Some HIV protease inhibitors alter lamin A/C maturation and stability, SREBP-1 nuclear localization and adipocyte differentiation

OBJECTIVES: To study whether HIV protease inhibitors could induce nuclear lamina alterations in adipocytes as observed in a genetic form of lipodystrophy due to lamin A/C mutation. DESIGN: We have previously observed that indinavir (IDV) impairs adipocyte differentiation and sterol regulatory element-binding protein-1 (SREBP-1) nuclear localization in 3T3-F442A adipocytes. We compared here the effects of IDV with that produced by two other PIs, nelfinavir (NFV) and amprenavir (APV) on adipose conversion, cellular localization of SREBP-1, nuclear morphology, and maturation and stability of the lamina network. RESULTS: IDV and NFV, but not APV, altered adipose cell differentiation, as shown by lipid staining and protein expression of SREBP-1, CAAAT/enhancer binding protein (C/EBP)alpha and fatty acid synthase (FAS). In IDV-treated cells, 50-60 % of the nuclei could not accumulate SREBP-1. Twenty percent of these SREBP-negative nuclei were grossly dysmorphic, with blebs and prominent herniations, and showed an altered distribution of lamin A/C and lamin B. In IDV-treated cells, nuclear fragilization was shown by the abnormal extractibility of lamina proteins and SREBP-1, and the accumulation of prelamin A. NFV similarly altered lamin A/C maturation whereas APV was almost ineffective. CONCLUSIONS: We show in an adipose cell line that IDV and NFV induced alterations at the nuclear level by promoting defects in lamin A/C maturation, organization and stability. We suggest that these lamina network alterations might be responsible for SREBP-1 nuclear mislocalization therefore resulting in altered adipocyte differentiation.     

Determination of indinavir and nelfinavir trough plasma concentration efficacy thresholds according to virological response in HIV-infected patients

BACKGROUND: There is evidence to suggest a pharmacokinetic-pharmacodynamic relationship in HIV-infected patients receiving protease inhibitor (PI)-containing highly active antiretroviral therapy (HAART); however, the effective trough PI plasma concentrations achieved have not been precisely determined. METHODS: The relationship between HIV viral load and concomitant PI trough plasma concentration (C(trough)) was evaluated in 101 patients receiving at least 4 months of thrice daily indinavir (IDV)-containing (n=68) or nelfinavir (NFV)-containing (n=33) HAART. The more discriminating C(trough) efficacy thresholds were determined statistically for each PI by using the raw C(trough) and the time-corrected C(trough), using the precise delay since the last PI intake and the half-life of each PI. RESULTS: For IDV (P=0.002) and NFV (P=0.019) median C(trough) levels were higher in patients with undetectable viral load [0.23 mg/L (n=30) and 2.3 mg/L (n=16) respectively] than in patients with detectable viral load [0.11 mg/L (n=38) and 0.6 mg/L (n=17) respectively]. C(trough) levels of IDV (r=-0.45; P<0.0001) and NFV (r=-0.43; P=0.011) were correlated with the concomitant viral load. The more discriminating C(trough) efficacy thresholds were estimated statistically as 0.12 mg/L for IDV and 0.5 mg/L for NFV. When C(trough) values were time-corrected, the C(trough) efficacy thresholds, 8 h after the last intake, were 0.15 mg/L for IDV and 0.65 mg/L for NFV. CONCLUSIONS: These results support the importance of achieving minimal effective C(trough) to improve the virological efficacy of PI-containing HAART, and specify the target concentrations for IDV and NFV.     

