Latent Warheads for Targeted Cancer Therapy: Design and Synthesis of pro-Pyrrolobenzodiazepines and Conjugates

Abstract

Pyrrolobenzodiazepines (PBDs) and their dimers (bis-PBDs) have emerged as some of the most potent chemotherapeutic compounds, and are currently under development as novel payloads in antibody-drug conjugates (ADCs). However, when used as stand-alone therapeutics or as warheads for small molecule drug conjugates (SMDCs), dose-limiting toxicities are often observed. As an elegant solution to this inherent problem, we designed diazepine-ring-opened conjugated prodrugs lacking the imine moiety. Once the prodrug (pro-PBD) conjugate enters a targeted cell, cleavage of the linker system triggers the generation of a reactive intermediate possessing an aldehyde and aromatic amine. An intramolecular ring-closing reaction subsequently takes place as the aromatic amine adds to the aldehyde with the loss of water to give the imine and, as a result, the diazepine ring. In our pro-PBDs, we mask the aldehyde as a hydrolytically sensitive oxazolidine moiety which in turn is a part of a reductively labile self-immolative linker system. To prove the range of applications for this new class of latent DNA-alkylators, we designed and synthesized several novel latent warheads: pro-PBD dimers and hybrids of pro-PBD with other sequence-selective DNA minor groove binders. Preliminary preclinical pharmacology studies showed excellent biological activity and specificity.

Octreotide Conjugates for Tumor Targeting and Imaging

“…Small molecule-mediated targeting represents advantages in terms of the straightforward organic synthesis of SMDCs and their uniform structure as compared to antibodies. Indeed, small molecule ligands have been used to efficiently target tumors expressing the folate receptor [7], prostate-specific membrane antigen (PSMA) [8], carbonic anhydrase IX (CAIX) [9,10,11], and somatostatin receptors (SSTRs) [12].…”

Pre-clinical studies of EC2629, a highly potent folate- receptor-targeted DNA crosslinking agent

“…It contains the tumor-targeting ligand, folate (in black), a hydrophilic peptide-polyethylene glycol (PEG) spacer consisting of L-Asp-L-Glu-L-Cys-PEG4 (in blue), a self-immolative penicillamine protected disulfide (in green) and the pro-drug form of the potent-PBD dimer (in red). Once the prodrug (pro-PBD) conjugate enters a targeted cell, cleavage of the linker system triggers the generation of a reactive intermediate which goes through an intramolecular ring-closing reaction to form the second diazepine ring of the cytotoxic PBD dimer, EC2491 24 .

Figure 1 Chemical structure and relative folate receptor binding affinity of EC2629.

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An Alternative Focus for Route Design for the Synthesis of Antibody–Drug Conjugate Payloads

“…The key structural features imparting cytotoxicity to the PBDs are well understood, 14 with a requirement for the A-, B-, and C-rings of the molecule to be in place. In the original synthesis of tesirine, high-containment facilities were required during construction of the two halves of the molecule and for the dimerization and subsequent stages (Figure 1).…”