Correlates of GLA family adjuvants' activities.

Lipopolysaccharide (LPS) is a well-defined agonist of Toll-like receptor (TLR) 4 that activates innate immune responses and influences the development of the adaptive response during infection with Gram-negative bacteria. Many years ago, Dr. Edgar Ribi separated the adjuvant activity of LPS from its toxic effects, an effort that led to the development of monophosphoryl lipid A (MPL). MPL, derived from Salmonella minnesota R595, has progressed through clinical development and is now used in various product-enabling formulations to support the generation of antigen-specific responses in several commercial and preclinical vaccines. We have generated several synthetic lipid A molecules, foremost glucopyranosyl lipid adjuvant (GLA) and second-generation lipid adjuvant (SLA), and have advanced these to clinical trial for various indications. In this review we summarize the potential and current positioning of TLR4-based adjuvant formulations in approved and emerging vaccines.

Leprosy - an overview of clinical features, diagnosis, and treatment.

Leprosy is a chronic infectious disease caused by Mycobacterium (M.) leprae. Worldwide, 210,758 new cases were diagnosed in 2015. The highest incidence is found in India, Brazil, and Indonesia. While the exact route of transmission remains unknown, nasal droplet infection is thought to be most likely. The pathogen primarily affects the skin and peripheral nervous system. The disease course is determined by individual host immunity. Clinically, multibacillary lepromatous variants are distinguished from paucibacillary tuberculoid forms. Apart from the various characteristic skin lesions, the condition is marked by damage to the peripheral nervous system. Advanced disease is characterized by disfiguring mutilations. Current treatment options are based on WHO recommendations. Early treatment frequently results in complete remission without sequelae. While paucibacillary forms are treated with rifampicin and dapsone for at least six months, multibacillary leprosy is treated for at least twelve months, additionally requiring clofazimine. Leprosy reactions during therapy may considerably aggravate the disease course. Besides individual treatment, WHO-supported preventive measures and strategies play a key role in endemic areas.

[Paradoxical reactions and responses during antibiotic treatment for Mycobacterium ulcerans infection (Buruli ulcer). Four cases from French Guiana]. / Réponses et réactions paradoxales au cours du traitement médicamenteux de l'infection à Mycobacterium ulcerans (ulcère de Buruli). Quatre observations en Guyane française.

BACKGROUND: In recent years, first-line therapy for Mycobacterium ulcerans infection in French Guiana has consisted of antibiotics active against this organism. Two regimens are used comprising rifampicin associated with clarithromycin or amikacin. PATIENTS AND METHODS: We describe four patients presenting apparent worsening of their lesions during treatment: ulceration of a nodular lesion in a 32-year-old woman and worsening of an ulcerated lesion in three patients aged 16, 27 and 79 years. DISCUSSION: In these 4 patients, we concluded that the symptoms were caused by a paradoxical response or a reaction, a phenomenon already described in tuberculosis and leprosy. Such worsening is transient and must not be misinterpreted as failure to respond to treatment. The most plausible pathophysiological hypothesis involves the re-emergence of potentially necrotizing cellular immunity secondary to the loss of mycolactone, a necrotizing and immunosuppressive toxin produced by M. ulcerans, resulting from the action of the antibiotics.

IL-2 and IL-12 act in synergy to overcome antigen-specific T cell unresponsiveness in mycobacterial disease.

IL-12 secretion by APC is critical for the development of protective Th1-type responses in mycobacterial (Mycobacterium avium and Mycobacterium tuberculosis) infections in mice. We have studied the role of IL-12 and IL-2 in the generation of Mycobacterium leprae-specific T cell responses in humans. Leprosy patients were defined as low/nonresponders or high responders based on the level of T cell proliferation in M. leprae-stimulated PBMC. In high responders, M. leprae-induced proliferation was markedly suppressed by neutralizing anti-IL-12 mAb (inhibition 55 +/- 6%). Neutralization of IL-2 activity resulted in an inhibition of 77 +/- 4%. Given the importance of endogenous IL-2 and IL-12 in M. leprae-induced responses, we investigated the ability of rIL-2 and rIL-12 to reverse T cell unresponsiveness in low/nonresponder patients. Interestingly, rIL-12 and rIL-2 strongly synergized in restoring both M. leprae-specific T cell proliferation and IFN-gamma secretion almost completely to the level of responder patients. A similar synergy between rIL-2 and rIL-12 was also observed in high responders when suboptimal M. leprae concentrations were used for T cell stimulation. Our data demonstrate a crucial role for endogenous IL-12 and IL-2 in M. leprae-induced T cell activation. Most importantly, we show that rIL-2 and rIL-12 act in synergy to overcome Ag-specific Th1 cell unresponsiveness. These findings may be applicable to the design of antimicrobial and antitumor vaccines.

Long-lasting T-cell reactivity to Mycobacterium leprae antigens in human volunteers vaccinated with killed M. leprae.

