Skip to main content
  • ASM
    • Antimicrobial Agents and Chemotherapy
    • Applied and Environmental Microbiology
    • Clinical Microbiology Reviews
    • Clinical and Vaccine Immunology
    • EcoSal Plus
    • Eukaryotic Cell
    • Infection and Immunity
    • Journal of Bacteriology
    • Journal of Clinical Microbiology
    • Journal of Microbiology & Biology Education
    • Journal of Virology
    • mBio
    • Microbiology and Molecular Biology Reviews
    • Microbiology Resource Announcements
    • Microbiology Spectrum
    • Molecular and Cellular Biology
    • mSphere
    • mSystems
  • Log in
  • My alerts
  • My Cart

Main menu

  • Home
  • Articles
    • Current Issue
    • Accepted Manuscripts
    • COVID-19 Special Collection
    • Archive
    • Minireviews
  • For Authors
    • Submit a Manuscript
    • Scope
    • Editorial Policy
    • Submission, Review, & Publication Processes
    • Organization and Format
    • Errata, Author Corrections, Retractions
    • Illustrations and Tables
    • Nomenclature
    • Abbreviations and Conventions
    • Publication Fees
    • Ethics Resources and Policies
  • About the Journal
    • About AAC
    • Editor in Chief
    • Editorial Board
    • For Reviewers
    • For the Media
    • For Librarians
    • For Advertisers
    • Alerts
    • AAC Podcast
    • RSS
    • FAQ
  • Subscribe
    • Members
    • Institutions
  • ASM
    • Antimicrobial Agents and Chemotherapy
    • Applied and Environmental Microbiology
    • Clinical Microbiology Reviews
    • Clinical and Vaccine Immunology
    • EcoSal Plus
    • Eukaryotic Cell
    • Infection and Immunity
    • Journal of Bacteriology
    • Journal of Clinical Microbiology
    • Journal of Microbiology & Biology Education
    • Journal of Virology
    • mBio
    • Microbiology and Molecular Biology Reviews
    • Microbiology Resource Announcements
    • Microbiology Spectrum
    • Molecular and Cellular Biology
    • mSphere
    • mSystems

User menu

  • Log in
  • My alerts
  • My Cart

Search

  • Advanced search
Antimicrobial Agents and Chemotherapy
publisher-logosite-logo

Advanced Search

  • Home
  • Articles
    • Current Issue
    • Accepted Manuscripts
    • COVID-19 Special Collection
    • Archive
    • Minireviews
  • For Authors
    • Submit a Manuscript
    • Scope
    • Editorial Policy
    • Submission, Review, & Publication Processes
    • Organization and Format
    • Errata, Author Corrections, Retractions
    • Illustrations and Tables
    • Nomenclature
    • Abbreviations and Conventions
    • Publication Fees
    • Ethics Resources and Policies
  • About the Journal
    • About AAC
    • Editor in Chief
    • Editorial Board
    • For Reviewers
    • For the Media
    • For Librarians
    • For Advertisers
    • Alerts
    • AAC Podcast
    • RSS
    • FAQ
  • Subscribe
    • Members
    • Institutions
Experimental Therapeutics

Exebacase in Addition to Daptomycin Is More Active than Daptomycin or Exebacase Alone in Methicillin-Resistant Staphylococcus aureus Osteomyelitis in Rats

