Secretion of CKAP4 with SEVs (Figure S1). Generation of CKAP4 KO mice (Figure S2). Determination of epitopes of anti-CKAP4 mAbs (Figure S3). Development of ELISA for the detection of human serum CKAP4 (Figure S4). Detection of human serum CKAP4 by the ELISA (Figure S5). Characterization of anti-CKAP4 mAbs (Figure S6). Inhibition of xenograft tumor formation by anti-CKAP4 mAbs (Figure S7).
Ministry of Education, Culture, Sports, Science and Technology
Cancer Research And Therapeutic Evolution
Japan Agency for Medical Research and Development
History
ARTICLE ABSTRACT
The survival rate of pancreatic ductal adenocarcinoma (PDAC) is poor; thus, novel molecularly targeted therapy and companion diagnostics are required. We asked whether cytoskeleton-associated protein 4 (CKAP4), a novel Dickkopf1 (DKK1) receptor, is a candidate for PDAC diagnosis and therapy.Experimental Design: Whether CKAP4 can be secreted with small extracellular vesicles (SEV) from PDAC cells was examined. It was also investigated whether CKAP4 can be detected in sera from patients with PDAC by ELISA using newly generated anti-CKAP4 mAbs and whether anti-CKAP4 mAbs can show antitumor activity in vivo.
CKAP4 was secreted with SEVs from PDAC cells, and the SEVs exhibited the characteristics of exosomes. The secretion of CKAP4-containing exosomes was mediated by DKK1-dependent endocytosis routes and required exosome biogenesis molecules. Two ELISAs capable of detecting tumor-secreted CKAP4 were developed. The serum CKAP4 levels were higher in patients with PDAC than healthy control individuals. CKAP4 was highly detected in the sera of pancreatic tumor-bearing xenografted mice and patients with PDAC, whereas CKAP4 was barely detectable in sera from normal mice and postoperative patients. Anti-CKAP4 mAbs with different epitopes demonstrated the inhibitory activities for the binding of DKK1 and CKAP4, AKT activity, and proliferation and migration of PDAC cells. Anti-CKAP4 mAbs also suppressed xenograft tumor formation in immunodeficient mice and extended the survival of mice receiving intraperitoneal or orthotopic injection of PDAC cells.
CKAP4 secreted in exosomes may represent a biomarker for PDAC. Anti-CKAP4 mAbs can contribute to the development of novel diagnostic methods and therapeutics.