Identification of novel chemotherapeutic strategies for metastatic uveal melanoma.

Melanoma of the uveal tract accounts for approximately 5% of all melanomas and represents the most common primary intraocular malignancy. Despite improvements in diagnosis and more effective local therapies for primary cancer, the rate of metastatic death has not changed in the past forty years. In the present study, we made use of bioinformatics to analyze the data obtained from three public available microarray datasets on uveal melanoma in an attempt to identify novel putative chemotherapeutic options for the liver metastatic disease. We have first carried out a meta-analysis of publicly available whole-genome datasets, that included data from 132 patients, comparing metastatic vs. non metastatic uveal melanomas, in order to identify the most relevant genes characterizing the spreading of tumor to the liver. Subsequently, the L1000CDS2 web-based utility was used to predict small molecules and drugs targeting the metastatic uveal melanoma gene signature. The most promising drugs were found to be Cinnarizine, an anti-histaminic drug used for motion sickness, Digitoxigenin, a precursor of cardiac glycosides, and Clofazimine, a fat-soluble iminophenazine used in leprosy. In vitro and in vivo validation studies will be needed to confirm the efficacy of these molecules for the prevention and treatment of metastatic uveal melanoma.

Transcriptional changes that characterize the immune reactions of leprosy.

BACKGROUND: Leprosy morbidity is increased by 2 pathologic immune reactions, reversal reaction (RR) and erythema nodosum leprosum (ENL). METHODS: To discover host factors related to immune reactions, global transcriptional profiles of peripheral blood mononuclear cells were compared between 11 RR, 11 ENL, and 19 matched control patients, with confirmation by quantitative polymerase chain reaction. Encoded proteins were investigated in skin biopsy specimens by means of immunohistochemistry. RESULTS: There were 275 genes differentially expressed in RR and 517 differentially expressed in ENL on the microarray. Pathway analysis showed immunity-related pathways represented in RR and ENL transcriptional profiles, with the "complement and coagulation" pathway common to both. Interferon γ was identified as a significant upstream regulator of the expression changes for RR and ENL. Immunohistochemical staining of skin lesions showed increased C1q in both RR and ENL. CONCLUSIONS: These data suggest a previously underrecognized role for complement in the pathogenesis of both RR and ENL, and we propose new hypotheses for reaction pathogenesis.

The Haemophilus ducreyi Fis protein is involved in controlling expression of the lspB-lspA2 operon and other virulence factors.

Expression of the lspB-lspA2 operon encoding a virulence-related two-partner secretion system in Haemophilus ducreyi 35000HP is directly regulated by the CpxRA regulatory system (M. Labandeira-Rey, J. R. Mock, and E. J. Hansen, Infect. Immun. 77:3402-3411, 2009). In the present study, we show that this secretion system is also regulated by the small nucleoid-associated protein Fis. Inactivation of the H. ducreyi fis gene resulted in a reduction in expression of both the H. ducreyi LspB and LspA2 proteins. DNA microarray experiments showed that a H. ducreyi fis deletion mutant exhibited altered expression levels of genes encoding other important H. ducreyi virulence factors, including DsrA and Flp1, suggesting a possible global role for Fis in the control of virulence in this obligate human pathogen. While the H. ducreyi Fis protein has a high degree of sequence and structural similarity to the Fis proteins of other bacteria, its temporal pattern of expression was very different from that of enterobacterial Fis proteins. The use of a lacZ-based transcriptional reporter provided evidence which indicated that the H. ducreyi Fis homolog is a positive regulator of gyrB, a gene that is negatively regulated by Fis in enteric bacteria. Taken together, the Fis protein expression data and the observed regulatory effects of Fis in H. ducreyi suggest that this small DNA binding protein has a regulatory role in H. ducreyi which may differ in substantial ways from that of other Fis proteins.

Characterization of the MSMEG_2631 gene (mmp) encoding a multidrug and toxic compound extrusion (MATE) family protein in Mycobacterium smegmatis and exploration of its polyspecific nature using biolog phenotype microarray.

In Mycobacterium, multidrug efflux pumps can be associated with intrinsic drug resistance. Comparison of putative mycobacterial transport genes revealed a single annotated open reading frame (ORF) for a multidrug and toxic compound extrusion (MATE) family efflux pump in all sequenced mycobacteria except Mycobacterium leprae. Since MATE efflux pumps function as multidrug efflux pumps by conferring resistance to structurally diverse antibiotics and DNA-damaging chemicals, we studied this gene (MSMEG_2631) in M. smegmatis mc(2)155 and determined that it encodes a MATE efflux system that contributes to intrinsic resistance of Mycobacterium. We propose that the MSMEG_2631 gene be named mmp, for mycobacterial MATE protein. Biolog Phenotype MicroArray data indicated that mmp deletion increased susceptibility for phleomycin, bleomycin, capreomycin, amikacin, kanamycin, cetylpyridinium chloride, and several sulfa drugs. MSMEG_2619 (efpA) and MSMEG_3563 mask the effect of mmp deletion due to overlapping efflux capabilities. We present evidence that mmp is a part of an MSMEG_2626-2628-2629-2630-2631 operon regulated by a strong constitutive promoter, initiated from a single transcription start site. All together, our results show that M. smegmatis constitutively encodes an Na(+)-dependent MATE multidrug efflux pump from mmp in an operon with putative genes encoding proteins for apparently unrelated functions.

Macrophage polarization and bacterial infections.

PURPOSE OF REVIEW: Macrophages are the first line of defense against pathogens, and the mode of their activation will determine the success or failure of the host response to pathogen aggression. Based on limited numbers of markers, activated macrophages can be classified as classically activated (M1) macrophages that support microbicidal activity or alternatively activated (M2) macrophages that are not competent to eliminate pathogens. The development of high-throughput gene expression methods affords a reappraisal of the concept of macrophage activation in human infectious diseases. RECENT FINDINGS: By combining microarray data and conventional approaches, it is becoming clear that the M1 polarization program is associated with gastrointestinal infections (e.g. typhoid fever and Helicobacter pylori gastritis) and active tuberculosis. An M2 signature is observed in lepromatous leprosy, Whipple's disease, and localized infections (keratitis, chronic rhinosinusitis). However, these findings could not be predicted from the analysis of the M1/M2 programs of macrophages stimulated in vitro. SUMMARY: The reappraisal of macrophage polarization by high-throughput methods is critical to understanding the role of macrophage polarization in infectious diseases. Only the identification of individual profiles will support promising therapeutic approaches based on target determination.