Melissa J. Karau, Suzannah M. Schmidt-Malan, Qun Yan, Kerryl E. Greenwood-Quaintance, Jayawant Mandrekar, Dario Lehoux, Raymond Schuch, Cara Cassino, Robin Patel
Melissa J. Karau
aDivision of Clinical Microbiology, Department of Laboratory Medicine and Pathology, Mayo Clinic, Rochester, Minnesota, USA
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Suzannah M. Schmidt-Malan
aDivision of Clinical Microbiology, Department of Laboratory Medicine and Pathology, Mayo Clinic, Rochester, Minnesota, USA
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Qun Yan
bDepartment of Clinical Laboratory, Xiangya Hospital of Central South University, Changsha, Hunan, China
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Kerryl E. Greenwood-Quaintance
aDivision of Clinical Microbiology, Department of Laboratory Medicine and Pathology, Mayo Clinic, Rochester, Minnesota, USA
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Jayawant Mandrekar
cDivision of Biomedical Statistics and Informatics, Department of Health Sciences Research, Mayo Clinic, Rochester, Minnesota, USA
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Dario Lehoux
dContraFect, Yonkers, New York, USA
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Raymond Schuch
dContraFect, Yonkers, New York, USA
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
  • ORCID record for Raymond Schuch
Cara Cassino
dContraFect, Yonkers, New York, USA
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Robin Patel
aDivision of Clinical Microbiology, Department of Laboratory Medicine and Pathology, Mayo Clinic, Rochester, Minnesota, USA
eDivision of Infectious Diseases, Department of Medicine, Mayo Clinic, Rochester, Minnesota, USA
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
  • ORCID record for Robin Patel
DOI: 10.1128/AAC.01235-19
  • Article
  • Figures & Data
  • Info & Metrics
  • PDF
Loading

ABSTRACT

Bacteriophage-derived lysins are being developed as anti-infective agents. In an acute osteomyelitis methicillin-resistant Staphylococcus aureus (MRSA) model, rats receiving no treatment or treatment with daptomycin, exebacase (CF-301), or daptomycin plus exebacase had means of 5.13, 4.09, 4.65, and 3.57 log10 CFU/gram of bone, respectively. All treated animals had fewer bacteria than did untreated animals (P ≤ 0.0001), with daptomycin plus exebacase being more active than daptomycin (P = 0.0042) or exebacase (P < 0.001) alone.

INTRODUCTION

Osteomyelitis is a devastating infection that can be challenging to treat and that may be associated with morbidity, including damage to bone tissue and metastatic infection. Staphylococcus aureus is the most common cause of osteomyelitis (1). S. aureus has the ability to form biofilms and to enter and survive within osteoblasts, both of which may allow it to evade the immune system and be resistant to many traditional antimicrobials (2). Current therapeutic options include surgical irrigation and debridement and long-term therapy with antimicrobials such as daptomycin or vancomycin (1, 3, 4). Compounding the challenge of the virulence of S. aureus itself is the increasing antimicrobial resistance of S. aureus. Methicillin-resistant S. aureus (MRSA) osteomyelitis is growing in frequency and has been associated with poorer outcomes than methicillin-susceptible S. aureus infection (5). There is therefore a need for new, more effective antimicrobial strategies for S. aureus, especially MRSA, bone and joint infections.

Bacteriophage-derived lysins offer a novel therapeutic approach using bacterial species-specific enzymes that hydrolyze the peptidoglycan of the bacterial cell wall (6, 7). As direct lytic agents, lysins do not require bacteria to be actively growing to exert their activity and act immediately upon contact with bacterial cells, thus rendering them as promising potential therapeutics for bone and joint infections (8). Exebacase (CF-301) is a recombinantly produced lytic enzyme derived within a Streptococcus suis prophage (6, 7), with a catalytic N-terminal domain linked to a cell wall-binding C-terminal domain with d-alanyl-l-glycyl endopeptidase activity (6, 7). Exebacase may have antibiofilm activity as a result of its causing bacteriolysis within biofilms as a result of its cleavage of peptidoglycan in the structural framework (8). In vitro, exebacase is rapidly bactericidal, shows minimal resistance development, and has synergistic activity with daptomycin and vancomycin against S. aureus (6, 8). In an experimental murine S. aureus bacteremia model, a single intravenous dose of exebacase with and without vancomycin or daptomycin increased survival (6). The results of a phase 2 clinical trial demonstrate a 43% higher clinical response rate with a single dose of exebacase used in addition to standard-of-care antimicrobials versus antimicrobials alone for MRSA bacteremia, including endocarditis (9, 10). We hypothesized that exebacase would be active against MRSA in experimental acute osteomyelitis in rats.

