High performance liquid chromatographic assay for the determination of Protease Inhibitors (PIs) and Non-nucleoside Reverse Transcriptase Inhibitors (NNRTIs) in human plasma.

Objective: To develop and validate a high performance liquid chromatography (HPLC) method for simultaneous quantitative determination of five HIV protease inhibitors (PIs): indinavir (IDV), lopinavir (LPV), nelfinavir (NFV), ritonavir (RTV), saquinavir (SQV), and two non-nucleoside reverse transcriptase inhibitors (NNRTIs): nevirapine (NVP), and efavirenz (EFV) in human plasma. Methods: A sample of 200 µL of plasma and an internal standard were extracted with tert-butyl methyl ether. The compounds were separated on a reversed-phase C18 column with gradient phase of 25 mM phosphate buffer (pH 4.9) and acetonitrile. The limit of quantation, accuracy, precision, specificity, stability and recovery were tested. Results: The lower limit of quantitation for all drugs was 75 ng/mL. The standard curve was linear in the range of 75 ng/mL to 20,000 ng/mL. Intra-day and inter-day variability ranged from 0.1% to 2.4% and 0.3% to 4.1%, respectively. Accuracy ranged from 98.4%-102.4% for three quality controls (75, 100, and 1,000 ng/mL) for all drugs measured. The extraction recovery ranged from 98.7%-101.3%. Conclusion: This method provides a simple, accurate, and precise method for monitoring of plasma concentrations of five PIs and two NNRTIs in the case of weak economy and out of date instrumental limitations.

Diagnostic accuracy of perfusion CT in differentiating brain abscess from necrotic tumor.

OBJECTIVE: To evaluate the diagnostic accuracy of perfusion computed tomography (CTP) in differentiating between brain abscess and necrotic tumor. MATERIAL AND METHOD: Prospective study was performed in patients suspected of a space taking lesion in the brain. CTP was done at the suspected levels and post-processing measurement of cerebral blood volume (CBV), cerebral blood flow (CBF), mean transit time (MTT), and permeability surface index (PS) were evaluated at ring enhanced area, central non-enhanced area, edema and contralateral normal brain. RESULTS: Seventeen patients with 21 lesions were studied. Of the 21 lesions, 12 were abscess and nine were tumors. By comparing means, only MTT at the ring enhanced area showed statistically significant difference between brain abscess and tumor (p = 0.009, 95% CI = 1.403 to 4.900). When ratio of CBV, CBF and MTT of the ring enhanced area and contralateral normal brain were analyzed (CBVr, CBFr, MTTr respectively), there were significant differences of CBVr and CBFr between the two groups (p = 0.003, 95% CI = -4.266 to -1.051 and p = 0.006, 95% CI = -9.934 to -1.969 respectively). With the threshold of CBVr more than or equal to 1.5 and CBFr more than or equal to 1, the sensitivity, specificity, positive predictive value, negative predictive value, and accuracy for diagnosis of tumor were 100%, 75%, 75%, 100%, and 85.7% respectively. CONCLUSION: The CTP was shown to be useful in differentiating brain abscess from tumor. With CBVr less than 1.5, tumor can be excluded.

Guidelines for antiretroviral therapy in HIV-1 infected adults and adolescents: the recommendations of the Thai AIDS Society (TAS) 2008.

BACKGROUND: More than 100,000 patients have been treated, since the implementation of the National Universal Coverage for antiretroviral therapy (ART) in Thailand Although there are several comprehensive guidelines available internationally, there is a need to have guidelines that can be implemented in Thailand. MATERIAL AND METHOD: The guidelines were developed by a panel of 17 members who are the experts on HIV research and/or HIV patient care and appointed without incentive by the Thai AIDS Society (TAS). The recommendations were based on evidences from the published studies and availability of antiretroviral agents. Published studies that are relevant and applicable to Thailand in particular have been taken into consideration. RESULTS: The recommendations include: when to start ART; what to start; how to monitor the therapy; adverse effects and its management; diagnosis of treatment failure; and antiretroviral treatment options in patients with treatment failure. ART in special circumstances, i.e., patients with co-infection of tuberculosis or hepatitis B virus, is also included Appropriate level of CD4+ T-cell count to start ART among Thai patients has been considered carefully. The authors recommend to start ART at CD4+ T-cell count < 200 cells/mm3. CONCLUSION: ART should be initiated in adults and adolescents HIV-1 infected patients with a history of HIV-related illness or AIDS or with a CD4+ T-cell count <200 cells/mm3. For treatment-naive patients, the preferred initial therapy is a non-nucleoside reverse transcriptase inhibitor (NNRTI)-based regimen. CD4' T-cell count and viral load should be monitored for at least twice and once a year, respectively. Proper management of antiretroviral-related toxicity and enhancement of adherence are crucial for the long-term success of ART.

Rapid determination of nevirapine in human plasma by high- performance liquid chromatography.

