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Open Forum Infectious Diseases PERSPECTIVES

Antibiotics and Innate Immunity: A Cooperative Effort Toward the Successful Treatment of Andrew Berti,1,2 Warren Rose,3, Victor Nizet,4,5 and George Sakoulas4 1Department of Pharmacy Practice, Wayne State University College of Pharmacy and Health Sciences, Detroit, Michigan, USA, 2Department of Biochemistry, Microbiology and Immunology, Wayne State University College of Medicine, Detroit, Michigan, USA, 3School of Pharmacy, University of Wisconsin-Madison, Madison, Wisconsin, USA, 4Collaborative to Halt Antimicrobial Resistant Microbes, University of California San Diego School of Medicine, La Jolla, California, USA, and 5Skaggs School of Pharmacy, University of California San Diego, La Jolla, California, USA

Despite the common ancestry of antimicrobial and immunological science, a divergence driven by artificially construed paradigms

in microbiology has placed limits on how we understand the mechanisms of antibiotics in vivo. We summarize recent updates on Downloaded from https://academic.oup.com/ofid/article/7/8/ofaa302/5873654 by guest on 26 August 2020 data that shed light on how antibiotics interact with components of innate immunity. Keywords. antibiotic; antimicrobial peptides; innate immunity, mechanism.

Despite common roots of antimicro- species, suggesting that antibiotics them- Institute. This assay consists of collecting bial therapy and immunology in the re- selves may represent primeval precursors patient blood samples, ideally during search efforts of legendary pioneers like of the immune systems of higher organ- serum peak and trough concentrations Louis Pasteur and Paul Ehrlich, our di- isms. Along those lines, the mechanisms of the antibiotic, combining equally with vergent and hyperspecialized evolution of action of some current “exogenous” bacterial growth medium and testing the of medicine has largely separated the natural product antibiotics used in clin- bactericidal titer concentration against pharmacology of antibiotics adminis- ical medicine overlap considerably with the patient’s bacterial isolate. A high SBT tered to patients from the functions of the “endogenous” antimicrobial pep- may indicate that the dose chosen for the patient’s [1]. tides of the mammalian innate immune the patient is sufficient for treatment [6]. Ehrlich’s work in immunology won him system [3]. This process has implications However, significant variability in the the 1908 Nobel Prize in Medicine; his for antimicrobial susceptibility, as the medium, diluent, inoculum, incubation, contributions to antimicrobial chemo- presence of host defense peptides in vivo and controls has impeded the widespread therapy with salvarsan, an arsphenamine during chronic is sufficient to clinical application of SBT testing. In ad- compound that was the first treatment select for cross-resistance to daptomycin, dition, use of only the serum excludes for syphilis, should not be forgotten [2]. even in the treatment-naïve patient [4, 5]. other critical components of the patient The great majority of current antibiotics innate immune system, for example, are natural products produced by actino- ENDOGENOUS AND EXOGENOUS leukocytes, and this may fail to recapit- ANTIBIOTICS mycetes, fungi, or other microorganisms ulate the full interaction of antibiotics used in niche competition against other There is abundant literature examining with host immune defenses. For example, the pharmacodynamic relationships of macrolide antibiotics induce formation exogenous antibiotics with one another, of extracellular traps (NETs)

Received 30 April 2020; editorial decision 6 July 2020; defining synergy, additivity, indifference, containing antimicrobial peptides and accepted 15 July 2020. and antagonism. Conversely, studies of histones that can ensnare bacteria and Correspondence: George Sakoulas, MD, Center for Immunity, Infection & , UCSD School of Medicine the pharmacodynamics of innate im- attract other immune cells [7], while Biomedical Research Facility II, Room 4114, 9500 Gilman mune components and antibiotics are beta-lactams sensitize bacteria to neutro- Drive, Mail Code 0760, La Jolla, CA 92093-0760 (gsakoulas@ scarce and have been difficult to imple- phils [8]. Anthropocentric debates about health.ucsd.edu). Open Forum Infectious Diseases® ment and replicate in the patient care set- “monotherapy” vs “combination therapy” © The Author(s) 2020. Published by Oxford University Press ting. Serum inhibitory and bactericidal in the treatment of infection fail to con- on behalf of Infectious Diseases Society of America. This titers (SBTs) were devised as an investi- sider the fact that the net antimicrobial is an Open Access article distributed under the terms of the Creative Commons Attribution-NonCommercial- gational test to evaluate the bactericidal effect in a patient receiving antibiotic NoDerivs licence (http://creativecommons.org/licenses/ properties of antibiotic therapy in a more pharmacotherapy is the collective sum- by-nc-nd/4.0/), which permits non-commercial reproduction and distribution of the work, in any medium, provided physiological context, namely the serum mation of exogenous pharmacotherapy the original work is not altered or transformed in any of the patient. Specifics for the perfor- and innate immune defense factors. way, and that the work is properly cited. For commercial re-use, please contact [email protected] mance of SBT testing have been published “Monotherapy” never really exists in a DOI: 10.1093/ofid/ofaa302 by the Clinical and Laboratory Standards pure form.

