Potential of adjunctive Mycobacterium w (MIP) immunotherapy in reducing the duration of standard chemotherapy against tuberculosis.

INTRODUCTION: The need to shorten the treatment duration in tuberculosis has always been felt. Immunotherapy in combination with chemotherapy has been considered a promising approach for this purpose into tuberculosis. We studied the adjuvant immunotherapeutic activity of Mycobacterium indicus pranii (MIP or Mw) in combination with conventional chemotherapy using guinea pig of pulmonary tuberculosis infected with Mycobacterium tuberculosis H37Rv via aerosol. METHODS: Experimental animals treated with standard chemotherapy and immunotherapy (MIP) separately and in combination of both. Guinea pig lungs evaluated following infection and subsequent therapy at predefine time point. Various cytokine mRNA expressions levels were quantified by quantitative reverse transcriptase PCR at the 4th, 8th and 12th week post-infection of M. tuberculosis. RESULTS: We determined the time required for bacterial clearance from guinea pig lungs. Standard chemotherapy (RvCh) compared to the animals where chemotherapy plus Mw immunotherpay (RvChMwT) was given. It took 12 weeks to achieve bacterial clearance in the RvCh group while this was achieved in 8 weeks in RvChMwT group. Pro-inflammatory cytokines (IFN-γ, IL-2, IL-12p35 and TNF-α) level were higher in RvCh, RvChMwT and RvMwT group, while the IL-10 and TGF-ß were suppressed. CONCLUSION: Cytokine expression level showed that Mw in conjunction with chemotherapy enhances the effect of pro-inflammatory cytokines (such as, IFN-γ, IL-2, IL-12 and TNF-α) and reduces the production and effect of anti-inflammatory cytokines (like IL-10 and TGF-ß) thereby restoring the pro-inflammatory / anti-inflammatory cytokines balance. Thus, the present study indicates that subject to rigorous testing by other parameters, Mw (MIP) as adjunct immunotherapy has potential for reducing treatment duration.

Expression profile of CXCL12 chemokine during M. tuberculosis infection with different therapeutic interventions in guinea pig.

Mycobacterium indicus pranii (MIP) already established as an immune-modulator in mycobacterial infections generates immune response by acting on CXC chemokines. In the present study, the immunomodulatory effect of MIP in conjunction with chemotherapy against M.tb infection was evaluated by colony forming units (CFUs) following aerosol infection to guinea pig and by measuring CXCL12 chemokine expression using q-PCR and in situ RT-PCR. Different experimental groups included, infection (Rv), immunoprophylaxis (RvMw), chemotherapy (RvCh) and combination of immunoprophylaxis+chemotherapy (RvChMw) group and normal healthy (NH) group. In the combination of immunoprophylaxis+chemotherapy (RvChMw) group, the CFU counts reduced significantly (p<0.001) at 4th week of infection as compared to other treated groups (RvMw and RvCh group). The expression of CXCL12 was recorded in all the treated groups of animals. The study demonstrated suppressed expression of CXCL 12 in both immunoprophylaxis as well as chemotherapy groups (6th and 8th week) that become elevated in immunoprophylaxis plus chemotherapy group (10th week), at which time point no CFUs were detected in RvCh and RvChMw group. The findings indicate that the expression of CXCL12 is associated with good response to anti - tubercular treatment. Thus, prior immunization with MIP appears to show good immunomodulatory effect to release CXCL12 chemokine during infection and also correlates with enhanced effect to chemotherapy.

Expression of CXCL10 (IP-10) and CXCL11 (I-TAC) chemokines during Mycobacterium tuberculosis infection and immunoprophylaxis with Mycobacterium indicus pranii (Mw) in guinea pig.

Mycobacterium indicus pranii (earlier known as Mycobacterium w) has been used as an immunmodulatory agent in leprosy and tuberculosis by mediating the release of various cytokines and chemokines. CXCL10 (IP-10) and CXCL11 (I-TAC) chemokines are involved in T-cell migration and stimulation of natural killer cells in Mycobacterium tuberculosis infection. In this study, the effect of heat killed M. indicus pranii (alone and in conjunction with chemotherapy) on disease progression was determined by colony forming units (CFUs) in guinea pig lung following their aerosol infection and the expression levels of CXCL10 and CXCL11 were studied by quantitative Reverse Transcriptase Polymerase Chain Reaction (qRT-PCR) and in situ RT-PCR. Four groups of animals included; infection only (Rv), immunoprophylaxis (RvMw), chemotherapy (RvCh) and combination of immunoprophylaxis with chemotherapy (RvChMw). In the group where immunoprophylaxis was given in combination with chemotherapy, the CFU counts reduced significantly at 4th week post-infection as compared to animals that received immunoprophylaxis or chemotherapy alone. At the same time, all groups of animals had elevated expression of CXCL 10 which was significantly high only in animals that received Mw with or without chemotherapy. Unlike to CXCL 10, study demonstrated suppressed expression CXCL 11 in both immunoprophylaxis as well as chemotherapy groups that became up-regulated in synergistic response of immunoprophylaxis and chemotherapy. Taken together, data indicates that the expression of CXCL10 and CXCL11 positively correlates with anti-tubercular treatment (at least with combination of immunoprophylaxis and chemotherapy). Therefore, prior immunization with Mw appears to be a good immunomodulator for release of chemokines and augments the effect of chemotherapy.

