Global COE Program Global Education and Research Center Aiming at the control of AIDS

Program Overview > Program Members > Nobuo, Sakaguchi

Faculty of Life Sciences, Professor
Immunology, M.D., Ph.D. Program Board member, Development of neutralizing antibody against HIV

Development of Novel Antiretroviral Drugs That Are Potent against HIV-1 and Resistance-Deferring and Study of In Vivo Dynamics of HIV-1 Virions and HIV-1-Infected Cells upon Initial HIV-1 Infection and Its Alteration by Antiretroviral Treatment

[Introduction]
The immune system is dependent largely on the acquired immunity against various exogenous pathogens and tumors existing or invading in the body. We have studied the molecular mechanism involved in activation of antigen specific B cells and in generation of high-affinity antibodies against T cell-dependent antigens (1, 2). The high-affinity antibodies produced after vaccination are particularly required for the defense from the viral infection. The eradication of virus is absolutely dependent on the success of designing the efficient vaccine to produce high-affinity antibodies against the virus. Nevertheless, the nature of virus often changes by the high-frequent mutation rate and escape from the first treatment with medicine and the antibodies generated in the patients. A major difficulty in the medical treatment and prevention of AIDS patient is associated with the hypermutation activity of HIV.

[Purpose]
Aim of our study is to develop a new immunomanipulation technique that may be applied for the prevention and hopefully for the therapy of HIV infected patients.

[Background]
The production of high-affinity antibody is initiated by the activation-induced cytidine deaminase (AID) that deaminates deoxycytidine (dC) into deoxyuridine (dU) of immunoglobulin (Ig) V-region genes. The attack of the single stranded Ig V-region DNA causes further alteration of the adjacent sequence, generating somatic hypermutation as a novel hypermutation mechanism by cytidine deamination followed by the other components including uracil-DNA glycosylase(UNG) and error prone DNA polymerases. In a screening of functional molecules involved in generation of somatic hypermutation, we identified the second molecule named germinal center-associated nuclear protein (GANP) that augments generation of AID-induced somatic hypermutation in Ig V-region gene during activation of antigen driven B cells.

[Study plan]
In this gCOE group of AIDS research, we study two major subjects. The first is regarding the molecular mechanism of GANP in regulation of cytidine deamination of AID/Apobec family members. Apobec members are recently known to be cytidine deaminase that deaminates RNA strand or DNA strand of various target genes in the mammalian cells. Recently, the infectivity and resistant activity of human cells to HIV is thought to be dependent on the balance of APOBEC 3G of host cells and Vif protein encoded by HIV genome. Apobec 3b is a cytidine deaminase that mutates dC of single stranded DNA in human cells and targets the HIV cDNA, causing the damage for the virus replication in the host cells. Vif recruits a Cul5-containing E3 ubiquitin ligases resulting in the polyubiqutination and destruction of APOBEC3G.
Therefore, the balance of both molecules may be critical for the clinical course of HIV infection. GANP might play a role in the protection of HIV infection in association with Apobec member protein. We would like to focus on the study of regulation of Apobec member protein particularly in the replication of HIV as collaboration with Dr. Koito, a member of gCOE group.
Secondly, we apply the new biotechnology based on the GANP function to prepare high-affinity monoclonal antibodies that can prevent HIV infection. We have so far demonstrated that the GANP-transgenic (GANP^Tg) mouse is a useful tool to establish extraordinarily high-affinity monoclonal antibodies against the V3 epitope of HIV (3). We have obtained the high-affinity monoclonal antibody that neutralize HIV infection with KD = 1 x 10^-11 M, which is extremely difficult to attain by the conventional method. GANP^Tg mice with international patent has been widely applied to establish the high-affinity antibodies to many kinds of antigens including the virus spike proteins, cancer-specific epitopes of surface membrane molecules, and the endogenous proteins. Using this technology, we will establish the effective monoclonal antibodies for the prevention of highly mutated HIV strains, which will be certainly useful for the treatment of HIV patients.

[Expected results]
Establishment of efficient preventive tool (monoclonal antibody and the associated chemical compounds) by GANP^Tg mice

♦Reference
1.	Kuwahara K, Tomiyasu S, Fujimura S, Nomura K, Xing Y, Nishiyama N, Ogawa M, Imajoh-Ohmi S, Izuta S, Sakaguchi N. Germinal center-associated nuclear protein (GANP) has a phosphorylation-dependent DNA-primase activity that is up-regulated in germinal center regions. Proc Natl Acad Sci USA 2001;98:10279-83.
2.	Kuwahara K, Fujimura S, Takahashi Y, Nakagata N, Takemori T, Aizawa S, Sakaguchi N. Germinal center-associated nuclear protein contributes to affinity maturation of B cell antigen receptor in T cell-dependent responses. Proc Natl Acad Sci USA 2004;101:1010-5.
3.	Sakaguchi N, Kimura T, Matsushita S, Fujimura S, Shibata J, Araki M, Sakamoto T, Minoda C, Kuwahara K. Generation of high-affinity antibody against T cell-dependent antigen in the ganp gene-transgenic mouse. J Immunol 2005;174:4485-94.