Innovative cancer-targeting antibody production strategy by using cancer-derived small Extracellular Vesicles (sEV)

Abstract Introduction: With all the advancements in the development of tumor-targeting agents, most of the research has showed that only less than 0.7% of targeted therapy arrives at the tumor site, suggesting high levels of off-target adverse reactions [1-3]. Therefore, developing a new tumor-targeting ligand that could enhance tumor-targeting efficacy seems inevitable. Here, we designed a novel tumor-targeting antibody production method using cancer-derived small Extracellular Vesicles (sEVs). Since sEVs inherit their structure and function from their parent cells [4] , the antibodies produced against sEVs could target both cancer cells and sEVs. Using this approach, antibody-decorated targeted systems may achieve superior tumor-targeting efficiency, potentially overcoming the suboptimal drug concentration in the tumor. Methods: The sEVs (150 ~200 nm) from ovarian cancer cell (OVCAR-8) were isolated and injected into mice. Once the specificity of the collected plasma from mice to the OVCAR-8 cells was confirmed, hybridoma cells were generated by fusing the mice’s B cells as the antibody-producing part and the myeloma cells. Among various monoclonal antibodies (mAbs) produced by different hybridoma cells, the best OVCAR-8-targeting mAb was isolated and purified. Next, the selected mAb was decorated to the surface of paclitaxel-encapsulated CD8+ T cell sEVs (AB-TSEV/P). Then the formulations including AB-TSEV/P were treated to the OVCAR-8-bearing mice and the changes in tumor volume and body weight was measured for 80 days. An in vivo biodistribution study of AB-TSEV loaded with IR-780 dye (AB-TSEV/IR) was then conducted using an IVIS Spectrum Imaging System to evaluate the accumulation of AB-TSEV/IR in the tumor site. Results: Based on the characterization results, the size of OVCAR-8-derived sEVs and CD8+ T cell sEVs were 150-200 nm. The plasma from mice after injection of sEVs showed specificity to OVCAR-8 cells through multifold cell killing efficacy over non-parent cancer and non-cancer cells. Based on our results from immunocytochemistry analysis, the selected monoclonal antibody had a significantly higher affinity to OVCAR-8 cells compared to non-cancerous cells. AB-TSEV/P decreased the tumor growth compared to formulations without mAb. Mice’s body weight was not changed significantly during the study. Also, in vivo biodistribution study showed that compared to control, AB-TSEV preferred a higher accumulation in the tumor. Conclusions: We developed a new strategy for antibody production using cancer cell sEVs. This antibody could target the sEV’s parent cells. Thus, a significant amount of the targeted formulation decorated with this antibody will potentially arrive at the tumor, overcoming the current suboptimal dose of cancer therapy. Although we only used ovarian cancer sEVs for this study, considering the potential of this approach, this technology can be applied to not only different cancers but also other diseases, where we can culture their cells and subsequently produce their sEVs.