The Plasma, Whole Blood and Intracellular Concentrations of Antiretroviral Agents in South African Children Receiving Combination Antiretroviral Therapy with and without Concomitant Antitubercular Treatment

 

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dc.contributor.advisor McIlleron, Helen en_ZA
dc.contributor.advisor Smith, PJ en_ZA
dc.contributor.author Ren, Yuan en_ZA
dc.date.accessioned 2014-07-28T18:20:14Z
dc.date.available 2014-07-28T18:20:14Z
dc.date.issued 2009 en_ZA
dc.identifier.citation Ren, Y. 2009. The Plasma, Whole Blood and Intracellular Concentrations of Antiretroviral Agents in South African Children Receiving Combination Antiretroviral Therapy with and without Concomitant Antitubercular Treatment. University of Cape Town. en_ZA
dc.identifier.uri http://hdl.handle.net/11427/3297
dc.description.abstract Background: Tuberculosis (TB) is the most common opportunistic infection in children with human immunodeficiency virus (HIV) infection in developing countries, and co-treatment for HIV infection and TB is frequently indicated. Efavirenz and lopinavir/ritonavir (ratio 1:1) as part of antiretroviral therapy are used in combination with rifampicin-based antitubercular treatment in South African TB/HIV co-infected children. Adult studies show that concomitant rifampicin significantly reduces efavirenz and lopinavir plasma concentrations. However, the pharmacokinetics (PK) of efavirenz and lopinavir/ritonavir are poorly characterized in children, especially African children and no study has evaluated the effect of rifampicin-based antitubercular treatment on efavirenz and lopinavir/ritonavir plasma concentrations in children. Although therapeutic drug monitoring (TDM) is recommended in selected patients (including young children and patients receiving concomitant antitubercular treatment), TDM is seldom available in resource-constrained countries. There is an urgent need to develop a field friendly method which requires small volumes of blood, and inexpensive processing and storage conditions. Furthermore, because HIV replicates in the cells, efavirenz and lopinavir need to penetrate into these infected cells to inhibit viral replication. Therefore, directly measurement of intracellular concentrations of these drugs in HIV-infected children could provide better understanding of drug exposure at the action site. It is also important to evaluate the effects of frequently co-administered drugs on intracellular accumulation of efavirenz and lopinavir. Objectives: 1) To evaluate efavirenz and lopinavir/ritonavir plasma concentrations and determine the effects of rifampicin on efavirenz and lopinavir/ritonavir PK in HIV-infected African children with and without rifampicin-based antitubercular treatment. 2) To develop and validate the dried blood spot (DBS) method as an alternative to conventional plasma methods of drug concentration measurement in TDM. 3) To evaluate in vivo intracellular concentrations of efavirenz and lopinavir/ritonavir in HIV-infected children with and without concomitant antitubercular treatment. 4) To determine the in vitro modulation effects on the intracellular accumulation of efavirenz IV and lopinavir in human peripheral blood mononuclear cells (PBMCs) by drug efflux protein inhibitors, as well as frequently co-administered rifampicin and ritonavir (at low dose; as pharmacoenhancer). Methods: 1) Plasma efavirenz and lopinavir/ritonavir concentrations were measured by validated liquid chromatography/tandem mass spectrometry (LC/MS/MS) method in TB/HIV co-infected children during and after rifampicin-based antitubercular treatment as well as in a group of controls (HIV-infected children without TB). Children in the efavirenz study (n= 30) were receiving standard doses of efavirenz as part of antiretroviral treatment. Trough concentrations (Cmin) of efavirenz were estimated by extrapolation of the log-linear concentration-time line to 24 hours after the previous dose. Children in the lopinavir/ritonavir study were receiving additional ritonavir (lopinavir: ritonavir ratio 1:1) during antitubercular treatment (n= 15), and standard doses of lopinavir/ritonavir (LPV/r; ratio 4:1) after antitubercular treatment, and in controls (n= 15). The PK of lopinavir and ritonavir were characterized from concentration-time curves using WinNonlin version 4.1 by non-compartmental analysis. 2) Aliquots of 50 μ L of whole blood from the efavirenz and lopinavir/ritonavir studies were dried onto filter paper. The drug concentrations were analyzed using validated LC/MS/MS method. The effects of high temperature and direct sunlight on the stabilities of these antiretroviral drugs in DBS samples were tested. 