The effects of cannabinoids on the pharmacokinetics of indinavir and nelfinavir

BACKGROUND AND OBJECTIVES: The use of cannabinoids for appetite stimulation and the management of wasting and antiretroviral side-effects has become a common practice in the care of HIV-infected individuals. We present pharmacokinetic data from a randomized placebo-controlled study designed to evaluate the metabolic effects of smoked marijuana and dronabinol in HIV-infected patients receiving indinavir (IDV) or nelfinavir (NFV). METHODS: Subjects on stable regimens containing IDV 800 mg every 8 h (n = 28) or NFV 750 mg three time a day (n = 34) were randomized to one of three treatment arms: 3.95% THC marijuana cigarettes, dronabinol 2.5 mg capsules or placebo capsules administered three times daily. Serial blood sampling was performed at baseline and on day 14 of treatment. The changes in NFV and IDV pharmacokinetics were measured as the median percentage change from baseline. RESULTS: At day 14, the 8-h area under the curve (AUC(8)) changed by -10.2% (P = 0.15), maximum concentration (C(max)) by -17.4% (P = 0.46), and minimum concentration (C(min)) by -12.2% (P = 0.28) for patients in the NFV marijuana arm (n = 11). Similar decreases had occurred by day 14 among patients in the IDV marijuana arm (n = 9): AUC8 had changed by -14.5% (P = 0.074), C(max) by -14.1% (P = 0.039), and C(min) by -33.7% (P = 0.65). CONCLUSION: Despite a statistically significant decrease in C(max) of IDV in the marijuana arm, the magnitude of changes in IDV and NFV pharmacokinetic parameters in the marijuana arm are likely to have no short-term clinical consequence. The use of marijuana or dronabinol is unlikely to impact antiretroviral efficacy.     

Regimen-dependent variations in adherence to therapy and virological suppression in patients initiating protease inhibitor-based highly active antiretroviral therapy

OBJECTIVE: To examine differences among four protease inhibitor (PI)-based drug regimens in adherence to therapy and rate of achievement of virological suppression in a cohort of antiretroviral-naive patients initiating highly active antiretroviral therapy (HAART). METHODS: Participants were antiretroviral-naive and were first dispensed combination therapy containing two nucleosides and a ritonavir (RTV)-boosted PI, or unboosted nelfinavir, between 1 January 2000 and 30 September 2003. Logistic regression analysis was used to examine associations between the prescribed PI and other baseline factors associated with being >90% adherent to therapy and then to determine the associations of prescribed drug regimen, adherence to therapy and baseline variables with the odds of achieving two consecutive viral loads of <500 HIV-1 RNA copies/mL. RESULTS A total of 385 subjects were available for analysis. Lopinavir (LPV)/RTV was prescribed for 168 patients (42% of total); 86 (22%) received indinavir (IDV)/RTV; 91 (24%) received nelfinavir (NFV) and 40 (10%) received saquinavir (SQV)/RTV. SQV/RTV-based HAART was associated with reduced adherence to therapy [odds ratio (OR)=0.40; 95% confidence interval (CI) 0.19-0.83]. In multivariate models, IDV/RTV (OR=0.45; 95% CI 0.22-0.92), SQV/RTV (OR=0.18; 95% CI 0.07-0.43) and NFV were associated with reduced odds of achieving virological suppression within 1 year in comparison to LPV/RTV-based therapy. For patients receiving NFV, adjusting for adherence (OR=0.73; 95% CI 0.36-1.47) rendered this association nonsignificant. CONCLUSION: Patients prescribed IDV/RTV, NFV or SQV/RTV were less likely to achieve virological suppression on their first regimen compared with patients prescribed LPV/RTV. Reduced adherence to these therapies only partly explained these observed differences.     

