Robust Reconstitution of Tuberculosis-Specific Polyfunctional CD4+ T-Cell Responses and Rising Systemic Interleukin 6 in Paradoxical Tuberculosis-Associated Immune Reconstitution Inflammatory Syndrome.

BACKGROUND: The immunopathogenesis of paradoxical tuberculosis-associated immune reconstitution inflammatory syndrome (TB-IRIS) remains unclear. We determined the association between pathogen-specific T-cell responses and development of paradoxical TB-IRIS on antiretroviral therapy (ART). METHODS: This study was nested within a prospective cohort study of HIV-infected patients with active pulmonary tuberculosis and baseline CD4 counts ≤125 cells/µL initiating ART. T-cell immune activation (CD38, HLA-DR, and PD-1 expression), phenotype, and polyfunctional pathogen-specific cellular immune responses prior to and 4 weeks after ART initiation were determined by flow cytometry. Patients with TB-IRIS were compared to non-IRIS controls using χ(2) and rank-sum tests and logistic regression. RESULTS: TB-IRIS patients and controls had similar CD4 counts, levels of T-cell-associated immune activation, frequencies of T-cell memory subsets, and frequencies of interferon gamma (IFN-γ(+))/interleukin 2 (IL-2(+))/tumor necrosis factor alpha (TNF-α(+)) CD4(+) T-cells prior to ART initiation. After ART initiation, cellular immune activation and T-cell subsets also were similar in TB-IRIS patients and controls. In contrast, TB-IRIS patients had significantly greater early increases in the frequency of tuberculosis-specific polyfunctional IFN-γ(+)/IL-2(+)/TNF-α(+) CD4(+) T-cells on ART (P = .02); each quartile increase in the percentage of these cells was independently associated with a 2.8-fold increased risk of TB-IRIS (95% confidence interval, 1.1 to 7.5-fold). In a secondary analysis, patients with TB-IRIS had rapid, concomitant increases in tuberculosis-specific adaptive immune responses and interleukin 6 (IL-6) levels, whereas controls with similarly rapid increases in cellular immune function had IL-6 levels that tended to decrease on ART. CONCLUSIONS: Rapid expansion of tuberculosis-specific polyfunctional CD4(+) T-cell responses, likely linked to increases in IL-6, is associated with development of paradoxical TB-IRIS.

Immunological profiling of tuberculosis-associated immune reconstitution inflammatory syndrome and non-immune reconstitution inflammatory syndrome death in HIV-infected adults with pulmonary tuberculosis starting antiretroviral therapy: a prospective observational cohort study.

BACKGROUND: Patients co-infected with advanced HIV and tuberculosis are at risk of tuberculosis-associated immune reconstitution inflammatory syndrome (IRIS) and death soon after initiation of antiretroviral therapy (ART). Tuberculosis-associated IRIS has been associated with quicker recovery of cellular immune responses after ART initiation and early mortality with slower recovery of these responses. We aimed to assess whether patients who have these outcomes have distinct immunological profiles before and after ART initiation. METHODS: We undertook this prospective cohort study at 22 public clinics and the main public hospital in Gaborone, Botswana, in ART-naive adults (aged ≥21 years) with advanced HIV (CD4 cell counts ≤125 cells per µL) and pulmonary tuberculosis. We obtained data for clinical variables and for levels of 29 plasma biomarkers, quantified by Luminex assay. We classified patients as having tuberculosis-associated IRIS, early mortality, or survival without a diagnosis of tuberculosis-associated IRIS (controls), on the basis of outcomes recorded in the 6 months after ART initiation. We used rank-sum or χ(2) tests, and logistic regression with odds ratios (OR) and 95% CIs, to assess the association between variables measured before and 4 weeks after ART initiation with death and tuberculosis-associated IRIS, compared with controls. FINDINGS: Between Nov 12, 2009, and July 3, 2013, we enrolled 201 participants. 31 (15%) patients left the study before ART initiation, leaving 170 (85%) patients for analysis. Patients with tuberculosis-associated IRIS had reduced pre-ART concentrations of several pro-inflammatory biomarkers, including interleukin (IL)-6 (adjusted OR per 1 log10 increase 0·40 [95% CI 0·18-0·89]). However, patients with early death had increased pre-ART concentrations of inflammatory biomarkers, including monocyte chemoattractant protein-1 (adjusted OR 9·0 [95% CI 1·0-80·0]) and tumour necrosis factor (TNF)α (7·8 [1·1-55·2]). At week 4 after ART initation, tuberculosis-associated IRIS was independently associated with greater increases in several inflammatory biomarkers, including IL-6 (adjusted OR 1·7 [95% CI 1·2-2·5]) and TNFα (1·5 [1·0-2·2]), versus controls. Death was likewise associated with greater increases in systemic inflammatory markers, including granulocyte colony-stimulating factor (adjusted OR 2·8 [95% CI 1·3-6·1]), IL-12p40 (1·8 [1·0-3·4]), and IL-15 (2·0 [1·1-3·7]), versus controls. However, changes in CD4 cell count during ART, which were similar between controls and patients with tuberculosis-associated IRIS (p=0·45), were substantially lower in patients who died (p=0·006). INTERPRETATION: Distinct immunological profiles before and after ART initiation characterise patients with advanced HIV and tuberculosis who have tuberculosis-associated IRIS and death. Interventions that decrease inflammation while promoting cellular immune recovery during ART should be considered in patients co-infected with HIV and tuberculosis. FUNDING: National Institutes of Health and the Penn Center for AIDS Research.

