Journal Article•
Novel peptide linkers for highly potent antibody-auristatin conjugates
TL;DR: In this article, the authors describe several other dipeptide sequences, many of which are not cathepsin B substrates, for the attachment of auristatins to mAb carriers.
Abstract: 4907 A great deal of interest has surrounded the use of monoclonal antibodies (mAbs) for the selective delivery of cytotoxic agents to tumor cells. We have previously demonstrated that mAb-auristatin conjugates are highly active, leading to cures and regressions of established tumors in nude mice. These antibody-drug conjugates (ADCs) contain a dipeptide valine-citrulline-p-aminobenzylcarbamate (vc-PABC) linker used to join the antimitotic drugs monomethylauristatin E or F (MMAE, MMAF) to a series of antitumor mAbs. This linker was selected for plasma stability and cleavage by lysosomal enzymes such as cathepsin B. Here, we describe several other dipeptide sequences, many of which are not cathepsin B substrates, for the attachment of auristatins to mAb carriers. More than 20 mAb-dipeptide-PABC-MMAF ADCs were prepared, all of which were active in vitro and in vivo independent of peptide sequence. There was no apparent improvement over the vc-PABC linker, since the PABC group facilitated lysosomal drug release. In contrast, a new series of mAb-MMAF conjugates was prepared in which the dipeptide linkers were directly attached to the C-terminal phenylalanine on the drug without using an intervening PABC spacer. Many of the dipeptide sequences contained unnatural amino acids in order to increase the stringency of drug release. Peptide sequences were identified within the new conjugates that led to both greater tolerability and higher potency compared to corresponding vc-PABC ADCs. The new linkers play an important role in ADC activity and tolerability, and mAb-auristatin conjugates derived from them are candidates for future development.
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TL;DR: Antibody-drug conjugates (ADCs) combine the specificity of monoclonal antibodies with the potency of cytotoxic molecules, thereby taking advantage of the best characteristics of both components.
612 citations
TL;DR: Genetically encoded unnatural amino acids with orthogonal chemical reactivity are used to synthesize homogeneous ADCs with precise control of conjugation site and stoichiometry to facilitate the optimization of ADCs for a host of therapeutic uses.
Abstract: Antibody-drug conjugates (ADCs) allow selective targeting of cytotoxic drugs to cancer cells presenting tumor-associated surface markers, thereby minimizing systemic toxicity. Traditionally, the drug is conjugated nonselectively to cysteine or lysine residues in the antibody. However, these strategies often lead to heterogeneous products, which make optimization of the biological, physical, and pharmacological properties of an ADC challenging. Here we demonstrate the use of genetically encoded unnatural amino acids with orthogonal chemical reactivity to synthesize homogeneous ADCs with precise control of conjugation site and stoichiometry. p-Acetylphenylalanine was site-specifically incorporated into an anti-Her2 antibody Fab fragment and full-length IgG in Escherichia coli and mammalian cells, respectively. The mutant protein was selectively and efficiently conjugated to an auristatin derivative through a stable oxime linkage. The resulting conjugates demonstrated excellent pharmacokinetics, potent in vitro cytotoxic activity against Her2+ cancer cells, and complete tumor regression in rodent xenograft treatment models. The synthesis and characterization of homogeneous ADCs with medicinal chemistry-like control over macromolecular structure should facilitate the optimization of ADCs for a host of therapeutic uses.
538 citations
TL;DR: This review summarizes the current knowledge and developments in the field of antibody drug conjugates.
Abstract: Over the past couple of decades, antibody–drug conjugates (ADCs) have revolutionized the field of cancer chemotherapy. Unlike conventional treatments that damage healthy tissues upon dose escalation, ADCs utilize monoclonal antibodies (mAbs) to specifically bind tumour-associated target antigens and deliver a highly potent cytotoxic agent. The synergistic combination of mAbs conjugated to small-molecule chemotherapeutics, via a stable linker, has given rise to an extremely efficacious class of anti-cancer drugs with an already large and rapidly growing clinical pipeline. The primary objective of this paper is to review current knowledge and latest developments in the field of ADCs. Upon intravenous administration, ADCs bind to their target antigens and are internalized through receptor-mediated endocytosis. This facilitates the subsequent release of the cytotoxin, which eventually leads to apoptotic cell death of the cancer cell. The three components of ADCs (mAb, linker and cytotoxin) affect the efficacy and toxicity of the conjugate. Optimizing each one, while enhancing the functionality of the ADC as a whole, has been one of the major considerations of ADC design and development. In addition to these, the choice of clinically relevant targets and the position and number of linkages have also been the key determinants of ADC efficacy. The only marketed ADCs, brentuximab vedotin and trastuzumab emtansine (T-DM1), have demonstrated their use against both haematological and solid malignancies respectively. The success of future ADCs relies on improving target selection, increasing cytotoxin potency, developing innovative linkers and overcoming drug resistance. As more research is conducted to tackle these issues, ADCs are likely to become part of the future of targeted cancer therapeutics.
316 citations
TL;DR: This review will focus on various targeted prodrug designs that have been developed to increase the selectivity of chemotherapy drugs.
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232 citations
References
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TL;DR: CAC10-vcMMAE was highly potent and selective against CD30+ tumor lines but was more than 300-fold less active on antigen-negative cells in SCID mouse xenograft models of anaplastic large cell lymphoma or Hodgkin disease, and was efficacious at doses as low as 1 mg/kg.
894 citations
TL;DR: Antibody-drug conjugates (ADCs) combine the specificity of monoclonal antibodies with the potency of cytotoxic molecules, thereby taking advantage of the best characteristics of both components.
612 citations
TL;DR: Genetically encoded unnatural amino acids with orthogonal chemical reactivity are used to synthesize homogeneous ADCs with precise control of conjugation site and stoichiometry to facilitate the optimization of ADCs for a host of therapeutic uses.
Abstract: Antibody-drug conjugates (ADCs) allow selective targeting of cytotoxic drugs to cancer cells presenting tumor-associated surface markers, thereby minimizing systemic toxicity. Traditionally, the drug is conjugated nonselectively to cysteine or lysine residues in the antibody. However, these strategies often lead to heterogeneous products, which make optimization of the biological, physical, and pharmacological properties of an ADC challenging. Here we demonstrate the use of genetically encoded unnatural amino acids with orthogonal chemical reactivity to synthesize homogeneous ADCs with precise control of conjugation site and stoichiometry. p-Acetylphenylalanine was site-specifically incorporated into an anti-Her2 antibody Fab fragment and full-length IgG in Escherichia coli and mammalian cells, respectively. The mutant protein was selectively and efficiently conjugated to an auristatin derivative through a stable oxime linkage. The resulting conjugates demonstrated excellent pharmacokinetics, potent in vitro cytotoxic activity against Her2+ cancer cells, and complete tumor regression in rodent xenograft treatment models. The synthesis and characterization of homogeneous ADCs with medicinal chemistry-like control over macromolecular structure should facilitate the optimization of ADCs for a host of therapeutic uses.
538 citations
TL;DR: Several new linkers were generated in which various components within the L1 linker were either altered or deleted to optimize the ADC, and one of the most promising linkers contained a noncleavable maleimidocaproyl (L4) spacer between the drug and the mAb.
527 citations
TL;DR: The data indicate that new linkers can be obtained with improved in vivo stability by replacing the maleimide with an acetamide, but the resulting ADCs had similar tolerability and activity profiles.
445 citations