3) Intracellular concentrations of efavirenz, lopinavir and ritonavir were measured in trough concentrations of 11 TB/HIV co-infected children using a validated LC/MS/MS method. Six children were receiving double dose of LPV/r (4:1) with concomitant rifampicin; 5 children were receiving standard doses of efavirenz with rifampicin-based antitubercular treatment, 3 of them had intracellular concentrations measured again after completing rifampicin-based antitubercular treatment. 4) in vitro intracellular accumulation of efavirenz and lopinavir were measured in human PBMCs in the absence and presence of P-glycoprotein inhibitors (verapamil at 50 μ M, V furosemide at 50 μ M and cyclosporine A at 20 μ M) and frequently co-administered drugs at levels representing the average concentrations found in patients (ritonavir at 5 mg/L and rifampicin at 4 mg/L). The concentrations of efavirenz and lopinavir in PBMCs were determined by LC/MS/MS. Results and Conclusions: 1) The co-administration of rifampicin did not significantly reduce efavirenz estimated Cmin concentrations. A high proportion of children with and without concomitant antitubercular treatment had sub-therapeutic efavirenz concentrations despite being correctly dosed according to the manufacturer's instructions, raising concerns about the adequacy of current efavirenz dosing recommendations in children. The lopinavir key PK parameter, Cmin, was not significantly different in same group of children during and after rifampicin-based antitubercular treatment or compared to HIV-infected children without tuberculosis. The recommended minimum therapeutic concentration was achieved in 87% of children during antitubercular treatment and in 92% without concomitant antitubercular treatment. Therefore, in the context of limited options, LPV/r with additional ritonavir (ratio 1:1) is an acceptable approach to treat young children receiving concomitant rifampicin-based antitubercular treatment, although safety remains a concern and hepatic alanine transaminase levels should be monitored regularly. 2) Plasma and DBS concentrations of efavirenz, lopinavir and ritonavir were strongly correlated. The median (interquartile range, IQR) DBS/plasma concentration ratios for efavirenz, lopinavir and ritonavir were 0.93 (IQR 0.83, 1.08), 0.73 (IQR 0.61, 0.90) and 1.05 (IQR 0.74, 1.21), respectively. PK parameters of efavirenz and ritonavir were closely similar between DBS and plasma; whereas lopinavir pre-dose and Cmin (at 12 hours after lopinavir intake) concentrations were 16% lower in DBS samples. The 3 antiretroviral drugs in DBS samples were stable at 37 deg C for 7 days and with exposure to direct sunlight for 2 hours. DBS can be used as an alternative field-friendly method for efavirenz, lopinavir and ritonavir concentration monitoring. However, pre-dose and Cmin concentrations of lopinavir in DBS samples need to be increased by 16% when used to predict plasma concentrations. VI 3) In vivo median intracellular/plasma concentration ratios for efavirenz, lopinavir and ritonavir amongst 11 TB/HIV co-infected children during antitubercular treatment were 0.91 (IQR 0.54, 1.19), 0.22 (IQR 0.09, 0.31) and 4.17 (IQR 1.30, 7.33), respectively. Two children had efavirenz intracellular/plasma concentration ratios during vs. after antitubercular treatment: 1.00 vs. 0.61 and 0.27 vs. 0.79. 4) Furosemide significantly increased efavirenz and lopinavir accumulation in healthy human PBMC samples by 1.2- 1.5 fold. Whereas, neither verapamil nor cyclosporin A had significant effects on efavirenz or lopinavir intracellular accumulation. Despite being an inducer of P-glycoprotein, rifampicin increased the accumulation of both efavirenz and lopinavir to different extents in all 3 PBMC samples. The low-dose ritonavir (at the concentration found in HIV-infected patients) had no effect on intracellular accumulation of efavirenz and lopinavir at therapeutic concentrations. en_ZA
dc.language.iso eng
dc.subject.other Clinical Pharmacology en_ZA
dc.title The Plasma, Whole Blood and Intracellular Concentrations of Antiretroviral Agents in South African Children Receiving Combination Antiretroviral Therapy with and without Concomitant Antitubercular Treatment en_ZA
dc.type Doctoral Thesis
uct.type.publication Research en_ZA
uct.type.resource Thesis en_ZA
dc.publisher.institution University of Cape Town
dc.publisher.faculty Faculty of Health Sciences en_ZA
dc.publisher.department Division of Clinical Pharmacology en_ZA
dc.type.qualificationlevel Doctoral
dc.type.qualificationname PhD en_ZA
uct.type.filetype Text
uct.type.filetype Image
dc.identifier.apacitation Ren, Y. (2009). <i>The Plasma, Whole Blood and Intracellular Concentrations of Antiretroviral Agents in South African Children Receiving Combination Antiretroviral Therapy with and without Concomitant Antitubercular Treatment</i>. (Thesis). University of Cape Town ,Faculty of Health Sciences ,Division of Clinical Pharmacology. Retrieved from http://hdl.handle.net/11427/3297 en_ZA
