Mucosal-associated invariant T (MAIT) cells are activated in the gastrointestinal tissue of patients with combination ipilimumab and nivolumab therapy-related colitis in a pathology distinct from ulcerative colitis.

The aim of this study was to investigate the pathogenesis of combination ipilimumab and nivolumab-associated colitis (IN-COL) by measuring gut-derived and peripheral blood mononuclear cell (GMNC; PBMC) profiles. We studied GMNC and PBMC from patients with IN-COL, IN-treated with no adverse-events (IN-NAE), ulcerative colitis (UC) and healthy volunteers using flow cytometry. In the gastrointestinal-derived cells we found high levels of activated CD8+ T cells and mucosal-associated invariant T (MAIT) cells in IN-COL, changes that were not evident in IN-NAE or UC. UC, but not IN-C, was associated with a high proportion of regulatory T cells (Treg ). We sought to determine if local tissue responses could be measured in peripheral blood. Peripherally, checkpoint inhibition instigated a rise in activated memory CD4+ and CD8+ T cells, regardless of colitis. Low circulating MAIT cells at baseline was associated with IN-COL patients compared with IN-NAE in one of two cohorts. UC, but not IN-COL, was associated with high levels of circulating plasmablasts. In summary, the alterations in T cell subsets measured in IN-COL-affected tissue, characterized by high levels of activated CD8+ T cells and MAIT cells and a low proportion of Treg , reflected a pathology distinct from UC. These tissue changes differed from the periphery, where T cell activation was a widespread on-treatment effect, and circulating MAIT cell count was low but not reliably predictive of colitis.

Hepatitis C-specific effector and regulatory CD4 T-cell responses are associated with the outcomes of primary infection.

Clearance of primary hepatitis C virus (HCV) infection has been associated with strong and broadly targeted cellular immune responses. This study aimed to characterize HCV-specific CD4+ effector and regulatory T-cell numbers and cytokine production during primary infection. Antigen-specific CD4+ T-cell responses were investigated in a longitudinal cohort of subjects from pre-infection to postoutcome, including subjects who cleared [n=12] or became chronically infected [n=17]. A cross-sectional cohort with previously cleared, or chronic infection [n=15 for each], was also studied. Peripheral blood mononuclear cells were incubated with HCV antigens and surface stained for T-effector (CD4+CD25high CD134+CD39-) and T-regulatory (CD4+CD25high CD134+CD39+) markers, and culture supernatants assayed for cytokine production. Contrary to expectations, the breadth and magnitude of the HCV-specific CD4+ T-cell responses were higher in subjects who became chronically infected. Subjects who cleared the virus had HCV-specific CD4+ T-cell responses dominated by effector T cells and produced higher levels of IFN-γ, in contrast to HCV-specific CD4+ T-cell responses dominated by regulatory T cells and more IL-10 production in those who became chronically infected. Better understanding of the role of antigen-specific CD4+ T-cell responses in primary HCV will further define pathogenesis and help guide development of a preventative vaccine.

Group 2 innate lymphoid cells (ILC2s) are increased in chronic rhinosinusitis with nasal polyps or eosinophilia.

BACKGROUND: Chronic rhinosinusitis (CRS) is a heterogeneous disease with an uncertain pathogenesis. Group 2 innate lymphoid cells (ILC2s) represent a recently discovered cell population which has been implicated in driving Th2 inflammation in CRS; however, their relationship with clinical disease characteristics has yet to be investigated. OBJECTIVE: The aim of this study was to identify ILC2s in sinus mucosa in patients with CRS and controls and compare ILC2s across characteristics of disease. METHODS: A cross-sectional study of patients with CRS undergoing endoscopic sinus surgery was conducted. Sinus mucosal biopsies were obtained during surgery and control tissue from patients undergoing pituitary tumour resection through transphenoidal approach. ILC2s were identified as CD45(+) Lin(-) CD127(+) CD4(-) CD8(-) CRTH2(CD294)(+) CD161(+) cells in single cell suspensions through flow cytometry. ILC2 frequencies, measured as a percentage of CD45(+) cells, were compared across CRS phenotype, endotype, inflammatory CRS subtype and other disease characteristics including blood eosinophils, serum IgE, asthma status and nasal symptom score. RESULTS: 35 patients (40% female, age 48 ± 17 years) including 13 with eosinophilic CRS (eCRS), 13 with non-eCRS and 9 controls were recruited. ILC2 frequencies were associated with the presence of nasal polyps (P = 0.002) as well as high tissue eosinophilia (P = 0.004) and eosinophil-dominant CRS (P = 0.001) (Mann-Whitney U). They were also associated with increased blood eosinophilia (P = 0.005). There were no significant associations found between ILC2s and serum total IgE and allergic disease. In the CRS with nasal polyps (CRSwNP) population, ILC2s were increased in patients with co-existing asthma (P = 0.03). ILC2s were also correlated with worsening nasal symptom score in CRS (P = 0.04). CONCLUSION AND CLINICAL RELEVANCE: As ILC2s are elevated in patients with CRSwNP, they may drive nasal polyp formation in CRS. ILC2s are also linked with high tissue and blood eosinophilia and have a potential role in the activation and survival of eosinophils during the Th2 immune response. The association of innate lymphoid cells in CRS provides insights into its pathogenesis.

