Linker-Dependent Folding Rationalizes PROTAC Cell Permeability

Abstract

Proteolysis-targeting chimeras (PROTACs) must be cell permeable to reach their target proteins. This is challenging as the bivalent structure of PROTACs puts them in chemical space at, or beyond, the outer limits of oral druggable space. We used NMR spectroscopy and molecular dynamics (MD) simulations independently to gain insights into the origin of the differences in cell permeability displayed by three flexible cereblon PROTACs having closely related structures. Both methods revealed that the propensity of the PROTACs to adopt folded conformations with a low solvent-accessible 3D polar surface area in an apolar environment is correlated to high cell permeability. The chemical nature and the flexibility of the linker were essential for the PROTACs to populate folded conformations stabilized by intramolecular hydrogen bonds, π–π interactions, and van der Waals interactions. We conclude that MD simulations may be used for the prospective ranking of cell permeability in the design of cereblon PROTACs.

Predictive Modeling of PROTAC Cell Permeability with Machine Learning

“…Recent studies have revealed that the propensity of PROTACs to adopt folded, non-polar conformations in an apolar environment correlates to high cell permeability. 17,18 We conclude that BCMs can be implemented to support decision-making in drug discovery projects so that PROTACs predicted to have high permeability are prioritized for synthesis and subsequent testing over those predicted to have low permeability. Such classification models could be of particular value for prioritization of compounds early in lead optimization before large numbers of PROTACs have been synthesized in the project.…”

“…One reason for this complex behavior may be the 3D folding of PROTACs. Recent studies have revealed that the propensity of PROTACs to adopt folded, non-polar conformations in an apolar environment correlates to high cell permeability. , …”

“…The ratio between the potencies for binding of a PROTAC to its E3 ligase in a cell-based and in a biochemical assay is the measure of cell permeability used in this investigation. 19 The potencies were determined as reported previously for binding to VHL 17 and CRBN, 18 respectively. Permeability class boundaries for the VHL and CRBN PROTACs were derived from the observation that the majority of compounds that display a strong drop-off, i.e., a high numerical value for the permeability ratio, show cellular IC 50 values where their biological usefulness is questionable.…”

“…Information about conformational preferences has been found to improve predictions of cell permeability for moderately flexible compounds in bRo5 spaces such as macrocycles. , Two recent NMR studies indicated that conformational preferences are also important for the permeability of PROTACs. , However, PROTACs are often highly flexible and inclusion of conformational sampling in structure–property models may be challenging, as found in a recent investigation of PROTAC solubility . The modular construction of PROTACs from three units allows high-throughput synthesis of libraries with large structural diversity .…”

Degrader–Antibody Conjugates: Emerging New Modality

“…Even TPD provided a promising potential to access undruggable protein degradation, but still, in general, the lack of discrimination between healthy cells and cancer cells could raise the issue of off-target toxicities. Additionally, their physicochemical characteristics are associated with generally poor DMPK properties and low bioavailability. − Therefore, as a drug delivery concept, antibody conjugates provide emerging strategies for the TPD area. Using antibody conjugates, degraders can be delivered specifically to the targeted cancer-associated cells.…”