The study strain, MRSA IDRL-6169, had MICs of 0.5 μg/ml for both exebacase and daptomycin, as determined by broth microdilution (11, 12). The minimum biofilm inhibitory concentrations and minimum biofilm bactericidal concentrations were 1 and 4 μg/ml for exebacase and 1 and 2 μg/ml for daptomycin, as determined using previously described methods (14). Exebacase testing was supplemented with 0.5 mM dl-dithiothreitol and 25% horse serum (6).

The studies described were approved by the Institutional Animal Care and Use Committee of the Mayo Clinic. Osteomyelitis was established in 64 male Sprague Dawley rats using a modification of Zak’s model of experimental osteomyelitis designed to mimic human infection (15). Animals were anesthetized with isoflurane and the left knee shaved and disinfected with chlorohexidine. To induce osteomyelitis, the knee joint was bent at a 45° angle to expose the top of the tibial process. A 1-ml syringe with a 21 G needle containing 10 μl arachidonic acid (50 μg/ml) and 50 μl of a 107 CFU suspension of IDRL-6169 in tryptic soy broth was inserted into the tibia. The bacterial suspension was slowly injected into the tibia, the needle removed, the knee joint straightened, and pressure placed on the injection site for 1 min. One week after establishing infection (day 8), rats were randomly assigned to one of four treatment arms, as follows: (i) no treatment, (ii) 60 mg/kg of body weight daptomycin intraperitoneally every 12 h (16) for 4 days, (iii) single-dose 40 mg/kg exebacase in the tail vein, or (iv) single-dose 40 mg/kg exebacase plus 60 mg/kg daptomycin intraperitoneally every 12 h for 4 days. Daptomycin was administered 15 min prior to exebacase injection. Exebacase, formulated for clinical use, was maintained on ice until injection. Rats were sacrificed 4 days after starting therapy (day 12). The left tibia from each animal was collected, weighed, and cryopulverized for quantitative bacterial culture (15). The results of quantitative cultures were compared using SAS software version 9.4 (SAS, Inc., Cary, NC) using the Kruskal-Wallis test as well as Bonferroni correction for multiple comparisons. Means and standard deviations (SD) were reported as log10 CFU/gram of bone. All tests were two sided; P values less than 0.05 were considered statistically significant.

Rats receiving no treatment had a mean (±SD) bacterial density of 5.13 (±0.34) log10 CFU/gram of bone. Rats in the daptomycin, exebacase, and daptomycin plus exebacase groups had mean (±SD) bacterial densities of 4.09 (±0.37), 4.65 (±0.65), and 3.57 (±0.48) log10 CFU/gram of bone, respectively (Fig. 1). The colony counts in all treatment groups were lower than those of the untreated rats (P ≤ 0.0001). Daptomycin combined with exebacase-treated animals had lower colony counts than did those treated with daptomycin (P = 0.0042) or exebacase (P < 0.0001) alone. The conclusions from the study remain unchanged after application of the Bonferroni correction for multiple comparisons.

FIG 1
  • Open in new tab
  • Download powerpoint
FIG 1

Results of quantitative cultures of the tibias after treatment (log10 CFU of MRSA/gram of bone). All treatment groups had lower bacterial loads than those with no treatment (P ≤ 0.0001), with exebacase combined with daptomycin resulting in lower bacterial loads than those with monotherapy with either exebacase (P < 0.0001) or daptomycin (P = 0.0042). The horizontal lines depict median values.

The animal model used here is one of acute MRSA osteomyelitis. S. aureus bone and joint infections are challenging to treat, partly due to the ability of S. aureus to survive in bone tissue (2). Antimicrobials such as vancomycin may not penetrate bone tissue well (17, 18), yet they are often used to treat osteomyelitis (4). It has been suggested that daptomycin has bone penetration (19). We found levels of exebacase in bone to be ∼15% of plasma levels after a single dose of 10 mg/kg. As determined by enzyme-linked immunosorbent assay (ELISA), the maximum concentration in serum (Cmax) and area under the concentration-time curve (AUC) in plasma and bone were 88 μg/ml and 12 μg/g and 28 μg·h/ml and 4 μg·h/g, respectively. Exebacase may therefore offer a strategy to target S. aureus bone and joint infections through its ability to lyse S. aureus at the site of infection. It was previously shown that exebacase is synergistic with daptomycin, possibly by increasing the ability of daptomycin to bind to its target (6). Thus, the use of exebacase plus daptomycin may offer a treatment option for acute osteomyelitis.

