Inhibition of mycobacterial alanine racemase activity and growth by thiadiazolidinones.

The genus Mycobacterium includes non-pathogenic species such as M. smegmatis, and pathogenic species such as M. tuberculosis, the causative agent of tuberculosis (TB). Treatment of TB requires a lengthy regimen of several antibiotics, whose effectiveness has been compromised by the emergence of resistant strains. New antibiotics that can shorten the treatment course and those that have not been compromised by bacterial resistance are needed. In this study, we report that thiadiazolidinones, a relatively little-studied heterocyclic class, inhibit the activity of mycobacterial alanine racemase, an essential enzyme that converts l-alanine to d-alanine for peptidoglycan synthesis. Twelve members of the thiadiazolidinone family were evaluated for inhibition of M. tuberculosis and M. smegmatis alanine racemase activity and bacterial growth. Thiadiazolidinones inhibited M. tuberculosis and M. smegmatis alanine racemases to different extents with 50% inhibitory concentrations (IC50) ranging from <0.03 to 28µM and 23 to >150µM, respectively. The compounds also inhibited the growth of these bacteria, including multidrug resistant strains of M. tuberculosis. The minimal inhibitory concentrations (MIC) for drug-susceptible M. tuberculosis and M. smegmatis ranged from 6.25µg/ml to 100µg/ml, and from 1.56 to 6.25µg/ml for drug-resistant M. tuberculosis. The in vitro activities of thiadiazolidinones suggest that this family of compounds might represent starting points for medicinal chemistry efforts aimed at developing novel antimycobacterial agents.

Thiadiazolidinones: a new class of alanine racemase inhibitors with antimicrobial activity against methicillin-resistant Staphylococcus aureus.

Methicillin-resistant Staphylococcus aureus (MRSA) is a human pathogen and a major cause of hospital-acquired infections. New antibacterial agents that have not been compromised by bacterial resistance are needed to treat MRSA-related infections. We chose the S. aureus cell wall synthesis enzyme, alanine racemase (Alr) as the target for a high-throughput screening effort to obtain novel enzyme inhibitors, which inhibit bacterial growth. Among the 'hits' identified was a thiadiazolidinone with chemical properties attractive for lead development. This study evaluated the mode of action, antimicrobial activities, and mammalian cell cytotoxicity of the thiadiazolidinone family in order to assess its potential for development as a therapeutic agent against MRSA. The thiadiazolidones inhibited Alr activity with 50% inhibitory concentrations (IC50) ranging from 0.36 to 6.4 µM, and they appear to inhibit the enzyme irreversibly. The series inhibited the growth of S. aureus, including MRSA strains, with minimal inhibitory concentrations (MICs) ranging from 6.25 to 100 µg/ml. The antimicrobial activity showed selectivity against Gram-positive bacteria and fungi, but not Gram-negative bacteria. The series inhibited human HeLa cell proliferation. Lead development centering on the thiadiazolidinone series would require additional medicinal chemistry efforts to enhance the antibacterial activity and minimize mammalian cell toxicity.

The inhibitory effect of minocycline on hypertrophic scarring.

BACKGROUND: Matrix metalloproteinases are enzymes that serve to degrade the extracellular matrix, giving them a central role in the inflammatory and wound-healing processes; they have been implicated in the pathophysiology of hypertrophic scarring. The purpose of this study was to examine the effect of minocycline, a matrix metalloproteinase inhibitor, on hypertrophic scarring. METHODS: Standardized wounds were created on the ears of eight New Zealand White rabbits. Half of the rabbits received daily injections of minocycline, whereas the other half received daily injections of saline (control). After 4 weeks, the resulting ear scars were harvested. Histologic slides were prepared from the thickest cross-sections of the scars, and from these slides the cross-sectional area of each scar was measured. A hypertrophic index was calculated by comparing the area of the scar to the baseline value of unwounded skin. Statistical analysis was performed using the SAS/STAT NESTED Procedure for hierarchical data. RESULTS: Among the rabbits treated with minocycline, the mean hypertrophic index was 1.08 +/- 0.01, compared with 1.54 +/- 0.03 in the control group (p = 0.03), representing an 85 percent reduction in hypertrophic area. CONCLUSIONS: Systemically administered minocycline significantly reduces the severity of hypertrophic scarring in a rabbit model. Although not directly examined in this study, matrix metalloproteinase inhibition is hypothesized to be responsible for this effect.

The pathology of extrapulmonary small cell carcinoma.

