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Poster No. E-1183
In Vitro Activity of the Novel Pleuromutilin BC-3781 Tested Against Bacterial Pathogens
Nabriva Therapeutics AG
Causing Sexually Transmitted Diseases (STD)
Paukner S.1, Gruss A.1, Fritsche T.R.,2 Ivezic-Schoenfeld Z.1, Jones R.N.2
Nabriva Therapeutics AG, Vienna, Austria; 2 JMI Laboratories, North Liberty, IA, USA;
[µg/mL] of BC-3781 and comparator antibiotics against
MIC [µg/mL] of BC-3781 and comparator antibiotics against Mycoplasma spp. and Ureaplasma spp.
C. trachomatis, N. gonorrhoeae, H. ducreyi and anaerobic organisms
AMENDED ABSTRACT
Azithro-
Erythro-
Reference
Background: STD are a significant health challenge in Europe and the USA. Chlamydia
n
floxacin
floxacin
C. trachomatis serovars L /434 (ATCC VR-902B), L /404 (ATCC VR-903), A/G-17, B/TW-5, Ba/AP-2 trachomatis infection and gonorrhea caused by Neisseria gonorrhoeae are the most (ATCC VR-347), E/UW-5, G/UW-57 (ATCC VR-878), SA f were cultured in McCoy cells (ATCC CRL- frequently reported sexually transmitted and reportable diseases in both areas with high 1696) and the serovars C/TW-3 (ATCC VR-578), D/UW-3 (ATCC VR-885), F/MRC 301, J/UW-36 (ATCC VR-886), K/UW-31 (ATCC VR-887) were cultured in HeLa 229 cells (ATCC CCL-2.1). MIC infection rates among young persons particularly women. Decreased susceptibility of determinations were performed essentially as described earlier6 on monolayers of the respective N. gonorrhoeae isolates to azithromycin, cefixime and ceftriaxone, recommended eukaryotic cells infected with C. trachomatis [102-103 inclusion forming units (IFU) per 3x105 cells] on therapies for gonorrhea, is extremely concerning. BC-3781 is a novel pleuromutilin for oral glass cover slips at drug concentrations ranging from 12.8-0.0003 µg/mL. After incubation at 35°C and intravenous administration as treatment of bacterial skin and respiratory infections. (5% CO ) for 48-72 h the cells were fixed in methanol and inclusions were stained with alcoholic iodine This study evaluated its in vitro activity against the most prevalent bacterial pathogens solution. The MIC was defined as the lowest amount of antibiotic at which no inclusion was observed. Susceptibility testing of M. genitalium (ATCC 33530), M. hominis (ATCC 23114), U. urealyticum (ATCC Methods: The antibacterial activity of BC-3781 and comparators was tested against
27814) was performed by broth microdilution as described earlier.7-9 Initial MICs were read when the change of color in the broth was first observed in the control wells (typically after 5-7 days). Susceptibility C. trachomatis (serovars A-K, L2,L3 and SA2f; n=15) in McCoy and HeLa229 cells and testing of the multi-drug resistant M. genitalium isolates was performed using the Vero cell culture and Mycoplasma genitalium (n=6), M. hominis (n=1), Ureaplasma urealyticum (n=1) by broth quantitative real-time PCR method as described earlier.10 Here, the MIC was expressed as the minimal microdilution methods. N. gonorrhoeae (n=24), the anaerobe Peptostreptococcus spp. concentration of the test-antibiotic causing a 99% inhibition of growth when compared to the mean of the (n=10) and Prevotella spp. (n=10) were tested by agar dilution methods (CLSI).
