A Polar Sulfamide Spacer Significantly Enhances the Manufacturability, Stability, and Therapeutic Index of Antibody–Drug Conjugates

Abstract

Despite tremendous efforts in the field of targeted cancer therapy with antibody-drug conjugates (ADCs), attrition rates have been high. Historically, the priority in ADC development has been the selection of target, antibody, and toxin, with little focus on the nature of the linker. We show here that a short and polar sulfamide spacer (HydraSpace™, Oss, The Netherlands) positively impacts ADC properties in various ways: (a) efficiency of conjugation; (b) stability; and (c) therapeutic index. Different ADC formats are explored in terms of drug-to-antibody ratios (DAR2, DAR4) and we describe the generation of a DAR4 ADC by site-specific attachment of a bivalent linker-payload construct to a single conjugation site in the antibody. A head-to-head comparison of HydraSpace™-containing DAR4 ADCs to marketed drugs, derived from the same antibody and toxic payload components, indicated a significant improvement in both the efficacy and safety of several vivo models, corroborated by in-depth pharmacokinetic analysis. Taken together, HydraSpace™ technology based on a polar sulfamide spacer provides significant improvement in manufacturability, stability, and ADC design, and is a powerful platform to enable next-generation ADCs with enhanced therapeutic index.

Enzymatic glycan remodeling–metal free click (GlycoConnect™) provides homogenous antibody-drug conjugates with improved stability and therapeutic index without sequence engineering

“…All four azido-antibodies were subsequently conjugated with either linker-drug 4 or linker-drug 5 (see Supplementary Figure S9–S11). Both linker-drugs comprised bicyclononyne (BCN) 26 for metal-free click conjugation, our earlier reported 8 polar spacer technology based on carbamoyl sulfamide (HydraSpace™, green), a short PEG spacer, and either maytansine or MMAE tubulin inhibitor payload ( Figure 4a ). Further, to maximize variability in drug-to-antibody ratio (DAR), linker-drug 4 contains a linear linker, while compound 5 contains a branched linker.…”

“…7 Another advantage of ADCs generated with GlycoConnect™ is the concomitant annihilation of binding to CD16/CD32 (Fc- γ receptor III and II) and significant reduction (<30% remaining) of binding to CD64 (Fc- γ receptor I), which is generally undesirable, given the potential Fc- γ receptor-mediated uptake in healthy tissue. We later demonstrated 8 how GlycoConnect™ can be empowered by introduction of a highly polar spacer unit (HydraSpace™) based on a carbamoyl sulfamide group ( Figure 1 ). Three GlycoConnect™ ADCs are currently in Phase 1 clinical trials (ADCT-601, XMT-1592, and MRG004a), with more than a dozen additional ADCs in various stages of preclinical development, 9 thereby rendering the GlycoConnect™ approach the most prevalent (chemo)enzymatic antibody modification technology in the clinic.…”

Targeting cancer with antibody-drug conjugates: Promises and challenges

“…Hydrophilic linkers and spacers, including cyclodextrins, polyethylene glycol, and other polymers, may play a role in improving the stability of circulation, potency toward the target cells, and overall pharmacokinetics of the conjugate. [148][149][150] In addition to selectively choosing a chemical linker, the method by which the payloads are conjugated to the antibody is essential in modulating the homogeneity and potency of the ADC. 151 Until recently, conventional methods relied upon lysine and interchain cysteines to conjugate cytotoxic molecules to the antibody.…”

Non-Genetic Generation of Antibody Conjugates Based on Chemoenzymatic Tyrosine Click Chemistry

“…To this end, a range of linker–payload constructs was synthesized (Scheme ) by covalent attachment of cytotoxic payloads PBD dimer ( 5 ) or MMAE ( 6 and 7 ) to BCN derivatives 4a via a Val-Cit-PABC cleavable linker (shown in red) or to 4b via Val-Ala-PABC. Notably, in each case we also inserted a carbamoyl-sulfamide unit (shown in green), a highly polar linker from medicinal chemistry programs and marketed as HydraSpace technology, in order to impart better solubility to the BCN-linker-drugs and to improve the putative therapeutic window. In addition, for MMAE we prepared both a linear linker-drug bearing one payload ( 6 ) as well as a linker-drug with a branching unit and two payloads ( 7 ), in order to generate the respective ADCs with drug-to-antibody ratio (DAR) of 2 or 4, respectively.…”