Development of a bridging ELISA for detection of antibodies against ZV0203 in cynomolgus monkey serum.

Immunogenicity has been a major concern in the safety evaluation of therapeutic proteins. The assessment of the unwanted immunogenicity of the therapeutic proteins performed in animals prior to clinical trials has been a regulatory requirement. In preclinical studies of therapeutic proteins, cynomolgus monkeys are usually the most relevant animal species. ZV0203, a recombinant humanized anti-human epidermal growth factor receptor 2 monoclonal antibody covalently bound to a cytotoxic drug (Duo-5), possesses a novel format of antibody drug conjugates. In this study, we reported the development, validation, and application of a bridging enzyme-linked immunosorbent assay for the detection of antibodies against ZV0203 in cynomolgus monkey serum. Drug interference at low positive control (18.0 ng/mL) and high positive control (130 ng/mL) of anti-ZV0203 antibodies was not observed when ZV0203 concentration is below 1.74 µg/mL and 1.49 µg/mL, respectively. In addition, no interference was found from mouse IgG1, but interference was observed with human IgG1. No effect of hemolysis was found on the analysis results of the testing samples present in 100% pooled rabbit serum containing 2% (V/V) erythrocyte hemolysates. Besides, spiked anti-ZV0203 antibody in rabbit serum was stable after 5 freeze/thaw cycles. The results showed that the method is suitable for the detection of anti-ZV0203 antibodies in cynomolgus monkey serum. The assay was also successfully applied in the repeated dose study of ZV0203.

Antitumor activity of a 5T4 targeting antibody drug conjugate with a novel payload derived from MMAF via C-Lock linker.

Antibody-drug conjugates (ADCs) belong to a promising class of biopharmaceuticals in which target-killing of tumor cells was achieved by marrying the potency of the cytotoxic payload with the tumor specificity of the antibody. Here we developed a novel ADC (ZV0508) that targets 5T4 oncofetal antigen, which is overexpressed in many carcinomas on both bulk tumor cells and cancer stem cells. A novel cytotoxic payload called Duostatin-5 (Duo-5) which was derived from monomethyl auristatin F (MMAF) was attached to a 5T4 targeting antibody (ZV05) by interchain cysteine cross-linking conjugation via a disubstituted C-Lock linker. We have investigated the antitumor efficacy of ZV0508 by in vitro and in vivo studies, and compared its antitumor activity with ZV05-mcMMAF (ZV0501), in which MMAF was linked via a conventional noncleavable maleimidocaproyl linker. As results, ZV0508 exhibited ideal antiproliferative effects through blocking cell cycle and inducing cell apoptosis. The in vivo studies revealed that both ZV0501 and ZV0508 exhibited excellent antitumor activities even at a single dose. Although ZV0508 was inferior to ZV0501 in vitro, it elicited more durable antitumor responses than ZV0501 in vivo. The superior in vivo activity of ZV0508 may be due to the combined use of the disubstituted C-Lock linker and the novel payload Duo-5, resulting in a more stable and potent ADC. Taken together, these data suggest ZV0508 is a worthy candidate for the treatment of 5T4 positive cancers.

TAK-960, a novel, orally available, selective inhibitor of polo-like kinase 1, shows broad-spectrum preclinical antitumor activity in multiple dosing regimens.

Polo-like kinase 1 (PLK1) is a serine/threonine protein kinase involved in key processes during mitosis. Human PLK1 has been shown to be overexpressed in various human cancers, and elevated levels of PLK1 have been associated with poor prognosis, making it an attractive target for anticancer therapy. TAK-960 [4-[(9-cyclopentyl-7,7-difluoro-5-methyl-6-oxo-6,7,8,9-tetrahydro-5H-pyrimido[4,5-b][1,4]diazepin-2-yl)amino]-2-fluoro-5-methoxy-N-(1-methylpiperidin-4-yl) benzamide] is a novel, investigational, orally bioavailable, potent, and selective PLK1 inhibitor that has shown activity in several tumor cell lines, including those that express multidrug-resistant protein 1 (MDR1). Consistent with PLK1 inhibition, TAK-960 treatment caused accumulation of G(2)-M cells, aberrant polo mitosis morphology, and increased phosphorylation of histone H3 (pHH3) in vitro and in vivo. TAK-960 inhibited proliferation of multiple cancer cell lines, with mean EC(50) values ranging from 8.4 to 46.9 nmol/L, but not in nondividing normal cells (EC(50) >1,000 nmol/L). The mutation status of TP53 or KRAS and MDR1 expression did not correlate with the potency of TAK-960 in the cell lines tested. In animal models, oral administration of TAK-960 increased pHH3 in a dose-dependent manner and significantly inhibited the growth of HT-29 colorectal cancer xenografts. Treatment with once daily TAK-960 exhibited significant efficacy against multiple tumor xenografts, including an adriamycin/paclitaxel-resistant xenograft model and a disseminated leukemia model. TAK-960 has entered clinical evaluation in patients with advanced cancers.