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A plant-biotechnology approach for producing highly potent anti-HIV antibodies for antiretroviral therapy consideration

Abstract

Despite a reduction in global HIV prevalence the development of a pipeline of new therapeutics or pre-exposure prophylaxis to control the HIV/AIDS epidemic are of high priority. Antibody-based therapies offer several advantages and have been shown to prevent HIV-infection. Plant-based production is efficient for several biologics, including antibodies. We provide a short review on the work by Singh et al., 2020 who demonstrated the transient production of potent CAP256-VRC26 broadly neutralizing antibodies. These antibodies have engineered posttranslational modifications, namely N-glycosylation in the fragment crystallizable region and O-sulfation of tyrosine residues in the complementary-determining region H3 loop. The glycoengineered Nicotiana benthamiana mutant (ΔXTFT) was used, with glycosylating structures lacking β1,2-xylose and/or α1,3-fucose residues, which is critical for enhanced effector activity. The CAP256-VRC26 antibody lineage targets the first and second variable region of the HIV-1 gp120 envelope glycoprotein. The high potency of this lineage is mediated by a protruding O-sulfated tyrosine in the CDR H3 loop. Nicotiana benthamiana lacks human tyrosyl protein sulfotransferase 1, the enzyme responsible for tyrosine O-sulfation. The transient coexpression of the CAP256-VRC26 antibodies with tyrosyl protein sulfotransferase 1 in planta had restored the efficacy of these antibodies through the incorporation of the O-sulfation modification. This approach demonstrates the strategic incorporation of posttranslational modifications in production systems, which may have not been previously considered. These plant-produced CAP256-VRC26 antibodies have therapeutic as well as topical and systemic pre-exposure prophylaxis potential in enabling the empowerment of young girls and women given that gender inequalities remain a major driver of the epidemic.

Introduction

Despite the global reduction in HIV prevalence, of the 38 million people living with human immunodeficiency virus/ acquired immunodeficiency syndrome (HIV/AIDS), only 25.4 million people are currently on antiretroviral treatment [1]. A compounding factor is gender inequality which remains a major social driver of the epidemic, with young women and adolescent girls accounting for one in four new infection in 2019 [1]. Thus, development of protective vaccines and a pipeline of new therapeutics or prophylaxes to control the HIV/AIDS epidemic remains a high priority [2, 3]. As an alternative or as a complement to small-molecule therapy such as highly active antiretroviral therapy (HAART) which utilizes small-molecule therapeutics (Table 1) in varying combination, antibody-based therapies have several advantages, such as safety and specificity [4]. The use of VRC01, a broadly neutralizing antibody (bNAb) has been shown to prevent HIV-infection in over 70% of people exposed to strains which are able to be neutralized by VRC01 [5].

Table 1 Food and Drug Administration (FDA)-approved small molecule HIV therapeutics

The production of protein-based biopharmaceuticals has been dominated by the use of mammalian cell culture-based approaches [6]. There are alternative systems to mammalian-based production for protein-based biopharmaceuticals production; however, the ability of these non-mammalian systems to produce monoclonal antibodies (mAbs) is limited by their inherent properties. These properties comprise of cellular machinery which influence the ability to correctly fold both monomeric and multimeric structures and incorporate the correct post-translational modifications (PTMs) at the correct amino acids [7]. In contrast to other developing regions, the local manufacturing of protein-based vaccines and biopharmaceuticals is limited or growing at a slow rate in Africa thereby contributing to the widening trade deficit and limited access to essential medicines by the underprivileged [8]. Cost of goods, which will influence mass roll-out of such products, is also a major consideration [9]. Plant-based production is efficient for a number of biologics, and is particularly suitable for cost-sensitive markets in Africa and other low- and middle-income countries (LMICs) (reviewed by [10, 11]). The Nicotiana benthamiana (N. benthamiana) plant-based system has been employed for the production of anti-HIV antibodies, such as 2G12, VRC01, and PG9 [12,13,14,15], and the efficacy of some plant-produced versions have already been tested in animal trials [16].