Concurrent nonavalent human papillomavirus (HPV) vaccination and immune stimulation with imiquimod to treat recalcitrant HPV-associated high grade vaginal intra-epithelial neoplasia.

This is the first report describing detailed T cell responses to viral-like proteins contained in an HPV specific vaccine given in combination with Imiquimod for treatment of persistent VAIN2/3. We postulate that stimulation of the innate immune system with Imiquimod and the specific CD4 and CD8T cell responses following HPV vaccination with Gardasil9@ combined to induce clinical remission in a woman with treatment-refractory disease.

Effects of primary HIV-1 infection on subsets of CD4+ and CD8+ T lymphocytes.

OBJECTIVE: To analyse changes in T-lymphocyte subsets in patients with primary HIV infection and to determine their specificity (and therefore their diagnostic utility) by comparing these changes with those seen in other acute illnesses as well as in HIV-uninfected patients. METHODS: T-lymphocyte subsets were analysed by two- and three-colour flow cytometry, and compared between HIV seroconverters (n = 16), HIV-infected (n = 18) and uninfected (n = 33) controls, patients with infectious mononucleosis (n = 7), and patients suspected clinically of having primary HIV infection but who were later found to be uninfected (referred to as HIV non-converters; n = 17). RESULTS: CD4+ lymphocyte counts were significantly lower in HIV seroconverters (mean, 444 x 10(6)/l) than in the HIV non-converters (793 x 10(6)/l; P = 0.003), HIV-seronegative controls (888 x 10(6)/l; P < 0.0001) and, to a lesser extent, those with mononucleosis (694 x 10(6)/l; P = 0.045). The reduction in CD4+ lymphocytes occurred in both the CD45RA+ (55%) and CD45R0+ (33%) subsets. CD8+ lymphocyte counts were significantly higher in HIV seroconverters (942 x 10(6)/l) than in HIV non-converters (570 x 10(6)/l; P = 0.003) and seronegative controls (467 x 10(6)/l; P < 0.0001), but significantly lower than in the mononucleosis group (3682 x 10(6)/l; P = 0.004). The CD8+ cells in the HIV seroconverters had increased coexpression of CD45R0, human leukocyte antigen (HLA)-DR, CD38 and CD11a/CD18. The mean CD4: CD8 ratio in the HIV seroconverters was 0.49, versus 1.52 in the non-converters (P < 0.0001), 2.08 in the seronegative patients (P < 0.0001) and 0.37 in the mononucleosis patients (P > 0.2). CONCLUSIONS: Primary HIV infection is characterized by a depletion of CD4+ lymphocytes, especially of the CD45RA+ phenotype, and by an increase in CD8+ lymphocytes with an activated phenotype; the latter was also seen in patients with infectious mononucleosis but not in HIV non-converters or HIV-seronegative patients. Patients suspected clinically of having primary HIV infection but with normal T-cell phenotype are less likely to have primary HIV infection. These phenotypic changes, as well as an inverted CD4: CD8 ratio, can readily distinguish patients with primary HIV infection from HIV-uninfected patients except those with infectious mononucleosis. Therefore, T-cell-subset enumeration may be useful in the diagnosis of primary HIV infection.

Long-term immunological response in HIV-1-infected subjects receiving potent antiretroviral therapy.