A trial with a candidate anti-leprosy vaccine based on killed Mycobacterium leprae was started in Norway in 1983 to evaluate its toxicity and efficacy to induce cell-mediated immunity (CMI) in BCG-vaccinated healthy volunteers. The vaccinated subjects were found to be free of unacceptable side-effects and their T cells showed elevated proliferative response to M. leprae up to 1 year postvaccination. When tested in 1991, 8 years after vaccination, peripheral blood mononuclear cells from the same volunteers showed a persistent high proliferative response to M. leprae. From a total of 147 T-cell clones established from these subjects, 26 clones were specific to M. leprae and the remaining T-cell clones responded to M. leprae as well as to BCG and other cultivable mycobacteria. The epitopes recognized by the M. leprae-specific T-cell clones were present on several protein antigens including the 18 kDa and the 65 kDa heat shock proteins. A dominant epitope, peptides 38-50 on the M. leprae 18 kDa heat shock protein, which was recognized by M. leprae-specific T cells 1 year after vaccination, was also recognized 8 years after vaccination by the same donor. This is the first report demonstrating the unique property of killed M. leprae with respect to the induction of long-lasting T-cell reactivity towards M. leprae antigens in humans.

Effect of cutaneous cell-mediated immune response to rIFN gamma on Mycobacterium leprae viability in the lesions of lepromatous leprosy.

1. Studies were carried out to determine the effect of intra-dermal injections of recombinant human interferon-gamma (rIFN gamma) on the viability of Mycobacterium leprae. Twenty-three untreated and 4 treated multibacillary patients, 12 with lepromatous leprosy (LL) and 15 with borderline lepromatous leprosy (BL), were selected for intradermal administration of rIFN gamma or PPD. Treated patients (LL and BL) had received multi-drug therapy according to the recommendations of the World Health Organization, i.e., rifampicin (600 mg/month), dapsone (100 mg/day) and clofazimine (50 mg/day and 300 mg/month) for 1-4 months. Three daily doses of 10 or 30 micrograms rIFN gamma induced local induration and mononuclear leucocyte accumulation. Bacteria isolated from a punch biopsy of the site 21 days after lymphokine administration were injected into mouse foot pads and evaluated for viability and growth. 2. The local response to rIFN gamma (specific activity 2 x 10(7) units/mg protein) induced a delay or total inhibition of M. leprae growth in the mouse foot pad, indicating that the cellular response to the antigen reduced local M. leprae viability. The extent of reduction in viability depended on the dose of rIFN gamma injected and the extent of local induration induced by the lymphokine. With a vigorous cell-mediated immune response growth was fully inhibited. 3. A similar but less extensive effect on M. leprae viability was observed in response to the local injection of 5 units in 0.1 ml of purified protein derivative of tuberculin (PPD).

M. leprae- and BCG-induced chemiluminescence response of monocytes from leprosy patients and healthy subjects: effects of gamma-interferon and GM-CSF.

Mycobacterium leprae, in contrast to BCG, failed to trigger any chemiluminescence (CL) response in mononuclear cells from either leprosy patients or healthy subjects, a deficit not reversed by either interferon-gamma or GM-CSF. Chemiluminescence responses induced without mycobacteria or with BCG were found to be lower in leprosy patients than in controls. M. leprae were also less well phagocytosed than BCG. However, there was a significant difference in phagocytosis between healthy and tuberculoid leprosy subjects. Phagocytosis was not altered by the addition of either lymphokine, and no major differences between healthy subjects and patients were observed. Preincubating mononuclear cells with anti-mycobacteria antibodies (lepromatous patients' sera) did not increase the CL response nor the phagocytosis of M. leprae or BCG.

Effects of cyclosporin A on bacterial growth and immunological responsiveness in BALB/c mice infected with Mycobacterium leprae.

When BALB/c mice were infected with Mycobacterium leprae and orally treated 6 times weekly with a dose of 8 mg/kg cyclosporin A (CsA) for 19 months, the number of organisms was slightly higher at 19 months as compared with mice in which the dose of CsA was gradually decreased after 6 months and discontinued at the 8th month (p less than 0.01 for the 15th and 19th months). Lymphocyte blast transformation (LBT) showed that spleen cells from CsA-treated mice 4 weeks after infection with M. leprae and 3 weeks after CsA treatment was stopped responded to the sonicated supernatant of M. leprae suspension (SS), M. leprae (Ml), and concanavalin A (ConA) less than those cells from mice not treated with CsA. This response was dose-dependent. At week 15, 14 weeks after CsA administration was stopped, the LBT response to SS and Ml by cells from M. leprae-infected mice exceeded that of mice without CsA treatment, and the response to ConA in M. leprae-infected mice was less than that in uninfected mice without CsA-treatment. Thus, if CsA was administered, the T-cell functions were suppressed. However, when CsA treatment was discontinued for longer periods, the T-cell function was activated. From these results, we speculate that M. leprae would have the capability of growing more abundantly in mice treated with CsA 100 mg/kg for 1 week every month.

[Immunologic indices in a varying course of lepromatous leprosy]. / Immunologicheskie pokazateli pri razlichnom techenii lepromatoznoi lepry.