We provide a short review on the paper entitled “Plant-based production of highly potent anti-HIV antibodies with engineered posttranslational modifications”. This publication reported the production of potent CAP256-VRC26 bNAbs with engineered PTMs in the antigen and fragment crystallizable (Fc) (receptor binding) region of the antibodies, respectively [17]. The ability to perform crucial N-glycosylation lacking β1,2-xylose and/or α1,3-fucose residues using glycoengineered N. benthamiana (ΔXTFT) plants complemented with the coexpression of the antibodies with human tyrosyl protein sulfotransferase 1 (hTPST1) had been demonstrated, thus enabling the O-sulfation of tyrosine residues in the complementary-determining region (CDR) H3 loop. The latter PTM is critical to the neutralization potency of the CAP256-VRC26 lineage of bNAbs.

Glycoengineering of CAP256-VRC26 bNAbs

Glycosylation of the Fc region of Abs can significantly impact antibody effector functions like antibody-dependent cell-mediated cytotoxicity (ADCC) and antibody-dependent, cell-mediated virus inactivation (ADCVI) [18,19,20]. Wild type N. benthamiana glycosylates proteins with glycan species which are very different compared to mammalian production systems; proteins are produced with β1,2-xylose and/or α1,3-fucose containing N-glycans residues in wild-type N. benthamiana [21]. These N-glycan residues influence the pharmacokinetics of a biopharmaceutical product and effector functions. mAbs produced in Lemna minor, engineered to produce mAbs which lack β1,2-xylose and/or α1,3-fucose containing N-glycans residues, had demonstrated enhanced effector activity when compared with their Chinese hamster ovary (CHO)-derived homologs [22]. It is highly desirable to produce mAbs in plants which lack these β1,2-xylose and/or α1,3-fucose containing N-glycans residues [23, 24]. CAP256-VRC26.08 and CAP256-VRC26.09 were produced using a double knockout N. benthamiana mutant (ΔXTFT) which has attenuated expression of xylosyl- and fucosyltransferase via downregulation by ribonucleic interference (RNAi) resulting in the transient production of mAbs with a predominantly mammalian GnGn glycan structure [23, 24].

Incomplete glycosylation of the produced CAP256 bNAbs was observed [17], and this has been previously reported for other transiently plant-produced Abs [14, 15, 23, 25]. Higher glycosylation levels were observed in plant-produced VRC01 [14], which had led to assumption that these innate plant oligosaccharyltransferase (OST) complexes may not recognize the Fc glycosylation sites in different Abs with equivalent efficiency, resulting in the observed variations in glycosylation [13]. Increased in planta N-glycosylation can be achieved through the coexpression of recombinant protein with foreign OST subunits [13]. Glycosylation of the light chains of the CAP256 bNAbs produced in both mammalian and plant cells was observed [17]. The glycosylation of the light chains of Abs is known to shorten the clearance time of the antibody from blood [16]. These glycosylation sites can be removed to allow for increased blood circulation.

In planta tyrosine O-sulfation of CAP256-VRC26 bNAbs

The CAP256-VRC26 antibody lineage targets the first and second variable region (V1V2) of the HIV-1 gp120 envelope glycoprotein. The high potency of this lineage alongside other V1V2 targeting mAbs, such as PG9, is mediated by a protruding O-sulfated tyrosine in the CDR H3 loop of the antigen-binding domain, a characteristic posttranslational modification (PTM) of V1V2 targeting antibodies [26,27,28]. The O-sulfated tyrosine of the CDR H3 loop facilitates tight binding of the gp120 envelope glycoprotein, in a manner which mimics the HIV-1 gp120 affinity for the sulfated chemokine receptor 5 (CCR5) [27]. Association of HIV-1 with the cluster of differentiation 4 (CD4) receptor and critically the CCR5 coreceptor, which has a sulfated tyrosine at the N-terminal end, is essential for HIV-1 gp120 binding and ultimately cell entry [27]. The absence of this modification in V1V2 targeting antibodies leads to a significant decrease in antigen-binding and results in loss of function [16]. The O-sulfation of tyrosine residues is carried out by hTPST1, an enzyme which N. benthamiana lacks. It was previously demonstrated that the transient coexpression of a V1V2 targeting antibody with hTPST1 in plants restored the efficacy of antibody through the proper incorporation of the O-sulfation modification [15]. The same transient coexpression strategy was used to incorporate the O-sulfation PTM to two tyrosine residues, Tyr112 and Tyr113, of which Tyr112 is critical to the efficacy of the CAP256-VRC26 bNAbs. Lower levels of sulfation were achieved in the plant-produced CAP256-VRC26 bNAbs with transient coexpression of hTPST1 when compared to the mammalian produced counterparts; however, despite the difference in sulfation, it was demonstrated that the plant-produced CAP256-VRC26 bNAbs with transient coexpression of hTPST1 have equivalent potency to that of their mammalian-produced counterparts. A similar level of sulfation was observed with PG9, which is indicative of transiently coexpressed hTPST1’s inability to efficiently sulfate tyrosines in the CDR H3 domain as the native machinery of the human embryonic kidney 293 (HEK293) cells [15]. O-sulfation levels may be improved through the further in vitro incubation of the CAP256-VRC26 bNAbs with hTPST1 and substrate, 3′-phosphoadenosine 5′-phosphosulfate (PAPS). However, this approach may not be viable at large scale. In any case, despite the difference in sulfation, equivalent efficacy between the plant-produced bNAbs with hTPST1 coexpression and mammalian-produced CAP256-VRC26 bNAbs counterparts.

