Functional native disulfide bridging enables delivery of a potent, stable and targeted antibody–drug conjugate (ADC)

Nunes, João P. M.; Morais, Maurício; Vassileva, Vessela; Robinson, Eric; Rajkumar, Vineeth; Smith, Mark E.; Pedley, R. Barbara; Caddick, Stephen; Baker, James R.; Chudasama, Vijay

doi:10.1039/c5cc03557k

Cited by 104 publications

(113 citation statements)

References 24 publications

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“…Several groups including Baker, Caddick, Chudasama, Jackson and Haddleton have elaborated on the inherent reactivity of maleimides toward thiols to design next‐generation maleimides for disulfide rebridging (Figure , II, III; Scheme A) . Baker and Caddick proposed that the fast reaction kinetics of the maleimides compared to the bis‐sulfone reagents will facilitate reaction in tandem with the reducing agent resulting in less protein loss due to aggregation or disulfide scrambling .…”

Section: Disulfide Rebridging Agents: Syntheses Features and Biocmentioning

confidence: 99%

“…Baker and Caddick proposed that the fast reaction kinetics of the maleimides compared to the bis‐sulfone reagents will facilitate reaction in tandem with the reducing agent resulting in less protein loss due to aggregation or disulfide scrambling . The earliest examples reported are disubstituted maleimides (DSM), which carry substituents in the 3‐ and 4‐position with good leaving groups that allow reaction with two nucleophilic thiol groups such as the two thiols of a reduced cystine to rebridge the disulfides through a rigid two‐carbon spacer to afford the bisthiomaleimide bioconjugates (Scheme B) …”

Section: Disulfide Rebridging Agents: Syntheses Features and Biocmentioning

confidence: 99%

“…A fluorescent tag was also incorporated site‐selectively into lysozyme, an enzyme which hydrolyses the β (l–4) glycosidic bonds, and the mono‐modified enzyme retains 89 % activity . On the other hand, DSMs and dibromopyridazinediones have been employed extensively in the modification of peptides and antibodies to introduce bioorthogonal handles, macroinitiators, imaging probes, spin labels and drugs …”

Section: Disulfide Rebridging Agents: Syntheses Features and Biocmentioning

confidence: 99%

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Site‐Selective Disulfide Modification of Proteins: Expanding Diversity beyond the Proteome

Kuan

Wang

Weil

2016

Chemistry A European J

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The synthetic transformation of polypeptides with molecular accuracy holds great promise for providing functional and structural diversity beyond the proteome. Consequently, the last decade has seen an exponential growth of site‐directed chemistry to install additional features into peptides and proteins even inside living cells. The disulfide rebridging strategy has emerged as a powerful tool for site‐selective modifications since most proteins contain disulfide bonds. In this Review, we present the chemical design, advantages and limitations of the disulfide rebridging reagents, while summarizing their relevance for synthetic customization of functional protein bioconjugates, as well as the resultant impact and advancement for biomedical applications.

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Section: Disulfide Rebridging Agents: Syntheses Features and Biocmentioning

confidence: 99%

Section: Disulfide Rebridging Agents: Syntheses Features and Biocmentioning

confidence: 99%

Section: Disulfide Rebridging Agents: Syntheses Features and Biocmentioning

confidence: 99%

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Site‐Selective Disulfide Modification of Proteins: Expanding Diversity beyond the Proteome

Kuan

Wang

Weil

2016

Chemistry A European J

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“…In addition, the structure of the antibody remains intact and the ligand-to-antibody ratio can be tightly controlled [48,49]. Researchers in the labs of Chudasama, Baker and Caddick describe two different bridging approaches whereby they developed both a novel class of maleimides, designated as next generation maleimides, and successfully generated ADCs with all the aforementioned advantages [50,51] in addition to using pyridazinedione derivatives to generate dual-modified ADCs [52]. This technology can be used to site-specifically attach two different functionalities onto an antibody while benefiting from the aforementioned advantages of bridging approaches.…”

Section: Native Cysteinesmentioning

confidence: 99%

Antibody Conjugates: From Heterogeneous Populations to Defined Reagents

2015

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Monoclonal antibodies (mAbs) and their derivatives are currently the fastest growing class of therapeutics. Even if naked antibodies have proven their value as successful biopharmaceuticals, they suffer from some limitations. To overcome suboptimal therapeutic efficacy, immunoglobulins are conjugated with toxic payloads to form antibody drug conjugates (ADCs) and with chelating systems bearing therapeutic radioisotopes to form radioimmunoconjugates (RICs). Besides their therapeutic applications, antibody conjugates are also extensively used for many in vitro assays. A broad variety of methods to functionalize antibodies with various payloads are currently available. The decision as to which conjugation method to use strongly depends on the final purpose of the antibody conjugate. Classical conjugation via amino acid residues is still the most common method to produce antibody conjugates and is suitable for most in vitro applications. In recent years, however, it has become evident that antibody conjugates, which are generated via site-specific conjugation techniques, possess distinct advantages with regard to in vivo properties. Here, we give a comprehensive overview on existing and emerging strategies for the production of covalent and non-covalent antibody conjugates.