control. N. gonorrhoeae testing was performed by agar dilution technique as described by CLSI (M07-09, Results: BC-3781 demonstrated potent activity against C. trachomatis including serovars 2012). N. gonorrhoeae ATCC 49226 was included as a QC reference strain with all results being within
≤0.008->4
established limits. Haemophilus ducreyi MIC determination was performed as described earlier.11,12 MIC 8->128
determination against the anaerobic organisms was performed by agar dilution as described by CLSI 0.01-0.04 µg/mL). This was comparable to the activity of moxifloxacin, clarithromycin and doxycycline and 5-fold more active than azithromycin or erythromycin (MIC CONCLUSIONS
µg/mL). BC-3781 was also active against N. gonorrhoeae (MIC ciprofloxacin- and tetracycline-resistant strains being inhibited by BC-3781. BC-3781 demonstrated also activity against Mycoplasma spp. including the macrolide- resistant The high potency of BC-3781 against N. gonorrhoeae, C. trachomatis, Mycoplasma species, C. trachomatis
M. hominis (0.04 µg/mL), multi-drug resistant M. genitalium (0.016-0.063 µg/mL) and H. ducreyi and anaerobic organisms suggest that BC-3781 could be a promising first-line antibiotic  BC-3781 exhibited potent activity against the intracellular C. trachomatis including against U. urealyticum (1.6 µg/mL). All Peptostreptococcus spp. and 80% of Prevotella for the treatment of STD such as gonorrhea, non-gonococcal urethritis, cervicitis, chancroid and serovars A, B, Ba, C causing ocular trachoma, serovars D, E, F, G, J, K causing spp. isolates were inhibited at BC-3781 concentrations of ≤2 µg/mL. Porphyromonas spp. pelvic inflammatory disease, especially in populations with high resistance rates to standard of care oculogenital infections and the LGV strains L , L and SA f causing lymphogranuloma was highly susceptible to BC-3781 with MICs of 0.03 µg/mL. Good BC-3781 activity was of 0.02/0.04 µg/mL (Table 1). This was comparable to also demonstrated against H. ducreyi (MIC ≤0.015-0.25 µg/mL).
doxycycline, moxifloxacin and clarithromycin and 5-fold more active than azithromycin or Conclusion: Overall, BC-3781 displayed potent activity against the most relevant
Sexually transmitted diseases represent a major public health crisis; there is multi-drug resistance bacterial pathogens causing STD warranting further investigations on the potential of present and adequate oral therapies are lacking.
N. gonorrhoeae
 As BC-3781 is available as an intravenous as well as oral formulation (tablet) and active against 0.12->32
Against N. gonorrhoeae BC-3781 displayed MIC multi-drug resistant bacterial isolates, further studies are warranted to explore the BC-3781 activity INTRODUCTION
active against fluoroquinolone-, tetracycline- and aminoglycoside-resistant isolates against larger collections of isolates and to demonstrate its activity in human clinical STD trials.
Sexually transmitted diseases (STD) are a significant health challenge in the USA and in H. ducreyi
Europe. Infections caused by the bacterium Chlamydia trachomatis accounted for  BC-3781 demonstrated potent activity against H. ducreyi causing chancroid, a sexually 1.4 million cases in 2011 in USA. The US Centers for Disease Control and Prevention transmitted infection common in Africa and SE Asia. It displayed a MIC range of ≤0.03->32
SELECTED REFERENCES
(CDC) reports gonorrhea (GC) (caused by Neisseria gonorrhoeae) as one of the most ≤ 0.015-0.25 µg/mL, which was comparable to the activity of tetracyclines, erythromycin common sexually transmitted diseases in the US, with more than 800,000 cases estimated and ß-lactam antibiotics (Table 1).
1 . CDC. http://www.cdc.gov/std/stats00/2000Chlamydia.htm to occur each year.1 Overall, chlamydia and GC are the most frequently reported sexually 2. ECDC. Annual Epidemiological Report. Reporting on 2010 Surveillance Data and 2011 Epidemic Anaerobic organisms
transmitted and reportable infections in both Europe and the USA with high occurrence  When tested against the anaerobic Peptostreptococcus spp., Prevotella spp., 3. WHO. Global Incidence and Prevalence of Selected Curable Sexually Transmitted Infections - 2008. (2012) rates among young persons, particularly women.1,2 WHO estimated ~105 million Porphyromonas spp., which are commonly involved in pelvic inflammatory disease 4. Whiley, D. M., Goire, N., Lahra, M. M., Donovan, B., Limnios, A. E., Nissen, M. D., et al. J. Antimicrob. C. trachomatis and ~106 million GC infections globally in 2008.3 Left untreated, these among other female genital tract infections, soft tissue infections, abscesses and Chemother. 67(9), 2059 (2012)
infections can cause serious health problems, particularly for women, including chronic infections of the oral cavity, BC-3781 displayed potent activity with all 5. Unemo, M. and Nicholas, R. A. Future. Microbiol. 7(12), 1401 (2012)
pelvic pain, life-threatening ectopic pregnancy, pelvic inflammatory disease and infertility.3 6. Roblin, P. M., Kohlhoff, S. A., Parker, C., Hammerschlag, M. R. Antimicrob. Agents Chemother. 54(3), 1358
Gonorrhea infection also increases a person’s risk of contracting and transmitting HIV.4 Peptostreptococcus spp. and Prevotella spp. being inhibited at BC-3781 concentration a, contains serovars A, B, Ba,C, D, E, F, G, J, K, L2, L3, SA2f (LGV lab strain), H, I; MIC test in HeLa229 and Control strategy relies on effective antibiotic therapy. STD organisms are progressively of ≤ 2 µg/mL and all Porphyromonas spp. strains at 0.03 µg/mL (Table 1).