Proteolytic bottleneck of plant production of Abs

It was also noted that the plant-produced CAP256-VRC26 bNAbs were prone to proteolytic degradation [17]. A challenge faced with the production of proteins in Nicotiana species, is the proteolytic degradation of these recombinantly produced proteins in planta [29, 30]. The peptidase database, MEROPS, (27/07/2021) lists 515 known or putative peptidases and 98 non-peptidase homologs in N. benthamiana which may be responsible for in planta degradation of some recombinantly produced protein [31]. Proteolytic degradation may not only reduce the purity and yields of recombinantly produced protein but also compromises the structural integrity of these proteins. Proteolytic degradation such as this can result in altered biological activity or no protein production at all, ultimately resulting in a bottleneck in the production of biopharmaceuticals [32,33,34]. The antibodies are targeted through the plant secretory pathway for PTMs making them prone to proteolytic degradation by proteases which are auto-catalytically matured in low-pH environments [35, 36]. Plant-produced CAP256-VRC26 bNAbs were structurally similar to that of the mammalian produced bNAbs, despite the observation of proteolytic degradation fragments in the plant-produced CAP256-VRC26 bNAbs samples under reducing conditions.

The identity of the cleavage site/s are unknown, however, it was noted that under non-reducing conditions, no proteolytic degradation band is observed. Importantly, despite the presence of protease degradation products in the plant-produced Abs, similar neutralization potency was observed for the sulfated plant-produced Abs when compared to the mammalian-produced Abs. This suggests that this cleavage site does not influence antigen-binding in vitro. However, in vivo effects of such degradation are still unknown and will be a topic of our further research. The plant-produced CAP256-VRC26 bNAbs were structurally similar to that of the mammalian produced bNAbs. However, it may be important to improve the quality and/or quantity of produced mAbs in future. To circumvent such proteolytic degradation, commonly used strategies involve the use of RNAi to downregulate protease genes, or either the coexpression of plant protease inhibitors or proton channels to inhibit their enzymatic activity [37,38,39,40].

Conclusion

Despite the success of current antiretroviral therapy, long term usage could introduce multiple drug-resistant escape mutants as a result of the high mutation rate and recombinant frequency of HIV [41]. Continuous development of HIV therapeutics which are both more effective and less toxic is essential [4]. The approach taken in our work [17] and others allows for the incorporation of strategic PTMs in production systems, which may have not been previously considered for the cost-effective production HIV antibody-based biotherapeutics. Gender inequalities remain a major driver of the epidemic with half of all new HIV infections in sub-Saharan Africa are among young people, with girls being two to three times more likely to be infected than boys [1]. Apart from the therapeutic potential of these plant-produced bNAbs, these bNAbs also have the potential to be used in topical and systemic pre-exposure prophylaxis (PrEP). It is thus prudent that work such as this be used in approaches which can enable the empowerment of young girls and women. The plant production system is likely to become increasingly important in enabling the production of antibodies and other proteins with therapeutic potential for mass roll-out in cost-sensitive markets where unequal access to resources, income opportunities, and social power drive high levels of HIV prevalence. The use of the plant-production system requires less capital expense/ investment and > 50% reduction in cost of goods than bioreactor-based processes, making it ideally suitable for LMICs [9]. This work has increased the likelihood that such plant-produced immunotherapeutic production strategies can be considered for adoption to prevent and treat HIV-1 infection in these markets, including sub-Saharan Africa.

