Evaluation of local, regional and abscopal effects of Boron Neutron Capture Therapy (BNCT) combined with immunotherapy in an ectopic colon cancer model.

OBJECTIVE: The aim of the present study was to evaluate the local and regional therapeutic efficacy and abscopal effect of BNCT mediated by boronophenyl-alanine, combined with Bacillus Calmette-Guerin (BCG) as an immunotherapy agent in this model. METHODS: The local effect of treatment was evaluated in terms of tumor response in the irradiated tumor-bearing right hind flank. Metastatic spread to tumor-draining lymph nodes was analyzed as an indicator of regional effect. The abscopal effect of treatment was assessed as tumor growth inhibition in the contralateral (non-irradiated) left hind flank inoculated with tumor cells 2 weeks post-irradiation. The experimental groups BNCT, BNCT + BCG, BCG, Beam only (BO), BO +BCG, SHAM (tumor-bearing, no treatment, same manipulation) were studied. RESULTS: BNCT and BNCT + BCG induced a highly significant local anti-tumor response, whereas BCG alone induced a weak local effect. BCG and BNCT + BCG induced a significant abscopal effect in the contralateral non-irradiated leg. The BNCT + BCG group showed significantly less metastatic spread to tumor-draining lymph nodes vs SHAM and vs BO. CONCLUSION: This study suggests that BNCT + BCG-immunotherapy would induce local, regional and abscopal effects in tumor-bearing animals. BNCT would be the main effector of the local anti-tumor effect whereas BCG would be the main effector of the abscopal effect. ADVANCES IN KNOWLEDGE: Although the local effect of BNCT has been widely evidenced, this is the first study to show the local, regional and abscopal effects of BNCT combined with immunotherapy, contributing to comprehensive cancer treatment with combined therapies.

Phenotypic and Genotypic Drug Susceptibility Assessment of Mycobacterium bovis Bacillus Calmette-Guérin Clinical Strains.

PURPOSE: Mycobacterium bovis Bacillus Calmette-Guérin (BCG) is the only vaccine licensed against tuberculosis. Despite the protection offered by the vaccine, in some circumstances children and immunocompromised individuals can develop associated infections, known as BCGitis. Drug susceptibility patterns of BCG clinical strains have rarely been described. We aimed to describe the susceptibility pattern of BCG clinical strains isolated in two different countries. METHODS: We performed culture-based drug susceptibility testing of thirty one BCG strains isolated from patients in Brazil (n=5, 16%) and Argentina (n=26, 84%) using the broth micro-dilution method (phenotypic method). Final antibiotic concentrations for susceptibility testing ranged from 0.5 to 16 mg/L for amikacin, 0.3125 to 10 mg/L for ethambutol, 0.05 to 1.6 mg/L for isoniazid and 0.25 to 8 mg/L for rifampicin, streptomycin and ofloxacin. Additionally, we compared the results with genetic data obtained by whole genome sequencing. RESULTS: By using the phenotypic method we detected one strain resistant to ethambutol, three strains resistant to rifampicin and one resistant to isoniazid. Additionally, two strains that were phenotypically resistant to both isoniazid and rifampicin carried mutations in the katG and rpoB genes simultaneously. CONCLUSION: There is evidence of the emergence of BCG-resistant strains isolated from vaccine-related complications. We recommend drug susceptibility testing of the BCG strain causing the infection in order to prevent treatment failure.

Newly sequenced genomes of four Bacillus Calmette Guerin vaccines.

Bacillus Calmette Guerin (BCG) vaccines comprise a family of related strains. Whole genome sequencing has allowed the better characterisation of the differences between many of the BCG vaccines. As sequencing technologies improve, updating of publicly available sequence data becomes common practice. We hereby announce the draft genome of four commonly used BCG vaccines in Brazil, Argentina and Venezuela.

Newly sequenced genomes of four Bacillus Calmette Guerin vaccines

Bacillus Calmette Guerin (BCG) vaccines comprise a family of related strains. Whole genome sequencing has allowed the better characterisation of the differences between many of the BCG vaccines. As sequencing technologies improve, updating of publicly available sequence data becomes common practice. We hereby announce the draft genome of four commonly used BCG vaccines in Brazil, Argentina and Venezuela.

Secretory leukocyte protease inhibitor: a secreted pattern recognition receptor for mycobacteria.

RATIONALE: Human secretory leukocyte protease inhibitor (SLPI) displays bactericidal activity against pathogens such as Escherichia coli and Streptococcus. Furthermore, it has been reported that murine SLPI shows potent antimycobacterial activity. OBJECTIVES: The aim of the present study was to investigate whether human recombinant SLPI not only kills mycobacteria but also acts as a pattern recognition receptor for the host immune system. METHODS: For the in vivo experiment, BALB/c mice were infected by intranasal instillation with Mycobacterium bovis BCG and viable BCG load in lung homogenates was later determined. For the in vitro experiments, SLPI was incubated overnight with a suspension of M. bovis BCG or the virulent strain Mycobacterium tuberculosis H37Rv, and the percentage survival as well as the binding of SLPI to mycobacteria was determined. Furthermore, bacteria phagocytosis was also determined by flow cytometry. MEASUREMENTS AND MAIN RESULTS: Intranasal SLPI treatment decreased the number of colony-forming units recovered from lung homogenates, indicating that SLPI interfered with M. bovis BCG infection. Moreover, SLPI decreased the viability of both M. bovis BCG and H37Rv. We demonstrated that SLPI attached to the surface of the mycobacteria by binding to pathogen-associated molecular pattern mannan-capped lipoarabinomannans and phosphatidylinositol mannoside. Furthermore, we found that in the sputum of patients with tuberculosis, mycobacteria were coated with endogenous SLPI. Finally, we showed that phagocytosis of SLPI-coated mycobacteria was faster than that of uncoated bacteria. CONCLUSIONS: The present results demonstrate for the first time that human SLPI kills mycobacteria and is a new pattern recognition receptor for them.

Cathepsin B is involved in the apoptosis intrinsic pathway induced by Bacillus Calmette-Guérin in transitional cancer cell lines.

Bacillus Calmette-Guérin (BCG) is the most effective treatment for superficial and in situ transitional bladder cancer. Although the complete mechanisms for its effect are not fully understood yet, both immunological and direct effects on tumor cells have been proposed. It has been proposed that apoptotic tumor cells could be better inducers of immunity than necrotic ones. Thus, apoptosis of bladder cancer cells could contribute to a global response to BCG. Lysosomal hydrolase cathepsin B (CB) is involved in the apoptotic process and has a key role in breast cancer cell programmed death through the activation of a pro-apoptotic protein BID. Truncated BID participates in the mitochondrial apoptotic pathway that involves the activation of pro-caspase 9. The possibility that CB can be involved in apoptosis of TCC line has not been explored yet. Therefore, we analyzed the participation of CB in BCG-induced apoptosis of human and murine TCC lines. Apoptosis was evaluated by a morphologic assay and CB activity by a substrate-specific colorimetric method. Expression of CB, BID and pro-caspase 9 was determined by Western blotting. BCG induced apoptosis of murine (MBT2, MB49) and human (T24) TCC lines. An increase in both CB activity and protein was also observed. The apoptosis of T24 and MB49 cell lines was mediated by activation of pro-caspase 9 and BID, both proteins are involved in mitochondrial apoptosis. Apoptosis and activation of pro-caspase 9 and BID were inhibited by CA-074Me (CA), a cell permeable CB inhibitor. Thus, CB is involved in BCG-induced apoptosis of TCC lines, using at least in part the mitochondrial pathway.