Genotypic and phenotypic cross-resistance patterns to lopinavir and amprenavir in protease inhibitor-experienced patients with HIV viremia

Genotypic correlates of reduced phenotypic susceptibility to amprenavir (APV) and lopinavir (LPV) were examined in 271 HIV isolates from 207 protease inhibitor (PI)-experienced subjects. All samples were from LPV-naive subjects; two were from APV-experienced subjects. Using a fold resistance (FR) of <2.5, 179 (66%) were APV susceptible. Using FRs of <2.5 and <10, 107 (39%) and 194 (72%), respectively, were LPV susceptible. The I84V mutation was the strongest APV resistance marker in PI-experienced subjects in both univariate and multivariate analyses, with an increased relative incidence (RI) of 6.9 with >2.5 FR. Mutations L10I (RI, 1.7), M46I (RI, 2.3), and L90M (RI, 1.9, but 65% linked with the I84V) were associated with decreased APV susceptibility in the univariate analysis (p < 0.001). Mutations L10I, G48V, I54T, I54V, and V82A were significantly associated with decreased LPV susceptibility (p < 0.001 for each) and had increased RIs of 2.2, 4.4, 13, 4.6, and 3.2, respectively. Decreased susceptibility to LPV (FR, >or=10) was significantly associated with prior exposure to the following PIs: ritonavir (RTV) (p < 0.001), saquinavir (SQV) (p < 0.001), nelfinavir (NFV) (p = 0.008), and indinavir (IDV) (p = 0.028). Decreased APV susceptibility (FR, >or=2.5) was significantly associated with prior exposure to RTV (p = 0.009), NFV (p = 0.003), and IDV (p = 0.021) but not with prior SQV (p = 0.103). These results suggest that APV and LPV have different cross-resistance mutation patterns that may help determine choice of PI therapy after therapy failure.     

Efficacy, tolerance, and pharmacokinetics of the combination of stavudine, nevirapine, nelfinavir, and saquinavir as salvage regimen after ritonavir or indinavir failure

The high rate of protease inhibitor treatment failure in clinical cohorts makes it necessary to define novel salvage therapies. In a prospective study of 31 HIV-infected patients included in a salvage regimen with stavudine, nevirapine, nelfinavir, and saquinavir, viral load decreased a median of 1.65 log(10) and 1.95 log(10) after 6 and 12 months of treatment, respectively, and 35 and 56% of patients had an HIV RNA level below 50 copies/ml at the same time points. At baseline, the mean number of mutations in the protease gene was 10 (2-19), and the V82A and L90M mutations were present in 54 and 21% of patients. The presence of the V82A mutation did not affect significantly the rate of response (36 vs. 38%), whereas the L90M mutation was associated with treatment failure (0 vs. 43%). Plasma trough levels of nelfinavir (NFV) and saquinavir (SQV), in a twice daily dosing regimen, were above the protein-corrected IC(95) in most patients despite the addition of an enzymatic inducer such as nevirapine, and peak levels were 2- and 5-fold increased with respect to standard doses. However, pharmacokinetics of saquinavir-hard gel capsule (SQV-hgc) did not improve significantly in the three times daily dosing regimen. In conclusion, the combination of stavudine, nevirapine, nelfinavir, and saquinavir increased plasma drug levels and produced an adequate virological response in patients who had failed indinavir or ritonavir therapy. This degree of response is not significantly decreased in the presence of genotypic mutations associated with indinavir/ritonavir (IDV/RTV) resistance.     

Protease inhibitor drug levels in the management of human immunodeficiency virus-1 antiretroviral therapy

In order to evaluate the relationship between protease inhibitor (PI) plasma concentrations and viral suppression in individuals receiving highly active antiretroviral therapy (HAART), plasma concentrations and area under the time concentration curve (AUC(0.5-4)) for 35 HIV-infected adults receiving their initial (or first salvage) nelfinavir- (NFV) or indinavir (IDV)-based HAART were studied. Two groups were evaluated: those who had achieved HIV-RNA suppression (HIV-RNA <500 copies/mL, group 1, n=21) and those who had achieved incomplete HIV-RNA suppression (HIV-RNA>500 copies/mL, group 2, n=14) at the time of study entry. NFV one-hour pre-dose concentrations were significantly higher in group 1 compared to group 2 (P=0.023). The NFV AUC(0.5-4) for group 1 approached significance (P=0.068). No significant differences in IDV concentrations or AUC(0.5-4) were found between group 1 and group 2. It is feasible to use PI drug level monitoring in the outpatient setting.     

