Genotyping and functional analysis of the D104N variant of human endostatin

Abstract

KEY WORDSendostatin, prostate cancer, angiogenesis, single nucleotide polymorphism ABBREVIATIONS AIPCandrogen independent prostate cancer HUVEC human umbilical vein endothelial cell HSPG heparin sulfate proteoglycan SNP single nucleotide polymorphism ACKNOWLEDGEMENTSWe wish to thank Andrada Tomoaia-Cotisel for technical assistance, Erin Lepper for predictive structural modeling and Martijn Gebbink for helpful correspondence. Research Paper Genotyping and Functional Analysis of the D104N Variant of Human Endostatin ABSTRACTEndostatin is an endogenous inhibitor of angiogenesis derived from the extracellular matrix protein collagen XVIII. It has been reported that a variation at the 104 position (D104N) of human endostatin is associated with an increased risk of prostate cancer, potentially indicating that this protein variant is less active as an anti-angiogenic agent. Herein we reported the results of genotyping 389 patients with androgen independent prostate cancer (AIPC) and 352 normal control individuals for D104N endostatin. There was no significant association between the frequency of 104N endostatin and the incidence of AIPC in either Caucasian or African American patients compared to controls (15% Caucasian AIPC versus 13.7% in Caucasian controls, p = 0.79; 7.4% African American AIPC versus 5.6% in African American controls, p = 0.64). Actuarial analysis revealed no statistically significant association between incidence of the DN heterozygous genotype and survival (p = 0.62 by logrank test). To study the functional significance of the D104N conversion, we have expressed and purified insoluble recombinant human 104D and 104N endostatin and compared their respective activities in human umbilical vein endothelial cell (HUVEC) tube formation assays. The 104N variant of human endostatin inhibited HUVEC tube formation at least as well as the wild-type form. We concluded that the D104N variation in human endostatin is neither clinically relevant nor suitable as a pharmacogenomic endpoint to assess the risk for developing AIPC.

Molecular Interplay between Endostatin, Integrins, and Heparan Sulfate

“…This suggests that this sequence change may not be responsible for the reported high risk of occurrence of sporadic breast cancer (25) and for the influence on the age of onset of acute myeloid leukemia (26). Our finding is supported by several studies showing that the D104N polymorphism does not appear to be associated with breast cancer susceptibility or severity (38), with increased risk to develop prostate cancer (39), or with lung cancer susceptibility (40). Alternatively, the D104N polymorphism may change the interactions of endostatin with other partners not yet identified.…”

A high risk of occurrence of sporadic breast cancer in individuals with the 104NN polymorphism of the COL18A1 gene

“…In contrast, similar risks for disease were found by Macpherson et al [11] in individuals with and without the polymorphism. The authors also expressed and purified human 104D and 104N ES and tested their function in human umbilical vein endothelial cell (HUVEC) tube formation assays.…”

Mutations in collagen 18A1 (COL18A1) and their relevance to the human phenotype

“…In a casecontrol study including 181 prostate cancer cases and 198 non-cancer individuals, we observed that the heterozygous N104 individuals have a 2.5 times increased chance of developing prostate cancer as compared with homozygous D104 subjects (Iughetti et al 2001). However, this association was not confirmed by us in another sample (data not published) as well as by others including different solid tumors (Ortega et al 2003, Liu et al 2003, Nascimento et al 2004, Macpherson et al 2004. It is possible that the variation of the genotypic frequencies observed in our original report just represent racial stratification.…”