Development of Bisphosphonate-Conjugated Antibiotics to Overcome Pharmacodynamic Limitations of Local Therapy: Initial Results with Carbamate Linked Sitafloxacin and Tedizolid

Abstract

The use of local antibiotics to treat bone infections has been questioned due to a lack of clinical efficacy and emerging information about Staphylococcus aureus colonization of the osteocyte-lacuno canalicular network (OLCN). Here we propose bisphosphonate-conjugated antibiotics (BCA) using a “target and release” approach to deliver antibiotics to bone infection sites. A fluorescent bisphosphonate probe was used to demonstrate bone surface labeling adjacent to bacteria in a S. aureus infected mouse tibiae model. Bisphosphonate and hydroxybisphosphonate conjugates of sitafloxacin and tedizolid (BCA) were synthesized using hydroxyphenyl and aminophenyl carbamate linkers, respectively. The conjugates were adequately stable in serum. Their cytolytic activity versus parent drug on MSSA and MRSA static biofilms grown on hydroxyapatite discs was established by scanning electron microscopy. Sitafloxacin O-phenyl carbamate BCA was effective in eradicating static biofilm: no colony formation units (CFU) were recovered following treatment with 800 mg/L of either the bisphosphonate or α-hydroxybisphosphonate conjugated drug (p < 0.001). In contrast, the less labile tedizolid N-phenyl carbamate linked BCA had limited efficacy against MSSA, and MRSA. CFU were recovered from all tedizolid BCA treatments. These results demonstrate the feasibility of BCA eradication of S. aureus biofilm on OLCN bone surfaces and support in vivo drug development of a sitafloxacin BCA.

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

“…This efficacy is hypothesized to be from the BCA killing of S. aureus biofilm bacteria within the OLCN, which is not achieved by sitafloxacin due to the aforementioned biodistribution limits. Additionally, since the nitrogen-containing bisphosphonates have great potency to inhibit osteoclasts, the bisphosphonates used as the bonetargeting component of BCA are designed to be innocuous or have minimal effects on bone cells (Adjei-Sowah et al, 2021), and the lack of efficacy of our "non-nitrogen-containing" bisphosphonate controls (HPHBP and HPBP) is consistent with our design and hypothesis. However, one unexplained finding in our study is that only HBCS reduces osteoclast number, and not BCS (Figure 3).…”

“…Based on this, we aimed to create a bone-targeted antibiotic conjugate that could achieve sustained concentrations of a drug well above the MEC at the site of bone infection. In support of this concept, we recently demonstrated that systemic administration of a BP-conjugated fluorophore (AF647-ZOL) specifically labels the cortical surface of the bone in immediate proximity to S. aureus bacteria during chronic osteomyelitis (Adjei-Sowah et al, 2021). We were also encouraged by our initial success with BP-conjugated ciprofloxacin for the oral indication (Sedghizadeh et al, 2017).…”

“…The most prevalent community-acquired MRSA strain of USA300LAC was chromosomally transduced with the bacterial luciferase gene lux to generate USA300 LAC::Lux in an in vivo imaging study (Thurlow et al, 2011). The USA300 LAC::Lux was cultured in tryptic soy broth (TSB) media at 37°C overnight as previously described (Kourbatova et al, 2005;Adjei-Sowah et al, 2021). The bioluminescent construct is stably integrated into the USA300 LAC::lux bacterial chromosome and constitutively emits a blue-green light with a maximal emission wavelength of 490 nm only in these live and metabolically active bacteria (Guo et al, 2013).…”

“…To overcome the biodistribution limits of SOC treatments, we developed bisphosphonate-conjugated antibiotics (BCA) using a "target and release" approach to deliver antibiotics to bone infection sites (Adjei-Sowah et al, 2021), the theory being that non-nitrogen-containing bisphosphonates, which have a high affinity to hydroxyapatite at eroded bone surfaces but little or no effects on bone cells, could be conjugated to SOC antibiotics via a semi-labile, serum-stable, chemical linker, which would specifically target the drug to the bone-bacteria interface and kill the bacteria following acid and/or enzymatic cleavage of the linker releasing an active antibiotic (Adjei-Sowah et al, 2021) in that compartment. As proof of concept, we demonstrated that systemic administration of fluorescent bisphosphonate labels the bone surface adjacent to bacteria in a murine model of S. aureus osteomyelitis (Adjei-Sowah et al, 2021).…”

The Rationale for Using Neridronate in Musculoskeletal Disorders: From Metabolic Bone Diseases to Musculoskeletal Pain

“…From a clinical point of view, the studies concerning the use of BPs in the treatment of some protozoal infections [48], in the modulation of the immune response [49], in the mechanisms of tissue repair [50], and generally in improving cell survival [51], could have a significant impact on the management of many diseases. Additionally, in the field of bone health, new research addressed the role of BPs as transporters of other pharmacological substances that would otherwise reach the bone tissue with difficulty and low doses [52][53][54][55].…”