OBJECTIVE: To determine the long-term T-lymphocyte response to highly active antiretroviral therapy (HAART) and to define predictors of the immunological response. DESIGN: Cohort study, including 135 HIV-1-infected subjects at a city general practice who commenced HAART between 1996 and 1998. METHODS: Collection of plasma HIV-1 RNA, CD4+ and CD8+ T-lymphocyte data at 3-6 monthly time intervals over 2 years. RESULTS: Seventy-three subjects (54%) achieved suppression of plasma HIV-1 RNA to levels below 400 copies/ml during the observation period, 31 individuals (23%) had detectable plasma HIV-1 RNA below 10,000 copies/ml and 31 subjects (23%) had virological failures with viral loads above 10,000 copies/mL. Median CD4+ T lymphocytes increased from 246 to 463 x 10(6) cells/l, showing a median rise of 20 x 10(6) cells/l per month in the first 3 months and 7 x 10(6) cells/l per month thereafter. The proportion of individuals who reached CD4+ cell counts above 500 x 10(6) cells/l increased from 8% at baseline to 54% at 2 years. Treatment-naïve individuals, subjects with a large reduction of HIV-1 RNA or a large early CD8+ increase had better early CD4+ responses. Long-term CD4+ T-cell increases were inversely correlated with mean plasma HIV-1 RNA levels. Baseline CD4+ T-cell count was the most important determinant of reaching CD4+ cell counts above 500 x 10(6) cells/l. Nineteen per cent of subjects had no further CD4+ T-cell increases in the second year of therapy despite undetectable viral load. CONCLUSIONS: Immune reconstitution is a slow process, showing a large individual variability. The virological response to HAART was the most important determinant of the immunological short- and long-term response.

Rapid restoration of CD4 T cell subsets in subjects receiving antiretroviral therapy during primary HIV-1 infection.

OBJECTIVE: To compare the effect of highly active antiretroviral therapy on immune reconstitution in subjects with acute and chronic HIV-1 infection. DESIGN: Prospective study including 58 treatment-naive subjects who commenced indinavir or nelfinavir and two nucleosides during primary (PHI; n = 28) or chronic HIV-1 infection (CHI; n = 30). METHODS: Naive (CD45RA+ 62L+), memory (CD45RA-) and activated (CD38+ HLA-DR+) T cell subsets were quantified at 1-2 monthly time intervals using 4-colour flow cytometry. RESULTS: At 1 year, HIV-1 RNA declined in both cohorts to undetectable levels (< 50 copies/ml), while median CD4 lymphocyte count increased from 470 to 758 x 10(6) cells/l in PHI and from 204 to 310 x 10(6) cells/l in CHI, reaching > 500 x 10(6) cells/l in 93% of PHI, but only in 37% of CHI subjects (P < 0.001). Naive CD4 lymphocytes increased from 106 to 176 x 10(6) cells/l in PHI and from 41 to 44 x 10(6) cells/l in CHI (PHI versus CHI at 12 months: P = 0.003), while memory cells rose from 368 to 573 x 10(6) cells/l in PHI and from 148 to 223 x 10(6) cells/l in CHI (P < 0.001). Early increases (< 3 months) of CD4 lymphocytes were larger in subjects with PHI, consisting of naive CD45RA+ CD62L+ as well as memory CD45RA- CD62L+ cells (P = 0.001). CD4 activation declined from 5 to 2% in PHI and from 13 to 6% in CHI (P = 0.001), while CD8 cell activation was reduced from 33 to 15% in PHI and from 42 to 19% in CHI (P = 0.02). CONCLUSION: Immune reconstitution was more complete, occurred earlier and comprised both naive and memory CD4 T lymphocytes in subjects who commenced antiretroviral therapy during primary HIV-1 infection.

Phenotypic analysis of CD8+ T lymphocytes in a cohort of HIV type 1-infected patients treated with saquinavir, ritonavir, and two nucleoside analogs for 1 year, and association with plasma HIV type 1 RNA.