Objective: To develop and validate a high-performance liquid chromatography (HPLC) method for the determination of nevirapine in human plasma. Methods: A plasma sample and an internal standard were extracted with tert.-butyl methyl ether and determined nevirapine concentration by HPLC method. The limit of quantitation (LOQ), accuracy, precision, specificity, stability and recovery were tested for method validation. Results: Standard curve was linear in the range 0.1 ตg/mL to 20 ตg/mL. The limit of quantitation was 0.1 ตg/ml. Coefficients of variation (CV) of intraday and inter-day precision were less than 4 %. Accuracy was range from 97-101%. The extraction recovery was range from 94-112 %. Conclusion: A rapid, sensitive and specific HPLC method was developed and can be used for determination of plasma nevirapine concentration in adult and pediatric patients infected with HIV.

Vaccination against hepatitis B virus: are Thai medical students sufficiently protected?

Medical students are frequently at risk of being infected by hepatitis B virus (HBV) via occupational exposure to infected blood or body fluids. In 2002, the Faculty of Medicine, Siriraj Hospital provided screening tests for HBV serology to all medical students for a vaccination campaign against the infection. There were 1,165 medical students tested. Eight hundred and eleven (69.6%) students had immunity by previous vaccination, but more importantly 212 (18.2%) had no immunity and required vaccination. Most of the students who needed to be vaccinated were in the pre-clinical year (82.5%). Moreover, the students in the pre-clinical year who had previous vaccination had a 2.2 times greater risk of having negative anti-HBs than the students in the clinical year (OR = 2.2, 95% CI = 1.4-3.5). This is because they might have been vaccinated when they were young and the antibody waned overtime.

Estimated economic losses of hospitalized AIDS patients at Siriraj Hospital from January 2003 to December 2003: time for aggressive voluntary counseling and HIV testing.

This retrospective study was performed to explore the pattern of adult HIV-infected patients admitted to Siriraj Hospital from January 2003 to December 2003 and estimated the economic losses of these patients. Two hundred and forty four medical records were available for review. The proportion of male to female was 2 to 1. Mean age of patients was 36.64 +/- 9.72 years. The mean CD4 count among 112 patients was 82.79 +/- 96.49 cell/mm3. One hundred and twenty four (50.82%) were newly diagnosed of HIV infection. The three most common opportunistic infections were Tuberculosis (42.62%), Pneumocystis carinii pneumonia (14.75%), and cryptococcosis (13.11%). The mean duration of admission was 15.72 +/- 15.11 days. The mean expense per admission was 38,194.58 +/- 32,354.86 Baht. Fifty four patients (22.13%) died during admission. The mean income of these patients was 3,903.5 +/- 3,841.42 baht per month. The estimated economic losses of 54 patients who died during admission including medical care expense and income losses due to premature death was 69,769,739.32 baht. However, the expected medical expense of antiretroviral medications in these 54 patients if they had been diagnosed earlier and their lives had been saved would have been 42,214,608 baht. Therefore, vigorous voluntary counseling and HIV testing in patients aged 13-70 years when they have any risk factors for HIV infection regardless of symptoms might be more cost effective than diagnosis when they get sick.

Safety and efficacy of a simplified fixed-dose combination of stavudine, lamivudine and nevirapine (GPO-VIR) for the treatment of advanced HIV-infected patients: a 24-week study.

OBJECTIVE: To determine the efficacy and safety of the fixed-dose combination of stavudine (d4T), lamivudine (3TC) and nevirapine (NVP) in the treatment of antiretroviral naive HIV-infected Thai adults. PATIENTS AND METHOD: An open-label, single arm trial was conducted Baseline clinical assessment and blood test was done on 10, antiretroviral naive HIV-infected patients, who then received a fixed dose combination of d4T, 3TC and NVP (GPO- VIR, Thai Government Pharmaceutical Organization, Bangkok, Thailand). Nevirapine was given as 200 mg once daily for the first 2 weeks. The patients were followed up at 2, 4, 8, 12 and 24 weeks. A CD4 cell count and HIV-RNA assay were done at 12 and 24 weeks. RESULTS: One hundred and one patients were enrolled The mean baseline CD4 cell count and mean HIV RNA were 58.7 (57.7) cells/mm3 and 5.3 (0.5) log10, copies/mL respectively. At week 24th, the mean decrease in log HIV RNA was 3.6 (0.7) log10 copies/mL [P < 0.001; 95% confidence interval (CI), 2.70-3.03]. Eighty one (80.2%) patients had HIV RNA < 400 copies/mL by intention-to-treat analysis (ITT) and 97.6% had HIV RNA < 400 copies/mL by on-treatment analysis (OT). Sixteen (84.2%) patients with baseline HIV RNA < or = 100,000 copies/mL and 65 (82.3%) patients with baseline HIV RNA > 100,000 copies/mL had viral load < 400 copies/mL by ITT (P = 0.842; 95% CI, -20.9%-16.2%). Sixteen (94.1%) patients with baseline HIV RNA < or = 100,000 copies/mL and 65 (98.5%) patients with baseline HIV RNA > 100,000 copies/mL had viral load < 400 copies/mL by OT (P = 0.295; 95% CI, -25.5%-3.8%). The mean CD4 cell count at week 24 was 155.1 (89.0) cells/mm3 (range 13-402). The mean increase in CD4 cell count from baseline was 96.5 (63.5) cells/ mm3 (P < 0.001). A total of 12% of the patients receiving d4T + 3TC + NVP developed skin rashes. Grade 3 or 4 hepatotoxicity was recognized in 7% of the patients. CONCLUSION: Fixed-dose combination of d4T + 3 TC + NVP (GPO- VIR) is safe, well tolerated and effective in increasing CD4 cell counts and suppression of HIV RNA at 24 weeks in advanced HIV-infected patients in Thailand.

