A stable IgG-like bispecific antibody targeting the epidermal growth factor receptor and the type I insulin-like growth factor receptor demonstrates superior anti-tumor activity

Dong, Jianying; Sereno, Arlene; Aivazian, Dikran; Langley, Emma; Miller, Brian; Snyder, William B.; Chan, Eric Y.; Cantele, Matt; Morena, Ronald; Joseph, Ingrid B.J.; Boccia, Antonio; Virata, Cyrus; Gámez, J.A. Muñoz; Yco, Grace; Favis, Michael; Wu, Xiufeng; Graff, Candace; Wang, Qin; Rohde, Ellen; Rennard, Rachel; Berquist, Lisa; Huang, Flora; Zhang, Ying; Gao, Sharon X.; Ho, Steffan N.; Demarest, Stephen J.; Reff, Mitchell E.; Hariharan, Kandasamy; Glaser, Scott

doi:10.4161/mabs.3.3.15188

Cited by 76 publications

(65 citation statements)

References 53 publications

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“…Another pair tested, M60-A02/C06, did not exhibit simultaneous binding, although a M60-A02/C06 bispecific was previously reported to bind both antigens simultaneously in a tetravalent bispecific format, with two C06 scFvs on the C-term of the Fc. 20 The lack of binding observed here is likely a result of steric hindrance caused by the asymmetric bispecific format we used to study EFab domain substitution.…”

Section: Discussionmentioning

confidence: 91%

“…20 The test molecules were tagged with green fluorescent protein (GFP) (30 kDa, light chain of C06) or human serum albumin (HSA) (66 kDa, heavy chain of Fab 1 or EFab, M60-A02) to enable simple differentiation of correct versus incorrect Fab pairs by migration on non-reducing SDS-PAGE (Figure 2, for reducing SDS-PAGE, see Figure S1). The proteins were generated by transient expression in Chinese hamster ovary (CHO) cells.…”

Section: Resultsmentioning

confidence: 99%

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EFab domain substitution as a solution to the light-chain pairing problem of bispecific antibodies

Cooke

Arndt

Quan

et al. 2018

mAbs

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Bispecific antibody therapeutics can expand the functionality of a conventional monoclonal antibody drug because they can bind multiple antigens. However, their great potential is counterbalanced by the challenges faced in their production. The classic asymmetric bispecific containing an Fc requires the expression of four unique chains – two light chains and two heavy chains; each light chain must pair with its correct heavy chain, which then must heterodimerize to form the full bispecific. The light-chain pairing problem has several solutions, some of which require engineering and optimization for each bispecific pair. Here, we introduce a technology called EFab Domain Substitution, which replaces the Cϵ2 of IgE for one of the CL/CH1 domains into one arm of an asymmetric bispecific to encourage the correct pairing of the light chains. EFab Domain Substitution provides very robust correct pairing while maintaining antibody function and is effective for many variable domains. We report its effect on the biophysical properties of an antibody and the crystal structure of the EFab domain substituted into the adalimumab Fab (PDB ID 6CR1).

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Section: Discussionmentioning

confidence: 91%

Section: Resultsmentioning

confidence: 99%

EFab domain substitution as a solution to the light-chain pairing problem of bispecific antibodies

Cooke

Arndt

Quan

et al. 2018

mAbs

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“…EGFR and IGF1R are cell surface receptor tyrosine kinases known to cooperate to promote tumor growth and progression. 36 Dual inhibitions of EGFR and IGF1R have been shown to improve anti-tumor activity and to overcome resistance to therapy against a single receptor inhibition in preclinical models. 36 In iMab-EI, the anti-EGFR light chain is connected to its heavy chain by a 104-residue peptide linker (linker L1 in Fig.…”

Section: Design Of An Imab Targeting Egfr and Igf1r (Imab-ei)mentioning

confidence: 99%

“…[26][27][28] Although these approaches generate IgG-Bs, the deviation from the IgG chain sequence and structure remains a substantial issue, [6][7][8][9][10][11][12] and novel antibody engineering solutions are thus still needed.…”

Section: Introductionmentioning

confidence: 99%

“…[3][4][5] Some of these BsAbs have inferior pharmacokinetic properties, including rapid clearance and shorter half-life relative to their IgG counterparts, thus limiting their therapeutic applications. [6][7][8][9][10][11][12] Several solutions have been described for the generation of BsAb with an IgG chain-like sequence and structure (IgG-Bs). 13 For example, IgG-Bs have been produced by expressing the IgGs separately and subsequently mixing them, 14,15 but these methods require both site-specific mutations and reduction and oxidation, which complicate downstream development.…”

Section: Introductionmentioning

confidence: 99%

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Guiding bispecific monovalent antibody formation through proteolysis of IgG1 single-chain

Dimasi¹,

Fleming²,

Sachsenmeier

et al. 2017

mAbs

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We developed an IgG1 domain-tethering approach to guide the correct assembly of 2 light and 2 heavy chains, derived from 2 different antibodies, to form bispecific monovalent antibodies in IgG1 format. We show here that assembling 2 different light and heavy chains by sequentially connecting them with protease-cleavable polypeptide linkers results in the generation of monovalent bispecific antibodies that have IgG1 sequence, structure and functional properties. This approach was used to generate a bispecific monovalent antibody targeting the epidermal growth factor receptor and the type I insulin-like growth factor receptor that: 1) can be produced and purified using standard IgG1 techniques; 2) exhibits stability and structural features comparable to IgG1; 3) binds both targets simultaneously; and 4) has potent anti-tumor activity. Our strategy provides new engineering opportunities for bispecific antibody applications, and, most importantly, overcomes some of the limitations (e.g., half-antibody and homodimer formation, light chains mispairing, multi-step purification), inherent with some of the previously described IgG1-based bispecific monovalent antibodies.

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Bispecifics

McCluskey

Ghayur

2017

Methods and Principles in Medicinal Chemistry

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A stable IgG-like bispecific antibody targeting the epidermal growth factor receptor and the type I insulin-like growth factor receptor demonstrates superior anti-tumor activity

Abstract: (2011) A stable IgG-like bispecific antibody targeting the epidermal growth factor receptor and the type I insulin-like growth factor receptor demonstrates superior anti-tumor activity, mAbs, 3:3, 273-288,

Cited by 76 publications

References 53 publications

EFab domain substitution as a solution to the light-chain pairing problem of bispecific antibodies

EFab domain substitution as a solution to the light-chain pairing problem of bispecific antibodies

Guiding bispecific monovalent antibody formation through proteolysis of IgG1 single-chain

Bispecifics

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