The phenotype of circulating CD8+ T lymphocytes and its association with plasma HIV-1 RNA were analyzed in 34 HIV-1-infected subjects, who were treated with saquinavir, ritonavir, and two nucleoside analogs (HAART) for 1 year. Four-color flow cytometry was applied to measure the expression of cell surface antigens CD38, HLA-DR, CD45RA, CD28, and CD62L on CD8+ T lymphocytes. The results were compared with data on 35 HIV-1-seronegative subjects, 18 untreated asymptomatic HIV-1-seropositive individuals, and 24 HIV-1-infected subjects receiving reverse transcriptase inhibitors (RTIs). Subjects receiving HAART showed a significantly elevated number and percentage of CD38- and HLA-DR-positive and CD28-negative CD8+ T lymphocytes as well as a lower percentage of naive (CD45RA+62L+) CD8+ T lymphocytes compared with HIV-1-uninfected controls. Even subjects with undetectable plasma HIV-1 RNA showed a persistent elevation of activated CD8+ T lymphocytes. However, fewer activated CD8+ T lymphocytes were observed in subjects receiving HAART than in untreated individuals and subjects administered RTIs. In individuals receiving RTIs, CD8+ cell activation was not significantly reduced compared with untreated subjects. Of all evaluated activation markers, the percentage of CD8+ T lymphocytes expressing CD38 and the combination of CD38 and HLA-DR showed the best correlation with plasma HIV-1 RNA. The persistence of CD8+ T lymphocyte activation in subjects receiving HAART strongly suggests ongoing viral activity, even in subjects with undetectable plasma HIV-1 RNA. A complete normalization of immunologic changes of CD8+ T lymphocytes would therefore require a more potent drug regimen or a longer duration of therapy.

Relative significance of different pathways of immune reconstitution in HIV type 1 infection as estimated by mathematical modeling.

A major goal of antiretroviral HIV-1 therapy is the reversal of HIV-1-associated immunological dysfunction. However, the pathogenetic mechanisms involved and their significance are largely unknown. On the basis of the life cycle of naive, activated, and memory CD4(+) T cell subsets, a mathematical model of immune reconstitution was developed and applied to data for T cell subsets in individuals with acute or chronic HIV-1 infection receiving antiretroviral therapy. The final model that most accurately fitted the data, and resulted in realistic estimates for CD4(+) T cell turnover, considered three pathways of immune reconstitution for naive cells, including thymic production, peripheral expansion, and redistribution of naive cells from lymphoid tissue. The reconstitution of the memory compartment was fitted through differentiation and expansion of naive cells and peripheral expansion of memory cells as well as redistribution of memory cells trapped in the lymphoid tissue. Estimated median half-lives for naive and memory CD4(+) T cells were 114 and 21 days, while total production rates were 9.1 x 10(7) and 2.4 x 10(9) cells/day, respectively. Peripheral expansion and thymic production contributed equally to the regeneration of naive cells, but peripheral expansion of memory cells was larger than production of these cells by differentiation of naive cells. A comparison of immune reconstitution in acute and chronic HIV-1 infection revealed that, after adjustment for age, the main difference was the more rapid release of a larger number of naive cells in treated acute HIV-1 infection. Thymic function and peripheral expansion rates, however, were similar in both cohorts.

Impact of HIV type 1 protease, reverse transcriptase, cleavage site, and p6 mutations on the virological response to quadruple therapy with saquinavir, ritonavir, and two nucleoside analogs.

Genotype alterations of HIV-1 protease, reverse transcriptase, cleavage sites p7/p1 and p1/p6, as well as p6(gag) and transframe protein p6* were studied in an observational cohort of 42 individuals who received antiretroviral therapy consisting of saquinavir, ritonavir, and two nucleoside analogs. In a multivariate logistic regression analysis, the prior protease inhibitor experience (odds ratio, 6.20; 95% CI, 1.22-31.38) and the presence of primary protease mutations (odds ratio, 9.99; 95% CI, 1.05-94.72) were independently associated with virological failure. Moreover, a trend was observed in that individuals with N-terminal amino acid insertions in the proline-rich motif of the p6(gag) protein were less likely to experience virological failure (OR, 0.17; 95% CI, 0.02-1.35; p = 0.09). In contrast, the presence of secondary protease, reverse transcriptase, or cleavage site mutations was not independently associated with treatment failure. However, mutations at cleavage site p7/p1 (p = 0.01) and C-terminal p6* mutations (p = 0.02) were both associated with primary protease mutations. In conclusion, the presence of primary protease mutations was the most important predictor of the subsequent virological response. Moreover, there is some evidence that insertions in the proline-rich area of the p6(gag) protein may affect the virological response. The relationship between mutations of cleavage sites or C-terminal p6* residues and protease mutations suggests that these alterations may serve a compensatory role, increasing viral fitness.

Novel deletion of HIV type 1 reverse transcriptase residue 69 conferring selective high-level resistance to nevirapine.

