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Efficacy of Bisphosphonate-Conjugated Sitafloxacin in a Murine Model of S. aureus Osteomyelitis: Evidence of “Target & Release” Kinetics and Killing of Bacteria Within Canaliculi

S. aureus infection of bone is difficult to eradicate due to its ability to colonize the osteocyte-lacuno-canalicular network (OLCN), rendering it resistant to standard-of-care (SOC) antibiotics. To overcome this, we proposed two bone-targeted bisphosphonate-conjugated antibiotics (BCA): bisphosphon...

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Autores principales: Ren, Youliang, Xue, Thomas, Rainbolt, Joshua, Bentley, Karen L. de Mesy, Galloway, Chad A., Liu, Yuting, Cherian, Philip, Neighbors, Jeffrey, Hofstee, Marloes I., Ebetino, Frank H., Moriarty, Thomas Fintan, Sun, Shuting, Schwarz, Edward M., Xie, Chao
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Frontiers Media S.A. 2022
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9263620/
https://www.ncbi.nlm.nih.gov/pubmed/35811672
http://dx.doi.org/10.3389/fcimb.2022.910970
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author Ren, Youliang
Xue, Thomas
Rainbolt, Joshua
Bentley, Karen L. de Mesy
Galloway, Chad A.
Liu, Yuting
Cherian, Philip
Neighbors, Jeffrey
Hofstee, Marloes I.
Ebetino, Frank H.
Moriarty, Thomas Fintan
Sun, Shuting
Schwarz, Edward M.
Xie, Chao
author_facet Ren, Youliang
Xue, Thomas
Rainbolt, Joshua
Bentley, Karen L. de Mesy
Galloway, Chad A.
Liu, Yuting
Cherian, Philip
Neighbors, Jeffrey
Hofstee, Marloes I.
Ebetino, Frank H.
Moriarty, Thomas Fintan
Sun, Shuting
Schwarz, Edward M.
Xie, Chao
author_sort Ren, Youliang
collection PubMed
description S. aureus infection of bone is difficult to eradicate due to its ability to colonize the osteocyte-lacuno-canalicular network (OLCN), rendering it resistant to standard-of-care (SOC) antibiotics. To overcome this, we proposed two bone-targeted bisphosphonate-conjugated antibiotics (BCA): bisphosphonate-conjugated sitafloxacin (BCS) and hydroxybisphosphonate-conjugate sitafloxacin (HBCS). Initial studies demonstrated that the BCA kills S. aureus in vitro. Here we demonstrate the in vivo efficacy of BCS and HBCS versus bisphosphonate, sitafloxacin, and vancomycin in mice with implant-associated osteomyelitis. Longitudinal bioluminescent imaging (BLI) confirmed the hypothesized “target and release”-type kinetics of BCS and HBCS. Micro-CT of the infected tibiae demonstrated that HBCS significantly inhibited peri-implant osteolysis versus placebo and free sitafloxacin (p < 0.05), which was not seen with the corresponding non-antibiotic-conjugated bisphosphonate control. TRAP-stained histology confirmed that HBCS significantly reduced peri-implant osteoclast numbers versus placebo and free sitafloxacin controls (p < 0.05). To confirm S. aureus killing, we compared the morphology of S. aureus autolysis within in vitro biofilm and infected tibiae via transmission electron microscopy (TEM). Live bacteria in vitro and in vivo presented as dense cocci ~1 μm in diameter. In vitro evidence of autolysis presented remnant cell walls of dead bacteria or “ghosts” and degenerating (non-dense) bacteria. These features of autolyzed bacteria were also present among the colonizing S. aureus within OLCN of infected tibiae from placebo-, vancomycin-, and sitafloxacin-treated mice, similar to placebo. However, most of the bacteria within OLCN of infected tibiae from BCA-treated mice were less dense and contained small vacuoles and holes >100 nm. Histomorphometry of the bacteria within the OLCN demonstrated that BCA significantly increased their diameter versus placebo and free antibiotic controls (p < 0.05). As these abnormal features are consistent with antibiotic-induced vacuolization, bacterial swelling, and necrotic phenotype, we interpret these findings to be the initial evidence of BCA-induced killing of S. aureus within the OLCN of infected bone. Collectively, these results support the bone targeting strategy of BCA to overcome the biodistribution limits of SOC antibiotics and warrant future studies to confirm the novel TEM phenotypes of bacteria within OLCN of S. aureus-infected bone of animals treated with BCS and HBCS.