S- to N-Acyl transfer in S-acylcysteine isopeptides via 9-, 10-, 12-, and 13-membered cyclic transition states.

S-Acyl cysteine peptides containing α-, ß- or γ-amino acid residues undergo long-range S- to N-acyl transfer to give analogs of native tripeptides and tetrapeptides containing additional carbon atoms in the chain. The ease of intramolecular S → N-acyl transfer relative to intermolecular transacylation is favored increasingly for 9 < 12 < 13 ~ 10-membered cyclic transition states; the observed order is explained on conformational and intermolecular interaction considerations.

Whole cell & culture filtrate proteins from prevalent genotypes of Mycobacterium tuberculosis provoke better antibody & T cell response than laboratory strain H 37 Rv.

BACKGROUND & OBJECTIVES: The immune responses to different antigens of Mycobacterium tuberculosis H 37 Rv vary from patient to patient with tuberculosis (TB). Therefore, significant difference might be documented between the H 37 Rv with long histories of passages and recent clinical isolates of M. tuberculosis. In the present study, immune response of TB patients and healthy controls against 39 clinical M. tuberculosis isolates was correlated with laboratory strain H 37 Rv. METHODS: The antibody response was studied coating whole cell extracts and culture filtrate proteins of M. tuberculosis isolates and laboratory strain H 37Rv by enzyme linked immunosorbent assay (ELISA). Lymphoproliferation was studied by incorporation of tritiated thymidine and cytokines (IFN-γ and IL-4) by using commercially available kits. RESULTS: Sero-reactivity to whole cell extract (WCE) of 11 clinical isolates was higher with pooled serum and individual's serum from tuberculosis patients showed significant reactivity (P<0.05) to ten of these isolates using ELISA. Of the WCE of 39 clinical isolates, 10 were found to be potent inducer of lymphoproliferation as well as cytokine secretion (P<0.05) in peripheral blood mononuclear cells from PPD+ healthy controls. Six culture filtrate proteins (CFPs) from these selected clinical isolates were also better inducers of antibody and T-cell response. INTERPRETATION & CONCLUSION: Overall, our results revealed that the clinical isolates belonging to prevalent genotypes; CAS1_Del (ST-26), East African-Indian (ST-11) and Beijing family (ST-1) induced better antibody and T cell responses compared to H 37 Rv laboratory strain. Further studies need to be done to purify and identify the dominant protein (s) using whole cell extract and culture filtrates from these immunologically relevant clinical M. tuberculosis isolates, which will be worthwhile to find out pathogenic factors, potential diagnostic markers and protective molecules for tuberculosis.

Long-range intramolecular S → N acyl migration: a study of the formation of native peptide analogues via 13-, 15-, and 16-membered cyclic transition states.

The intramolecular long-range S → N acyl migration via 13-, 15-, and 16-membered cyclic transition states to form native tetra- and pentapeptide analogues was studied on S-acylcysteine peptides containing ß- or γ-amino acids. The pH-dependency study of the acyl migration via a 15-membered cyclic transition state indicated that the reaction is favored at a pH range from 7.0 to 7.6. Experimental observations are supported by structural and computational investigations.

Bile acid-based cyclic bisbenzimidazolium receptors for anion recognition: highly improved receptors for fluoride and chloride ions.

The anion binding properties of bile acid-based cyclic bisbenzimidazolium receptors 6-8 bridged with m-xylene, p-xylene, and 2,6-dimethylpyridine have been studied. Receptors 6 and 7 exhibit much higher binding affinity for fluoride and chloride ions, respectively, as compared to the imidazolium receptors 1 and 2. Receptor 8, however, shows high selectivity but very low binding affinity for anions due to the presence of pyridyl nitrogen. The single-crystal X-ray structure of imidazolium receptor 10-(Br)2 containing pyridyl spacer reveals the binding pattern.