HIV infection/acquired immunodeficiency syndrome at Siriraj Hospital, 2002: time for secondary prevention.

The authors retrospectively reviewed the medical records of HIV/AIDS patients who were admitted to the medical service, Siriraj Hospital from January 1, 2002 through December 31, 2002. Demographics, CD4 lymphocyte counts, discharge diagnoses, the incidence of Pneumocystis carinii pneumonia (PCP), cerebral toxoplasmosis and cryptococcosis in patients who received and did not receive appropriate chemoprophylaxis against those opportunistic infections when indicated, and outcome of the patients were collected. Three hundred medical records of 286 HIV/AIDS patients were available for review. One hundred and seventy two patients (60.1%) were male. Mean age of the patients was 36.8 +/- 9.91 years (range 14-74). The mean CD4 lymphocyte count that was determined in 165 patients was 74.7 +/- 134.21 cells/mm3 (range 0-894). Of the 300 admissions, 36 per cent were newly diagnosed HIV infection. Only 23 (7.7%) patients had received antiretroviral drugs at the time of hospitalization. The leading HIV-related diseases were tuberculosis (29.3%), Pneumocystis carinii pneumonia (18.7%), and cryptococcosis (15.7%). The rest of them included cytomegalovirus diseases (6.3%), lymphoma (6.3%), Salmonella bacteremia (6%), cerebral toxoplasmosis (5.7%), cryptosporidiosis (5.3%), disseminated Mycobacterium avium complex infection (1.0%), extrapulmonary histoplasmosis (1.0%), Candida esophagitis (1.0%), progressive multifocal leukoencephalopathy (1.0%), and rhodococcosis (0.7%). Among those for whom HIV infection was established and chemoprophylaxis for PCP, cerebral toxoplasmosis and cryptococcosis were indicated, 9.8 per cent vs 28.2 per cent, 3.6 per cent vs 5.1 per cent, and 10 per cent vs 15.2 per cent of whom received and did not receive the appropriate chemoprophylaxis developed PCP, cerebral toxoplasmosis and cryptococcosis respectively. One hundred and ninety (63.3%) patients were alive at discharge, 84 (28.0%) had died, 21 (7%) were referred to other hospitals, and 5 (1.7%) left hospital against medical advice. The mortality rate in newly diagnosed HIV and in known HIV without antiretroviral treatment were comparable but much lower in known HIV-infected patients who received antiretroviral therapy. Secondary prevention by detection of HIV-infected patients while they are asymptomatic and providing them with appropriate chemoprophylaxis against specific opportunistic infections as well as appropriate antiretroviral treatment would decrease morbidity, mortality, and improve the quality of life of HIV-infected patients in Thailand.

Multidrug resistant mycobacterium tuberculosis infections in patients with acquired immunodeficiency syndrome at Siriraj Hospital.

Bachground : The increase in the prevalence of multidrug-resistant tuberculosis (MDR-TB) has been alarming globally. HIV and Mycobacterium tuberculosis co-infection is not uncommon. The mortality rate of MDR-TB it self is generally high. We describe 21 patients with multidrug-resistant Mycobacterium tuberculosis infection (MDR-TB), who had acquired immunodeficiency syndrome (AIDS). Methods: Information concerning all patients with AIDS and M. tuberculosis co-infection treated at Siriraj Hospital was collected from January 1, 1998, through June 30, 2001 which included clinical presentation, laboratory data, chest x-ray findings, treatment and outcome. Results: Six hundred and forty nine isolates of M. tuberculosis were tested for antimycobacterial susceptibility. 58 isolates (8.9%) were multidrug-resistant. Twenty one patients (36%) had AIDS. Of these, seven patients (33.3%) had primary drug resistant tuberculosis and 15 (66.7%) had secondary drug resistant tuberculosis. The mortality rate of MDR-TB with AIDS was 71.4%. The most common clinical presentations were fever (61.9%), cough (47.6%), and peripheral lymphadenopathy (33.3%). Twenty four percent had pulmonary disease only, 52% had both pulmonary and extrapulmonary diseases. The mean CD4 lymphocyte count was 38.8+45.77 cells/?L. Thirty three percent of the isolates were resistant to isoniazid and rifampicin only, 43% were resistant to 3 drugs and 24% to 4 drugs. Conclusion: The prevalence of MDR-TB was 8.9% ( 58 of 649 cases) in Siriraj Hospital from January 1, 1998 through June 30, 2001. Twenty-one of these patients (36%) had AIDS. Patients with MDR-TB were more likely to have disseminated disease and low CD4 lymphocyte counts. The mortality rate was high. All isolates of M. tuberculosis should be tested for antimycobacterial susceptibility to guide treatment and improve the outcome.