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spelling pubmed-92636202022-07-09 Efficacy of Bisphosphonate-Conjugated Sitafloxacin in a Murine Model of S. aureus Osteomyelitis: Evidence of “Target & Release” Kinetics and Killing of Bacteria Within Canaliculi Ren, Youliang Xue, Thomas Rainbolt, Joshua Bentley, Karen L. de Mesy Galloway, Chad A. Liu, Yuting Cherian, Philip Neighbors, Jeffrey Hofstee, Marloes I. Ebetino, Frank H. Moriarty, Thomas Fintan Sun, Shuting Schwarz, Edward M. Xie, Chao Front Cell Infect Microbiol Cellular and Infection Microbiology S. aureus infection of bone is difficult to eradicate due to its ability to colonize the osteocyte-lacuno-canalicular network (OLCN), rendering it resistant to standard-of-care (SOC) antibiotics. To overcome this, we proposed two bone-targeted bisphosphonate-conjugated antibiotics (BCA): bisphosphonate-conjugated sitafloxacin (BCS) and hydroxybisphosphonate-conjugate sitafloxacin (HBCS). Initial studies demonstrated that the BCA kills S. aureus in vitro. Here we demonstrate the in vivo efficacy of BCS and HBCS versus bisphosphonate, sitafloxacin, and vancomycin in mice with implant-associated osteomyelitis. Longitudinal bioluminescent imaging (BLI) confirmed the hypothesized “target and release”-type kinetics of BCS and HBCS. Micro-CT of the infected tibiae demonstrated that HBCS significantly inhibited peri-implant osteolysis versus placebo and free sitafloxacin (p < 0.05), which was not seen with the corresponding non-antibiotic-conjugated bisphosphonate control. TRAP-stained histology confirmed that HBCS significantly reduced peri-implant osteoclast numbers versus placebo and free sitafloxacin controls (p < 0.05). To confirm S. aureus killing, we compared the morphology of S. aureus autolysis within in vitro biofilm and infected tibiae via transmission electron microscopy (TEM). Live bacteria in vitro and in vivo presented as dense cocci ~1 μm in diameter. In vitro evidence of autolysis presented remnant cell walls of dead bacteria or “ghosts” and degenerating (non-dense) bacteria. These features of autolyzed bacteria were also present among the colonizing S. aureus within OLCN of infected tibiae from placebo-, vancomycin-, and sitafloxacin-treated mice, similar to placebo. However, most of the bacteria within OLCN of infected tibiae from BCA-treated mice were less dense and contained small vacuoles and holes >100 nm. Histomorphometry of the bacteria within the OLCN demonstrated that BCA significantly increased their diameter versus placebo and free antibiotic controls (p < 0.05). As these abnormal features are consistent with antibiotic-induced vacuolization, bacterial swelling, and necrotic phenotype, we interpret these findings to be the initial evidence of BCA-induced killing of S. aureus within the OLCN of infected bone. Collectively, these results support the bone targeting strategy of BCA to overcome the biodistribution limits of SOC antibiotics and warrant future studies to confirm the novel TEM phenotypes of bacteria within OLCN of S. aureus-infected bone of animals treated with BCS and HBCS. Frontiers Media S.A. 2022-06-24 /pmc/articles/PMC9263620/ /pubmed/35811672 http://dx.doi.org/10.3389/fcimb.2022.910970 Text en Copyright © 2022 Ren, Xue, Rainbolt, Bentley, Galloway, Liu, Cherian, Neighbors, Hofstee, Ebetino, Moriarty, Sun, Schwarz and Xie https://creativecommons.org/licenses/by/4.0/This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.
spellingShingle Cellular and Infection Microbiology
Ren, Youliang
Xue, Thomas
Rainbolt, Joshua
Bentley, Karen L. de Mesy
Galloway, Chad A.
Liu, Yuting
Cherian, Philip
Neighbors, Jeffrey
Hofstee, Marloes I.
Ebetino, Frank H.
Moriarty, Thomas Fintan
Sun, Shuting
Schwarz, Edward M.
Xie, Chao
Efficacy of Bisphosphonate-Conjugated Sitafloxacin in a Murine Model of S. aureus Osteomyelitis: Evidence of “Target & Release” Kinetics and Killing of Bacteria Within Canaliculi
title Efficacy of Bisphosphonate-Conjugated Sitafloxacin in a Murine Model of S. aureus Osteomyelitis: Evidence of “Target & Release” Kinetics and Killing of Bacteria Within Canaliculi
title_full Efficacy of Bisphosphonate-Conjugated Sitafloxacin in a Murine Model of S. aureus Osteomyelitis: Evidence of “Target & Release” Kinetics and Killing of Bacteria Within Canaliculi
title_fullStr Efficacy of Bisphosphonate-Conjugated Sitafloxacin in a Murine Model of S. aureus Osteomyelitis: Evidence of “Target & Release” Kinetics and Killing of Bacteria Within Canaliculi
title_full_unstemmed Efficacy of Bisphosphonate-Conjugated Sitafloxacin in a Murine Model of S. aureus Osteomyelitis: Evidence of “Target & Release” Kinetics and Killing of Bacteria Within Canaliculi
title_short Efficacy of Bisphosphonate-Conjugated Sitafloxacin in a Murine Model of S. aureus Osteomyelitis: Evidence of “Target & Release” Kinetics and Killing of Bacteria Within Canaliculi
title_sort efficacy of bisphosphonate-conjugated sitafloxacin in a murine model of s. aureus osteomyelitis: evidence of “target & release” kinetics and killing of bacteria within canaliculi
topic Cellular and Infection Microbiology
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9263620/
https://www.ncbi.nlm.nih.gov/pubmed/35811672
http://dx.doi.org/10.3389/fcimb.2022.910970
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