Development of 177Lu-labeled antibody-drug conjugate for improving treatment efficacy of radioimmunotherapy for solid tumors

Introduction: To improve the treatment efficacy of radioimmunotherapy against solid tumors, we construct and evaluate radiolabeled antibody-drug conjugates (ADCs) equipped with lutetium-177 (¹⁷⁷Lu) and monomethyl auristatin E (MMAE), an antimitotic agent known to radiosensitize tumors.

Methods: An aglycosylated N297A monoclonal antibody (mAb) targeting trophoblast cell-surface antigen 2 (TROP2) was used in this study. Chelator-conjugated anti-TROP2 ADCs and homogeneous mAb-chelator conjugate were constructed via enzyme-mediated site-specific linker conjugation, followed by strain-promoted click reactions for chelator/payload installation. The heterogeneous mAb-chelator conjugate was also prepared by conventional lysine coupling. In vitro cytotoxicity of non-radioactive Lu-labeled conjugates was tested using human breast cancer cell lines. Chelator-conjugated ADC and mAb-chelator conjugates were radiolabeled with ¹⁷⁷Lu and G25 spin-column purified to produce ¹⁷⁷Lu-labeled ADC, ¹⁷⁷Lu-labeled homogenous radioimmunoconjugate (homo-RIC), and ¹⁷⁷Lu-labeled conventional RIC (hetero-RIC). Radiochemical yield and radiochemical purity were measured by iTLC. Biodistribution studies with ¹⁷⁷Lu-labeled ADC, homo-RIC, and hetero RIC were performed on JIMT-1 orthotopic breast tumor-bearing mice. In the treatment study, the tumor-bearing mice were randomly allocated to treatment groups as follows: no treatment (n=4), ¹⁷⁷Lu-labeled ADC (3MBq; 0.7 mg/kg, n=5), homo-RIC (3MBq, n=4), hetero-RIC (3MBq, n=5), and MMAE ADC (non-radioactive, 0.7 mg/kg, n=5). To further evaluate the therapeutic potential of ¹⁷⁷Lu-labeled ADC, the second therapeutic study was performed with increased radioactivity and ADC doses (5 MBq; 1.5 mg/kg). A single dose of each conjugate was administered intravenously, and tumor volume and body weight were monitored.

Results: Anti-TROP2 ADCs and homogeneous antibody-chelator conjugate were obtained with high homogeneity and defined chelator/payload-to-antibody ratios. Non-radioactive Lu-labeled ADC showed in vitro potency comparable to that of the original ADC. ¹⁷⁷Lu-labeled ADC and RICs were obtained in high radiochemical purity (>98%). The homogeneous conjugates, ¹⁷⁷Lu-labeled ADC, and homo-RIC showed significantly improved radioactivity accumulation in the tumor compared to hetero-RIC (p<0.01 at 72 h). In the therapeutic study, a single injection of radiolabeled ADC (3MBq; 0.7 mg/kg) delayed tumor growth significantly more than did ADC alone (0.7 mg/kg) or hetero-RIC (3MBq). Homo-RIC showed an antitumor effect comparable to that of hetero-RIC at the dose of 3 MBq/head, indicating homogeneity of the RIC itself did not significantly improve therapeutic efficacy. Administration of ¹⁷⁷Lu-labeled ADC (5MBq; 1.5 mg/kg) suppressed tumor growth for over 50 days. In contrast, tumors started to regrow at 30 days post-treatment of homo-RIC (5MBq).

Conclusions: We developed a highly homogeneous ¹⁷⁷Lu-labeled anti-TROP2 ADC. ¹⁷⁷Lu-labeled anti-TROP2 ADC showed higher tumor uptake and greater treatment efficacy than conventional RIC. ¹⁷⁷Lu-labeled anti-TROP2 ADC (5MBq; 1.5 mg/kg) suppressed tumor growth significantly longer than did homogeneous RIC (5 MBq). These results suggest that ¹⁷⁷Lu-labeled anti-TROP2 ADCs are promising for improving the treatment efficacy of beta-radioimmunotherapy for solid tumors.