In Vitro Selection and In Vivo Efficacy of Piperazine- and Alkanediamide-Linked Bisbenzamidines against Pneumocystis Pneumonia in Mice

Abstract

Bisbenzamidines, such as pentamidine isethionate, are aromatic dicationic compounds that are active against Pneumocystis and other microbes but are oftentimes toxic to the host. To identify potential antiPneumocystis agents, we synthesized bisbenzamidine derivatives in which the parent compound pentamidine was modified by a 1,4-piperazinediyl, alkanediamide, or 1,3-phenylenediamide moiety as the central linker. Several of the compounds were more active against P. carinii and less toxic than pentamidine in cytotoxicity assays. For this study, we evaluated nine bisbenzamidine derivatives representing a range of in vitro activities, from highly active to inactive, for the treatment of pneumocystosis in an immunosuppressed mouse model. Six of these in vitro-active compounds, 01, 02, 04, 06, 100, and 101, exhibited marked efficacies against infection at a dose of 10 mg/kg of body weight, and four compounds, 01, 04, 100, and 101, showed significant increases in survival versus that of untreated infected control mice. Compound 100 was highly efficacious against the infection at 20 mg/kg and 40 mg/kg, with >1,000-fold reductions in burden, and resulted in improved survival curves versus those for pentamidine-treated mice (at the same doses). All six bisbenzamidine compounds that exhibited high in vitro activity significantly decreased the infection in vivo; two compounds, 12 and 102, with marked to moderate in vitro activities had slight or no activity in vivo, while compound 31 was inactive in vitro and was also inactive in vivo. Thus, the selection of highly active compounds from in vitro cytotoxicity assays was predictive of activity in the mouse model of Pneumocystis pneumonia. We conclude that a number of these bisbenzamidine compounds, especially compound 100, may show promise as new anti-Pneumocystis drugs.

Biofilm Formation by Pneumocystis spp

“…Pneumocystis carinii (from rats) and P. murina (from mice) were used immediately after isolation and purification from infected rodent lungs or from cryopreserved aliquots. Infected lungs were removed from rats or mice after approximately 8 weeks of chronic administration of dexamethasone as an immunosuppressant (14). Organisms were released from lung tissue by homogenization followed by washing, hypotonic lysis (to reduce erythrocytes and other host cells), and then centrifugation at 1,000 ϫ g. The organisms were enumerated by tinctorial methods using a rapid Giemsa stain, Hema3 (Fisher Diagnostics, Middletown, VA), and expressed as the number of Pneumocystis nuclei/ml (13).…”

Novel pentamidine derivatives: Synthesis, anti-tumor properties and polynucleotide-binding activities

“…Amidinophenyl substituted benzo and pyridine dicarboxamides were prepared and tested for their antimicrobial activity or cytotoxic activity against several human cancer cell lines [12][13][14][15]. Although they showed moderate activity against P. carinii and Plasmodium falciparum, their poor antitumor activity was poor [6,14,16].…”

Pneumocystis Workshop: 10th Anniversary Summary

“…The reductions were comparable to those achieved with trimethoprim-sulfamethoxazole, suggesting that this drug class may have promise for treating clinical infections. Interestingly, in vitro studies from the same laboratory that evaluated the echinocandins in the mouse model of pneumocystosis showed little to no effect at concentrations of 100 g/ml or below in an established short-term drug screening assay based on ATP levels (23). Consideration of recent reports of the effects of sera on MICs of echinocandins in assays of Candida and Aspergillus (107) resulted in a serum concentration study with anidulafungin, caspofungin, and micafungin against P. carinii planktonic and biofilm cultures (16).…”

“…It was recently reported that blockade of Hsp90 in several yeast erg3 mutants reduced their resistance to azoles. Because P. carinii appears to lack the Erg3 gene, actively transcribes Hsp90, and has a functional Erg11 gene (the target of azoles), the effects of the combination of geldanamycin (GDA), an Hsp90 inhibitor, and various sterol pathway inhibitors were evaluated in an ATP assay system (17,23). A range of results were observed.…”