TM4SF1 as a promising ADC target for KRAS-mutated cancers

Transmembrane 4 L six family 1 (TM4SF1) is a protein with four transmembrane domains belonging to the transmembrane 4 L six family members. TM4SF1 interacts with tetraspanins, integrins, and various receptor tyrosine kinases. TM4SF1 promotes cancer cell migration and invasion and is overexpressed in many epithelial-derived malignancies, including prostate, breast, pancreatic, lung, colon, and gastric cancers, while found at low concentrations in normal human tissues like endothelium, skin, lung, and germ cells. In this study, we evaluated TM4SF1 as an ADC (antibody-drug conjugate) target and investigated its connection with KRAS-mutated cancer.To evaluate the potential of TM4SF1 as an ADC target, we employed both in silico analysis and in vitro assays. In silico analysis included data from TCGA/GTEx, single-cell RNA sequencing, and DEPMAP. In vitro assays comprised flow cytometry, internalization assays using IncuCyte, colony formation assays after gene knockdown, and cell viability assays using an ADC which was conjugated by ADC kit (CellMosaic, Inc., MA, USA).The key findings suggest that TM4SF1 is highly expressed in various solid tumors, particularly in KRAS-mutated tumors, indicating it is a promising ADC target. In silico analysis results revealed that TM4SF1 expression was significantly higher in lung cancer, pancreatic cancer, colorectal cancer, and bile duct cancer tumors compared to normal tissues; specifically, following our in-house scRNA sequencing data, expression was increased in EGFR wild-type and post-TKI treated lung tumors, and viability decreased upon gene knockout in many types of solid tumor. In vitro analysis confirmed TM4SF1 expression levels suitable for ADC targeting in 86% to 100% of various cancer cell lines (lung, pancreatic, colorectal, liver, gastric, etc.). Functional analysis demonstrated that TM4SF1 was internalized efficiently (comparable to or greater than the positive control CD71), and lung cancer cell line with high expression and KRAS mutation responded more sensitively to TM4SF1-targeting ADCs compare to low expression and without KRAS mutation or normal cell line. Our findings demonstrate that TM4SF1 is highly expressed in various solid tumors, particularly in specific genetic subsets such as those with KRAS mutated tumors. The observed high expression levels, efficient internalization kinetics (comparable to the positive control CD71), and robust in vitro efficacy of TM4SF1-targeting ADCs strongly suggest that TM4SF1 is a promising therapeutic target. These results provide a strong rationale for the clinical development of TM4SF1-targeting ADCs as a novel precision oncology treatment strategy across multiple cancer types.