Association of thymidylate synthase gene polymorphisms with stavudine triphosphate intracellular levels and lipodystrophy.

The antiviral activity and toxicity of stavudine (d4T) depend on its triphosphate metabolite, stavudine triphosphate (d4T-TP). Therefore, modifications in intracellular levels of d4T-TP may change the toxicity profile of stavudine. d4T-TP intracellular levels in peripheral blood mononuclear cells were determined with a prominence liquid chromatograph connected to a triple-quadruple mass spectrometer. Polymorphisms in the thymidylate synthase (TS), methylenetetrahydrofolate reductase (MTHFR), dihydrofolate reductase (DHFR), reduced folate carrier 1 (RFC1; SLC19A1), and cyclin D1 (CCND1) genes were determined by direct sequencing using an ABI Prism 3100 genetic analyzer or Fluidigm's Biomark system. The Mann-Whitney test, rank analysis of variance (with Bonferroni's adjusted post hoc comparisons), and logistic regression were used for the inferential analyses. Thirty-three stavudine-treated patients were enrolled in this cross-sectional study. d4T-TP intracellular levels were 11.50 fmol/10(6) cells (interquartile range [IQR] = 8.12 to 13.87 fmol/10(6) cells) in patients with a high-expression TS genotype (2/3G, 3C/3G, and 3G/3G), whereas in those with a low-expression TS genotype (2/2, 2/3C, and 3C/3C), they were 21.40 fmol/10(6) cells (IQR = 18.90 to 27.0 fmol/10(6) cells) (P < 0.0001). Polymorphisms in the MTHFR, DHFR, RFC1, and CCND1 genes did not influence the intracellular concentration of d4T-TP. d4T-TP levels were independently associated with the TS genotype (low versus high expression; odds ratio [OR] = 86.22; 95% confidence interval [CI] = 8.48 to nonestimable; P = 0.0023). The low-expression TS genotype was associated with the development of HIV/highly active antiretroviral therapy-associated lypodystrophy syndrome (HALS) (OR = 14.0; 95% CI = 2.09 to 108.0; P = 0.0032). Our preliminary data show that polymorphisms in the thymidylate synthase gene are strongly associated with d4T-TP intracellular levels and with development of HALS.

Characterization of L-cysteine capped CdTe quantum dots and application to test Cu(II) deficiency in biological samples from critically ill patients.

The catalytic activity of copper ion gives, from the physiological point of view, a central role in many biological processes. Variations in the composition and location of cellular copper have been addressed given their physiological and pathological consequences. In this paper L-cysteine capped CdTe quantum dots is used for the fluorimetric determination of Cu(II) in biological samples from healthy individuals and patients admitted to the Intensive Care Units (ICU). An acceptable homogeneity in the CdTe QDs size has been obtained with an average value of 3 nm. No significant alterations in the spectral properties were observed for 2 months when stored in vacutainers at 6°C and a concentration of approximately 2 µM. Data from oxidative stress markers such superoxide dismutase, total antioxidant capacity and DNA damage can be correlated with a Cu(II) deficiency for the ICU patients as measured by flame-atomic absorption spectroscopy (FAAS) and inductively coupled plasma source mass spectrometry (ICP-MS). Aqueous solutions 0.3 µM of L-cysteine capped CdTe QDs in MOPS buffer (6 mM, pH 7.4) used at 21°C in the range 15-60 min after preparation of the sample for the measurements of fluorescence gives contents in Cu(II) for erythrocytes in good agreement with those obtained in FAAS and ICP-MS but the comparative ease of use makes the fluorimetric technique more suitable than the other two techniques for routine analysis.