Site-specific Antibody-Nitric Oxide Conjugate HN02 Possesses Improved Antineoplastic and Safety Properties
Tianyue Cheng1, Jiajun Xie1, Xun Yuan2,3
1Antibody Engineering Laboratory, School of Life Science & Technology, China Pharmaceutical University, Nanjing, China.
Abstract:
Antibody-drug conjugates (ADCs) combine the high specificity of antibodies with the cytotoxicity of payloads and have great potential in pan-cancer immunotherapy. However, the current payloads for clinical uses have limited the therapeutic window due to their uncontrollable off-site toxicity. There is unmet needs to develop more potent ADC payloads with better safety and efficacy profiles. Nitric oxide (NO) is a special molecule that has low toxicity itself, which can kill tumor cells effectively when highly concentrated, has broad application prospects. Previously, we prepared for the first time an antibody-nitric oxide conjugate (ANC)-HN01, which showed inhibitory activity against hepatocellular carcinoma. However, the random conjugation method made HN01 highly heterogeneous and unstable. Here, we used site-specific conjugation-based engineered cysteine sites (CL-V211C) of anti-CD24 antibody to prepare a second-generation ANC with a drug-to-antibody ratio of 2. The homogeneous ANC, HN02 was stable in human plasma, shown in vitro bystander effect to neighboring cells and antiproliferative activity to CD24-targeted tumor cells. Compared with HN01, HN02 significantly prolonged the survival of tumor-bearing mice. In summary, we developed a stable and homogeneous site-specific conjugated ANC, which showed good antitumor activity and improved safety profile both in vitro and in vivo. This study provides new insight into the development of next generation of ADC candidates.
Insights
Researchers developed a new antibody-nitric oxide conjugate (ANC) for cancer therapy. This improved ANC demonstrates enhanced stability, targeted tumor cell killing, and better survival rates in mice compared to previous versions.
Area of Science:
- Oncology
- Immunotherapy
- Bioconjugation Chemistry
Background:
- Antibody-drug conjugates (ADCs) show promise in cancer immunotherapy but face limitations due to off-target toxicity of current payloads.
- Nitric oxide (NO) offers a potent, low-toxicity payload for ADCs when concentrated at the tumor site.
- Previous antibody-nitric oxide conjugates (ANCs) suffered from heterogeneity and instability due to random conjugation.
Purpose of the Study:
- To develop a more stable and homogeneous antibody-nitric oxide conjugate (ANC) using site-specific conjugation.
- To evaluate the in vitro and in vivo efficacy and safety of the novel ANC.
Main Methods:
- Engineered cysteine sites (CL-V211C) on an anti-CD24 antibody were used for site-specific conjugation.
- A second-generation homogeneous ANC (HN02) with a drug-to-antibody ratio of 2 was prepared.
- In vitro assays assessed stability, bystander effect, and antiproliferative activity; in vivo studies evaluated survival in tumor-bearing mice.
Main Results:
- The homogeneous ANC (HN02) exhibited stability in human plasma.
- HN02 demonstrated a bystander effect on neighboring cells and antiproliferative activity against CD24-targeted tumor cells.
- HN02 significantly improved survival rates in tumor-bearing mice compared to the first-generation ANC (HN01).
Conclusions:
- Site-specific conjugation yielded a stable and homogeneous ANC with improved antitumor activity and safety.
- This next-generation ANC candidate offers a promising advancement over traditional ADCs with random conjugation.
- The study provides valuable insights for developing novel ADC therapeutics with enhanced efficacy and safety profiles.
