An in Vitro Assay Using Cultured Kupffer Cells Can Predict the Impact of Drug Conjugation on in Vivo Antibody Pharmacokinetics.

While antibody-drug conjugates (ADCs) are advancing through clinical testing and receiving new marketing approvals, improvements to the technology continue to be developed in both academic and industrial laboratories. Among the key ADC attributes that can be improved upon with new technology are their biodistribution and pharmacokinetic properties. During the course of ADC development, it has become apparent that conjugation of drugs to the surface of a monoclonal antibody can alter its physicochemical characteristics in a manner that results in increased nonspecific interactions and more rapid elimination from plasma.

Native size-exclusion chromatography-mass spectrometry: suitability for antibody-drug conjugate drug-to-antibody ratio quantitation across a range of chemotypes and drug-loading levels.

Native size-exclusion chromatography-mass spectrometry (nSEC-MS) is an analytical methodology that is appropriate for accurately quantitating the drug-to-antibody ratio (DAR) on a wide variety of interchain cysteine-linked antibody-drug conjugates (ADCs), irrespective of chemotype. In the current preclinical environment, novel ADCs conjugated with unique drug-linkers need to progress toward the clinic as quickly as possible. Platform analytical approaches can reduce time-to-clinic because key process development and optimization activities can be decoupled from the development of bespoke, molecule-specific analytical methods.

Mouse Strains Influence Clearance and Efficacy of Antibody and Antibody-Drug Conjugate Via Fc-FcγR Interaction.

To provide a better understanding of the pharmacokinetics-pharmacodynamics relationships of antibody-based drugs, we analyzed several chimeric and humanized monoclonal antibodies or antibody-drug conjugates (ADC) for PK and efficacy among four strains of mice. Notably, antibodies and ADCs displayed a dose-dependent drug disposition profile in the plasma of NSG mice. The increased clearance rate in NSG mice resulted in the reduction of antitumor activity of ADCs.

Optimization of a PEGylated Glucuronide-Monomethylauristatin E Linker for Antibody-Drug Conjugates.

The emergence of antibody-drug conjugates (ADC), such as brentuximab vedotin and ado-trastuzumab emtansine, has led to increased efforts to identify new payloads and develop improved drug-linker technologies. Most antibody payloads impart significant hydrophobicity to the ADC, resulting in accelerated plasma clearance and suboptimal in vivo activity, particularly for conjugates with high drug-to-antibody ratios (DAR). We recently reported on the incorporation of a discrete PEG polymer as a side chain in a β-glucuronidase-cleavable monomethylauristatin E (MMAE) linker to provide homogeneous DAR 8 conjugates with decreased plasma clearance and increased antitumor activity in xenograft models relative to a non-PEGylated control.

Orthogonal Cysteine Protection Enables Homogeneous Multi-Drug Antibody-Drug Conjugates.

A strategy for the preparation of homogeneous antibody-drug conjugates (ADCs) containing multiple payloads has been developed. This approach utilizes sequential unmasking of cysteine residues with orthogonal protection to enable site-specific conjugation of each drug. In addition, because the approach utilizes conjugation to native antibody cysteine residues, it is widely applicable and enables high drug loading for improved ADC potency.

Development of Novel Quaternary Ammonium Linkers for Antibody-Drug Conjugates.

A quaternary ammonium-based drug-linker has been developed to expand the scope of antibody-drug conjugate (ADC) payloads to include tertiary amines, a functional group commonly present in biologically active compounds. The linker strategy was exemplified with a β-glucuronidase-cleavable auristatin E construct. The drug-linker was found to efficiently release free auristatin E (AE) in the presence of β-glucuronidase and provide ADCs that were highly stable in plasma.

Reducing hydrophobicity of homogeneous antibody-drug conjugates improves pharmacokinetics and therapeutic index.

The in vitro potency of antibody-drug conjugates (ADCs) increases with the drug-to-antibody ratio (DAR); however, ADC plasma clearance also increases with DAR, reducing exposure and in vivo efficacy. Here we show that accelerated clearance arises from ADC hydrophobicity, which can be modulated through drug-linker design. We exemplify this using hydrophilic auristatin drug linkers and PEGylated ADCs that yield uniform, high-DAR ADCs with superior in vivo performance.

SGN-LIV1A: a novel antibody-drug conjugate targeting LIV-1 for the treatment of metastatic breast cancer.

In this article, we describe a novel antibody-drug conjugate (ADC; SGN-LIV1A), targeting the zinc transporter LIV-1 (SLC39A6) for the treatment of metastatic breast cancer. LIV-1 was previously known to be expressed by estrogen receptor-positive breast cancers. In this study, we show that LIV-1 expression is maintained after hormonal therapy in primary and metastatic sites and is also upregulated in triple-negative breast cancers.

A potent anti-CD70 antibody-drug conjugate combining a dimeric pyrrolobenzodiazepine drug with site-specific conjugation technology.

A highly cytotoxic DNA cross-linking pyrrolobenzodiazepine (PBD) dimer with a valine-alanine dipeptide linker was conjugated to the anti-CD70 h1F6 mAb either through endogenous interchain cysteines or, site-specifically, through engineered cysteines at position 239 of the heavy chains. The h1F6239C-PBD conjugation strategy proved to be superior to interchain cysteine conjugation, affording an antibody-drug conjugate (ADC) with high uniformity in drug-loading and low levels of aggregation. In vitro cytotoxicity experiments demonstrated that the h1F6239C-PBD was potent and immunologically specific on CD70-positive renal cell carcinoma (RCC) and non-Hodgkin lymphoma (NHL) cell lines.

Catalytic and ligand-binding characteristics of Plasmodium falciparum serine hydroxymethyltransferase.

The plant-like, bifunctional dihydrofolate reductase-thymidylate synthase (DHFR-TS) from malaria parasites has been a good target for drug development. Dihydrofolate reductase (DHFR) is inhibited by clinically established antimalarials, pyrimethamine and cycloguanil. Thymidylate synthase (TS) is the target of potent experimental antimalarials such as 5-fluoroorotate and 1843U89.

Kinetics and ligand-binding preferences of Mycobacterium tuberculosis thymidylate synthases, ThyA and ThyX.

Background: Mycobacterium tuberculosis kills approximately 2 million people each year and presents an urgent need to identify new targets and new antitubercular drugs. Thymidylate synthase (TS) enzymes from other species offer good targets for drug development and the M. tuberculosis genome contains two putative TS enzymes, a conventional ThyA and a flavin-based ThyX.