Development of novel ADCs: conjugation of tubulysin analogues to trastuzumab monitored by dual radiolabeling.

Abstract

Tubulysins are highly toxic tubulin-targeting agents with a narrow therapeutic window that are interesting for application in antibody-drug conjugates (ADC). For full control over drug-antibody ratio (DAR) and the effect thereof on pharmacokinetics and tumor targeting, a dual-labeling approach was developed, wherein the drug, tubulysin variants, and the antibody, the anti-HER2 monoclonal antibody (mAb) trastuzumab, are radiolabeled. (131)I-radioiodination of two synthetic tubulysin A analogues, the less potent TUB-OH (IC50 > 100 nmol/L) and the potent TUB-OMOM (IC50, ~1 nmol/L), and their direct covalent conjugation to (89)Zr-trastuzumab were established. Radioiodination of tubulysins was 92% to 98% efficient and conversion to N-hydroxysuccinimide (NHS) esters more than 99%; esters were isolated in an overall yield of 68% ± 5% with radiochemical purity of more than 99.5%. Conjugation of (131)I-tubulysin-NHS esters to (89)Zr-trastuzumab was 45% to 55% efficient, resulting in ADCs with 96% to 98% radiochemical purity after size-exclusion chromatography. ADCs were evaluated for their tumor-targeting potential and antitumor effects in nude mice with tumors that were sensitive or resistant to trastuzumab, using ado-trastuzumab emtansine as a reference. ADCs appeared stable in vivo. An average DAR of 2 and 4 conferred pharmacokinetics and tumor-targeting behavior similar to parental trastuzumab. Efficacy studies using single-dose TUB-OMOM-trastuzumab (DAR 4) showed dose-dependent antitumor effects, including complete tumor eradications in trastuzumab-sensitive tumors in vivo. TUB-OMOM-trastuzumab (60 mg/kg) displayed efficacy similar to ado-trastuzumab emtansine (15 mg/kg) yet more effective than trastuzumab. Our findings illustrate the potential of synthetic tubulysins in ADCs for cancer treatment.

• Authors: Ruth Cohen, Danielle J Vugts, Gerard W M Visser, Marijke Stigter-van Walsum, Marije Bolijn, Marco Spiga, Paolo Lazzari, Sreejith Shankar, Monica Sani, Matteo Zanda, Guus A M S van Dongen
• Journal: Cancer research (Cancer Res) Vol. 74 Issue 20 Pg. 5700-10 (Oct 15 2014) ISSN: 1538-7445 [Electronic] United States
• PMID: 25145670 (Publication Type: Journal Article)
• Copyright: ©2014 American Association for Cancer Research.

Chemical References

• Antibodies, Monoclonal, Humanized
• Antineoplastic Agents
• Oligopeptides
• TUB-OH-trastuzumab
• TUB-OMOM-trastuzumab
• Trastuzumab

Topics

• Animals
• Antibodies, Monoclonal, Humanized (chemistry, pharmacology, therapeutic use)
• Antineoplastic Agents (chemical synthesis, pharmacology, therapeutic use)
• Cell Line, Tumor
• Cell Survival (drug effects)
• Drug Stability
• Female
• Humans
• Maximum Tolerated Dose
• Mice, Nude
• Oligopeptides
• Tissue Distribution
• Trastuzumab
• Xenograft Model Antitumor Assays