Early immunologic failure is associated with early mortality among advanced HIV-infected adults initiating antiretroviral therapy with active tuberculosis.

BACKGROUND: The relationship between antiretroviral therapy (ART) response and early mortality after ART initiation is unknown. We hypothesized that early mortality is associated with decreased early immunologic response to ART. METHODS: We prospectively determined the association between changes in plasma human immunodeficiency virus type 1 (HIV-1) RNA and CD4(+) T-cell counts (CD4 count) after 4 weeks of ART and early mortality in adults with pulmonary tuberculosis and pre-ART CD4 counts ≤ 125 cells/µL. Purified protein derivative (PPD)-specific immune recovery was determined by interferon-γ enzyme-linked immunosorbent spot assays. Levels of interleukin 6, C-reactive protein, and soluble CD14 were assessed. Patients with CD4 count and viral load values at baseline and week 4 were analyzed using multiple logistic regression. RESULTS: Early immunologic response, but not pre-ART CD4 counts or virologic response, was related to early mortality (8 [interquartile range {IQR}, -18 to 43] vs 68 [IQR, 24-131] cells/µL, P = .002). In a logistic regression model, every 20 cells/µL increase in the CD4 count from baseline to week 4 was independently associated with a 40% reduction in the odds of death (odds ratio, 0.59 [95% confidence interval, .41-.87]). PPD-specific immune recovery was lower, whereas levels of immune activation were higher, among deaths. CONCLUSIONS: Early immunologic failure despite virologic suppression is associated with early mortality after ART initiation in advanced HIV/tuberculosis.

Isoniazid-resistant tuberculous meningitis, United States, 1993-2005.

To determine patient characteristics associated with isoniazid resistance in cases of tuberculous meningitis, we conducted a cross-sectional study by using data from the US National Tuberculosis Surveillance System during 1993-2005. Foreign-born patients were more likely to be infected with an isoniazid-resistant strain.

Isoniazid resistance and death in patients with tuberculous meningitis: retrospective cohort study.

OBJECTIVE: To determine whether initial isoniazid resistance is associated with death during the treatment of tuberculous meningitis. DESIGN: Retrospective cohort study. SETTING: National Tuberculosis Surveillance System at the Centers for Disease Control in the United States. PARTICIPANTS: Patients with a clinical diagnosis of tuberculous meningitis, reported to the National Tuberculosis Surveillance System between 1 January 1993 and 31 December 2005. MAIN OUTCOME MEASURE: All cause mortality during antituberculous treatment. RESULTS: Between 1993 and 2005, 1896 patients had a clinical diagnosis of tuberculous meningitis and positive cultures from any site. In 123 (6%) of these patients, isoniazid resistance was present on initial susceptibility testing. The unadjusted association between initial isoniazid resistance and subsequent death among these 1896 patients did not reach statistical significance (odds ratio 1.38, 95% confidence interval 0.94 to 2.02). However, among 1614 patients with positive cerebrospinal fluid cultures, a significant unadjusted association was found between initial isoniazid resistance and subsequent death (odds ratio 1.61, 1.08 to 2.40). This association increased after adjustment for age, race, sex, and HIV status (odds ratio 2.07, 1.30 to 3.29). CONCLUSIONS: Isoniazid resistance on initial susceptibility testing was associated with subsequent death among cases of tuberculous meningitis with positive cerebrospinal fluid cultures. Randomised controlled trials are needed to evaluate the optimal empirical regimen for treating patients with tuberculous meningitis who are at high risk for both initial isoniazid resistance and poor clinical outcomes.

