Enhancing natural killer cell function with gp41-targeting bispecific antibodies to combat HIV infection.
Enhancing natural killer cell function with gp41-targeting bispecific antibodies to combat HIV infection.
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DOI：
10.1097/qad.0000000000002543
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发表时间：
2020-07-15
期刊：
影响因子：
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通讯作者：
Blish CA
中科院分区：
文献类型：
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作者：
Ramadoss NS;Zhao NQ;Richardson BA;Grant PM;Kim PS;Blish CA
目的研制并评价双特异性抗体(BsAbs)增强NK细胞抗体依赖性细胞毒(ADCC)对HIV感染细胞的杀伤活性。这些抗体是基于患者来源的针对HIV Env保守的gp41残基的抗体，还包括一个针对NK细胞上激活受体CD16的高亲和力单链可变片段(ScFv)。总体而言，我们预计bsAbs将提供比其相应的单抗更高的亲和力和亲和力，允许改善ADCC对表达Env的靶细胞的活性。在293T细胞中表达并纯化了bsAbs及其对应的mAbs。使用生物层干涉计量法以及基于体外感染细胞的流式细胞术结合分析来确定bsAbs和mAbs与其预期靶标的结合。采用体外共培养实验、流式细胞仪检测NK细胞脱颗粒、钙调素释放检测NK细胞对HIV感染细胞的杀伤作用。BsAbs与gp41结合的亲和力与其对应的mAb相似，与CD16的亲和力增强。BsAbs还与体外感染了两种不同HIV毒株的原代CD4T细胞结合。此外，bsAbs还能诱导NK细胞脱颗粒，并对自身感染HIV的CD4T细胞产生杀伤作用。基于它们的体外杀伤效应，bsAbs可能为在HIV感染过程中提高NK介导的免疫靶向性提供了一种有前途的策略。
To develop and evaluate the activity of bispecific antibodies (bsAbs) to enhance NK cell antibody-dependent cellular cytotoxicity (ADCC) against HIV-infected cells. These bsAbs are based on patient-derived antibodies targeting the conserved gp41 stump of HIV Env, and also incorporate a high affinity single chain variable fragment (scFv) targeting the activating receptor CD16 on NK cells. Overall, we expect the bsAbs to provide increased affinity and avidity over their corresponding monoclonal antibodies, allowing for improved ADCC activity against Env-expressing target cells. bsAbs and their corresponding mAbs were expressed in 293T cells and purified. The binding of bsAbs and mAbs to their intended targets was determined using Bio-Layer Interferometry, as well as flow cytometry-based binding assays on in vitro infected cells. The ability of these bsAbs to improve NK cell activity against HIV-infected cells was tested using in vitro co-culture assays, using flow cytometry and calcein release to analyze NK cell degranulation and target cell killing, respectively. The bsAbs bound gp41 with similar affinity to their corresponding mAbs, and had increased affinity for CD16. The bsAbs also bound to primary CD4 T cells infected in vitro with two different strains of HIV. In addition, the bsAbs induce increased NK cell degranulation and killing of autologous HIV-infected CD4 T cells. Based on their in vitro killing efficacy, bsAbs may provide a promising strategy to improve NK-mediated immune targeting of infected cells during HIV infection.