Availability of data and materials

Not applicable.

Abbreviations

HIV/AIDS:

Human immunodeficiency virus/acquired immunodeficiency syndrome

HAART:

Highly active antiretroviral therapy

bNAb:

Broadly neutralizing antibody

LMICs:

Low- and middle-income countries

N. benthamiana :

Nicotiana benthamiana

Fc:

Fragment crystallizable

ΔXTFT:

Glycoengineered N. benthamiana

hTPST1:

Human tyrosyl protein sulfotransferase 1

CDR H3:

Complementary-determining region H3

PTM:

Posttranslational modification

ADCC:

Antibody-dependent cell-mediated cytotoxicity

ADCVI:

Antibody-dependent, cell-mediated virus inactivation

CHO:

Chinese hamster ovary

RNAi:

Ribonucleic interference

OST:

Oligosaccharyltransferase

V1V2:

First and second variable region

CCR5:

Chemokine receptor 5

CD4:

Cluster of differentiation 4

Tyr:

Tyrosine

HEK293:

Human embryonic kidney 293

PAPS:

3′-phosphoadenosine 5′-phosphosulfate

References

  1. 1.

    UNAIDS (2020) UNAIDS DATA 2020. United Nations, Geneva

  2. 2.

    Burton DR, Ahmed R, Barouch DH, Butera ST, Crotty S, Godzik A, Kaufmann DE, McElrath JM, Nussenzweig MC, Pulendran B, Scanlan CN, Schief WR, Silvestri G, Streeck H, Walker BD, Walker LM, Ward AB, Wilson IA, Wyatt R (2012) A blueprint for HIV vaccine discovery. Cell Host Microbe 12:396–407. https://doi.org/10.1016/j.chom.2012.09.008.A

    Article  Google Scholar 

  3. 3.

    Mascola JR, Haynes BF (2013) HIV-1 neutralizing antibodies: understanding nature’s pathways. Immunol Rev 254:225–244. https://doi.org/10.1111/imr.12075

    Article  Google Scholar 

  4. 4.

    Salazar G, Zhang N, Fu TM, An Z (2017) Antibody therapies for the prevention and treatment of viral infections. NPJ Vaccines 2:1–12. https://doi.org/10.1038/s41541-017-0019-3

    Article  Google Scholar 

  5. 5.

    Why broadly neutralising antibodies might be the next big thing in HIV | Health24. https://www.news24.com/health24/medical/hiv-aids/why-broadly-neutralising-antibodies-might-be-the-next-big-thing-in-hiv-20210208. Accessed 5 May 2021

  6. 6.

    Tripathi NK, Shrivastava A (2019) Recent Developments in Bioprocessing of Recombinant Proteins: Expression Hosts and Process Development. Front Bioeng Biotechnol 7:1–35. https://doi.org/10.3389/fbioe.2019.00420

    Article  Google Scholar 

  7. 7.

    Frenzel A, Hust M, Schirrmann T (2013) Expression of recombinant antibodies. Front Immunol 4:1–20. https://doi.org/10.3389/fimmu.2013.00217

    Article  Google Scholar 

  8. 8.

    Mackintosh M, Banda G, Tibandebage P, Wamae W (2015) Making medicines in Africa. Palgrave Macmillan

    Book  Google Scholar 

  9. 9.

    Nandi S, Kwong AT, Holtz BR, Erwin RL, Marcel S, McDonald KA (2016) Techno-economic analysis of a transient plant-based platform for monoclonal antibody production. MAbs 8:1456–1466. https://doi.org/10.1080/19420862.2016.1227901

    Article  Google Scholar 

  10. 10.

    Tsekoa TL, Singh AA, Buthelezi SG (2020) Molecular farming for therapies and vaccines in Africa. Curr Opin Biotechnol 61:89–95. https://doi.org/10.1016/j.copbio.2019.11.005

    Article  Google Scholar 

  11. 11.

