-ß2m-/-/IAß-/-/Rag2-/-/IL2rγ-/-/Perf-/- mice), which expressed human HLA molecules instead of mouse MHC molecules (H-2), and whose immuno-deficient status was reversed by transferring functional HLA-matched PBMCs thus producing mice with an immuno-competent status with a functional human immune system. We showed that in this HLA-matched context, the hPBMC-transfer led to high lymphocytes engraftment rates without GvHD over three months in this novel mouse model. Furthermore, to evaluate the utility of the hPBMC-HUMAMICE, we immunized them with commercial vaccine of Hepatitis B virus (HBsAg, Hepvac@) which resulted in robust and reproducible production of high levels of HBsAg-specific antibodies, implying that both transferred T and B lymphocytes were functional in HUMAMICE. These responses are comparable to those observed in human clinical trials with this identical vaccine. In conclusion, these findings indicated that the HLA-matched-hPBMC-HUMAMICE represents a promising model for dissecting human immune responses in various human diseases, including infectious diseases, cancers and tumors, and to facilitate the development of novel vaccines and cellular therapies.

Development of a humanized HLA-A2.1/DP4 transgenic mouse model and the use of this model to map HLA-DP4-restricted epitopes of HBV envelope protein.

A new homozygous humanized transgenic mouse strain, HLA-A2.1(+/+)HLA-DP4(+/+) hCD4(+/+)mCD4(-/-)IAß(-/-)ß2m(-/-) (HLA-A2/DP4), was obtained by crossing the previously characterized HLA-A2(+/+)ß2m(-/-) (A2) mouse and our previously created HLA-DP4(+/+) hCD4(+/+)mCD4(-/-)IAß(-/-) (DP4) mouse. We confirmed that the transgenes (HLA-A2, HLA-DP4, hCD4) inherited from the parental A2 and DP4 mice are functional in the HLA-A2/DP4 mice. After immunizing HLA-A2/DP4 mice with a hepatitis B DNA vaccine, hepatitis B virus-specific antibodies, HLA-A2-restricted and HLA-DP4-restricted responses were observed to be similar to those in naturally infected humans. Therefore, the present study demonstrated that HLA-A2/DP4 transgenic mice can faithfully mimic human cellular responses. Furthermore, we reported four new HLA-DP4-restricted epitopes derived from HBsAg that were identified in both vaccinated HLA-A2/DP4 mice and HLA-DP4-positive human individuals. The HLA-A2/DP4 mouse model is a promising preclinical animal model carrying alleles present to more than a quarter of the human population. This model should facilitate the identification of novel HLA-A2- and HLA-DP4-restricted epitopes and vaccine development as well as the characterization of HLA-DP4-restricted responses against infection in humans.

An H5N1 M2e-based multiple antigenic peptide vaccine confers heterosubtypic protection from lethal infection with pandemic 2009 H1N1 virus.

BACKGROUND: A 2009 global influenza pandemic caused by a novel swine-origin H1N1 influenza A virus has posted an increasing threat of a potential pandemic by the highly pathogenic avian influenza (HPAI) H5N1 virus, driving us to develop an influenza vaccine which confers cross-protection against both H5N1 and H1N1 viruses. Previously, we have shown that a tetra-branched multiple antigenic peptide (MAP) vaccine based on the extracellular domain of M2 protein (M2e) from H5N1 virus (H5N1-M2e-MAP) induced strong immune responses and cross-protection against different clades of HPAI H5N1 viruses. In this report, we investigated whether such M2e-MAP presenting the H5N1-M2e consensus sequence can afford heterosubtypic protection from lethal challenge with the pandemic 2009 H1N1 virus. RESULTS: Our results demonstrated that H5N1-M2e-MAP plus Freund's or aluminum adjuvant induced strong cross-reactive IgG antibody responses against M2e of the pandemic H1N1 virus which contains one amino acid variation with M2e of H5N1 at position 13. These cross-reactive antibodies may maintain for 6 months and bounced back quickly to the previous high level after the 2nd boost administered 2 weeks before virus challenge. H5N1-M2e-MAP could afford heterosubtypic protection against lethal challenge with pandemic H1N1 virus, showing significant decrease of viral replications and obvious alleviation of histopathological damages in the challenged mouse lungs. 100% and 80% of the H5N1-M2e-MAP-vaccinated mice with Freund's and aluminum adjuvant, respectively, survived the lethal challenge with pandemic H1N1 virus. CONCLUSIONS: Our results suggest that H5N1-M2e-MAP has a great potential to prevent the threat from re-emergence of pandemic H1N1 influenza and possible novel influenza pandemic due to the reassortment of HPAI H5N1 virus with the 2009 swine-origin H1N1 influenza virus.