In this study, while treatment with daptomycin or exebacase alone showed a reduction in bacterial counts, exebacase in addition to daptomycin showed a more pronounced effect.

ACKNOWLEDGMENTS

This study was sponsored by ContraFect and a grant from the Department of Defense Congressionally Directed Medical Research Program (award W81XWH-16-1-0245).

FOOTNOTES

    • Received 19 June 2019.
    • Returned for modification 27 June 2019.
    • Accepted 24 July 2019.
    • Accepted manuscript posted online 29 July 2019.
  • Copyright © 2019 American Society for Microbiology.

All Rights Reserved.

REFERENCES

  1. 1.↵
    1. Birt MC,
    2. Anderson DW,
    3. Toby EB,
    4. Wang J
    . 2017. Osteomyelitis: recent advances in pathophysiology and therapeutic strategies. J Orthop 14:45–52. doi:10.1016/j.jor.2016.10.004.
    OpenUrlCrossRef
  2. 2.↵
    1. Kavanagh N,
    2. Ryan EJ,
    3. Widaa A,
    4. Sexton G,
    5. Fennell J,
    6. O’Rourke S,
    7. Cahill KC,
    8. Kearney CJ,
    9. O’Brien FJ,
    10. Kerrigan SW
    . 2018. Staphylococcal osteomyelitis: Disease progression, treatment challenges, and future directions. Clin Microbiol Rev 31:e00084-17. doi:10.1128/CMR.00084-17.
    OpenUrlAbstract/FREE Full Text
  3. 3.↵
    1. Liu C,
    2. Chambers HF,
    3. Kaplan SL,
    4. Karchmer AW,
    5. Levine DP,
    6. Rybak MJ,
    7. Murray BE,
    8. Bayer A,
    9. Talan DA,
    10. Cosgrove SE,
    11. Daum RS,
    12. Fridkin SK,
    13. Gorwitz RJ
    , Infectious Diseases Society of America. 2011. Clinical practice guidelines by the Infectious Diseases Society of America for the treatment of methicillin-resistant Staphylococcus aureus infections in adults and children. Clin Infect Dis 52:e18–e55. doi:10.1093/cid/ciq146.
    OpenUrlCrossRefPubMedWeb of Science
  4. 4.↵
    1. Fraimow HS
    . 2009. Systemic antimicrobial therapy in osteomyelitis. Semin Plastic Surg 23:90–99. doi:10.1055/s-0029-1214161.
    OpenUrlCrossRef
  5. 5.↵
    1. Davis WT,
    2. Gilbert SR
    . 2018. Comparison of methicillin-resistant versus susceptible Staphylococcus aureus pediatric osteomyelitis. J Ped Orthop 38:e285–e291. doi:10.1097/BPO.0000000000001152.
    OpenUrlCrossRef
  6. 6.↵
    1. Schuch R,
    2. Lee HM,
    3. Schneider BC,
    4. Sauve KL,
    5. Law C,
    6. Khan BK,
    7. Rotolo JA,
    8. Horiuchi Y,
    9. Couto DE,
    10. Raz A,
    11. Fischetti V,
    12. Huang DB,
    13. Nowinski RC,
    14. Wittekind M
    . 2014. Combination therapy with lysin CF-301 and antibiotic is superior to antibiotic alone for treating methicillin-resistant Staphylococcus aureus–induced murine bacteremia. J Infect Dis 209:1469–1478. doi:10.1093/infdis/jit637.
    OpenUrlCrossRefPubMed
  7. 7.↵
    1. Lood R,
    2. Molina H,
    3. Fischetti VA
    . 2017. Determining bacteriophage endopeptidase activity using either fluorophore-quencher labeled peptides combined with liquid chromatography-mass spectrometry (LC-MS) or Förster resonance energy transfer (FRET) assays. PLoS One 12:e0173919. doi:10.1371/journal.pone.0173919.
    OpenUrlCrossRef
  8. 8.↵
    1. Schuch R,
    2. Khan BK,
    3. Raz A,
    4. Rotolo JA,
    5. Wittekind M
    . 2017. Bacteriophage lysin CF-301: a potent anti-staphylococcal biofilm agent. Antimicrob Agents Chemother 61:02666-16. doi:10.1128/AAC.02666-16.
    OpenUrlAbstract/FREE Full Text
  9. 9.↵