Safety and immunogenicity of a live attenuated varicella vaccine in VZV-seropositive HIV-infected adults.

BACKGROUND: Herpes-zoster is common in HIV-infected patients in spite of antiretroviral therapy. We evaluated the safety and immunogenicity of a live attenuated varicella-zoster virus (VZV) vaccine as a candidate for protecting HIV-infected adults against herpes-zoster. RESULTS: Sixty-seven HIV-infected and 15 uninfected subjects, 18 to 65 years old, were enrolled. Adverse events were minor and similar in HIV-infected vaccine and placebo recipients. At 12 weeks after the 2(nd) dose of vaccine the magnitude of each measure of VZV CMI increased significantly in healthy controls. In HIV-infected vaccinees, VZV RCF significantly increased and ELISPOT showed a positive trend. None of VZV CMI measures significantly increased in HIV-infected placebo recipients. The immunogenicity of the vaccine did not correlate with the nadir CD4 cells of HIV-infected subjects. METHODS: HIV-infected adults with CD4 > or = 400 cells/microL and plasma HIV RNA <1,000/mL were randomly assigned to receive two doses of VZV vaccine or placebo 12 weeks apart. HIV-uninfected age-matched controls also received two doses of vaccine. VZV-specific cell-mediated immunity (CMI) was measured at baseline and after vaccination using responder cell frequency (RCF), lymphocyte proliferation, and IFNgamma ELISPOT. CONCLUSIONS: Two doses of varicella vaccine were safe in HIV-infected subjects with CD4 > or = 400 cells/microL, but were only modestly immunogenic.

Tuberculous meningitis in HIV-infected individuals.

HIV-infected individuals are at increased risk for all forms of extrapulmonary tuberculosis, including tuberculous meningitis. This risk is increased at more advanced levels of immunosuppression. The time interval between onset of symptoms and presentation to medical care may vary widely, and consequently individuals may present with acute or chronic meningitis. The clinical presentation of tuberculous meningitis in HIV-infected individuals is more likely to include an altered level of consciousness, cranial imaging is more likely to show cerebral infarctions, and the yield of culture of cerebrospinal fluid may also be greater. Given that delayed initiation of therapy is a strong predictor of mortality in cases of tuberculous meningitis, clinicians must consider tuberculosis in the differential diagnosis of the HIV-infected individual with acute or chronic lymphocytic meningitis. Additional treatment considerations for HIV-infected individuals include the timing of initiation of antiretroviral therapy, the potential for drug-drug interactions, and the role of adjunctive corticosteroid therapy.

Cryptococcus and lymphocytic meningitis in Botswana.

We retrospectively reviewed microbiological data from a tertiary care hospital in Botswana, and found that Cryptococcus neoformans was cultured from 15% (193/1307) of all cerebrospinal fluid (CSF) specimens submitted for analysis, making it the most common diagnosed cause of meningitis in this population. Moreover, almost 70% of CSF samples with significant lymphocytosis did not yield a pathogen, suggesting that many causes of lymphocytic meningitis go undiagnosed.

Serological evidence of HIV-associated infection among HIV-1-infected adults in Botswana.

In industrialized countries, it is recommended that adults with human immunodeficiency virus type 1 (HIV-1) infection undergo baseline screening for pathogens that might cause latent or active infections, such as syphilis, hepatitis B, hepatitis C, infection due to Toxoplasma gondii, and cytomegalovirus infection. A paucity of data exist from sub-Saharan Africa describing the prevalence of these pathogens. We report data for HIV-1-infected adults referred for initiation of highly active antiretroviral therapy in Botswana.