    Murad S, Fuller S, Menary J, Moore C, Pinneh E, Szeto T, Hitzeroth I, Freire M, Taychakhoonavudh S, Phoolcharoen W, Ma JKC (2020) Molecular Pharming for low and middle income countries. Curr Opin Biotechnol 61:53–59. https://doi.org/10.1016/j.copbio.2019.10.005

    Article  Google Scholar 

  12. 12.

    Schähs M, Strasser R, Stadlmann J, Kunert R, Rademacher T, Steinkellner H (2007) Production of a monoclonal antibody in plants with a humanized N-glycosylation pattern. Plant Biotechnol J 5:657–663. https://doi.org/10.1111/j.1467-7652.2007.00273.x

    Article  Google Scholar 

  13. 13.

    Castilho A, Beihammer G, Pfeiffer C, Göritzer K, Montero-Morales L, Vavra U, Maresch D, Grünwald-Gruber C, Altmann F, Steinkellner H, Strasser R (2018) An oligosaccharyltransferase from Leishmania major increases the N-glycan occupancy on recombinant glycoproteins produced in Nicotiana benthamiana. Plant Biotechnol J 16:1700–1709. https://doi.org/10.1111/pbi.12906

    Article  Google Scholar 

  14. 14.

    Teh AYH, Maresch D, Klein K, Ma JKC (2014) Characterization of VRC01, a potent and broadly neutralizing anti-HIV mAb, produced in transiently and stably transformed tobacco. Plant Biotechnol J 12:300–311. https://doi.org/10.1111/pbi.12137

    Article  Google Scholar 

  15. 15.

    Loos A, Gach JS, Hackl T, Maresch D, Henkel T, Porodko A, Bui-Minh D, Sommeregger W, Wozniak-Knopp G, Forthal DN, Altmann F, Steinkellner H, Mach L (2015) Glycan modulation and sulfoengineering of anti–HIV-1 monoclonal antibody PG9 in plants. Proc Natl Acad Sci 112:12675–12680. https://doi.org/10.1073/pnas.1509090112

    Article  Google Scholar 

  16. 16.

    Rosenberg Y, Sack M, Montefiori D, Labranche C, Lewis M, Urban L, Mao L, Fischer R, Jiang X (2015) Pharmacokinetics and immunogenicity of broadly neutralizing HIV monoclonal antibodies in macaques. PLoS One 10:1–15. https://doi.org/10.1371/journal.pone.0120451

    Article  Google Scholar 

  17. 17.

    Singh AA, Pooe O, Kwezi L, Lotter-Stark T, Stoychev SH, Alexandra K, Gerber I, Bhiman JN, Vorster J, Pauly M, Zeitlin L, Whaley K, Mach L, Steinkellner H, Morris L, Tsekoa TL, Chikwamba R (2020) Plant-based production of highly potent anti-HIV antibodies with engineered posttranslational modifications. Sci Rep 10:1–9. https://doi.org/10.1038/s41598-020-63052-1

    Article  Google Scholar 

  18. 18.

    Lewis GK (2013) Qualitative and quantitative variables that affect the potency of Fc- mediated effector function in vitro and in vivo: considerations for passive immunization using non-neutralizing antibodies. Curr HIV Res 11:354–364. https://doi.org/10.2174/1570162x113116660060

    Article  Google Scholar 

  19. 19.

    Baum LL, Cassutt KJ, Knigge K, Khattri R, Margolick J, Rinaldo C, Kleeberger CA, Nishanian P, Henrard DR, Phair J (1996) HIV-1 gp120-specific antibody-dependent cell-mediated cytotoxicity correlates with rate of disease progression. J Immunol 157:2168–2173

    Google Scholar 

  20. 20.

    Shields RL, Lai J, Keck R, O’Connell LY, Hong K, Meng YG, Weikert SHA, Presta LG (2002) Lack of fucose on human IgG1 N-linked oligosaccharide improves binding to human FcγRIII and antibody-dependent cellular toxicity. J Biol Chem 277:26733–26740. https://doi.org/10.1074/jbc.M202069200

    Article  Google Scholar 

  21. 21.

    Whaley KJ, Hiatt A, Zeitlin L (2011) Emerging antibody products and Nicotiana manufacturing. Hum Vaccin 7:349–356. https://doi.org/10.4161/hv.7.3.14266

    Article  Google Scholar 

  22. 22.