MT1-MMP、MMP-2和TIMP-2基因在去卵巢大鼠成骨细胞中的表达

目的　通过观察去卵巢 (OVX)骨质疏松模型大鼠成骨细胞膜型基质金属蛋白酶 1(MT1 MMP)基因的表达 ,探讨绝经后骨质疏松的发病机制。　方法　对OVX大鼠与假手术大鼠进行骨密度和骨组织形态计量学测定。股骨远端行原位杂交检测骨组织MT1 MMP、MMP 2和TIMP 2mRNA表达 ,免疫组化检测骨组织MT1 MMP、MMP 2和TIMP 2蛋白质表达。　结果　OVX大鼠第 3、4腰椎骨密度 (0 14 4± 0 0 11) g cm2 、(0 14 3± 0 0 15 ) g cm2 较对照组 (0 15 8± 0 0 18)g cm2 、(0 16 2± 0 0 17) g cm2 明显减少 ,OVX组骨小梁面积 (9 5 8± 3 39) %、厚度 (40 85± 8 0 4 ) μm和数目 (2 30± 0 4 8)个 mm分别较对照组 (2 0 6 3± 4 8) %、(44 73± 6 8) μm、(4 6± 0 7)个 mm明显下降 ,OVX组骨小梁间隔 (5 85 8± 115 1) μm较对照组 (2 5 4 6± 4 8 0 ) μm明显增宽 ;成骨细胞MT1 MMPmRNA与蛋白质表达下调 ,而MMP 2和TIMP 2之间表达无差异。　结论　雌激素不足可使成骨细胞MT1 MMP基因表达减少 ,可能为绝经后骨质疏松的发病机制之一。     

Activation of p38 mitogen-activated protein kinase is required for osteoblast differentiation

p38 MAPK is a conserved subfamily of MAPKs involved in inflammatory response, stress response, cell growth and survival, as well as differentiation of a variety of cell types. In this report we demonstrated that p38 MAPK played an important role in osteoblast differentiation using primary calvarial osteoblast, bone marrow osteoprecursor culture, and a murine cell line, MC3T3-E1. We found that p38 MAPK was activated as calvarial osteoblast differentiates along with extracellular signal-regulated kinases (ERKs). When p38 MAPK is inhibited with a specific inhibitor, the expression of differentiation markers, such as alkaline phosphatase and mineral deposition, were significantly reduced. MC3T3-E1 cells expressing dominant negative p38 MAPK also displayed signs of delay in ALP and mineral deposition. Differentiation of the bone marrow osteoprecursors was also impeded by the p38 MAPK inhibitor, justified by the same markers. Yet the inhibitory effects observed in calvarial osteoblasts and bone marrow osteoprogenitor cells could be partially prevailed by bone morphogenetic protein-2. Inhibition of ERKs with a specific drug did not significantly affect osteoblast differentiation even though ERK1/2 were also activated during osteoblast differentiation. These results taken together indicate that p38 MAPK, but not ERKs, is necessary for osteoblast differentiation.     

A randomized trial of 2 different 4-drug antiretroviral regimens versus a 3-drug regimen,in advanced human immunodeficiency virus disease

To compare long-term virologic benefits of antiretroviral regimens in persons with advanced human immunodeficiency virus (HIV) disease, a randomized, open-label study was conducted of 517 subjects with no or limited previous experience with antiretroviral therapy. Subjects received lamivudine plus zidovudine and indinavir (indinavir group), efavirenz plus indinavir (efavirenz + indinavir group), or nelfinavir plus indinavir (nelfinavir + indinavir group) and were monitored for 2.1 years. Virologic failure was lower in the efavirenz + indinavir group (P=.04) and higher in the nelfinavir + indinavir group (P=.006), compared with that in the indinavir group. No difference in grade 3 or 4 adverse event rates in the efavirenz + indinavir group (P=.97) and a trend toward an increased rate in the nelfinavir + indinavir group (P=.07), compared with the indinavir group, were noted. A 4-drug regimen containing efavirenz plus indinavir resulted in a superior virologic response, whereas one containing nelfinavir plus indinavir resulted in an inferior response and a greater likelihood of toxicity.     