A novel deletion of residue 69 of the HIV-1 reverse transcriptase (RT) gene was detected in combination with mutations V75I/V and F77L/F in a patient with partial virological response to several antiretroviral drug regimens, including stavudine (D4T), didanosine (DDI), lamivudine (3TC), saquinavir (SQV), and nevirapine (NVP). Longitudinal analysis of samples revealed that this deletion emerged upon reinitiation DDI/D4T therapy following a toxicity-induced short discontinuation of all antiretrovirals. Analysis of the resistance phenotype showed a greater than 62-fold increase of the IC50 of NVP, but no significant change in sensitivity to other single nonnucleoside reverse transcriptase inhibitors (NNRTIs). The mutated virus showed only a moderately reduced sensitivity to DDI (6.7-fold) and D4T (4.8 fold). In a subsequent sample 3 months later additional RT mutations were found, including A62V, Y188L, and Q151M, conferring high-level cross-resistance to multiple nucleoside analogs. Our findings provide evidence that the deletion of RT residue 69 selectively confers high-level NVP resistance.

Effect of long-term infection with nef-defective attenuated HIV type 1 on CD4+ and CD8+ T lymphocytes: increased CD45RO+CD4+ T lymphocytes and limited activation of CD8+ T lymphocytes.

Members of the Sydney Blood Bank Cohort (SBBC) have been infected with an attenuated strain of HIV-1 with a natural nef/LTR mutation and have maintained relatively stable CD4+ T lymphocyte counts for 14-18 years. Flow cytometric analysis was used to examine the phenotype of CD4+ and CD8+ T lymphocytes in these subjects, including the immunologically important naive (CD45RA+CD62L+), primed (CD45RO+), and activated (CD38+HLA-DR+ and CD28-) subsets. The median values were compared between the SBBC and control groups, comprising age-, sex-, and transfusion-matched HIV-1-uninfected subjects; transfusion-acquired HIV-1-positive LTNPs; and sexually acquired HIV-1-positive LTNPs. Members of the SBBC not only had normal levels of naive CD4+ and CD8+ T lymphocytes, but had primed CD45RO+ CD4+ T lymphocytes at or above normal levels. Furthermore, these primed cells expressed markers suggesting recent exposure to specific antigen. SBBC members exhibited variable activation of CD8+ T lymphocytes. In particular, SBBC members with undetectable plasma HIV-1 RNA had normal levels of activated CD8+ T lymphocytes. Therefore, the result of long-term infection with natural nef/LTR mutant HIV-1 in these subjects suggests a decreased cytopathic effect of attenuated HIV-1 on susceptible activated CD4+ T lymphocyte subsets in vivo, and minimal activation of CD8+ T lymphocytes.

An examination of signs of disease progression in survivors of the Sydney Blood Bank Cohort (SBBC).

BACKGROUND: The Sydney Blood Bank Cohort (SBBC) was infected between 1981 and 1984 with a nef/LTR defective strain of HIV-1. Different responses to HIV-1 infection have emerged between cohort members in the last 5 years. Three recipients (C135, C64 and C49) remain asymptomatic, have normal CD4 T cell counts, below detection (BD) viral loads (VL), remain therapy naive and are termed long-term non-progressors (LTNP). The donor (D36) and the two recipients (C98 and C54) have significantly declining CD4 T cell counts, detectable VL and are now long-term survivors (LTS). In contrast, in the SA cohort, comparison study group for the SBBC, five of 24 remain therapy naïve after 15 years infection with HIV-1 and all have detectable VL. OBJECTIVES: This paper examines different outcomes to long-term infection with HIV-1 in the SBBC and provides a brief overview of the therapy naïve in a comparison study group, the SA cohort. STUDY DESIGN: Retrospective epidemiological follow-up of the SBBC and the SA cohort has been conducted for >15 years. Analysis of CD4 T cell counts, VL and intermittent monitoring of HIV-specific proliferative responses are reviewed. Viral sequence changes in the SBBC will be considered. RESULTS: Prior to therapy D36 had a CD4 T cell count of 160/mm(3) and plasma VL of 9900 copies/ml while C98 had a CD4 T cell count of 387/mm(3) and plasma VL of 11491 copies/ml. After 1 month of therapy, plasma VL was BD (<400 copies/ml) and both showed significant increase in CD4 T cell counts. Molecular changes have occurred in D36 and C98 viral strains, the most recently evolved quasispecies have larger deletions in the nef/LTR region. CONCLUSIONS: Infection with nef/LTR deleted HIV-1 has resulted in slower disease progression for the SBBC. The three LTNP have maintained normal low levels of activated CD8 T cells and strong HIV-specific proliferative responses to HIV-1 p24, which are associated with control of viral replication.