    ClinicalTrials.gov. 2017. Safety, efficacy and pharmacokinetics of CF-301 vs. placebo in addition to antibacterial therapy for treatment of S. aureus bacteremia, NCT 03163446. https://clinicaltrials.gov/ct2/show/NCT03163446. Accessed 18 March 2019.
  10. 10.↵
    1. Fowler VG,
    2. Das A,
    3. Lipka J,
    4. Schuch R,
    5. Cassino C
    . 2019. Exebacase (lysin CF-301) improved clinical responder rates in methicillin-resistant Staphylococcus aureus bacteremia and endocarditis compared to standard of care antibiotics alone in a first-in-patient phase 2 study. 29th European Congress of Clinical Microbiology and Infectious Diseases Annual Meeting, 13 to 16 April, 2019, Amsterdam, the Netherlands.
  11. 11.↵
    CLSI. 2018. Methods for dilution antimicrobial susceptibility tests for bacteria that grow aerobically, 11th ed. Clinical and Laboratory Standards Institute, Wayne, PA.
  12. 12.↵
    CLSI. 2019. Performance standards for antimicrobial susceptibility testing, 29th ed. Clinical and Laboratory Standards Institute, Wayne, PA.
  13. 13.
    Reference deleted.
  14. 14.↵
    1. Schmidt-Malan SM,
    2. Greenwood Quaintance KE,
    3. Karau MJ,
    4. Patel R
    . 2016. In vitro activity of tedizolid against staphylococci isolated from prosthetic joint infections. Diagn Microbiol Infect Dis 85:77–79. doi:10.1016/j.diagmicrobio.2016.01.008.
    OpenUrlCrossRefPubMed
  15. 15.↵
    1. O’Reilly T,
    2. Mader J
    . 1999. Rat model of bacterial osteomyelitis of the tibia, p 561–575. In Zak O, Sande M (ed), Handbook of animal models of infection. Academic Press, San Diego, CA.
  16. 16.↵
    1. Rouse MS,
    2. Piper KE,
    3. Jacobson M,
    4. Jacofsky DJ,
    5. Steckelberg JM,
    6. Patel R
    . 2006. Daptomycin treatment of Staphylococcus aureus experimental chronic osteomyelitis. J Antimicrob Chemother 57:301–305. doi:10.1093/jac/dki435.
    OpenUrlCrossRefPubMedWeb of Science
  17. 17.↵
    1. Albayati ZAF,
    2. Sunkara M,
    3. Schmidt-Malan SM,
    4. Karau MJ,
    5. Morris AJ,
    6. Steckelberg JM,
    7. Patel R,
    8. Breen PJ,
    9. Smeltzer MS,
    10. Taylor KG,
    11. Merten KE,
    12. Pierce WM,
    13. Crooks PA
    . 2016. Novel bone-targeting agent for enhanced delivery of vancomycin to bone. Antimicrob Agents Chemother 60:1865–1868. doi:10.1128/AAC.01609-15.
    OpenUrlAbstract/FREE Full Text
  18. 18.↵
    1. Karau MJ,
    2. Schmidt-Malan SM,
    3. Greenwood Quaintance KE,
    4. Mandrekar J,
    5. Cai J,
    6. Pierce WM,
    7. Merten K,
    8. Patel R
    . 2013. Treatment of methicillin-resistant Staphylococcus aureus experimental osteomyelitis with bone-targeted vancomycin. Springerplus 2:329. doi:10.1186/2193-1801-2-329.
    OpenUrlCrossRefPubMed
  19. 19.↵
    1. Montange D,
    2. Berthier F,
    3. Leclerc G,
    4. Serre A,
    5. Jeunet L,
    6. Berard M,
    7. Muret P,
    8. Vettoretti L,
    9. Leroy J,
    10. Hoen B,
    11. Chirouze C
    . 2014. Penetration of daptomycin into bone and synovial fluid in joint replacement. Antimicrob Agents Chemother 58:3991–3996. doi:10.1128/AAC.02344-14.
    OpenUrlAbstract/FREE Full Text
PreviousNext
Back to top
Download PDF
Citation Tools
Exebacase in Addition to Daptomycin Is More Active than Daptomycin or Exebacase Alone in Methicillin-Resistant Staphylococcus aureus Osteomyelitis in Rats
Melissa J. Karau, Suzannah M. Schmidt-Malan, Qun Yan, Kerryl E. Greenwood-Quaintance, Jayawant Mandrekar, Dario Lehoux, Raymond Schuch, Cara Cassino, Robin Patel
Antimicrobial Agents and Chemotherapy Sep 2019, 63 (10) e01235-19; DOI: 10.1128/AAC.01235-19