Membrane-type matrix metalloproteinase-1 (MT1-MMP) is down-regulated in estrogen-deficient rat osteoblast in vivo

Our previous study showed that estrogen stimulates membrane-type matrix metalloproteinases-1 (MT1-MMP) production in osteoblastic cells culture, but has no effect on MMP-2 and TIMP-2 synthesis. Osteoblast-derived MT1-MMP have been recently implied to play a role in bone metabolism by degrading tumor necrosis factor-alpha (TNF-alpha), resolving extracellular matrix and activating proMMP-2, which requires the process of activation mediated by MT1-MMP/tissue inhibitor of metalloproteinase (TIMP-2) complex on the cell surface. To investigate the mechanism of bone loss following estrogen deficiency, we examined the effects of estrogen on osteoblast synthesis of MT1-MMP, MMP-2 and TIMP-2. In situ hybridization and immunohistochemistry of rat bone samples were used to document the synthesis of MT1-MMP, MMP-2, and TIMP-2 mRNA and protein. Osteoblasts from distal femoral head showed an increase in the pattern of MT1-MMP mRNA and protein production in sham-operated controls and 17beta-estradiol (E2)-treated rats, compared with the ovariectomized group; the synthesis of MMP-2 and TIMP-2 mRNA and protein was unaffected. Our data show a down-regulation of MT1-MMP synthesis by osteoblast in vivo following estrogen withdrawal, and treatment with E2 resulted in induced MT1-MMP expression in vivo. There is evidence suggesting a role for MT1-MMP in the process of bone loss during the pathogenesis of osteoporosis.     

A randomized, comparative study of lamivudine plus stavudine, with indinavir or nelfinavir, in treatment-experienced HIV-infected patients

OBJECTIVE: To compare adherence and clinical outcome with two modalities of highly active antiretroviral therapy (HAART), in HIV-infected patients. DESIGN: Randomized, open-label, prospective study. SETTING: Tertiary care centre in Spain. PATIENTS: A total of 112 non-naive HIV-infected patients, recruited from March 1998 through August 1998, were studied. INTERVENTIONS: Triple drug therapy with stavudine and lamivudine, plus indinavir or nelfinavir. MAIN OUTCOME MEASURES: Adherence, side-effects, and immunological, virological, and clinical efficacy of treatment were assessed at 3-month intervals. RESULTS: After a median follow-up of 9 months, 32% of patients in the indinavir group versus 50% of those in the nelfinavir group showed adequate adherence in all clinical appointments (P= 0.0559). Adherence was superior in the nelfinavir group in every visit. After 6 months of treatment 48% of subjects in the indinavir group and 70% of those in the nelfinavir group exhibited adequate adherence (P= 0.0311). After 9 months 35% of patients in the indinavir group and 59% of those in the nelfinavir group showed adequate adherence (P= 0.0291). Side-effects provoked discontinuation of treatment in 34% of patients in the indinavir group and 12% of patients in the nelfinavir group (P= 0.0073). Immunological and virological efficacy were similar in both groups. CONCLUSIONS: Adherence to a HAART regimen with stavudine plus lamivudine plus nelfinavir was superior to a regimen with stavudine plus lamivudine plus indinavir. Side-effects provoked more discontinuation of treatment in the indinavir group than in the nelfinavir group.     