Gene transfer in humans using a conditionally replicating lentiviral vector.

We report findings from a clinical evaluation of lentiviral vectors in a phase I open-label nonrandomized clinical trial for HIV. This trial evaluated the safety of a conditionally replicating HIV-1-derived vector expressing an antisense gene against the HIV envelope. Five subjects with chronic HIV infection who had failed to respond to at least two antiviral regimens were enrolled. A single i.v. infusion of gene-modified autologous CD4 T cells was well tolerated in all patients. Viral loads were stable, and one subject exhibited a sustained decrease in viral load. CD4 counts remained steady or increased in four subjects, and sustained gene transfer was observed. Self-limiting mobilization of the vector was observed in four of five patients. There is no evidence for insertional mutagenesis after 21-36 months of observation. Immune function improved in four subjects. Lentiviral vectors appear promising for gene transfer to humans.

Management of potential neurocysticercosis in patients with HIV infection.

In patients with human immunodeficiency virus, the diagnosis of neurocysticercosis can be complex, and the current diagnostic criteria may not apply. We report 3 cases and suggest including the CD4+ T lymphocyte count as an important factor in the proper diagnosis and treatment of patients with human immunodeficiency virus and potential neurocysticercosis.

IL-13 acutely augments HIV-specific and recall responses from HIV-1-infected subjects in vitro by modulating monocytes.

We show in this study that acute exposure of PBMCs derived from HIV-infected subjects to IL-13 results in increased recall T cell lymphoproliferative responses against HIV-1 p24 (n = 30, p < 0.0001) and other recall Ags (influenza, n = 43, p < 0.0001; purified protein derivative tuberculin, n = 6, p = 0.0299). This effect is due to a mechanism that acutely targets APC function in the adherent monocyte subset, as shown by the expansion of CD4(+) T cell responses following coculture of IL-13-treated enriched CD14(+) monocytes with donor-matched enriched CD4(+) T cells and Ag. Exposure to IL-13 over 18-72 h resulted in a significant enhancement of monocyte endocytosis (n = 11, p = 0.0005), CD86 expression (n = 12, p = 0.001), and a significant decrease in spontaneous apoptosis (n = 8, p = 0.008). Moreover, IL-13 exposure induced a significant decrease of significantly elevated constitutive levels of PBMC-secreted TNF-alpha (n = 14, p < 0.001) and IL-10 (n = 29, p < 0.001) within 18 h of exposure ex vivo, also reflected by decreased gene expression in the adherent cell population. Our data show that IL-13 is able to acutely enhance the function of the CD14(+) cell subset toward supporting Ag-specific cell-mediated responses in chronic HIV-1 infection.

Clinical, microbiological, and immunological characteristics in HIV-infected subjects at risk for disseminated Mycobacterium avium complex disease: an AACTG study.

The clinical, microbiologic, and immunologic parameters in HIV-infected subjects first presenting with disseminated Mycobacterium avium complex (DMAC) were determined. Four HIV-positive groups not yet on DMAC treatment were enrolled: 19 subjects with CD4 lymphocyte counts < or =50/microl thought to have DMAC on clinical grounds; 18 subjects newly found to have a positive blood culture for MAC; 25 asymptomatic controls (CD4 cell counts < or =50); and 25 asymptomatic controls (CD4 counts 100-250/microl). Outcome measures include comparisons between groups for clinical characteristics; results of cultures from blood, marrow, and gastrointestinal and respiratory tracts; immunological markers from staining of marrow and flow cytometry of circulating lymphocytes; and cytokine production of PBMCs. Only 21% of the 19 patients entered on suspicion of having DMAC grew MAC from blood or marrow. Neither clinical presentation nor laboratory tests differentiated those culture-positive from those culture-negative patients. However, prior PCP or multiple other opportunistic infections were more common in the DMAC group. MAC was isolated from 82% of marrow and 50% of blood specimens from the DMAC group. Respiratory or gastrointestinal colonization was present in 36% of DMAC subjects, but only 5% of non-DMAC subjects with CD4 counts <50 cells/microl. CD8+ cells were more frequent in bone marrow, and CD4 cells recognizing MAC antigen were more frequent in blood from DMAC subjects vs. controls. Results suggest an early stage of tissue dissemination preceding persistent bacteremia, and mucosal entry without persistence of colonization. MAC-specific T cell responses apparently develop and persist during DMAC, but are dysfunctional or too infrequent to prevent persistence.

