The instrument for this application is a state-of-the-art mass spectrometer, an API5000 from Applied Biosystems, for accurate, reproducible, and sensitive measurement of drugs and small biological molecules in support of NIH-funded clinical pharmacology research programs mainly investigating the treatment of human immunodeficiency virus (HIV). Multiple researchers from the University of Colorado at Denver and Health Sciences Center and National Jewish Research and Medical Center are proposed to share the instrument. The primary justification for this instrument is an urgent need for the highest level of analytical sensitivity possible. This need arises from a commonality among the various research projects, reproducible quantitation of femtomolar concentrations of intracellular nucleoside-analog-triphosphate concentrations in cells from patients taking these agents. ? ? The investigators in this application possess two important characteristics that assure a positive and productive environment for the shared instrument into the foreseeable future. 1. The researchers from the Antiviral Pharmacology Laboratory, where the instrument is proposed to be housed, bring significant and direct technical and managerial experiences to the operation of such an instrument, and 2. All investigators have ties to an NIH-funded University of Colorado-Center for AIDS research (UC-CFAR) program grant. The primary function of the UC-CFAR is to provide infrastructure and resources for a sharing and collaborative AIDS research environment. The UC-CFAR views this application favorably as an opportunity to build the pharmacology resource for the UC-CFAR investigators. The UC-CFAR has committed significant funds for personnel to manage the instrument. Further institutional funds have been committed by the Department of Pharmaceutical Sciences for a long-term maintenance contract. With these commitments, three years of personnel and service contracts are fully covered by the institution. A cost recovery fee plan guarantees the financial viability for the instrument after the three years covered by the institution. ? ? In this revised application we show that the API5000 instrument is 17- to 61-fold more sensitive than the instruments currently available to the shared users for our research compounds. We show that, without this sensitivity increase, we are impeded from moving forward with new research initiatives for the study intracellular nucleoside analog-triphosphate in patients. These novel and innovative research initiatives include measurement of intracellular nucleoside analog triphosphates within CD4 purified cells (the cells infected by HIV) and lymph tissue cells (the tissue where HIV resides) in HIV infected patients. These advances would address our short-term and long-term objectives of elucidating the disposition of nucleoside analogs in vivo to enable the most informed and rational utilization of this drug class in patients. ? ? ?
| Zheng, Jia-Hua; Rower, Caitlin; McAllister, Kevin et al. (2016) Application of an intracellular assay for determination of tenofovir-diphosphate and emtricitabine-triphosphate from erythrocytes using dried blood spots. J Pharm Biomed Anal 122:16-20 |
| Zheng, Jia-Hua; Guida, Louis A; Rower, Caitlin et al. (2014) Quantitation of tenofovir and emtricitabine in dried blood spots (DBS) with LC-MS/MS. J Pharm Biomed Anal 88:144-51 |
| Bushman, Lane R; Kiser, Jennifer J; Rower, Joseph E et al. (2011) Determination of nucleoside analog mono-, di-, and tri-phosphates in cellular matrix by solid phase extraction and ultra-sensitive LC-MS/MS detection. J Pharm Biomed Anal 56:390-401 |
