Hydrazino-Pictet-Spengler Ligation as a Biocompatible Method for the Generation of Stable Protein Conjugates

Agarwal, Paresh; Kudirka, Romas; Albers, Aaron E.; Barfield, Robyn M.; Hart, Gregory W. de; Drake, Penelope M.; Jones, Lesley C.; Rabuka, David

doi:10.1021/bc400042a

Cited by 149 publications

(155 citation statements)

References 56 publications

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“…The rate of oxime formation is known to be dependent on acidity of the reaction medium. A solution pH 25,26 Aniline and its derivatives are frequently used to accelerate the reaction. 26,27 The ligation between 2 and 14a was attempted at pH 4.6 with aniline, but we did not observe the formation of peptide oxime 15a (Figure 3a).…”

Section: Acs Chemical Biologymentioning

confidence: 99%

Critical Evaluation and Rate Constants of Chemoselective Ligation Reactions for Stoichiometric Conjugations in Water

2015

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Chemoselective ligation reactions have contributed immensely to the development of organic synthesis and chemical biology. However, the ligation of stoichiometric amounts of large molecules for applications such as protein-protein conjugates is still challenging. Conjugation reactions need to be fast enough to proceed under dilute conditions and chemoselective in the presence of unprotected functional groups; the starting materials and products must be stable under the reaction conditions. To compare known ligation reactions for their suitability under these conditions, we determined the second-order rate constants of ligation reactions using peptide substrates with unprotected functional groups. The reaction conditions, the chemoselectivity of the reactions, and the stability of the starting materials and products were carefully evaluated. In some cases, the stability could be improved by modifying the substrate structure. These data obtained under the ligation conditions provide a useful guide to choose an appropriate ligation reaction for synthesis of large molecules by covalent ligation reactions of unprotected substrates in water.

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Section: Acs Chemical Biologymentioning

confidence: 99%

Critical Evaluation and Rate Constants of Chemoselective Ligation Reactions for Stoichiometric Conjugations in Water

2015

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“…Redwood Biosciences disclosed a new reaction, the hydrazino-Pictet-Spengler (HIPS) ligation to produce ADCs with aldehyde-tagged proteins ( Table 2, 5iii). The HIPS ligation is a quick reaction carried out at neutral pH to produce stable conjugates similar to the oxime-linked conjugate [69]. Trastuzumab with an aldehyde tag at three different sites was conjugated with five different linker HIPS-maytansinoids to produce a panel of ADCs to explore the impact of small changes in linker design on ADC potency and stability [70].…”

Section: Enzyme Modification Conjugationsmentioning

confidence: 99%

Emerging formats for next-generation antibody drug conjugates

Deonarain¹,

Yahioglu²,

Stamati³

et al. 2015

Expert Opinion on Drug Discovery

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“…Thus, the Pictet-Spengler reaction 52,53,55,56 or recently reported trapped Knoevenagel condensation could become viable alternatives. 57 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 To further functionalize the Fc fragment carrying fluorescent cargo at the N-terminus (1o), we coupled PNA 2 using eSrtA-mediated ligation towards 11.…”

Section: Internalization Of Hybrid Constructs By C-metoverexpressing mentioning

confidence: 99%

Self-Assembled Hybrid Aptamer-Fc Conjugates for Targeted Delivery: A Modular Chemoenzymatic Approach

et al. 2015

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Over the past decade, DNA and RNA aptamers have attracted keen research interest due to their ability to specifically bind targets of therapeutic relevance. However, their application is often hampered by a short serum half-life and missing effector functions. Conjugation of aptamers to antibody Fc fragments could improve pharmacokinetics, enable immune effector mechanisms, and provide an option for the introduction of desired payloads (e.g., toxins or fluorescent dyes). We developed a modular scaffold-supported system based on human IgG1 Fc fragments, which allows for its dual functionalization with moieties of interest. In our approach, two bioorthogonal, enzyme-mediated reactions were used in combination with oxime ligation and self-assembly based on PNA-DNA base pairing. Thus, an engineered synthetic peptide nucleic acid (PNA) oligomer was coupled to the C-termini of the Fc dimer upon sequence-specific sortase A-mediated transpeptidation. Hybridization of the resulting Fc-PNA conjugate with a tailored DNA aptamer that binds cancer-related hepatocyte growth factor receptor (c-MET) led to a hybrid construct which showed strong and specific binding to c-MET and was readily internalized by c-MET-overexpressing cells. To install an additional orthogonally addressable site, aldehyde tag technology was applied followed by oxime ligation with an aminooxy-bearing fluorescent dye as model cargo. Delivery of fluorescent probe specifically to c-MET-overexpressing cells was confirmed by flow cytometry. Our approach can provide access to engineered aptamer-Fc conjugates with desired target specificity and cytotoxic payloads.

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Hydrazino-Pictet-Spengler Ligation as a Biocompatible Method for the Generation of Stable Protein Conjugates

Cited by 149 publications

References 56 publications

Critical Evaluation and Rate Constants of Chemoselective Ligation Reactions for Stoichiometric Conjugations in Water

Critical Evaluation and Rate Constants of Chemoselective Ligation Reactions for Stoichiometric Conjugations in Water

Emerging formats for next-generation antibody drug conjugates

Self-Assembled Hybrid Aptamer-Fc Conjugates for Targeted Delivery: A Modular Chemoenzymatic Approach

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