Chamow S M, Zhang D Z, Tan X Y, Mhatre S M, Marsters S A, Peers D H, Byrn R A, Ashkenazi A, Junghans R P
Department of Recovery Sciences, Genentech Inc., South San Francisco, CA 94080.
J Immunol. 1994 Nov 1;153(9):4268-80.
HIV infection depletes the immune system of the coordinating functions of CD4+ T cells and APCs, whereas the population of CD8+ CTLs remains largely intact: functional but undirected. We have developed a humanized bispecific immunoadhesin-antibody (BIA) that redirects these remaining T cells to kill HIV-infected cells. This BIA expresses effector cell retargeting via a targeting activity that exploits the natural affinity of CD4 for gp120, and a recruiting activity that employs an anti-CD3 moiety to engage CTLs. The resultant molecule is 97% human in origin. In functional tests, this BIA mediated killing of HIV-infected cells using either pure CTL preparations, or whole PBL fractions that additionally include Fc gamma receptor-bearing large granular lymphocyte effectors. In contrast, a human anti-gp120 Ab induced target lysis via Ab-dependent cellular cytotoxicity (ADCC) only with large granular lymphocyte-containing fractions and not with CTLs. ADCC with this Ab was blocked in human serum, whereas BIA-mediated effector cell retargeting lysis of HIV-infected cells by CTLs was preserved. The affinity of the BIA for HIV-gp120 on infected cells and for CD3 epsilon on CTLs was derived in a flow cytometric Scatchard procedure. Relative to the bivalent parent molecules, CD4/gp120 affinity on cells was unchanged in the BIA (Ka 7 x 10(7) M-1), whereas the anti-CD3 affinity was diminished 50-fold (Ka 2 x 10(6) M-1 vs 1 x 10(8) M-1). Physical association of CD3+ effectors and gp120-expressing targets was confirmed by fluorescence microscopy and was dependent upon the presence of BIA and expression of target gp120. The unimpaired cytocidal activity of the BIA in the presence of serum highlights a potentially important advantage of this type of construct over native Abs for HIV-directed therapy.
HIV感染会使免疫系统丧失CD4+ T细胞和抗原呈递细胞（APC）的协调功能，而CD8+ 细胞毒性T淋巴细胞（CTL）群体在很大程度上保持完整：有功能但无定向作用。我们研发了一种人源化双特异性免疫粘附素抗体（BIA），它能重新引导这些剩余的T细胞来杀死HIV感染的细胞。这种BIA通过一种靶向活性来实现效应细胞重定向，该靶向活性利用了CD4对gp120的天然亲和力，以及一种招募活性，该活性采用抗CD3部分来结合CTL。所得分子97%源自人类。在功能测试中，这种BIA使用纯CTL制剂或全外周血淋巴细胞（PBL）组分介导对HIV感染细胞的杀伤，全外周血淋巴细胞组分中还额外包括带有Fcγ受体的大颗粒淋巴细胞效应细胞。相比之下，一种人抗gp120抗体仅通过含大颗粒淋巴细胞的组分，而非CTL，诱导通过抗体依赖性细胞毒性（ADCC）实现靶细胞裂解。用这种抗体进行的ADCC在人血清中被阻断，而BIA介导的CTL对HIV感染细胞的效应细胞重定向裂解得以保留。BIA对感染细胞上的HIV - gp120以及CTL上的CD3ε的亲和力是通过流式细胞术的斯卡查德程序得出的。相对于二价亲本分子，BIA中细胞上的CD4/gp120亲和力未变（Ka 7×10⁷ M⁻¹），而抗CD3亲和力降低了50倍（Ka 2×10⁶ M⁻¹对1×10⁸ M⁻¹）。荧光显微镜证实了CD3+效应细胞与表达gp120的靶细胞的物理结合，且这种结合依赖于BIA的存在和靶标gp120的表达。BIA在血清存在下不受损害的杀细胞活性突出了这种构建体相对于天然抗体在HIV导向治疗方面潜在的重要优势。
