Measurement of the critical DNA lesions produced by antibody-directed enzyme prodrug therapy (ADEPT) in vitro, in vivo and in clinical material

Abstract

Summary An antibody-directed enzyme prodrug therapy (ADEPT) system against CEA-positive tumours is currently in phase I clinical trials. It consists of a prodrug, 4-[N,N-bis(2-iodoethyl) amino] phenoxycarbonyl L-glutamic acid (ZD2767P) and a conjugate of the F(ab') 2 anti-CEA antibody A5B7 and the bacterial enzyme carboxypeptidase G2 (CPG2). ZD2767P is converted by antibody-targeted CPG2 into an active bifunctional alkylating drug (ZD2767) at the tumour site. The IC 50 value of the prodrug against the human colorectal tumour LS174T cell line was 55 ± 9 µM following a 1 h exposure. In contrast, co-incubation of ZD2767P with CPG2 resulted in 229-fold increase in activity. Using a modified comet assay, DNA interstrand cross links (ISC) were detected within 1 h of ZD2767P + CPG2 treatment and were repaired by 24 h. A clear dose-response was seen between the level of ISC, growth inhibition and ZD2767 concentration. Administration of a therapeutic dose of ZD2767P 72 h after the F(ab′) 2 A5B7 conjugate to mice bearing LS147T xenografts resulted in extensive ISC in the tumour after 1 h; repair was seen at 24 h. Tumour biopsies and peripheral lymphocytes were studied in 5 patients on the ADEPT phase I clinical trial. In 4 patients no ISC were detected. These patients also demonstrated poor localization of conjugate and no tumour response was seen. However a significant level of ISC was detected in one tumour biopsy, which also showed evidence of conjugate localization and clinical response. These studies demonstrate the application of the comet assay in the measurement of ISC in vitro and in clinical material and confirm that activation of ZD2767P results in the formation of DNA crosslinks.

<i>In Vitro</i> Investigation of DNA Damage Induced by the DNA Cross-Linking Agents Oxaliplatin and Satraplatin in Lymphocytes of Colorectal Cancer Patients

“…Oxaliplatin also forms interstrand crosslinks, which represent 10% of the total number of adducts [53]. The mode of action of many chemotherapeutic drugs of importance in cancer treatment is through production of DNA interstrand crosslinks detectible by the comet assay in cells exposed to the drug [54]. It has recently been demonstrated that oxaliplatin-induced crosslinks were detected by the alkaline Comet assay using a H460 tumour cell line and patients' lymphocytes.…”

Involvement of the HER2 pathway in repair of DNA damage produced by chemotherapeutic agents

“…It is now increasingly used to monitor ICL formation and unhooking in clinical samples (30,40) as a pharmacodynamic end point in clinical trials (41,42), and following ex vivo treatment (39,43).…”

Repair of DNA interstrand crosslinks as a mechanism of clinical resistance to melphalan in multiple myeloma

“…It is also more sensitive than other techniques used to measure ICL such as alkaline elution and allows the formation and repair of ICL to be studied in clinical material at pharmacologically relevant doses. 20 For example, initial ICL formation and repair studies using in vitro cell lines and human tumor xenograft models were extended to tumor biopsies from patients undergoing antibody-directed enzyme prodrug therapy phase 1 clinical trials, 21 and in vivo formation of ICL has also been followed in lymphocytes taken from patients receiving ifosfamide therapy. 20 In the present study we have used the Comet assay to examine formation and repair of melphalan-induced ICL in plasma cells from patients with MM following ex vivo treatment with melphalan.…”