The use of anti-beta 1 integrin monoclonal antibody in lung cancer treatment has proven beneficial. Here, we developed a novel monoclonal antibody (mAb), called P5, by immunizing mice with human peripheral blood mononuclear cells (PBMC). Its anti-tumor effect is now being tested, in a clinical phase III trial, in combinatorial treatments with various chemical drugs. To confirm that P5 indeed binds to beta 1 integrin, cell lysates were immunoprecipitated with commercial anti-beta 1 integrin mAb (TS2/16) and immunoblotted against P5 to reveal a 140 kDa molecular weight band, as expected. Immunoprecipitation with P5 followed by LC/MS protein sequence analysis further verified P5 antigen to be beta 1 integrin. Cisplatin treatment upregulated cell surface expression of beta 1 integrin in A549 cells, while causing inhibition of cell growth. When cells were co-treated with different concentrations of P5 mAb, the cisplatin-mediated inhibitory effect was enhanced in a dose-dependent manner. Our findings show that a combinatorial treatment of P5 mAb and cisplatin in A549 cells resulted in a 30% increase in apoptosis, compared to baseline, and significantly more when compared to either the cisplatin or P5 alone group. The entire peptide sequences in CDR from variable region of Ig heavy and light chain gene for P5 mAb are also disclosed. Together, these results provide evidence of the beneficial effect of P5 mAb in combinatorial treatment of human lung adenocarcinoma.

JL1, a specific epitope on CD43, is a potential biomarker for the diagnosis of acute leukemia. Although qualitative assays for detecting leukemia-specific CD43 exist, there is a need to develop quantitative assays for the same. Here, we developed two novel monoclonal antibodies (mAbs), 2C8 and 8E10, recognizing different epitopes on CD43. These clones are capable of pairing with YG5, another mAb against JL1 epitope, because they were selectively obtained using sandwich ELISA. Antigens recognized by 2C8 and 8E10 were confirmed as CD43 by western blotting using the CD43-hFC recombinant protein. When expression on various leukemic cell lines was investigated, 2C8 and 8E10 displayed a disparity in the distribution of the epitope. Enzyme assays revealed that these mAbs recognized a sialic acid-dependent epitope on CD43. Using normal thymus and lymph node paraffin-embedded tissues, we confirmed a difference in the epitopes recognized by the two mAbs that was predicted based on the maturity of the cells in the tissue. In summary, we developed and characterized two mAbs, 2C8 and 8E10, which can be used with YG5 in a sandwich ELISA for detecting leukemia-specific CD43.

No abstract

Canine natural killer (NK) cells are large, granular lymphocytes that are neither B lymphocytes nor T lymphocytes. However, it has been reported that canine NK cells share some of the phenotypic characteristics of T lymphocytes, such as CD3 and CD5. Studies are needed to assess the safety of canine NK cells for immunotherapy, especially because the safety of using allogeneic NK cells as an immunotherapy for dogs has yet to be shown. In this study, the safety of cultured canine NK cells was assessed using a xenogeneic mouse model of graft‐versus‐host disease (GVHD). Mice were injected with either canine peripheral blood mononuclear cells (PBMCs) or cultured NK cells for 2 or 3 weeks. Data were then collected on changes in mice body weights, disease severity scores, and survival rates. Histopathological and immunohistochemical evaluations were also performed. All mice injected with canine PBMCs died within 45 days after injection. Severe clinical signs were caused by GVHD. The histopathological and immunohistochemical evaluations showed that mice injected with canine PBMCs had multiple lesions, including necrosis in their lungs, livers, kidneys, and stomachs, and the injected cells were present around the lesions. By contrast, no mice injected with cultured NK cells without removing the CD3+TCR– cells exhibited any clinical abnormalities. Moreover, they all survived the 90‐day experimental period without exhibiting any histopathological changes. Accordingly, the results of this study suggest that canine NK cells do not cause significant side effects such as GVHD and allogeneic NK cells can safely be used for cancer immunotherapy in dogs.