7 . Hannan, P. C., Windsor, H. M., Ripley, P. H. Res. Vet. Sci. 63(2), 157 (1997)
developing resistance to the antibiotic drugs prescribed to treat them: sulfonilamides, , includes: Finegoldia magna (3), P. anaerobius (1), P. asaccharolyticus (2), P. micros (2), and P. tetradius (2) Mycoplasma genitalium and other Mycoplasma/Ureaplasma spp.
c, includes: P. bivia (4), P. intermedia (2), P. melaninogenica (2) and P. oralis (2) 8. Lorian, V. Antibiotics in laboratory medicine, 5th Ed. Lippincott Williams & Wilkins, Philadelphia, USA (2005) penicillin, cephalosporins, azithromycin, tetracycline, and ciprofloxacin. GC resistant to all d, includes: Porphyromonas gingivalis (9), and unspeciated Porphyromonas (1)  BC-3781 showed good activity against the tested Mycoplasma species with all isolates . Ridgway, G. Antimicrobial susceptibility testing of intracellular and cell-associated pathogens. EUCAST e, Criteria as published by the CLSI [2013] of these antibiotics have been reported globally.5 New antibacterials overcoming those Discussion Document E. Dis 6.1 (2001)
being inhibited at a BC-3781 concentration of ≤ 0.06 µg/mL. When compared with infections, preferably available as oral formulations, are therefore urgently needed.
10.Hamasuna, R., Osada, Y., Jensen, J. S. Antimicrob. Agents Chemother. 49(12), 4993 (2005)
standard of care medications, BC-3781 was among the most active compounds ACKNOWLEDGMENTS
1 1 . Fortney, K., Totten, P. A., Lehrer, R. I., Spinola, S. M. Antimicrob. Agents Chemother. 42(10), 2690 (1998)
BC-3781 is a novel pleuromutilin antibiotic in development for oral and intravenous 12.Knapp, J. S., Back, A. F., Babst, A. F., Taylor, D., Rice, R. J. Antimicrob. Agents Chemother. 37(7), 1552
administration in treatment of bacterial skin and respiratory infections. The presented study  When tested against very recent multi-drug resistant clinical isolates from patients failing We thank J.S. Jensen from the Statens Serum Institute (Denmark) for demonstrated the activity of BC-3781 against the most prevalent bacterial pathogens treatment with high doses of azithromycin, moxifloxacin, and doxycycline, promt testing of multi-drug resistant M. genitalium isolates and kind 13.Sader, H. S., Paukner, S., Ivezic-Schoenfeld, Z., Biedenbach, D. J., Schmitz, F. J., Jones, R. N. J. causing STD. This includes C. trachomatis, gonococci, mycoplasmas, ureaplasmas and BC-3781 displayed potent activity with MICs ranging from 0.016-0.063 µg/mL.
provision of results for this poster.
Antimicrob. Chemother. 67(5), 1170 (2012)
anaerobic cocci and bacilli causing gonorrhea, non-gonococcal urethritis, cervicitis and pelvic inflammatory disease. 53rd Interscience Conference on Antimicrobial Agents and Chemotherapy, Denver, CO, September 10-13, 2013

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