L-DOS-47

L-DOS47, a targeted urease-anti-CEACAM6 immunoconjugate, alters the acidity of the tumor microenvironment by increasing local ammonia production. In vitro, the cytotoxic effects of L-DOS47 were additive when combined with pemetrexed and carboplatin.

This phase I, open-label, dose-escalation study evaluated the safety and tolerability of up to four cycles of L-DOS47 (administered on days 1, 8, and 15 of each cycle at doses ranging from 0.59 to 9.0 μg/kg) combined with pemetrexed and carboplatin in patients with stage IV nonsquamous NSCLC. Continued L-DOS47 treatment after the fourth cycle was allowed at the treating physicians' discretion.

A total of 14 patients received at least one dose of L-DOS47. Overall, L-DOS47 was well tolerated. Grade greater than or equal to 3 adverse events (AEs) were typically neutropenia related. Two grade greater than or equal to 3 AEs and no serious AEs were considered at least possibly related to L-DOS47. No dose-limiting toxicities were reported, so the maximum tolerated dose was not reached. The objective response rate was 41.7% with a median duration of response of 187 days. Clinical benefit was observed in 75.0% of the patients. After the first dose, L-DOS47 systemic exposure increased in a generally dose-proportional manner but decreased substantially with repeat dosing. Anti-L-DOS47 antibodies were detectable in 13 of 14 patients by cycle 2 with titers typically increasing with continued treatment. There was an apparent association between best overall response rate and highest anti-L-DOS47 antibody titer measured.

L-DOS47 combined with standard pemetrexed and carboplatin chemotherapy is well tolerated in patients with recurrent or metastatic nonsquamous NSCLC at doses up to 9.0 μg/kg.

A novel immunoconjugate (L-DOS47) was developed and characterized as a therapeutic agent for tumors expressing CEACAM6. The single domain antibody AFAIKL2, which targets CEACAM6, was expressed in the Escherichia coli BL21 (DE3) pT7-7 system. High purity urease (HPU) was extracted and purified from Jack bean meal. AFAIKL2 was activated using N-succinimidyl [4-iodoacetyl] aminobenzoate (SIAB) as the cross-linker and then conjugated to urease. The activation and conjugation reactions were controlled by altering pH. Under these conditions, the material ratio achieved conjugation ratios of 8-11 antibodies per urease molecule, the residual free urease content was practically negligible (<2%), and high purity (>95%) L-DOS47 conjugate was produced using only ultradiafiltration to remove unreacted antibody and hydrolyzed cross-linker. L-DOS47 was characterized by a panel of analytical techniques including SEC, IEC, Western blot, ELISA, and LC-MS(E) peptide mapping. As the antibody-urease conjugate ratio increased, a higher binding signal was observed. The specificity and cytotoxicity of L-DOS47 was confirmed by screening in four cell lines (BxPC-3, A549, MCF7, and CEACAM6-transfected H23). BxPC-3, a CEACAM6-expressing cell line was found to be most susceptible to L-DOS47. L-DOS47 is being investigated as a potential therapeutic agent in human phase I clinical studies for nonsmall cell lung cancer.