The glycosylated Rv1860 protein of Mycobacterium tuberculosis inhibits dendritic cell mediated TH1 and TH17 polarization of T cells and abrogates protective immunity conferred by BCG.

We previously reported interferon gamma secretion by human CD4⁺ and CD8⁺ T cells in response to recombinant E. coli-expressed Rv1860 protein of Mycobacterium tuberculosis (MTB) as well as protection of guinea pigs against a challenge with virulent MTB following prime-boost immunization with DNA vaccine and poxvirus expressing Rv1860. In contrast, a Statens Serum Institute Mycobacterium bovis BCG (BCG-SSI) recombinant expressing MTB Rv1860 (BCG-TB1860) showed loss of protective ability compared to the parent BCG strain expressing the control GFP protein (BCG-GFP). Since Rv1860 is a secreted mannosylated protein of MTB and BCG, we investigated the effect of BCG-TB1860 on innate immunity. Relative to BCG-GFP, BCG-TB1860 effected a significant near total reduction both in secretion of cytokines IL-2, IL-12p40, IL-12p70, TNF-α, IL-6 and IL-10, and up regulation of co-stimulatory molecules MHC-II, CD40, CD54, CD80 and CD86 by infected bone marrow derived dendritic cells (BMDC), while leaving secreted levels of TGF-ß unchanged. These effects were mimicked by BCG-TB1860His which carried a 6-Histidine tag at the C-terminus of Rv1860, killed sonicated preparations of BCG-TB1860 and purified H37Rv-derived Rv1860 glycoprotein added to BCG-GFP, but not by E. coli-expressed recombinant Rv1860. Most importantly, BMDC exposed to BCG-TB1860 failed to polarize allogeneic as well as syngeneic T cells to secrete IFN-γ and IL-17 relative to BCG-GFP. Splenocytes from mice infected with BCG-SSI showed significantly less proliferation and secretion of IL-2, IFN-γ and IL-17, but secreted higher levels of IL-10 in response to in vitro restimulation with BCG-TB1860 compared to BCG-GFP. Spleens from mice infected with BCG-TB1860 also harboured significantly fewer DC expressing MHC-II, IL-12, IL-2 and TNF-α compared to mice infected with BCG-GFP. Glycoproteins of MTB, through their deleterious effects on DC may thus contribute to suppress the generation of a TH1- and TH17-dominated adaptive immune response that is vital for protection against tuberculosis.

Consequence of prior exposure to environmental mycobacteria on BCG vaccination and diagnosis of tuberculosis infection.

The protective efficacy of Mycobacterium bovis-bacille Calmette Guérin (BCG) against tuberculosis (TB) is variable in both humans and cattle. Exposure to environmental mycobacteria is thought to result in inappropriate priming of host immune responses. To investigate the impact of environmental mycobacteria on BCG efficacy, cattle were infected with M. avium, vaccinated with BCG, challenged with M. bovis and skin tested prior to necropsy. Elevated levels of IFNgamma were evident in M. avium-exposed animals before and after BCG vaccination with a bias towards avian purified protein derivative (PPD-A), suggesting that M. avium primed host immune responses. Exposure to M. avium also resulted in a higher frequency of circulatory IFNgamma-producing cells in response to PPD antigens at the time of M. bovis challenge. After M. bovis inoculation, the IFNgamma response to bovine PPD (PPD-B) increased compared to pre-challenge levels, indicating that all animals had been exposed to M. bovis. Skin test responses indicated 2/6 M. avium-BCG-M. bovis animals as reactors and 2/6 as inconclusive compared with 6/6 BCG-M. bovis animals as reactors. M. avium-exposed animals also had fewer lesions and the number of tissues containing viable M. bovis at post-mortem was significantly lower (P<0.02 compared with BCG-M. bovis animals), with two of the animals described as skin test negative with no visible lesions or viable bacteria. Thus, exposure of cattle to environmental mycobacteria such as M. avium prior to BCG vaccination did not dampen BCG-specific immune responses and resulted in lower TB pathology. However, the PPD-A bias associated with M. avium exposure is likely to undermine current TB diagnostic tests and the IFNgamma test in cattle.

Do bacteriostatic urethral lubricants affect the clinical efficacy of intravesical bacillus Calmette-Guérin therapy?

