Abstract
Broadly neutralising antibodies (bNAbs) against human immunodeficiency virus type 1 (HIV-1), such as CAP256-VRC26 are being developed for HIV prevention and treatment. These Abs carry a unique but crucial post-translational modification (PTM), namely O-sulfated tyrosine in the heavy chain complementarity determining region (CDR) H3 loop. Several studies have demonstrated that plants are suitable hosts for the generation of highly active anti-HIV-1 antibodies with the potential to engineer PTMs. Here we report the expression and characterisation of CAP256-VRC26 bNAbs with posttranslational modifications (PTM). Two variants, CAP256-VRC26 (08 and 09) were expressed in glycoengineered Nicotiana benthamiana plants. By in planta co-expression of tyrosyl protein sulfotransferase 1, we installed O-sulfated tyrosine in CDR H3 of both bNAbs. These exhibited similar structural folding to the mammalian cell produced bNAbs, but non-sulfated versions showed loss of neutralisation breadth and potency. In contrast, tyrosine sulfated versions displayed equivalent neutralising activity to mammalian produced antibodies retaining exceptional potency against some subtype C viruses. Together, the data demonstrate the enormous potential of plant-based systems for multiple posttranslational engineering and production of fully active bNAbs for application in passive immunisation or as an alternative for current HIV/AIDS antiretroviral therapy regimens.
Highlights
Neutralising antibodies against human immunodeficiency virus type 1 (HIV-1), such as CAP256-VRC26 are being developed for Human Immunodeficiency Virus (HIV) prevention and treatment
This study demonstrates the efficient production of functional anti-HIV bNAbs, CAP256-VRC26 (08 and 09) in N. benthamiana (ΔXTFT)
The presence and functional impact of O-sulfated tyrosine in the antigen-binding domains of Abs have so far only been reported for anti-HIV bNAbs that target the V1V2 region of the HIV envelope trimer
Summary
Neutralising antibodies (bNAbs) against human immunodeficiency virus type 1 (HIV-1), such as CAP256-VRC26 are being developed for HIV prevention and treatment These Abs carry a unique but crucial post-translational modification (PTM), namely O-sulfated tyrosine in the heavy chain complementarity determining region (CDR) H3 loop. The absence of tyrosine sulfation leads to a significant decrease in antigen binding and subsequent loss of function[7] This typical human type PTM is catalysed by tyrosyl protein sulfotransferases (TPSTs)[8,9], which makes the expression of such an antibody version restricted to mammalian cell systems which are generally costly and cumbersome[10]. In planta CDR H3 tyrosine O-sulfation of PG9 and PG16 bNAbs that, like CAP256-VRC26, require this PTM for antigen binding, was reported This was achieved by the overexpression of human TPST1 (hTPST1) in N. benthamiana[14]. The presence of engineered Fc glycans allowed for the production of PG9 and PG16 versions with increased effector functions compared to the mammalian cell-derived variants[14]
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