Dataset Information

Enhancing natural killer cell function with gp41-targeting bispecific antibodies to combat HIV infection.

ABSTRACT:

Objective(s)

The aim of this study was to develop and evaluate the activity of bispecific antibodies (bsAbs) to enhance natural killer (NK) cell antibody-dependent cellular cytotoxicity (ADCC) against HIV-infected cells.

Design

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 mAbs, allowing for improved ADCC activity against Env-expressing target cells.

Methods

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 analyse NK cell degranulation and target cell killing, respectively.

Results

The bsAbs-bound gp41 with similar affinity to their corresponding mAbs 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.

Conclusion

On the basis of their in-vitro killing efficacy, bsAbs may provide a promising strategy to improve NK-mediated immune targeting of infected cells during HIV infection.

SUBMITTER: Ramadoss NS

PROVIDER: S-EPMC7595577 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:ObjectivesHIV-exposed uninfected (HEU) infants have higher rates of severe and fatal infections compared with HIV-unexposed (HUU) infants, likely due to immune perturbations. We hypothesized that alterations in natural killer (NK) cell activity might occur in HEU infants and predispose them to severe infections.DesignCase-control study using cryopreserved peripheral blood mononuclear cells (PBMCs) at birth and 6 months from HEU infants enrolled from 2002 to 2009 and HUU infants enrolled from 2011 to 2013.MethodsNK cell phenotype and function were assessed by flow cytometry after 20-h incubation with and without K562 cells.ResultsThe proportion of NK cells among PBMCs was lower at birth in 12 HEU vs. 22 HUU (1.68 vs. 10.30%, p < 0.0001) and at 6 months in 52 HEU vs. 72 HUU (3.09 vs. 4.65%, p = 0.0005). At birth, HEU NK cells demonstrated increased killing of K562 target cells (p < 0.0001) and increased expression of CD107a (21.65 vs. 12.70%, p = 0.047), but these differences resolved by 6 months. Stimulated HEU NK cells produced less interferon (IFN)γ at birth (0.77 vs. 2.64%, p = 0.008) and at 6 months (4.12 vs. 8.39%, p = 0.001), and showed reduced perforin staining at 6 months (66.95 vs. 77.30%, p = 0.0008). Analysis of cell culture supernatants indicated that lower NK cell activity in HEU was associated with reduced interleukin (IL)-12, IL-15, and IL-18. Addition of recombinant human IL-12 to stimulated HEU PBMCs restored IFNγ production to that seen in stimulated HUU cultures.ConclusionNK cell proportion, phenotype, and function are altered in HEU infants. NK cell cytotoxicity and degranulation are increased in HEU at birth, but HEU NK cells have reduced IFNγ and perforin production, suggesting an adequate initial response, but decreased functional reserve. NK cell function improved with addition of exogenous IL-12, implicating impaired production of IL-12 by accessory cells. Alterations in NK cell and accessory cell function may contribute to the increased susceptibility to infection in HEU infants.