The use of calcium carbonate in nelfinavir-associated diarrhoea in HIV-1-infected patients

Objectives

To evaluate the efficacy of oral calcium supplements in HIV‐infected patients with nelfinavir (NFV)‐associated diarrhoea, and to investigate the influence on the pharmacokinetics of nelfinavir and the active metabolite M8.

Methods

An open‐label prospective trial with enrolment of 15 patients with NFV‐associated diarrhoea. Study subjects received either calcium carbonate or calcium gluconate/calcium carbonate in addition to highly active antiretroviral therapy (HAART), and were randomized to (i) calcium supplements for 14 days followed by 14 without calcium supplements, or (ii) 14 days without calcium supplements followed by calcium supplements for 14 days. Clinical endpoint was the severity of diarrhoea, graded and summarized for the specific 14 day‐period. In the pharmacokinetic evaluation concentrations of NFV and M8 were measured before morning dosing, and 3 h after dosing.

Results

Nine patients were treated with calcium carbonate, and six with calcium gluconate/calcium carbonate. In the paired analysis, neither of the groups had a significant improvement in diarrhoea score when treated with calcium supplements (P = 0.34 and 0.46, respectively). We found no significant differences in the concentrations of NFV and M8 between the calcium and control periods.

Conclusions

Oral calcium supplements did not significantly improve nelfinavir‐associated diarrhoea. In the pharmacokinetic analysis calcium supplements did not induce major alterations in plasma concentrations of NFV and M8.

     

Long-term effects of HIV-1 protease inhibitors on insulin secretion and insulin signaling in INS-1 beta cells

The mechanism by which chronic treatment with HIV (human immunodeficiency virus)-1 protease inhibitors leads to a deterioration of glucose metabolism appears to involve insulin resistance, and may also involve impaired insulin secretion. Here we investigated the long-term effects of HIV-1 protease inhibitors on glucose-stimulated insulin secretion from beta cells and explored whether altered insulin secretion might be related to altered insulin signaling. INS-1 cells were incubated for 48 h with different concentrations of amprenavir, indinavir, nelfinavir, ritonavir or saquinavir, stimulated with 20 mM d-glucose, and insulin determined in the supernatant. To evaluate insulin signaling, cells were stimulated with 100 nM insulin for 2 min, and insulin-receptor substrate (IRS)-1, -2 and Akt phosphorylation determined. Incubation for 48 h with ritonavir, nelfinavir and saquinavir resulted in impaired glucose-induced insulin secretion at 2.5, 5 and 5 microM respectively, whereas amprenavir or indinavir had no effects even at 20 and 100 microM respectively. The impaired insulin secretion by ritonavir, nelfinavir and saquinavir was associated with decreased insulin-stimulated IRS-2 phosphorylation, and, for nelfinavir and saquinavir, with decreased insulin-stimulated IRS-1 and Thr308-Akt phosphorylation. No such effects on signaling were observed with amprenavir or indinavir. In conclusion, certain HIV-1 protease inhibitors, such as ritonavir, nelfinavir and saquinavir, not only induce peripheral insulin resistance, but also impair glucose-stimulated insulin secretion from beta cells. With respect to the long-term effect on beta-cell function there appear to be differences between the protease inhibitors that may be clinically relevant. Finally, these effects on insulin secretion after a 48 h incubation with protease inhibitor were associated with a reduction of the insulin-stimulated phosphorylation of insulin signaling parameters, particularly IRS-2, suggesting that protease inhibitor-induced alterations in the insulin signaling pathway may contribute to the impaired beta-cell function.     

