Mutation of Threonine 34 in Mouse Podoplanin-Fc Reduces CLEC-2 Binding and Toxicity in Vivo While Retaining Anti-lymphangiogenic Activity

Abstract

Background: Podoplanin-Fc inhibits lymphangiogenesis, but also causes a bleeding disorder by binding to CLEC-2 expressed on platelets. Results: Mutation of threonine 34 in mouse Pdpn-Fc reduces 30-fold the binding to CLEC-2 and does not hamper its antilymphangiogenic activity. Conclusion: PdpnT34A-Fc is an active lymphangiogenesis inhibitor with a better tolerability. Significance: Mutagenesis of Pdpn-Fc is a valid approach to improve this tool for anti-lymphangiogenic therapy.

New Insights into the Role of Podoplanin in Epithelial–Mesenchymal Transition

“…These acidic residues interact with a series of Arg residues in CLEC-2. These data are consistent with the results of Bianchi et al (2014), which showed that in mouse podoplanin, O-glycosylation of Thr-34 (localized in PLAG1…”

Postnatal Deletion of Podoplanin in Lymphatic Endothelium Results in Blood Filling of the Lymphatic System and Impairs Dendritic Cell Migration to Lymph Nodes

“…20 In addition, blocking of podoplanin function by administration of podoplanin-Fc during late embryonic development resulted in a less complex diaphragmatic lymphatic network in pups. 41 Thus, it is likely that, early in development, podoplanin functionally contributes not only to the separation of the blood vascular and the lymphatic system but also to LV patterning. However, our data indicate that after birth, podoplanin is dispensable for proper LV maintenance and drainage function.…”

Podoplanin in Inflammation and Cancer

“…The transgenic overexpression of soluble PDPN-Fc in mouse skin provoked platelet activation via CLEC-2, causing disseminated intravascular coagulation and thrombocytopenia [121]. This event required O -glycosylated T34, as mutation of this residue, considered to be essential for the binding of podoplanin to CLEC-2, reduced platelet activation and its side effects [122]. However, the mutant soluble PDPN-Fc retained the ability to inhibit lymphangiogenesis in vitro and in vivo [122], suggesting that the function of podoplanin in LECs is independent of binding to CLEC-2.…”

“…This event required O -glycosylated T34, as mutation of this residue, considered to be essential for the binding of podoplanin to CLEC-2, reduced platelet activation and its side effects [122]. However, the mutant soluble PDPN-Fc retained the ability to inhibit lymphangiogenesis in vitro and in vivo [122], suggesting that the function of podoplanin in LECs is independent of binding to CLEC-2. The above conclusion is contradictory with earlier reports showing that platelets inhibit lymphangiogenesis [58,123].…”