    Cox KM, Sterling JD, Regan JT, Gasdaska JR, Frantz KK, Peele CG, Black A, Passmore D, Moldovan-Loomis C, Srinivasan M, Cuison S, Cardarelli PM, Dickey LF (2006) Glycan optimization of a human monoclonal antibody in the aquatic plant Lemna minor. Nat Biotechnol 24:1591–1597. https://doi.org/10.1038/nbt1260

    Article  Google Scholar 

  23. 23.

    Strasser R, Stadlmann J, Schähs M, Stiegler G, Quendler H, Mach L, Glössl J, Weterings K, Pabst M, Steinkellner H (2008) Generation of glyco-engineered Nicotiana benthamiana for the production of monoclonal antibodies with a homogeneous human-like N-glycan structure. Plant Biotechnol J 6:392–402. https://doi.org/10.1111/j.1467-7652.2008.00330.x

    Article  Google Scholar 

  24. 24.

    Strasser R, Castilho A, Stadlmann J, Kunert R, Quendler H, Gattinger P, Jez J, Rademacher T, Altmann F, Mach L, Steinkellner H (2009) Improved virus neutralization by plant-produced anti-HIV antibodies with a homogeneous 1,4-galactosylated N-glycan profile. J Biol Chem 284:20479–20485. https://doi.org/10.1074/jbc.M109.014126

    Article  Google Scholar 

  25. 25.

    Bendandi M, Marillonnet S, Kandzia R, Thieme F, Nickstadt A, Herz S, Fröde R, Inogés S, Lòpez-Dìaz de Cerio A, Soria E, Villanueva H, Vancanneyt G, McCormick A, Tusé D, Lenz J, Butler-Ransohoff J-E, Klimyuk V, Gleba Y (2010) Rapid, high-yield production in plants of individualized idiotype vaccines for non-Hodgkin’s lymphoma. Ann Oncol 21:2420–2427. https://doi.org/10.1093/annonc/mdq256

    Article  Google Scholar 

  26. 26.

    Doria-Rose NA, Schramm CA, Gorman J, Moore PL, Bhiman JN, DeKosky BJ, Ernandes MJ, Georgiev IS, Kim HJ, Pancera M, Staupe RP, Altae-Tran HR, Bailer RT, Crooks ET, Cupo A, Druz A, Garrett NJ, Hoi KH, Kong R, Louder MK, Longo NS, McKee K, Nonyane M, O’Dell S, Roark RS, Rudicell RS, Schmidt SD, Sheward DJ, Soto C, Wibmer CK, Yang Y, Zhang Z, NISC comparative sequencing, Mullikin JC, Binley JM, Sanders RW, Wilson IA, Moore JP, Ward AB, Georgiou G, Williamson C, Abdool Karim SS, Morris L, Kwong PD, Shapiro L, Mascola JR (2014) Developmental pathway for potent V1V2-directed HIV-neutralizing antibodies. Nature 509:55–62. https://doi.org/10.1038/nature13036

    Article  Google Scholar 

  27. 27.

    Choe H, Li W, Wright PL, Vasilieva N, Venturi M, Huang C-C, Grundner C, Dorfman T, Zwick MB, Wang L, Rosenberg ES, Kwong PD, Burton DR, Robinson JE, Sodroski JG, Farzan M (2003) Tyrosine sulfation of human antibodies contributes to recognition of the CCR5 binding region of HIV-1 gp120. Cell 114:161–170. https://doi.org/10.1016/s0092-8674(03)00508-7

    Article  Google Scholar 

  28. 28.

    Lin H, Du J, Jiang H (2008) Modifications to regulate protein function

    Google Scholar 

  29. 29.

    Doran PM (2006) Foreign protein degradation and instability in plants and plant tissue cultures. Trends Biotechnol 24:426–432. https://doi.org/10.1016/j.tibtech.2006.06.012

    Article  Google Scholar 

  30. 30.

    Benchabane M, Goulet C, Rivard D, Faye L, Gomord V, Michaud D (2008) Preventing unintended proteolysis in plant protein biofactories. Plant Biotechnol J 6:633–648. https://doi.org/10.1111/j.1467-7652.2008.00344.x

    Article  Google Scholar 

  31. 31.