The study of the proliferative and regulatory functions of lymphocytes in patients with lepra of the lepromatous type has shown that at the active stage of the disease both the response of lymphocytes to mitogens and their suppressor functions are decreased. During the regression of the disease these characteristics are restored to the normal level only in patients with the relapse-free course of the disease, while patients with relapses in their medical history retain the low level of such characteristics. It is expedient to use these cell-mediated immunity characteristics as signs permitting the formation of risk groups of patients who may expect the relapse of the disease.

[Immunocorrective properties of new antileprotic preparations]. / Immunokorrigiruiushchie svoistva novykh protivoleproznykh preparatov.

Antimycobacterial and immunotropic characteristics of new antileprosy drugs, made in this country, have been studied and compared with those of dapsone. Animal experiments have demonstrated a high immunostimulating activity of the new drugs; this calls for clinical trials of these agents, for they may improve the therapy efficacy, help prevent the disease recurrences, and cut down the length of treatment for leprosy.

Occurrence of reversal reactions in BT patients during who paucibacillary leprosy MDT (1982).

55 B.T. patients were treated with WHO Paucibacillary MDT (1982). The patients suffered from reversal reaction neither at the time of initiation of MDT nor prior to that. During the 6 months period of MDT, one patient developed reversal reaction of a skin patch, and another patient developed neuritis of a peripheral nerve trunk.

Avaliaçäo do levamizole como imuno-estimulador na hanseníase / Levamizole assessment as immuno-stimulator in leprosy

Pacientes hansenianos classificados como portadores da forma clínica virchoviana foram inicialmente submetidos a avaliaçäo imulógica específica (Mitsudina) e inespecífica (PPD, tricofitina, DNCB e determinaçäo do percentual de linfócitos T no sangue periférico). A seguir, foram estimulados com levamizole (150 mg/dia, 2 vezes por semana, durante 2 meses) e novamente submetidos a avaliaçäo imunológica, visando determinar o efeito deste modulador sobre o defeito imunológico descrito nestes pacientes

In situ and in vitro characterization of the cellular immune response in erythema nodosum leprosum.

We sought to evaluate cell-mediated immune responses in erythema nodosum leprosum (ENL), a reactional state occurring in lepromatous leprosy. Skin biopsies from patients with leprosy were studied with monoclonal antibodies against T lymphocyte antigenic determinants, interleukin 2 (IL 2), and IL 2 receptors (Tac) by using immunoperoxidase staining of frozen sections. Peripheral blood lymphocytes from 18 ENL patients were tested in vitro for lepromin-induced suppression of Con A stimulation. Serial studies of seven lepromatous patients who developed ENL during the course of the study showed increases in both the Leu-3a:Leu-2a ratio and the number of IL 2-positive cells. IL 2-positive cells comprised 0.3% of the cells in all of the ENL lesions studied as compared with the 0.03% found in nonreactional lepromatous lesions (P less than 0.001). Lepromin-induced suppression of the Con A response, present in nonreactional lepromatous patients, significantly decreased in patients developing the ENL reaction, but returned after recovery from ENL. These changes in tissues and peripheral blood suggest that the pathogenesis of ENL is related to cell-mediated immune processes. Despite these immunologic changes, however, ENL patients do not recover antigen-specific skin tests or eliminate Mycobacterium leprae.

Negative observations on immunological side effects of rifampin and dapsone in mice.

The in vivo effects of rifampin and dapsone on immunological responses were investigated using mice immunized with sheep erythrocytes. The number of cells producing antibody was not affected by a clinical (1 CD) or a threefold excess dose (3 CD) of the drugs administered for ten days. A similar result was obtained in an experiment using a mouse strain known to be low responders to the antigen. Induction of suppressor cells acting on antibody production was not influenced by 3 CD or 6 CD of the drugs. Neither delayed-type hypersensitivity nor induction of the suppressor cell population acting on delayed hypersensitivity was affected by 3 CD of the drugs. Phagocytosis of sheep erythrocytes by peritoneal cells and the growth of a tumor were not altered by 6 CD of the drugs.

Immunomodulation with corticosteroids and levamisole in leprosy as gauged by in vivo lepromin and in vitro CMI responses.

Corticosteroids and Levamisole are known to be immuno suppressive and immuno stimulating agents respectively. Their effects on polar types of leprosy, tuberculoid and lepromatous have been studied using in vivo lepromin and in vitro lymphocyte count, rosette formation, L.T.T. and L.M.I.T. parameters. Immunosuppressive effect of corticosteroids on tuberculoid leprosy is marked with reduced and negative lepromin sensitivity but same does not hold true with other in vitro C.M.I. tests. Similar results are obtained with levamisole exhibiting its ineffectiveness in lepromin conversion in lepromatous cases although some improvement is observed in other in vitro C.M.I. tests. Evaluation of the results showed: lack of correlation between in vivo lepromin and in vitro other C.M.I. parameters with corticosteroids and levamisole lepromin sensitivity has some unknown influence other than thymic factors, prolonged corticosteroid therapy may produce permanent immunosuppression in tuberculoid cases making them more vulnerable towards lepromatous pole and lepromin sensitivity is more reliable, stable and easy to perform.