Primary HIV-1 infection: a review of clinical manifestations, immunologic and virologic changes.

In the past few years, major advances have been made in the field of primary HIV-1 infection. Several studies have reevaluated the clinical syndrome. The emergence of new molecular laboratory techniques has permitted a detailed analysis of viral dynamics and subsequent immunologic changes. Measurements of subsets of T-lymphocytes have allowed greater insight into the early pathogenesis of HIV-1 disease. There is now evidence that HIV-1-specific cytotoxic T-lymphocytes occur early during primary HIV-1 infection and are probably the most important immune defense against HIV-1. However, HIV-1 immune escape mutants have been identified during primary infection, which may be one reason for the failure of the immune system to completely eradicate the virus. Cytokines have been shown to play a role in primary HIV-1 infection, and the therapy of primary infection has gained more interest due to the introduction of potent triple combinations, including protease inhibitors.

Characterization of antigens recognized by natural killer cells in cell-culture supernatants.

Inhibitors of natural killer (NK) cell activity in cell-culture supernatants, believed to be antigens recognized by NK cells, were defined by their ability to inhibit NK cells in 51Cr-release cytotoxic assays. Supernatants from cultures of melanoma cells and Chang cells were used as the source of the antigen. Partial characterization by a number of sequential separation procedures suggested that the antigens were glycoproteins in the size range 120-140,000 daltons which had affinity for both concanavalin A and wheat germ lectin. Inhibitory activity was destroyed by trypsin digestion, but was resistant to neuraminidase and a number of physical procedures. Addition of supernatants to NK assays against a number of different target cells indicated that inhibition was restricted to certain target cells. This indicated that the inhibition of NK cells was not non-specific, and that the antigens were not expressed on all target cells. These studies provide a basis for further analysis of antigens recognized by NK cells, and allow investigation of their role in vivo in tumour-bearing hosts.

Immune reconstitution in HIV-1 infected subjects treated with potent antiretroviral therapy.

The introduction of potent antiretroviral drug regimens contributed to a decline in HIV-1 associated morbidity and mortality. Clinical observations of spontaneous remission of previously untreatable opportunistic infections in subjects on highly active antiretroviral therapy (HAART) reflect the substantial degree of immune reconstitution which can be achieved by those therapies. A biphasic increase of CD4+ T lymphocytes has been reported including naive (CD45RA+) and memory (CD45RO+) cell subsets. Proliferative lymphocyte responses to recall antigens and mitogens are enhanced over time, while T lymphocyte activation is largely reduced and T cell receptor (TCR) repertoires are partly restored. Proliferative lymphocyte responses specific to HIV-1 antigens, in contrast, remain weak. A complete normalisation of HIV-1 associated immunological alterations has not been reported so far, but the observation period of subjects on potent antiretroviral therapies is still relatively short.

Alterations in the immune response of human immunodeficiency virus (HIV)-infected subjects treated with an HIV-specific protease inhibitor, ritonavir.

Effects of a human immunodeficiency virus (HIV) type 1 protease inhibitor, ritonavir, were evaluated in 21 patients enrolled in a phase I/II study. The magnitude and rates of CD4 and CD8 lymphocyte increase, changes in subsets of CD4 and CD8 lymphocytes, and proliferative responses to mitogen and antigens were analyzed. Significant increases were noted in CD4 and CD8 lymphocyte counts; numbers of CD4CD45RO lymphocytes increased significantly by week 1 of therapy. Increases in the CD4CD45RA subset were observed at week 4. Reductions in the percentage of CD4 and CD8 lymphocytes expressing CD38 were noted. Increases in proliferative responses to phytohemagglutinin were noted in 6 of 7 patients and correlated with duration of virus load suppression. Increased responses to recall antigens and to HIV-specific proteins were observed. Treatment with ritonavir produced alterations in the immune system that included changes in T cell subset distribution and increases in CD4 and CD8 lymphocyte numbers and of lymphocyte function.

Characterization of the phenotypic and lymphokine profile associated with strong CD8+ anti-HIV-1 suppressor activity (CASA).