PERSPECTIVES • ofid • 1 CONVENTIONAL BACTERIOLOGIC treated in animal models of endocar- beta-lactams like nafcillin and oxacillin MEDIA: AN OBSTACLE TO OUR ditis with beta-lactam monotherapy [11, has been successfully deployed as adjunc- COMPLETE UNDERSTANDING OF ANTIMICROBIAL ACTIVITY 13]. Azithromycin efficacy against mul- tive salvage therapy to successfully clear tiple gram-negative bacterial species refractory MRSA bacteremia [18, 19]. The conceptual divergence of antibiotic (eg, Acinetobacter baumannii, Klebsiella Ampicillin has been used in an analogous pharmacology and immunology dis- pneumoniae) depends on bicarbonate, fashion to clear persistent bacteremia ciplines may have been aggravated by and this activity translates to in vivo ef- due to ampicillin-resistant, vancomycin- the historical choice of media used to ficacy in murine infection models [14, resistant Enterococcus faecium (VRE) cultivate and examine microorganism 15]. MHB lacks bicarbonate buffer; thus [20]. In both cases, concentrations well susceptibility and resistance. From the standard clinical AST testing declares below the MIC rendered MRSA and VRE beginnings of the human antibiotic era, azithromycin inactive against these hypersusceptible to cationic antimicro- antimicrobial susceptibility testing (AST) pathogens. Another recent study shows bial peptides, a property completely of pathogens has focused exclusively on how hyperphysiological concentrations missed in standard AST testing, which Downloaded from https://academic.oup.com/ofid/article/7/8/ofaa302/5873654 by guest on 26 August 2020 media optimized for microbial growth, of zinc in MHB render metallo-beta- may serve a critical role in the resolution with the addition of antibiotics up to the lactamase-producing Enterobacteriaceae of severe infection. Synergy with anti- point of growth inhibition defining the resistant to carbapenems in such media. microbial peptides has also been shown “minimum inhibitory concentration” However, these organisms are susceptible for ceftaroline against Streptococcus (MIC), which serves as the centerpiece to carbapenems in vivo, where nutritional pneumoniae [21] and ceftriaxone against of clinical microbiology [9]. AST para- immunity restricts zinc concentrations to Salmonella enterica [22]. The cationic de- digms have somewhat arbitrarily con- much lower levels. Removal of zinc from fense peptide cathelicidin is a key host verged on Mueller-Hinton broth (MHB) MHB broth more reliably predicts in vivo defense against systemic infection and for most assays in the clinical microbi- activity of carbapenems against these or- bacterial meningitis. Antimicrobial syn- ology lab. This medium, first developed ganisms [16]. Rigid continuation of AST ergy with cathelicidin may be an im- 1941 for isolating pathogenic strains of paradigms utilizing only standard bac- portant factor driving better clinical Neisseria spp. [10], does not recapitulate teriological media overlooks potential outcomes [22]. in vitro the in vivo environment relevant useful activities of the currently available This enhancement between antibiotics for infection, nor is it a medium in which antibiotics against multidrug-resistant and immune response extends beyond host antimicrobial peptide activity can bacteria [14], but it also hinders the tran- beta-lactams. Azithromycin, a macro- be reliably assayed [9]. Recent work has sition of pharmacodynamic analysis of lide antibiotic, demonstrates synergy shown considerable differences in MICs innate immunity synergy with antibiotics with host antimicrobial peptides against between standard bacteriological media to a more clinical mainstream. Pseudomonas and Acinetobacter [14]. and more physiological media such as Even certain beta-lactamase inhibitors, mammalian tissue culture media [11, 12]. BACTERICIDAL VS conceptually deployed to inhibit beta- Concordance between AST performed BACTERIOSTATIC IN THE lactam hydrolysis by beta-lactamase en- CONTEXT OF INNATE IMMUNITY on these medias is about 60%; however, zymes, can themselves act synergistically 10%–20% of the time, an MIC difference The few studies on the pharmacody- with endogenous antimicrobial peptides of ≥8× is noted. Importantly, antimicro- namic interactions between innate im- or peptide antibiotics like daptomycin bial activity in vivo has been shown to be munity and exogenous antibiotics have and colistin [23, 24]. All of these prop- more concordant with activity in physi- revealed differences in the exposure–re- erties require the use of physiological ological media compared with standard sponse relationship. Some antibiotics, (eg, tissue culture) media to evaluate, as bacteriological media [12]. Additional including those that have been tradi- host antimicrobial peptides are gener- studies have shown that buffering with tionally labeled “bacteriostatic,” such ally not active in bacteriological media. bicarbonate, the major anionic buffer in as chloramphenicol and erythromycin, Mechanisms for synergy between host in- mammalian physiology, is a major con- may antagonize the activity of endoge- nate immunity and antibiotics are certain tributor to the superior predictability of nous host defense peptides [17]. In con- to be multifactorial. This complexity may physiological media in vitro antibiotic trast, beta-lactam antibiotics, which are be one unrecognized pharmacodynamic susceptibility to in vivo efficacy [11]. For often touted for their overall bactericidal faculty helping to explain the differences example, some strains of methicillin- activities, have been shown to further in clinical efficacy of different antibiotic resistant Staphylococcus aureus (MRSA) synergize with cationic antimicrobial classes irrespective of MIC in standard demonstrate a methicillin-susceptible peptides produced by innate immu- AST testing. For example, vancomycin is phenotype in bicarbonate-buffered nity [8]. This synergy is so profound a far inferior agent to beta-lactams clin- media, and such strains can be effectively that the addition of antistaphylococcal ically against S. aureus, despite showing

