No abstract

This study represents a platform to produce fully human antibodies against emerging infectious diseases in a timely and safe manner.

When expressed in mammalian cells, the nucleocapsid (N) and membrane (M) proteins of the severe acute respiratory syndrome coronavirus (SARS-CoV) are sufficient to form pseudoparticles. To identify region(s) of the N molecule required for pseudoparticle formation, we performed biochemical analysis of the interaction of N mutants and M in HEK293 cells. Using a peptide library derived from N, we found that amino acids 101-115 constituted a novel binding site for M. We examined the ability of N mutants to interact with M and form pseudoparticles, and our observations indicated that M bound to NDelta(101-115), N1-150, N151-300, and N301-422, but not to N1-150Delta(101-115). However, pseudoparticles were formed when NDelta(101-115) or N301-422, but not N1-150 or N151-300, were expressed with M in HEK293 cells. These results indicated that the minimum portion of N required for the interaction with M and pseudoparticle formation consists of amino acids 301-422.

It is well known that trastuzumab (TTZ) is molecular target drug for breast cancer overexpressing human epidermal growth factor receptor 2 (HER2). Novel immunotherapy by human peripheral blood mononuclear cells (PBMCs) activated with TTZ were examined. Proliferation of lymphocytes after adding of TTZ was obtained. Furthermore, lymphocytes activated with TTZ inhibited growth of breast cancer cells in vitro. It is noteworthy that remarkably high cellular cytotoxicity in lymphocytes activated with TTZ compared with that of CD3-and lymphokine (interleukin (IL)2)-activated killer (CD3-LAK) cells commonly used in immunotherapy were revealed.Key words trastuzumab; immunotherapy; human epidermal growth factor receptor 2The human epidermal growth factor receptor 2 (HER2) is overexpressed in 20 to 30% of breast cancers, increasing the aggressiveness of the tumor. The trastuzumab (TTZ) that targets the extracellular domain of HER-2 has a remarkable antitumor effect and is currently used worldwide for the treatment of breast cancer patients overexpressing HER2.1,2) Although the mechanisms underlying the action of TTZ are still not fully understood, inhibitory effects of TTZ by cell cycle arrest in G1 and induction of apoptosis on the growth of HER2-positive breast cancer cells in vitro have been reported. 1,3) In addition, it has been reported that antibody-dependent cellular cytotoxicity (ADCC) plays an important role in the antitumor activity of TTZ.3) The combination chemotherapy with TTZ has been shown to improve survival for women with HER2-overexpressing metastatic breast cancer patients. [4][5][6][7] On the other hand, immunotherapy using lymphocyte stimulated with immunomodulators such as cytokines was noted in cancer therapy as attractive approach, since there is no serious side effects in immunotherapy, and significant prolongation in survival and disease free period in group of immunotherapy compared with no-treatment group have been reported. 8,9) Although the TTZ inhibits growth of human breast cancer cells overexpressing HER2 in vitro, the immunological effects of TTZ for peripheral blood mononuclear cells (PBMCs) have not yet been reported.Yoshida et al. have studied metastasis mechanism of cancer cells 10,11) and for clinical applications of immunotherapy using CD3-and lymphokine (interleukin (IL)2)-activated killer (CD3-LAK) cells. On the other hand, Ueoka et al. have investigated inhibitory effects of hybrid liposomes 12,13) composed of phosphatidylcholine and polyoxyethylenealkyl ether on the growth of tumor cells in vitro, in vivo, and for clinical applications. [14][15][16] On the basis of these studies for cancer, we have tried to develop new immunotherapy using TTZ which is molecular target drug. In this study, we investigated the cellular cytotoxicity of lymphocytes activated with TTZ on growth of breast cancer cells in vitro.First, we examined viability of PBMCs treated with TTZ on the basis of WST-8 assay.17) The results are shown in Fig. 1. No change in the number of PBMCs after adding TTZ solu...