    Rawlings ND, Barrett AJ, Thomas PD, Huang X, Bateman A, Finn RD (2018) The MEROPS database of proteolytic enzymes, their substrates and inhibitors in 2017 and a comparison with peptidases in the PANTHER database. Nucleic Acids Res 46:D624–D632. https://doi.org/10.1093/nar/gkx1134

    Article  Google Scholar 

  32. 32.

    Mandal MK, Ahvari H, Schillberg S, Schiermeyer A (2016) Tackling unwanted proteolysis in plant production hosts used for molecular farming. Front Plant Sci 7:1–6. https://doi.org/10.3389/fpls.2016.00267

    Article  Google Scholar 

  33. 33.

    Castilho A, Windwarder M, Gattinger P, Mach L, Strasser R, Altmann F, Steinkellner H (2014) Proteolytic and N-glycan processing of human α1-antitrypsin expressed in Nicotiana benthamiana. Plant Physiol 166:1839–1851. https://doi.org/10.1104/pp.114.250720

    Article  Google Scholar 

  34. 34.

    Faye L, Boulaflous A, Benchabane M, Gomord V, Michaud D (2005) Protein modifications in the plant secretory pathway: Current status and practical implications in molecular pharming. Vaccine 23:1770–1778. https://doi.org/10.1016/j.vaccine.2004.11.003

    Article  Google Scholar 

  35. 35.

    Zauner FB, Dall E, Regl C, Grassi L, Huber CG, Cabrele C, Brandstetter H (2018) Crystal structure of plant legumain reveals a unique two-chain state with pH-dependent activity regulation. Plant Cell 30:686–699. https://doi.org/10.1105/tpc.17.00963

    Article  Google Scholar 

  36. 36.

    Gu C, Shabab M, Strasser R, Wolters PJ, Shindo T, Niemer M, Kaschani F, Mach L, van der Hoorn RAL (2012) Post-translational regulation and trafficking of the granulin-containing protease rd21 of arabidopsis thaliana. PLoS One 7:1–11. https://doi.org/10.1371/journal.pone.0032422

    Article  Google Scholar 

  37. 37.

    Mandal MK, Fischer R, Schillberg S, Schiermeyer A (2014) Inhibition of protease activity by antisense RNA improves recombinant protein production in Nicotiana tabacum cv. Bright Yellow 2 (BY-2) suspension cells. Biotechnol J 9:1065–1073. https://doi.org/10.1002/biot.201300424

    Article  Google Scholar 

  38. 38.

    Duwadi K, Chen L, Menassa R, Dhaubhadel S (2015) Identification, characterization and down-regulation of cysteine protease genes in tobacco for use in recombinant protein production. PLoS One 10:1–19. https://doi.org/10.1371/journal.pone.0130556

    Article  Google Scholar 

  39. 39.

    Pillay P, Kibido T, Du Plessis M, Van Der Vyver C, Beyene G, Vorster BJ, Kunert KJ, Schlüter U (2012) Use of transgenic oryzacystatin-i-expressing plants enhances recombinant protein production. Appl Biochem Biotechnol 168:1608–1620. https://doi.org/10.1007/s12010-012-9882-6

    Article  Google Scholar 

  40. 40.

    Girard C, Rivard D, Kiggundu A, Kunert K, Gleddie SC, Cloutier C, Michaud D (2007) A multicomponent, elicitor-inducible cystatin complex in tomato, Solanum lycopersicum. New Phytol 173:841–851. https://doi.org/10.1111/j.1469-8137.2007.01968.x

    Article  Google Scholar 

  41. 41.

    Iyidogan P, Anderson KS (2014) Current perspectives on HIV-1 antiretroviral drug resistance. Viruses 6:4095–4139. https://doi.org/10.3390/v6104095

    Article  Google Scholar 

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This work was funded by the Council for Scientific and Industrial Research: Young Researcher Establishment Fund

(Grant No.: YREF 2022 13).

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AAS wrote the manuscript with contribution from PP. PP, LK and TLT revised the manuscript. All authors have read and approved the submission of this manuscript.

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Singh, A.A., Pillay, P., Kwezi, L. et al. A plant-biotechnology approach for producing highly potent anti-HIV antibodies for antiretroviral therapy consideration. J Genet Eng Biotechnol 19, 180 (2021). https://doi.org/10.1186/s43141-021-00279-z

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