A tale of 2 epidemics: the intersection between obesity and HIV infection in Philadelphia.

BACKGROUND: Obesity and HIV infection are ongoing epidemics in the United States. Obesity predisposes to diabetes and cardiovascular disease, which are complications also associated with HIV and/or its treatment. OBJECTIVE: To determine the prevalence and risk factors for overweight and obesity in HIV-infected individuals. DESIGN AND SETTING: Retrospective cross-sectional study in which 1689 patients enrolled in the University of Pennsylvania Center for AIDS Research Adult/Adolescent Database at 1 university hospital clinic, 2 affiliated practices, and 1 Veterans Administration clinic in Philadelphia had demographic, social, and medical data collected prospectively since 1999. PARTICIPANTS: Body mass index (BMI) data were available for 1669 HIV-infected subjects: 78% were men, and 60% were African American. The median CD4 count was 381 cells/microL, 47% of subjects had a viral load <400 copies/mL, and 9% of subjects were treatment naive. MAIN OUTCOME MEASURES: The prevalence and risk factors for overweight (BMI: 25-29.9 kg/m) and obesity (BMI>or=30 kg/m) in HIV-infected subjects. RESULTS: Obesity and overweight were more prevalent than wasting (14%, 31%, and 9%, respectively; P<0.0005), but they were not more common than in the general population. Although women and men were equally overweight (30% vs. 31%), women were more obese than men (28% vs. 11%; P<0.001). Among women, African American race (odds ratio [OR]=1.8, 95% confidence interval [CI]: 1.1-2.9) and a CD4 count>or=200 cells/microL (OR=2.8, 95% CI: 1.6-4.9) were associated with overweight and obesity. Among men, only a CD4 count>or=200 cells/microL (OR=1.6, 95% CI: 1.04-2.4) was associated with increased BMI. In men and women, smoking was associated with decreased obesity and overweight (OR=0.59, 95% CI: 0.47-0.74 and OR=0.65, 95% CI: 0.43-0.98, respectively). Age, income, employment, education, past or current intravenous drugs, being on HIV treatment, and viral load were not associated with obesity in the multivariate model. There was a positive correlation between BMI and total cholesterol, triglycerides, and glucose. CONCLUSION: Obesity is more common than wasting in this therapeutic era. Women, particularly those of African American race, are at high risk. Obesity might add to metabolic abnormalities associated with HIV or its treatment and contribute to morbidity, as patients with HIV live longer.

Stavudine entry into cerebrospinal fluid after single and multiple doses in patients infected with human immunodeficiency virus.

STUDY OBJECTIVE: To establish the pharmacokinetics of stavudine within the cerebrospinal fluid (CSF) of patients infected with human immunodeficiency virus (HIV). DESIGN: Pharmacokinetic study. SETTING: General clinical research center. PATIENTS: Thirty-six patients infected with HIV; 21 were receiving long-term stavudine therapy, 15 were not (single-dose treatment group). INTERVENTION: After an overnight fast, all patients received a single dose of stavudine 40 mg. Fifteen patients in the long-term treatment group and all 15 patients in the single-dose treatment group were randomized to undergo lumbar puncture 2, 4, or 6 hours after dosing (five patients for each time point from each group). The six other patients in the long-term treatment group underwent lumbar puncture 0 or 8 hours after dosing. MEASUREMENTS AND MAIN RESULTS: Serum stavudine concentrations were obtained just before dosing, 1 hour after dosing (approximate peak), and at the time of lumbar puncture. The CSF was also analyzed for cell counts, protein, and glucose levels. The mean peak serum stavudine concentration in the long-term treatment group was estimated to be 580.7 ng/ml (2.59 micromol/L), occurring approximately 1.3 hours after dosing. The CSF concentrations over 0-8 hours were 0.0-109.9 ng/ml (0.00-0.49 micromol/L) with an overall mean of 51.6 ng/ml (0.23 micromol/L). Mean peak CSF concentration was estimated to be 62.8 ng/ml (0.28 micromol/L), occurring 4.7 hours after dosing. For the 15 patients not taking stavudine, both the serum and the CSF estimated peaks were significantly lower than those of the long-term group: 475.3 ng/ml (2.12 micromol/L) and 40.4 ng/ml (0.18 micromol/L), respectively. However, time to peak was similar at 1.2 hours and 5.0 hours, respectively. In both groups, no correlation was found between CSF and baseline or peak serum stavudine concentrations, CSF white blood cell count, baseline CD4 + lymphocyte count, or plasma viral load. CONCLUSION: Mean CSF stavudine concentrations equaled or exceeded the mean concentration producing 50% of the maximal effect in vivo (EC 50 ) for HIV. The CSF concentrations were higher in the stavudine-experienced patients, indicating that concentrations rise with progressive doses until steady state is reached.