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“…Chemoenzymatic methods have been used for ADC synthesis such as glycan modication, [15][16][17] glutamine amidation 18 and cysteine oxidation to aldehyde tags. 19 Site-selective disulde modica-tion with small molecules has been achieved using nextgeneration maleimides (NGMs), [20][21][22][23][24][25][26][27] pyridazinediones (PDs) [28][29][30][31][32] and bis-sulfones. 33,34 Site-specic reactivity has also been accomplished through incorporation of non-natural aminoacids into proteins for bioorthogonal conjugation.…”

Section: 914mentioning

confidence: 99%

Use of a next generation maleimide in combination with THIOMAB™ antibody technology delivers a highly stable, potent and near homogeneous THIOMAB™ antibody-drug conjugate (TDC)

et al. 2017

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Herein we demonstrate that conjugation of a next generation maleimide (NGM) to engineered cysteines in a THIOMAB™ antibody delivers a THIOMAB™ antibody-drug conjugate (TDC) with a drug loading of ca. 2. This TDC is highly stable in blood serum conditions, selective and potent towards HER2 expressing cell lines and meets the current criteria for optimised antibody-drug conjugates (ADCs).Antibody-drug conjugates (ADCs) have emerged as targeted therapeutics against cancer by combining the selectivity of the antibody component with the potency of cytotoxic drugs. There are two FDA-approved ADCs already on the market (Adcetris™ and Kadcyla™) and over 40 additional ADCs currently undergoing clinical trials. 1-4Classical methods of conjugation for producing ADCs include lysine modication 5 and cysteine modication via reduction of solvent accessible disulde bonds followed by reaction with maleimides.6 It has been recognised that both methods generate heterogeneous mixtures of ADCs with different drug loadings and varied sites of attachment, resulting in sub-optimal therapeutic indices and higher clearance rates.7-13 Formation of higher drug-to-antibody ratio species (DAR > 4) have narrower therapeutic indices. 1,7,9,12,14 The latter method also results in the loss of structural disulde bonds which can lead to poor stability. 7,9,14The next generation of conjugation methods has aimed to reduce heterogeneity by focusing on site-selectivity. Chemoenzymatic methods have been used for ADC synthesis such as glycan modication, 15-17 glutamine amidation 18 and cysteine oxidation to aldehyde tags.19 Site-selective disulde modica-tion with small molecules has been achieved using nextgeneration maleimides (NGMs), 20-27 pyridazinediones (PDs)28-32 and bis-sulfones. 33,34 Site-specic reactivity has also been accomplished through incorporation of non-natural aminoacids into proteins for bioorthogonal conjugation. 10,35-39The THIOMAB™ antibody technology platform uses sitedirected mutagenesis to incorporate cysteines into the antibody. Conjugation to a classical maleimide bearing the drug of choice affords a THIOMAB™ antibody-drug conjugate (TDC) with full control over site reactivity and a precise drug-toantibody ratio (DAR).12,40-42 The technology also avoids issues with some chemoenzymatic methods that can result in over oxidation of aminoacids 43 and aldehyde tag conversion to unreactive gem-diols.44 TDCs have been demonstrated to improve therapeutic indices and pharmacodynamics over ADCs prepared by classical cysteine and lysine modication. 11,12,42Maleimides are rapid thiol-selective conjugation reagents.45,46 However, the succinimide conjugates thus formed can undergo retro-Michael reactions and scavenging with albumin, 47 resulting in premature drug release during blood circulation.10,41,48 N-Alkyl succinimide conjugates can be stabilised against retro-Michael reactions by hydrolysis to succinamic acid derivatives, 49-51 which resulted in ADCs with improved pharmacokinetics and in vivo efficacy. 41,51,52 Most Nalkyl t...

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Functional native disulfide bridging enables delivery of a potent, stable and targeted antibody–drug conjugate (ADC)

Cited by 104 publications

References 24 publications

Site‐Selective Disulfide Modification of Proteins: Expanding Diversity beyond the Proteome

Site‐Selective Disulfide Modification of Proteins: Expanding Diversity beyond the Proteome

Antibody Conjugates: From Heterogeneous Populations to Defined Reagents

Use of a next generation maleimide in combination with THIOMAB™ antibody technology delivers a highly stable, potent and near homogeneous THIOMAB™ antibody-drug conjugate (TDC)

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