OBJECTIVES: To investigate the effect of a bacteriostatic urethral lubricant on the clinical efficacy of intravesical bacillus Calmette-Guérin (BCG) therapy. METHODS: Between July 1987 and August 1999, 389 patients with superficial urothelial carcinoma of the bladder (pTa [multilocular, size greater than 2 cm, recurrent tumor], pT1, pTis) were treated adjuvantly with BCG in a 6-week intravesical cycle after complete transurethral tumor resection of the bladder. Within the framework of a prospective study on recurrence and progression of superficial urothelial carcinoma of the bladder after transurethral resection and BCG therapy, we retrospectively studied the clinical value of the observation that the use of bacteriostatic urethral lubricants reduces the viability and thus the efficacy of BCG. If a lubricant induces a clinically relevant reduction in the viability of BCG, instillation without lubricant should lead to a lower rate of recurrence and possibly a lower rate of progression. Lubricant (11 mL) was used during catheterization in every male patient in our population (group 1, n = 317, 81.5%); no lubricant was used in the women serving as the control group (group 2, n = 72, 18.5%). The two groups were similar with respect to age, distribution of tumor stage and grade, multifocality, and frequency of previous bladder carcinoma (Levene test, P = 0.008). The median follow-up was 54 months (range 4 to 143). RESULTS: Of the 389 patients, 90 (23.1%) developed recurrence during the follow-up period: 73 (23%) in group 1 and 17 (23.6%) in group 2 (P value not significant). Progression occurred in 14 patients in group 1 (4.4%) and in 8 patients in group 2 (11%) (P = 0.043). In groups 1 and 2, 19.2% and 47.1%, respectively, of the recurrences were progressive (P = 0.026). CONCLUSIONS: The use of bacteriostatic lubricants in the usual dose before BCG instillation had no detectable adverse effect on the clinical efficacy of intravesical BCG immunotherapy. To avoid traumatic catheterization with possible systemic BCG administration, we therefore recommend, especially in men, the additional use of a sufficient quantity of urethral lubricant.

Intravesical bacille Calmette-Guérin in the treatment of superficial transitional cell carcinoma of the bladder.

OBJECTIVE: To determine the efficacy of intravesical bacillus Calmette-Guérin (BCG) in the treatment of patients with superficial transitional cell carcinoma (TCC) of the bladder, and to assess the impact of fibrin clot inhibitors. PATIENTS AND METHODS: A retrospective review of 56 patients with superficial TCC of the bladder, treated with intravesical BCG after initial transurethral resection (TUR) of raised or papillary lesions or cold cup biopsy of areas of carcinoma in situ (CIS), was performed. Patient drug histories were reviewed for evidence of ingestion of medication known to inhibit fibrin clot formation. The impact of such medication was assessed using the Chi-square test. RESULTS: Fifty-six patients were treated between 1987 and 1991 of whom 52 were evaluable. Eighteen patients (35%) had a complete response with a mean follow-up of 19 months. Six patients (60%) in the group with CIS had a complete response rate with a mean follow-up of 28 months. Seven patients (13%) developed local progression and required cystectomy or external beam radiotherapy. Six patients (12%) died from metastatic disease. Three patients (6%) had significant complications. The adverse impact of fibrin clot inhibitors was found to be significant. CONCLUSION: Intravesical BCG is effective in the treatment of superficial TCC, especially CIS. A careful drug history is important to identify fibrin clot inhibitors so that, if possible, they may be withdrawn prior to intravesical BCG treatment.

The adverse effect of fibrin-clot inhibiting drugs on intravesical bacillus Calmette-Guérin efficacy for superficial bladder cancer.

Bacillus Calmette-Guérin (BCG) is currently thought to act as a biological immune modifier in effecting antitumour activity. Recent evidence suggests that BCG binding to fibronectin (FN), a tissue glycoprotein, may be a prerequisite step in initiating this response. Drugs inhibiting the availability of exposed FN in the bladder after urothelial disruption may adversely affect the efficacy of BCG. Data are presented of 45 patients with tumour limited to mucosa (pTa) or carcinoma in situ (CIS) given intravesical BCG therapy, with (group 1) or without (group 2) fibrin clot-inhibiting drugs concurrently during treatment. The success rate of 11.1% for group 1 (1/9) patients was significantly less than that of 69.4% for group 2 (25/36), (chi 2 = 7.79, P < 0.01 Fisher's exact test) supporting the suggestion that the concurrent administration of fibrin-clot inhibiting drugs may adversely affect the outcome of BCG therapy.

Adverse impact of fibrin clot inhibitors on intravesical bacillus Calmette-Guerin therapy for superficial bladder tumors.

Although intravesical bacillus Calmette-Guerin therapy has proved to be efficacious in the treatment and prophylaxis against tumor recurrence of superficial bladder tumors, its mechanism of action has not been fully elucidated. Previous work has suggested that bacillus Calmette-Guerin organisms attach to the matrix protein, fibronectin, during fibrin clot formation at sites of urothelial disruption and that this attachment was required for the antitumor effect of bacillus Calmette-Guerin to be expressed. Furthermore, drugs inhibiting clot formation were found to abrogate the antitumor effect of intravesical bacillus Calmette-Guerin therapy in a murine bladder tumor model. To examine the effect of inhibitors of fibrin clot formation on the results of intravesical bacillus Calmette-Guerin therapy, a retrospective analysis of 149 evaluable patients receiving intravesical bacillus Calmette-Guerin for superficial bladder tumors was performed. The over-all response rate free of tumor for 29 patients who concomitantly received inhibitors of fibrin clot formation with bacillus Calmette-Guerin therapy was 48%, as compared with 67% for 120 patients who were not receiving these medications (p = 0.0655, chi-square). The most striking difference was noted for patients who failed with recurrent superficial disease. Of the patients who received fibrin clot inhibitors during intravesical bacillus Calmette-Guerin therapy 35% had recurrent superficial tumors compared to only 8% of those who did not receive these drugs during a mean followup of 29.8 plus or minus 11 months (p = 0.005, chi-square). Our study suggests that inhibitors of fibrin clot formation may have an adverse influence on the results of intravesical bacillus Calmette-Guerin therapy for superficial bladder tumors.