2 • ofid • PERSPECTIVES potent activity in vitro [25, 26]. These ef- made through the use of immunologic meningococcus. Proc Soc Exp Biol Med 1941; 48:330–3. fects may unfortunately contribute to the and T-cell engineering therapies [35]. In 11. Ersoy SC, Abdelhady W, Li L, Chambers HF, poorer outcomes of patient who are de- infectious diseases, more objective exam- Xiong YQ, Bayer AS. Bicarbonate resensitization of methicillin-resistant Staphylococcus aureus to beta- nied beta-lactam drugs due to penicillin inations of the immune system—antibi- lactam antibiotics. Antimicrob Agents Chemother “allergies” listed in their medical records otic cooperativity through mathematical 2019; 63:e00496-19. [27]. modeling and computer simulation— 12. Ersoy SC, Heithoff DM, Barnes L 5th, et al. Correcting a fundamental flaw in the paradigm for The interaction of antibiotics and in- have already been undertaken by some antimicrobial susceptibility testing. EBioMedicine nate immunity can be mediated through groups [36], which will hopefully help 2017; 20:173–81. 13. Rose WE, Bienvenida AM, Xiong YQ, additional mechanisms, including al- transition this field into a more formal- Chambers HF, Bayer AS, Ersoy SC. Ability of bi- teration of virulence factor expression, ized pharmacological discipline. Steps carbonate supplementation to sensitize selected methicillin-resistant Staphylococcus aureus (MRSA) which in turn may influence host cy- must be taken to reunite the under- strains to beta-lactam antibiotics in an ex vivo sim- tokine expression. For example, beta- standing of innate immunity–antibiotic ulated endocardial vegetation model. Antimicrob Agents Chemother 2020; 64(3):e02072-19. lactams upregulate alpha-toxin and other relationships to improve treatments, slow 14. Lin L, Nonejuie P, Munguia J, et al. Azithromycin Downloaded from https://academic.oup.com/ofid/article/7/8/ofaa302/5873654 by guest on 26 August 2020 exotoxin expression in S. aureus [28], the development of resistance, and dis- synergizes with cationic antimicrobial peptides to which in turn stimulates a host inter- cover new therapeutic approaches. exert bactericidal and therapeutic activity against highly multidrug-resistant gram-negative bacterial leukin (IL) 1–beta response [29], which pathogens. EBioMedicine 2015; 2:690–8. is important in host recognition and Acknowledgments 15. Dillon N, Holland M, Tsunemoto H, et al. Surprising synergy of dual translation inhibition clearance of bacteremia [30, 31]. Beta- Financial support. No outside funding was received for this work vs. Acinetobacter baumannii and other multidrug- lactam therapy is more effective than Potential conflicts of interest. The authors resistant bacterial pathogens. EBioMedicine 2019; 46:193–201. declare no financial disclosures relevant to this vancomycin in eliciting an IL1-beta re- 16. Asempa TE, Abdelraouf K, Nicolau DP. Metallo- sponse in patients with S. aureus bacte- work. All authors have submitted the ICMJE beta-lactamase resistance in Enterobacteriaceae Form for Disclosure of Potential Conflicts of remia, including when used as adjunctive is an artefact of currently utilized antimicro- Interest. Conflicts that the editors consider rele- bial susceptibility testing methods. J Antimicrob therapy for MRSA [32]. Conversely, bac- vant to the content of the manuscript have been Chemother 2020; 75:997–1005. teriostatic antibiotics inhibit this process disclosed. 17. Kristian SA, Timmer AM, Liu GY, et al. Impairment of innate immune killing mechanisms by bacterio- [28]. Interestingly, the MRSA cell wall is static antibiotics. FASEB J 2007; 21:1107–16. References less cross-linked in the presence of beta- 18. Dhand A, Bayer AS, Pogliano J, et al. Use of 1. Gould K. Antibiotics: from prehistory to the present antistaphylococcal beta-lactams to increase lactam and induces a more potent IL1- day. J Antimicrob Chemother 2016; 71:572–5. daptomycin activity in eradicating persistent bac- beta response as well [33]. Reduction in 2. Bosch F, Rosich L. The contributions of Paul teremia due to methicillin-resistant Staphylococcus Ehrlich to pharmacology: a tribute on the occasion aureus: role of enhanced daptomycin binding. Clin peptidoglycan O-acetylation induced by of the centenary of his Nobel Prize. Pharmacology Infect Dis 2011; 53:158–63. beta-lactams has a similar effect [34]. 