Abstract 4783: Development and characterization of an antibody-drug conjugate (ADC) utilizing PB-223, a novel monoclonal antibody (mAb) specifically targeting core 2 O-glycans on human carcinomas

Abstract Introduction: The three essential components of an ADC include the mAb, the linker, and the cytotoxic payload. The mAb is designated to target specific tumor-associated antigens that are overexpressed on the surface of cancer cells. The mAb PB-223 was developed through the affinity maturation of mAb NEO-102, a chimeric human IgG1 mAb that specifically targets truncated Core-2 O-glycans, found in colorectal and pancreatic cancers. The binding affinity of PB-223 for its target was improved, compared to NEO-102, by optimizing its VH and VL sequences through Fast Screening for Expression Biophysical Properties and Affinity. PB-223 demonstrated a binding affinity (KD) at least 4-fold lower than NEO-102, indicating stronger tumor binding. IHC analysis also revealed that PB-223 binds to a wider spectrum of tumor tissues compared to NEO-102, but not to normal tissues. PB-223 can be internalized into human cancer cell lines expressing its target. The objective of this study is to develop an ADC utilizing PB-223 for treatment of various human malignancies. Experimental Design: PB-223 was used for the development of the ADC. Monomethyl auristatin E (MMAE) was used as payload, with mc-vc-PABc serving as the linker. PB-223 was conjugated to the linker-payload through a cysteine-based conjugation method. Drug-to-antibody ratio (DAR) was measured using Size Exclusion Chromatography (SEC)-MS. Flow cytometry was used for binding assessment of the ADC using the ovarian cancer cell line OV90 as the target. Cytotoxicity of the ADC was evaluated through cell viability assays using CellTiter-Glo cell viability Kit using OV90.as target. Results: Three ADCs were developed: PB-MMAE-2, PB-MMAE-5 and PB-MMAE-6. The DAR for these ADCs were 3.72, 3.92 and 4.15 respectively. We analyzed their target binding capabilities in comparison to PB-223. The results indicated that all three ADCs exhibit similar binding affinity to PB-223. The cytotoxicity of the ADCs was evaluated using the breast cancer cell line SK-BR-3 with Trastuzumab deruxtecan ADC (DS-8201) as positive control. At a concentration of 10nM, DS-8201 inhibited cell proliferation by 82.39%. All three PB-223 ADC effectively killed OV90 cells. At a concentration of 333nM, the percentage of cell killing for PB-MMAE-2, PB-MMAE-5 and PB-MMAE-6 was 82.91%, 84.04% and 83.16%, respectively. In contrast, both human IgG1 and naked PB-223 mAb showed no killing of OV90 cells. Conclusions: In this study, we developed PB-223 ADCs and evaluated their anti-tumor efficacy using an in vitro model. We assessed three PB-223 ADCs: PB-MMAE-2, PB-MMAE-5 and PB-MMAE-6. Our findings highlight the potential of these ADCs as innovative options for treating human carcinomas expressing core 2 O-glycans. Further In vivo studies to evaluate the pharmacokinetics and pharmacodynamics are ongoing. Citation Format: Kwong Y. Tsang, Massimo Fantini, Anjum Zaki, Sharon A. Mavroukakis, Philip M. Arlen. Development and characterization of an antibody-drug conjugate (ADC) utilizing PB-223, a novel monoclonal antibody (mAb) specifically targeting core 2 O-glycans on human carcinomas [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 4783.