A panel of 22 CD8+ T cell lines, with a broad range of CD8+ anti-HIV-1 suppressor activity (CASA) were generated from a single patient with HIV-1 infection. CD8+ T cell lines with either strong or weak CASA were examined and compared for cell surface and intracellular markers, constitutive chemokine and lymphokine mRNA levels and inducible lymphokine expression. Strong CASA significantly correlated with CD8+ T cell lines that highly coexpressed the molecule CD28+ (r=0.52, P=0.01) and Ki67+ (r=0.88, P=0.02), with strong CASA CD8+ T cell lines demonstrating significantly higher (P < 0.05) expression of CD8+CD28+ and CD8+Ki67+ compared to those with weak activity. No such correlations or findings were observed for the markers CD38, HLA-DR, CD57 or perforin. The Th1 cytokines were expressed at greater levels than the Th2 cytokines, with strong CASA significantly associated with an increased inducible level of IL-2 production (P=0.05). Constitutive RANTES, IP-10 and I-309 mRNA expression were significantly (P < 0.05) elevated in CD8+ T cell lines exhibiting strong CASA compared to those with weak CASA. There was no significant difference in the mRNA expression of the lymphokines IL-2, 4, 5, 8, 9, 10, 14, 15, or chemokines MIP-1alpha, MIP-1beta, MCP-1, and Ltn. Strong CASA was therefore associated with rapidly replicating CD8+ T cells of the phenotype CD8+CD28+Ki67+ that expressed greater levels of IL-2 and the ligands RANTES and I-309.

A monoclonal antibody-based radioimmunoassay for the in vitro production of IgE by lymphocyte cultures.

Very sensitive radioimmunoassay systems have been described for the measurement of IgE produced in cultures of human peripheral blood mononuclear cells. However, differing results have been reported when cultures from non-atopic donors are stimulated with pokeweed mitogen, which may be due to cross-reactivity of anti-IgE antibodies with IgG. A monoclonal antibody specific for the Fc region of human IgE, and two polyclonal affinity-purified antibodies to IgE were tested for binding to 125I-labelled IgE myeloma proteins and polyclonal IgG in a sensitive double antibody precipitation assay. The monoclonal antibody and one of the polyclonal antibodies bound only IgE, whereas the other polyclonal antibody bound a significant proportion of labelled IgG. A solid phase radioimmunoassay was developed which combined the specificity of the monoclonal antibody with the sensitivity of the first polyclonal antibody as radioactive tracer. A second assay system was also tested using the cross-reacting antibody as tracer. Supernatants of pokeweed mitogen-stimulated peripheral blood mononuclear cell cultures from non-atopic donors were examined for IgE synthesis using both assays. The assay based on the monoclonal antibody did not detect IgE synthesis, whilst the second assay, based on the cross-reacting antibody indicated that spurious IgE had been produced in the same cultures. This study shows that protein-binding assays provide a simple means for checking the specificity of antibodies in solid phase radioimmunoassays, and confirms that pokeweed mitogen does not stimulate IgE production by cells from non-atopic donors when measured by a specific radioimmunoassay.

Phase I clinical trial of a human idiotypic p53 vaccine in patients with advanced malignancy.

BACKGROUND: The purpose of this study was to induce immunity to p53 by using an idiotypic vaccine, composed of a pool of eight peptides derived from the complimentarity determining regions (CDRs) of human anti-p53 antibodies. PATIENTS AND METHODS: Subjects with advanced malignancy received up to four, monthly intradermal injections of pooled peptides (500 microg of each) admixed with granulocyte-macrophage colony-stimulating factor (GM-CSF; 100 microg). In addition, two sheep and two rabbits were also vaccinated with the pooled peptides. RESULTS: Fourteen subjects were enrolled into the study and six of these completed the vaccination schedule. The vaccine was well tolerated by all subjects and no major adverse events were attributable to the vaccine. All subjects mounted in vivo delayed type hypersensitivity (DTH) responses to two or more of the individual vaccine peptides. Vaccine-induced antibodies specific for peptides 2, 5 or 8 were detected in four of six subjects, and two of these had vaccine-specific, cell-mediated responses. Increasing titers of p53-specific antibodies were found in one patient. No T-cell response to p53 was observed in any of the subjects. All animals developed humoral immunity to the peptides and one of the sheep developed rising serum titers of anti-p53 antibodies. CONCLUSIONS: Vaccination with human antibody CDR regions represents a novel method for inducing human antibodies, which may in turn serve as immunological mimics of p53.