dc.identifier.chicagocitation Ren, Yuan. <i>"The Plasma, Whole Blood and Intracellular Concentrations of Antiretroviral Agents in South African Children Receiving Combination Antiretroviral Therapy with and without Concomitant Antitubercular Treatment."</i> Thesis., University of Cape Town ,Faculty of Health Sciences ,Division of Clinical Pharmacology, 2009. http://hdl.handle.net/11427/3297 en_ZA
dc.identifier.vancouvercitation Ren Y. The Plasma, Whole Blood and Intracellular Concentrations of Antiretroviral Agents in South African Children Receiving Combination Antiretroviral Therapy with and without Concomitant Antitubercular Treatment. [Thesis]. University of Cape Town ,Faculty of Health Sciences ,Division of Clinical Pharmacology, 2009 [cited yyyy month dd]. Available from: http://hdl.handle.net/11427/3297 en_ZA
dc.identifier.ris TY - Thesis / Dissertation AU - Ren, Yuan AB - Background: Tuberculosis (TB) is the most common opportunistic infection in children with human immunodeficiency virus (HIV) infection in developing countries, and co-treatment for HIV infection and TB is frequently indicated. Efavirenz and lopinavir/ritonavir (ratio 1:1) as part of antiretroviral therapy are used in combination with rifampicin-based antitubercular treatment in South African TB/HIV co-infected children. Adult studies show that concomitant rifampicin significantly reduces efavirenz and lopinavir plasma concentrations. However, the pharmacokinetics (PK) of efavirenz and lopinavir/ritonavir are poorly characterized in children, especially African children and no study has evaluated the effect of rifampicin-based antitubercular treatment on efavirenz and lopinavir/ritonavir plasma concentrations in children. Although therapeutic drug monitoring (TDM) is recommended in selected patients (including young children and patients receiving concomitant antitubercular treatment), TDM is seldom available in resource-constrained countries. There is an urgent need to develop a field friendly method which requires small volumes of blood, and inexpensive processing and storage conditions. Furthermore, because HIV replicates in the cells, efavirenz and lopinavir need to penetrate into these infected cells to inhibit viral replication. Therefore, directly measurement of intracellular concentrations of these drugs in HIV-infected children could provide better understanding of drug exposure at the action site. It is also important to evaluate the effects of frequently co-administered drugs on intracellular accumulation of efavirenz and lopinavir. Objectives: 1) To evaluate efavirenz and lopinavir/ritonavir plasma concentrations and determine the effects of rifampicin on efavirenz and lopinavir/ritonavir PK in HIV-infected African children with and without rifampicin-based antitubercular treatment. 2) To develop and validate the dried blood spot (DBS) method as an alternative to conventional plasma methods of drug concentration measurement in TDM. 3) To evaluate in vivo intracellular concentrations of efavirenz and lopinavir/ritonavir in HIV-infected children with and without concomitant antitubercular treatment. 4) To determine the in vitro modulation effects on the intracellular accumulation of efavirenz IV and lopinavir in human peripheral blood mononuclear cells (PBMCs) by drug efflux protein inhibitors, as well as frequently co-administered rifampicin and ritonavir (at low dose; as pharmacoenhancer). Methods: 1) Plasma efavirenz and lopinavir/ritonavir concentrations were measured by validated liquid chromatography/tandem mass spectrometry (LC/MS/MS) method in TB/HIV co-infected children during and after rifampicin-based antitubercular treatment as well as in a group of controls (HIV-infected children without TB). Children in the efavirenz study (n= 30) were receiving standard doses of efavirenz as part of antiretroviral treatment. Trough concentrations (Cmin) of efavirenz were estimated by extrapolation of the log-linear concentration-time line to 24 hours after the previous dose. Children in the lopinavir/ritonavir study were receiving additional ritonavir (lopinavir: ritonavir ratio 1:1) during antitubercular treatment (n= 15), and standard doses of lopinavir/ritonavir (LPV/r; ratio 4:1) after antitubercular treatment, and in controls (n= 15). The PK of lopinavir and ritonavir were characterized from concentration-time curves using WinNonlin version 4.1 by non-compartmental analysis. 2) Aliquots of 50 &Icirc;¼ L of whole blood from the efavirenz and lopinavir/ritonavir studies were dried onto filter paper. The drug concentrations were analyzed using validated LC/MS/MS method. The effects of high temperature and direct sunlight on the stabilities of these antiretroviral drugs in DBS samples were tested. 