Plasmid vaccination of stable HIV-positive subjects on antiviral treatment results in enhanced CD8 T-cell immunity and increased control of viral "blips".

Antiviral therapy prolongs suppression of viral replication and allows for significant immune reconstitution but has not been effective in eradicating reservoirs of virus, which produce resurgent viremia when highly active antiretroviral therapy (HAART) is discontinued. Immune-based therapy may provide an additional antiviral effect. We vaccinated stable HIV-positive patients on HAART with an HIV plasmid vaccine to determine safety, immunogenicity, and therapeutic potential. Volunteers received a combination of two HIV DNA plasmid constructs, which drive expression of env/rev and gag/pol genes. The vaccine was well tolerated with no toxicity. CD4 and CD8 lymphocyte counts did not change significantly among volunteers. CD8 MHC class I-restricted responses to HIV antigens were assayed. Eight of 13 vaccinees responded after vaccination with detectable ELISpot result. Importantly, we observed a difference in viral detection events in vaccinated compared to control patients. Three out of the five placebo recipients had "viral blips" (transient elevations of HIV RNA) during follow-up (10/49 assays) while these were only present in one of 13 vaccinees on one occasion (1/130 assays; p<0.04). The decrease in the frequency of transient viremia and failure suggests that DNA immunization with CD8-generating vaccines in HAART-controlled HIV-positive subjects may have therapeutic potential.

Regulatory considerations for novel gene therapy products: a review of the process leading to the first clinical lentiviral vector.

This review is intended to exemplify the roles and responsibilities of the two agencies under the Department of Health and Human Services, the National Institutes of Health and the Food and Drug Administration, that have oversight for human gene transfer clinical protocols, as seen through our experience of bringing a first-in-its-class lentiviral vector to clinical trials. In response to the changing circumstances in gene therapy research between 1999 and 2002, the concerns of these agencies regarding gene therapy have been evolving. This review provides an overview of the major safety concerns regarding insertional oncogenesis, the generation of a replication- competent lentivirus (RCL), and vector mobilization thought to be related to lentiviral vectors, which had to be addressed during the regulatory review process before initiating the clinical trial. Specific monitoring assays to address these concerns were established to test for RCL generation, vector mobilization, persistence of vector-modified cells, and abnormal clonal expansion of modified cells. We hope to provide a basic understanding and appreciation of the regulatory process and major safety concerns, toward providing useful insight to those presently embarking on the development of clinical application of lentiviral vectors.

Differential virulence of Mycobacterium avium strains isolated from HIV-infected patients with disseminated M. avium complex disease.

The role that colonization with Mycobacterium avium plays in the development of disseminated disease is unclear. In this study, we determined whether all M. avium strains isolated from the portals of M. avium infection are capable of crossing the mucosal border and causing infection. The patients in this study were enrolled in AIDS Clinical Trial Group protocol 341. The patients were divided into 3 groups; 2 groups differed in their immunological and clinical risk for M. avium disease. A third group (n=22 patients) had culture-documented disseminated M. avium complex disease at the time of entry in the study. Eight of 22 patients had M. avium isolated from both a colonized site and blood or bone marrow specimens. All 8 patients had distinct M. avium strains; 2 patients had a polyclonal infection. The virulence properties of 13 strains were determined, including invasion of gastrointestinal cells and replication in macrophages. There were significant differences in the virulence properties, and these differences may provide insight into the interplay between microbial pathogenesis and host defense.