Extrapulmonary small cell carcinomas (EPSCCs) are uncommon malignant neoplasms with a reported incidence of 0.1% to 0.4% in the United States. Since their first description in 1930, they have been seen in nearly every organ system. Like their more common pulmonary counterparts, EPSCCs are thought to arise from a multipotential stem cell. However, there is recent molecular evidence that small cell elements may arise as a late-stage phenomenon in the genetic progression of more organ-typical carcinomas. The morphologic, immunohistochemical, and ultrastructural features are similar to those described in pulmonary small cell carcinomas (PSCCs). The differential diagnosis of EPSCC includes PSCC, other neuroendocrine tumors, small round blue cell tumors, metastatic melanoma, lymphoma, and poorly differentiated non-small cell carcinomas. Molecular alterations reported to occur in EPSCCs include abnormalities described in PSCC and changes found in carcinomas more typically encountered in the organ from which they arise. In this article we discuss the pathology of EPSCC with a review of theories of histogenesis, sites of occurrence, diagnostic features, differential diagnosis, molecular alterations, and clinical behavior.

1D5 and 6F11: An immunohistochemical comparison of two monoclonal antibodies for the evaluation of estrogen receptor status in primary breast carcinoma.

The optimal monoclonal antibody to examine steroid hormone receptor status of primary breast carcinoma has yet to be defined. Estrogen receptor status was evaluated in 592 cases using routinely prepared paraffin-embedded tissue samples from primary breast carcinomas with the 1D5 (DAKO, Carpinteria, CA) and 6F11 (Novocastra, Newcastle upon Tyne, England) monoclonal antibodies. The stains were compared, assessing the percentage of positive cells stained and their intensity. They also were examined for nonspecific cytoplasmic staining and fixation artifact. In addition, a cost analysis for their production was performed. Overall, 1D5 and 6F11 showed a 97.5% concordance rate. 6F11 stained a significantly higher percentage of cells (P < .0001), more intensely (P < .0001), with less nonspecific cytoplasmic staining (P < .0001). There was no significant difference in fixation artifact between the 2 clones. The cost of antibody used for preparing a 1D5-stained slide was 86% more than for preparing a 6F11-stained slide (dollars 14.27 vs dollars 7.67).

Melanosis of the uterine cervix: a case report.

BACKGROUND: Melanosis of the uterine cervix is a rare, benign, melanocytic lesion of the cervix. A review of the English-language literature revealed 11 other reported cases. CASE: A 52-year-old African American woman who presented for evaluation and treatment of menorrhagia was found to have an irregular, pigmented cervical lesion. The ectocervical biopsy showed melanosis characterized by hyperkeratosis, acanthosis and prominent elongation of rete with abundant basilar pigmentation. Immunohistochemical staining showed scattered, basally situated, S-100 protein-positive melanocytes. CONCLUSION: The existence of this entity is further evidence of a spectrum of melanocytic lesions, including blue nevi and primary melanomas, known to occur within the cervix proper.

Villous adenocarcinoma of the colon and rectum: a clinicopathologic study of 36 cases.

Some colorectal adenocarcinomas show villous architecture with morphologic similarities to tubulovillous or villous adenomas. We reviewed 420 consecutive colorectal adenocarcinoma resection specimens and found that 95 tumors (23%) showed areas of villous architecture. Thirty-six tumors (8.6%) in 35 patients showed more than 50% villous architecture and were designated villous adenocarcinomas. Only 42% of the villous adenocarcinomas showed severe atypia and only 44% of the available pre-resection biopsies of these tumors were diagnosed as adenocarcinoma. Epithelial islands in desmoplastic stroma (EIDS) may be helpful in the diagnosis of these tumors. EIDS were found in 97% of the resection specimens for villous adenocarcinomas and none of 62 resection specimens for tubulovillous or villous adenomas. The presence of EIDS showed a 67% sensitivity, 100% specificity, and 100% predictive value in the diagnosis of villous adenocarcinoma in a blinded review of villous tumors. On review of the pre-resection biopsies of villous adenocarcinoma without a final diagnosis of adenocarcinoma, 40% showed EIDS. Clinical follow-up of the 35 patients with villous adenocarcinoma showed that only one died of colorectal adenocarcinoma (median follow-up, 46 months). This sole patient dying of colorectal adenocarcinoma showed a synchronous advanced stage of nonvillous adenocarcinoma at the time of diagnosis. Villous adenocarcinoma is a diagnostically challenging subset of colorectal adenocarcinoma, which appears to be associated with a favorable prognosis. Classifying these tumors as a special type of colorectal cancer may facilitate the development of diagnostic adjuncts and optimal treatment protocols.