NOG-hIL-4-Tg, a new humanized mouse model for producing tumor antigen-specific IgG antibody by peptide vaccination

<div><p>Immunodeficient mice transplanted with human peripheral blood mononuclear cells (PBMCs) are promising tools to evaluate human immune responses to vaccines. However, these mice usually develop severe graft-versus-host disease (GVHD), which makes estimation of antigen-specific IgG production after antigen immunization difficult. To evaluate antigen-specific IgG responses in PBMC-transplanted immunodeficient mice, we developed a novel NOD/Shi-scid-IL2r<b>γ</b>^null (NOG) mouse strain that systemically expresses the human IL-4 gene (NOG-hIL-4-Tg). After human PBMC transplantation, GVHD symptoms were significantly suppressed in NOG-hIL-4-Tg compared to conventional NOG mice. In kinetic analyses of human leukocytes, long-term engraftment of human T cells has been observed in peripheral blood of NOG-hIL-4-Tg, followed by dominant CD4+ T rather than CD8+ T cell proliferation. Furthermore, these CD4+ T cells shifted to type 2 helper (Th2) cells, resulting in long-term suppression of GVHD. Most of the human B cells detected in the transplanted mice had a plasmablast phenotype. Vaccination with HER2 multiple antigen peptide (CH401MAP) or keyhole limpet hemocyanin (KLH) successfully induced antigen-specific IgG production in PBMC-transplanted NOG-hIL-4-Tg. The HLA haplotype of donor PBMCs might not be relevant to the antibody secretion ability after immunization. These results suggest that the human PBMC-transplanted NOG-hIL-4-Tg mouse is an effective tool to evaluate the production of antigen-specific IgG antibodies.</p></div

Generation of NOG-hIL-4-Tg mice.

<p><b>A,</b> Genotyping of NOG-hIL-4-Tg mice. Human IL-4-specific bands (561 bp) were detected along with the internal control (151 bp). PC: positive control. These results confirmed the gene transfer. NC: negative control. Conventional NOG mice were used. <b>B,</b> ELISA of NOG-IL-4-Tg plasma corresponding to the mice submitted for genotyping. The mice producing more than 100 pg/ml hIL-4 (higher than the broken line) were selected and used for the assays. A representative assay is shown. <b>C,</b> Comparison of hIL-4 protein levels in HD plasma (n = 12) and NOG-hIL-4-Tg mice (n = 57). The broken line shows the 100 pg/ml level. The high hIL-4 (n = 47) and low hIL-4 groups (n = 10) are shown. <b>D</b>, Tissue-specific expression of hIL-4 mRNA. Representative data of 1 NOG and 2 NOG-hIL-4-Tg mice are shown. Human IL-4 specific bands (449 bp) were detected along with β-actin (569 bp).</p

The list of classical HLA types of HDs.

<p>The list of classical HLA types of HDs.</p

Comparison of engrafted human lymphocytes in NOG-hIL-4-Tg and conventional NOG mice.

<p><b>A,</b> Cellularity in HD PBMC-transferred NOG or NOG-hIL-4-Tg mice. NOG-hIL-4-Tg and NOG mice were transplanted with PBMCs (5x10⁶) from the same HD. After 4 weeks, the ratios of each T or B cell subset in BM or spleen obtained from PBMC-NOG (n = 6) and PBMC-NOG-hIL-4-Tg mice (n = 3) were analyzed by FCM. HD PBMCs (n = 20) are shown as a control. CD4-N; naïve CD4+ T cells, CD8-N; naïve CD8+ T cells, CD4-M; memory CD4+ T cells, CD8-M; memory CD8+ T cells, B-T; transitional B cells, B-N; naïve B cells, B-M; memory B cells, B-P; plasmablast/plasma cells. The mean ± SD is shown with the percentage score above each bar. <b>B.</b> Human naïve CD4+ T cells were purified and transferred to both NOG mice (n = 5) and NOG-hIL-4-Tg mice (n = 8). The left 4 panels show the typical expression profiles of CD45RA, CD45RO, CD3 and CD4 among purified naïve CD4+ T cells. MNCs were obtained from PBMC-NOG or PBMC-NOG-hIL-4-Tg mice, and the human CD4+ T cells were analyzed by FCM as shown in the 4 right panels. The lower panels were all gated on the upper gates surrounded by the lines. <b>C,</b> Isolated human CD4+ T cells were stimulated with PMA and ionomycin, and the expression levels of human IL-4 and IFN-γ were analyzed by FCM. Typical patterns are shown.</p