2008; 82:171–9. 19. Dhand A, Sakoulas G. Daptomycin in combination 3. Bayer AS, Mishra NN, Sakoulas G, et al. Hence, the functional determination for with other antibiotics for the treatment of compli- Heterogeneity of mprF sequences in methicillin- cated methicillin-resistant Staphylococcus aureus bacteriostatic vs bactericidal antibiotics resistant Staphylococcus aureus clinical isolates: role bacteremia. Clin Ther 2014; 36:1303–16. in bacteremia may diverge from their po- in cross-resistance between daptomycin and host 20. Sakoulas G, Bayer AS, Pogliano J, et al. Ampicillin defense antimicrobial peptides. Antimicrob Agents enhances daptomycin- and cationic host de- tency in AST testing in bacteriological Chemother 2014; 58:7462–7. fense peptide-mediated killing of ampicillin- and media; it may reflect immune response 4. Mishra NN, Yang SJ, Chen L, et al. Emergence of vancomycin-resistant Enterococcus faecium. daptomycin resistance in daptomycin-naïve rabbits Antimicrob Agents Chemother 2012; 56:838–44. factors that synergize with innate im- with methicillin-resistant Staphylococcus aureus 21. Sakoulas G, Nonejuie P, Kullar R, et al. Examining munity or induce protective cytokine prosthetic joint infection is associated with resist- the use of ceftaroline in the treatment of ance to host defense cationic peptides and mprF Streptococcus pneumoniae meningitis with refer- responses. polymorphisms. PLoS One 2013; 8:e71151. ence to human cathelicidin LL-37. Antimicrob 5. Kullar R, McKinnell JA, Sakoulas G. Avoiding the Agents Chemother 2015; 59:2428–31. perfect storm: the biologic and clinical case for 22. Sakoulas G, Kumaraswamy M, Kousha A, Nizet V. CONCLUSIONS reevaluating the 7-day expectation for methicillin- Interaction of antibiotics with innate host defense resistant Staphylococcus aureus bacteremia be- factors against Salmonella enterica serotype new- Looking toward the future, our sepa- fore switching therapy. Clin Infect Dis 2014; port. mSphere 2017; 2(6):e00410-17. 59:1455–61. 23. Sakoulas G, Rose W, Berti A, et al. Classical ration of antimicrobial pharmacology 6. Jenkins SG, Schuetz AN. Current concepts in labo- beta-lactamase inhibitors potentiate the ac- and immunology has resulted in para- ratory testing to guide antimicrobial therapy. Mayo tivity of daptomycin against methicillin-resistant Clin Proc 2012; 87:290–308. Staphylococcus aureus and colistin against digms that limit our understanding of 7. Papayannopoulos V. Neutrophil extracellular traps Acinetobacter baumannii. Antimicrob Agents antimicrobial action in an era of ever- in immunity and disease. Nat Rev Immunol 2018; Chemother 2017; 61(2):e01745-16. 18:134–47. 24. Ulloa ER, Dillon N, Tsunemoto H, et al. Avibactam expanding resistance. The oncology field 8. Sakoulas G, Okumura CY, Thienphrapa W, et al. sensitizes carbapenem-resistant NDM-1- has made much greater strides recog- Nafcillin enhances innate immune-mediated producing Klebsiella pneumoniae to innate immune killing of methicillin-resistant Staphylococcus au- nizing the importance of host immune clearance. J Infect Dis 2019; 220:484–93. reus. J Mol Med (Berl) 2014; 92:139–49. 25. Schweizer ML, Furuno JP, Harris AD, et al. cooperativity in treating certain cancers, 9. Nizet V. The accidental orthodoxy of Drs. Mueller Comparative effectiveness of nafcillin or cefazolin and significant advances toward cure and Hinton. EBioMedicine 2017; 22:26–7. versus vancomycin in methicillin-susceptible 10. Mueller JH, Hinton J. A protein-free medium Staphylococcus aureus bacteremia. BMC Infect Dis of nearly untreatable cancers have been for primary isolation of the gonococcus and 2011; 11:279.

