ROR1 NbTAC(ShanghaiTech University)

Receptor tyrosine kinase-like orphan receptor 1 (ROR1) plays a key role in noncanonical Wnt signaling. It is frequently overexpressed in tumors, and high ROR1 level is associated with poor patient survival. ROR1 has emerged as a potential therapeutic target for cancer therapy. Here, we report the first nanobody-based proteolysis-targeting chimeras (NbTAC) to degrade ROR1, by bringing the membrane-anchored E3 ubiquitin ligase RING finger protein 149 (RNF149) and membrane-bound ROR1 to proximity. We used phage display approach, discovered specific nanobodies for the extracellular domains (ECD) of RNF149 and ROR1, respectively. A number of bispecific NbTACs, formed by fusing an anti-RNF149 nanobody with an anti-ROR1 nanobody, were tested for their activities to degrade ROR1 in the cell. Among the NbTACs, N16-R14 exhibited specific and efficient elimination of ROR1 and proliferation inhibition of triple-negative breast cancer (TNBC) cell lines at nanomolar concentration. This ROR1 NbTAC may serve as a potential therapeutic agent to block ROR1-associated cancer progress in various indications. This RNF149-induced target degradation strategy may also be adapted to other membrane target proteins for broad applications.