Citation Manager Formats

  • BibTeX
  • Bookends
  • EasyBib
  • EndNote (tagged)
  • EndNote 8 (xml)
  • Medlars
  • Mendeley
  • Papers
  • RefWorks Tagged
  • Ref Manager
  • RIS
  • Zotero
Print

Alerts
Sign In to Email Alerts with your Email Address
Email

Thank you for sharing this Antimicrobial Agents and Chemotherapy article.

NOTE: We request your email address only to inform the recipient that it was you who recommended this article, and that it is not junk mail. We do not retain these email addresses.

Enter multiple addresses on separate lines or separate them with commas.
Exebacase in Addition to Daptomycin Is More Active than Daptomycin or Exebacase Alone in Methicillin-Resistant Staphylococcus aureus Osteomyelitis in Rats
(Your Name) has forwarded a page to you from Antimicrobial Agents and Chemotherapy
(Your Name) thought you would be interested in this article in Antimicrobial Agents and Chemotherapy.
CAPTCHA
This question is for testing whether or not you are a human visitor and to prevent automated spam submissions.
Share
Exebacase in Addition to Daptomycin Is More Active than Daptomycin or Exebacase Alone in Methicillin-Resistant Staphylococcus aureus Osteomyelitis in Rats
Melissa J. Karau, Suzannah M. Schmidt-Malan, Qun Yan, Kerryl E. Greenwood-Quaintance, Jayawant Mandrekar, Dario Lehoux, Raymond Schuch, Cara Cassino, Robin Patel
Antimicrobial Agents and Chemotherapy Sep 2019, 63 (10) e01235-19; DOI: 10.1128/AAC.01235-19
del.icio.us logo Digg logo Reddit logo Twitter logo CiteULike logo Facebook logo Google logo Mendeley logo
  • Top
  • Article
    • ABSTRACT
    • INTRODUCTION
    • ACKNOWLEDGMENTS
    • FOOTNOTES
    • REFERENCES
  • Figures & Data
  • Info & Metrics
  • PDF

KEYWORDS

CF-301
daptomycin
exebacase
methicillin-resistant Staphylococcus aureus
osteomyelitis

Related Articles

Cited By...

About

  • About AAC
  • Editor in Chief
  • Editorial Board
  • Policies
  • For Reviewers
  • For the Media
  • For Librarians
  • For Advertisers
  • Alerts
  • AAC Podcast
  • RSS
  • FAQ
  • Permissions
  • Journal Announcements

Authors

  • ASM Author Center
  • Submit a Manuscript
  • Article Types
  • Ethics
  • Contact Us

Follow #AACJournal

@ASMicrobiology

       

ASM Journals

ASM journals are the most prominent publications in the field, delivering up-to-date and authoritative coverage of both basic and clinical microbiology.

About ASM | Contact Us | Press Room

 

ASM is a member of

Scientific Society Publisher Alliance

 

American Society for Microbiology
1752 N St. NW
Washington, DC 20036
Phone: (202) 737-3600

Copyright © 2021 American Society for Microbiology | Privacy Policy | Website feedback

Print ISSN: 0066-4804; Online ISSN: 1098-6596