Leucine-rich repeat-containing G-protein-coupled receptor (LGR5) and LGR6 mark epithelial stem cells in normal tissues and tumors. They are expressed by stem cells in the ovarian surface and fallopian tube epithelia from which ovarian cancer arises. High-grade serous ovarian cancer is unique in expressing unusually high levels of LGR5 and LGR6 mRNA. R-spondins are the natural ligands for LGR5 and LGR6 to which they bind with nanomolar affinity. To target stem cells in ovarian cancer, we used the sortase reaction to site-specifically conjugate the potent cytotoxin monomethyl auristatin E (MMAE) via a protease sensitive linker to the two furin-like domains of RSPO1 (Fu₁-Fu₂) that mediate its binding to LGR5 and LGR6 and their co-receptors Zinc And Ring Finger 3 and Ring Finger Protein 43 via a protease-cleavable linker. An immunoglobulin Fc domain added to the N-terminal end served to dimerize the receptor-binding domains so that each molecule carries two MMAE. The resulting molecule, FcF2-MMAE, demonstrated: 1) selective LGR5-dependent low nanomolar cytotoxicity against ovarian cancer cells in vitro; 2) selectivity that was dependent on binding to both the LGR receptors and ubiquitin ligase co-receptors; 3) favorable stability and plasma pharmacokinetic properties when administered intravenously with an elimination half-life of 29.7 hours; 4) selective inhibition of LGR5-rich as opposed to isogenic LGR5-poor tumors in vivo; and, 5) therapeutic efficacy in three aggressive wild-type human ovarian cancer xenograft models. These results demonstrate the successful use of the Fu₁-Fu₂ domain of RSPO1 as a drug carrier and the ability of FcF2-MMAE to target cells in tumors that express stem cell markers. SIGNIFICANCE STATEMENT: FcF2-MMAE is a novel cancer therapeutic that exploits the high-affinity binding domains of RSPO1 to target monomethyl auristatin E to tumor stem cells that express LGR5. FcF2-MMAE has low nanomolar LGR5-dependent cytotoxicity in vitro, favorable pharmacokinetics, and differential efficacy in an isogenic LGR5-poor versus LGR5-rich ovarian cancer xenograft model when given on a weekly schedule.