Effects of dihydrotestrone on osteoblast gene expression in osteopenic ovariectomized rats

Androgens stimulate bone formation, however, the precise mechanism of androgen action on osteoblasts remains to be elucidated. In this study, we defined the expression profile of osteoblast genes in ovariectomized rats with established osteopenia and their response to treatment with dihydrotestosterone (DHT). Twenty-four, 8-month-old female Sprague-Dawley rats were ovariectomized (ovx) and were administered vehicle, 40 mg, 80 mg, or 160 mg/kg body weight DHT at 15-weeks post-ovariectomy for 14 weeks. Alkaline phosphatase (ALP) messenger ribonucleic acid (mRNA) levels were increased at 29-weeks post-ovariectomy compared with preoperative rats (P < 0.05). In contrast, osteopontin and osteocalcin mRNA levels were unchanged. Treatment of osteopenic ovx rats with DHT for 14 weeks suppressed the ovariectomy-induced increase in ALP (P < 0.05) mRNA levels, independent of dose. These data suggest that androgens may act to inhibit the stimulation of the early stages of osteoblast development that occurs in the absence of estrogen and in states of low bone turnover.     

糖尿病大鼠钙化血管中Msx2和Wnt3a的表达

目的 研究糖尿病对血管钙化的影响及Msx2和wnt3a基因在钙化血管中的表达变化.方法 将48只雄性wistar大鼠随机分为四组:维生素D3和尼古丁诱导的单纯血管钙化组(n=12)、链脲佐菌素诱导的单纯糖尿病组(n=12)、链脲佐菌素联合维生素D3和尼古丁诱导的糖尿病合并血管钙化组(n=12)和正常雄性Wistar大鼠为正常对照组(n=12).测定大鼠血糖、血清胰岛素、总胆固醇和甘油三酯水平,以血管von Kossa染色、血管钙含量和碱性磷酸酶活性作为判断血管钙化程度的指标,测定大鼠血管中Msx2和wnt3a mRNA 的表达.结果 与正常对照组相比,单纯血管钙化组大鼠血管中Msx2和wnt3a mRNA 相对表达量有所升高(P<0.05),但血管钙含量和碱性磷酸酶活性无明显变化.与正常对照组及单纯血管钙化组相比,糖尿病合并血管钙化组大鼠的血管中,可见沿中膜弹力层内广泛分布的钙盐沉积,大鼠血管中钙含量和碱性磷酸酶活性以及血管内Msx2和wnt3amRNA 相对表达量明显增高(P<0.05).结论 糖尿病可以明显加速血管钙化的发生和发展.在糖尿病大鼠钙化血管中,骨形成过程中的转录因子Msx2和Wnt3a表达增高,提示血管钙化是一个类似于骨形成的过程,Msx2和Wnt3a 参与血管钙化病变的发生.     

Therapeutic drug monitoring of nelfinavir and indinavir in treatment-naive HIV-1-infected individuals

BACKGROUND: Both virological failure and the toxicity of HIV protease inhibitors have been related to interindividual variability of plasma drug concentrations. Therapeutic drug monitoring (TDM) offers the possibility to detect patients with drug concentrations outside therapeutic ranges, who can subsequently benefit from dose modifications. METHODS: ATHENA was a randomized controlled clinical trial. Subjects were randomly assigned to either a TDM group, in which the results of drug concentration measurements plus advice were reported to their treating physician, or to a control group for whom TDM results were not reported. This analysis refers to treatment-naive patients who started a regimen containing indinavir or nelfinavir before November 1999. FINDINGS: A total of 147 patients were randomly assigned: 92 to nelfinavir, 55 to indinavir. After one year of follow-up significantly fewer patients in the TDM group had discontinued nelfinavir or indinavir than in the control group: 17.4 versus 39.7%. This was mainly driven by a significantly lower rate of discontinuation because of virological failure in nelfinavir patients: 2.4% in the TDM group versus 17.6% in the control group, and by a non-significant difference in the rate of discontinuation because of toxicity in indinavir patients: 14.3% in the TDM group versus 29.6% in the control group. In a non-completer equals failure analysis of all randomized patients, the TDM group showed a significantly higher proportion of patients with a viral load below 500 copies after 12 months of treatment (78.2 versus 55.1%). INTERPRETATION: TDM of nelfinavir and indinavir in treatment-naive patients improves treatment response.