PERSPECTIVES • ofid • 3 26. Le J, Dam Q, Schweizer M, et al. Effects of van- 30. Rose WE, Eickhoff JC, Shukla SK, et al. Elevated 33. Müller S, Wolf AJ, Iliev ID, et al. Poorly cross- comycin versus nafcillin in enhancing killing of serum interleukin-10 at time of hospital admis- linked peptidoglycan in MRSA due to mecA methicillin-susceptible Staphylococcus aureus sion is predictive of mortality in patients with induction activates the inflammasome and exacer- causing bacteremia by human cathelicidin LL-37. Staphylococcus aureus bacteremia. J Infect Dis bates immunopathology. Cell Host Microbe 2015; Eur J Clin Microbiol Infect Dis 2016; 35:1441–7. 2012; 206:1604–11. 18:604–12. 27. Sakoulas G, Geriak M, Nizet V. Is a reported pen- 31. Rose WE, Shukla SK, Berti AD, et al. Increased 34. Sanchez M, Kolar SL, Muller S, et al. O-acetylation icillin allergy sufficient grounds to forgo the mul- endovascular Staphylococcus aureus inoculum of peptidoglycan limits helper T cell priming and tidimensional antimicrobial benefits of β-lactam is the link between elevated serum interleukin permits Staphylococcus aureus reinfection. Cell antibiotics? Clin Infect Dis 2019; 68:157–64. 10 concentrations and mortality in patients Host Microbe 2017; 22:543–51, e4 28. Hodille E, Rose W, Diep BA, et al. The role of anti- with bacteremia. Clin Infect Dis 2017; 64: 35. Majzner RG, Mackall CL. Clinical lessons learned biotics in modulating virulence in Staphylococcus 1406–12. from the first leg of the CAR T cell journey. Nat aureus. Clin Microbiol Rev 2017; 30:887–917. 32. Volk CF, Burgdorf S, Edwardson G, Nizet V, Med 2019; 25:1341–55. 29. Craven RR, Gao X, Allen IC, et al. Staphylococcus Sakoulas G, Rose WE. IL-1beta and IL-10 host re- 36. Ankomah P, Levin BR. Exploring the collaboration aureus alpha-hemolysin activates the NLRP3- sponses in patients with Staphylococcus aureus bac- between antibiotics and the immune response in inflammasome in human and mouse monocytic teremia determined by antimicrobial therapy. Clin the treatment of acute, self-limiting infections. Proc cells. PLoS One 2009; 4:e7446. Infect Dis 2020 70:2634–40. Natl Acad Sci U S A 2014; 111:8331–8. Downloaded from https://academic.oup.com/ofid/article/7/8/ofaa302/5873654 by guest on 26 August 2020

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