Endocrine Abstracts

JOINT3299

Introduction: Turner syndrome (TS) is a common aneuploidy affecting 1 in 2500 girls and has a multi-systemic presentation including short stature, pubertal delay, congenital anomalies of the heart, kidney, low performance-IQ, high risk for cardiometabolic diseases and osteoporosis etc. It is diagnosed by peripheral-blood karyotyping only after overt clinical manifestations develop, thus losing valuable time for early growth hormone therapy, pubertal induction and screening for anomalies. Till date, is no point-of-care test for a rapid and accurate diagnosis of TS has been in use.

Objective: To assess the sensitivity and specificity of qPCR in diagnosing karyotypic variants of TS and develop a POC test for TS by developing a covalent organic framework (COF) based dual mode sensor for DNA samples.

Methods: In the first part, we extracted genomic DNA from peripheral blood samples of 50 girls with TS (45, X = 23, 45, X/46, XX = 10, Isochromosome Xq = 12, 45, X/46, XY mosaics = 5), 25 normal females(46, XX) and 5 normal males(46, XY). Using real-time PCR, we conducted qPCR with four genes - two Xp genes [SHOX and ARSE] and two Xq genes [VAMP7 and XIST] and a housekeeping autosomal gene HBB. The ΔΔ CT method was used for gene dose calculation and ROC curves were constructed to determine cut-offs for the different genes. Using the genes with best ROCs (SHOX and ARSE), we designed two pairs of complementary oligonucleotides decorated on a covalent organic framework (COF). Due to different doses of the target genes, the frameworks get arranged in a unique fashion, leading to quantitative changes in emission and electrochemical responses of the COF.

Results: qPCR could distinguish TS from normal females with >85 % sensitivity and specificity. SHOX could discriminate between TS from female controls with 86 % sensitivity and 86. 4 % specificity at a cut-off of 0. 75 while ARSE had a sensitivity of 82. 6% and specificity of 83. 6% at a cut-off of 0. 86. The ARSE gene was particularly useful to diagnose non-classic TS. The dual mode sensor developed using COF for SHOX and ARSE genes yielded the diagnosis in all the samples tested. However, it could not diagnose ring chromosomes and very low levels of mosaicism.

Conclusion: We have developed a novel dual-mode COF sensor for the diagnosis of Turner syndrome. This might be used for neonatal or community-based screening programmes to detect TS.