3) Intracellular concentrations of efavirenz, lopinavir and ritonavir were measured in trough concentrations of 11 TB/HIV co-infected children using a validated LC/MS/MS method. Six children were receiving double dose of LPV/r (4:1) with concomitant rifampicin; 5 children were receiving standard doses of efavirenz with rifampicin-based antitubercular treatment, 3 of them had intracellular concentrations measured again after completing rifampicin-based antitubercular treatment. 4) in vitro intracellular accumulation of efavirenz and lopinavir were measured in human PBMCs in the absence and presence of P-glycoprotein inhibitors (verapamil at 50 &Icirc;¼ M, V furosemide at 50 &Icirc;¼ M and cyclosporine A at 20 &Icirc;¼ M) and frequently co-administered drugs at levels representing the average concentrations found in patients (ritonavir at 5 mg/L and rifampicin at 4 mg/L). The concentrations of efavirenz and lopinavir in PBMCs were determined by LC/MS/MS. Results and Conclusions: 1) The co-administration of rifampicin did not significantly reduce efavirenz estimated Cmin concentrations. A high proportion of children with and without concomitant antitubercular treatment had sub-therapeutic efavirenz concentrations despite being correctly dosed according to the manufacturer's instructions, raising concerns about the adequacy of current efavirenz dosing recommendations in children. The lopinavir key PK parameter, Cmin, was not significantly different in same group of children during and after rifampicin-based antitubercular treatment or compared to HIV-infected children without tuberculosis. The recommended minimum therapeutic concentration was achieved in 87% of children during antitubercular treatment and in 92% without concomitant antitubercular treatment. Therefore, in the context of limited options, LPV/r with additional ritonavir (ratio 1:1) is an acceptable approach to treat young children receiving concomitant rifampicin-based antitubercular treatment, although safety remains a concern and hepatic alanine transaminase levels should be monitored regularly. 2) Plasma and DBS concentrations of efavirenz, lopinavir and ritonavir were strongly correlated. The median (interquartile range, IQR) DBS/plasma concentration ratios for efavirenz, lopinavir and ritonavir were 0.93 (IQR 0.83, 1.08), 0.73 (IQR 0.61, 0.90) and 1.05 (IQR 0.74, 1.21), respectively. PK parameters of efavirenz and ritonavir were closely similar between DBS and plasma; whereas lopinavir pre-dose and Cmin (at 12 hours after lopinavir intake) concentrations were 16% lower in DBS samples. The 3 antiretroviral drugs in DBS samples were stable at 37 deg C for 7 days and with exposure to direct sunlight for 2 hours. DBS can be used as an alternative field-friendly method for efavirenz, lopinavir and ritonavir concentration monitoring. However, pre-dose and Cmin concentrations of lopinavir in DBS samples need to be increased by 16% when used to predict plasma concentrations. VI 3) In vivo median intracellular/plasma concentration ratios for efavirenz, lopinavir and ritonavir amongst 11 TB/HIV co-infected children during antitubercular treatment were 0.91 (IQR 0.54, 1.19), 0.22 (IQR 0.09, 0.31) and 4.17 (IQR 1.30, 7.33), respectively. Two children had efavirenz intracellular/plasma concentration ratios during vs. after antitubercular treatment: 1.00 vs. 0.61 and 0.27 vs. 0.79. 4) Furosemide significantly increased efavirenz and lopinavir accumulation in healthy human PBMC samples by 1.2- 1.5 fold. Whereas, neither verapamil nor cyclosporin A had significant effects on efavirenz or lopinavir intracellular accumulation. Despite being an inducer of P-glycoprotein, rifampicin increased the accumulation of both efavirenz and lopinavir to different extents in all 3 PBMC samples. The low-dose ritonavir (at the concentration found in HIV-infected patients) had no effect on intracellular accumulation of efavirenz and lopinavir at therapeutic concentrations. DA - 2009 DB - OpenUCT DP - University of Cape Town LK - https://open.uct.ac.za PB - University of Cape Town PY - 2009 T1 - The Plasma, Whole Blood and Intracellular Concentrations of Antiretroviral Agents in South African Children Receiving Combination Antiretroviral Therapy with and without Concomitant Antitubercular Treatment TI - The Plasma, Whole Blood and Intracellular Concentrations of Antiretroviral Agents in South African Children Receiving Combination Antiretroviral Therapy with and without Concomitant Antitubercular Treatment UR - http://hdl.handle.net/11427/3297 ER - en_ZA


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