Effect and mechanism of HER2 antibody-drug conjugate combined with anti-PD-1 antibody in mouse bladder cancer models

To investigate the synergistic therapeutic effects of HER2 antibody-drug conjugate (HER2-ADC) combined with anti-PD-1 antibody (anti-PD-1) on HER2-expressing bladder cancer and elucidate its regulatory mechanisms on the tumor immune microenvironment.

Orthotopic tumor models were established in 40 female C57BL/6 mice (6~8 weeks old, body mass 18~22 g) using MB49 bladder cancer cells overexpressing human HER2. When tumors reached 100 mm³, the mice were randomized into (n=10) control (intraperitoneal injection of 1.0 mL PBS), anti-PD-1 (200 μg per mouse every 3 d), HER2-ADC (2.5 mg/kg once weekly), and combination groups (same regimens as above monotherapy). Tumor volume and body mass were measured every 3 d during 28-day treatment. Tumor growth kinetics and survival rates were analyzed. Post-treatment survival was monitored until natural death to determine median survival time (n=5). At day 28, blood and tumor samples (n=5) were collected to detect myeloid-derived suppressor cells (MDSCs; CD11b⁺Gr1⁺) and regulatory T cells (Tregs; CD4⁺CD25⁺FOXP3⁺) with flow cytometry, tumor-infiltrating CD3⁺T, CD8⁺T, and FOXP3⁺T cells with immunohistochemical assasy, and liver/kidney functions [alanine aminotransferase (ALT), aspartate aminotransferase (AST), blood urea nitrogen (BUN),creatinine (CRE)] and tissue damage indicators [lactate dehydrogenase isoenzyme (LDH-L)].

In 28 d after treatment, the combination group obtained significantly smallest tumor volume than the control group and the 2 monotherapy groups (all P<0.01). The longest median survival was observed in the combination group (65 d, P<0.01), followed by the HER2-ADC group (55 d), anti-PD-1 group (53 d) and control group (41 d). After 28 d of treatment, the combination group exhibited obviously the smallest peripheral proportions of MDSCs/Tregs, most tumor-infiltrating CD3⁺T/CD8⁺T cells, and less FOXP3⁺T cells when compared with the 2 monotherapy groups and control group (all P<0.05). While, the 2 monotherapy groups had smaller MDSCs/Tregs proportions than the control group (P<0.05). No significant differences were observed among the 4 groups in serum ALT, AST, BUN, CRE, or LDH-L levels, and all of them were within normal ranges.

HER2-ADC combined with anti-PD-1 suppresses the growth of orthotopic bladder tumor, probably through their synergic effects on down-regulating MDSCs/Treg and enhancing CD8⁺T cell infiltration.