Pharmacokinetic and Pharmacodynamic Studies of a Novel Spectinamide Series of Antituberculosis Agents

<p>Spectinamides are novel amide derivatives of the antibiotic spectinomycin that have emerged as a new class of agents to treat tuberculosis. These agents showed potent <em>in vitro </em>activity against <em>Mycobacterium tuberculosis</em> (MTB)<em> </em>compared to spectinomycin and in a preliminary <em>in vivo </em>study in interferon gamma (IFN‑γ) knockout mice, spectinamide Lee1329 reduced the lung bacillary load of TB comparable to streptomycin. We hypothesized that the application of an iterative pharmacokinetics and pharmacodynamics (PK/PD) guided approach would facilitate the optimization of these lead compounds suitable for further development.</p> <p>A series of <em>in vitro </em>experiments including parallel artificial membrane permeability assay (PAMPA), microsomal metabolic stability using rat liver microsomes and protein binding assay were designed to characterize the <em>in vivo</em> biopharmaceutic and pharmacokinetic behavior. Drug uptake studies into <em>Mycobacterium bovis</em> BCG and into J774 murine macrophages were performed to understand reasons for improved activity of spectinamides and to evaluate their potential to target primary TB infection that resides in macrophages. <em>In vivo </em>pharmacokinetic studies were carried out in rats by intravenous (10 mg/Kg) and oral administration (100 mg/Kg) of the compounds. LC‑MS/MS assays were developed to quantify concentrations in test samples obtained from these studies.</p> <p>Spectinamides exhibit low to intermediate plasma protein binding and were found to be metabolically stable. Following intravenous administration, spectinamides were relatively widely distributed (0.36 to 1.15 L/Kg) with short half‑lives (0.43 to 0.62 hr). Mean systemic clearance ranged between 0.36 and 0.89 L/hr/Kg with a significant fraction of drug eliminated unchanged in urine (0.46 to 1.0). Spectinamides exhibited a renal excretion ratio greater than one indicating filtration and active secretion as the net renal elimination process. In the uptake experiments, the spectinamides exhibited 3‑4 times higher uptake into murine macrophages compared to streptomycin and showed nearly four times higher uptake into <em>M. bovis </em>BCG compared to spectinomycin, which may in part explain their increased activity compared to spectinomycin.</p> <p>In a previously reported <em>in vitro</em> PK/PD system, PK concentration‑time profiles were simulated on the basis of the <em>in vivo</em> clearance of rats obtained from the PK studies and different daily doses of 0.4, 2, 10 and 50 mg/Kg/day of Lee1445 were added as QD, BID or TID regimens. The time‑kill effect of these regimens was studied on the growing <em>M. bovis</em> BCG present in the system. A semi‑mechanistic model incorporating a logistic function for growth of mycobacteria in the absence of drug and an inhibitory sigmoidal E_max model with a delay function for the time‑kill effect of the drug was fit to the data and the <em>in vitro </em>PK/PD parameters were determined. Since the <em>in vivo</em> efficacy model of tuberculosis infection was the gamma knock‑out (GKO) mice, a PK bridging study was performed in mice following intraperitoneal administration of 20 mg/Kg Lee1445. Simulations for various dosing regimens were performed using the obtained mouse PK parameters and the <em>in vitro </em>PK/PD parameters that were incorporated into a combined PK/PD model to predict the efficacy in terms of reduction of bacterial counts measured in log CFU/mL. Based on the results of the simulations, an optimal dose was chosen for the <em>in vivo</em> efficacy study.</p> <p>The results from the <em>in vitro</em> PK/PD studies showed that QD was marginally effective. The same total daily dose administered BID showed a marked reduction in mycobacterial counts. TID dosing did not show a significant difference in time‑kill compared to BID regimen. The maximum growth rate constant (K₀) was estimated as 0.0274 hr^-1 which corresponds to a maximum doubling time of 25.3 hr that is consistent with the commonly observed <em>in vitro</em> doubling times of 20‑24 hr. A maximum bacterial kill rate (I_max) was calculated as 0.0566 hr^-1. The free drug concentration required to produce half‑maximum inhibition (IC₅₀) was calculated as 2.62 mg/L. The delay rate constant for the initial kill was found to be 0.0245 hr^-1. The PK bridging study showed that the Lee1445 half‑life was shorter in mice (0.25 hr) compared to rats (0.43 hr). Simulations based on the parameters obtained from the mouse PK bridging study and the <em>in vitro</em> PK/PD model suggested that a total daily dose of 200 mg/Kg/day and 400 mg/Kg/day administered BID would be optimal and result in approximately 2 and 3 log reduction in bacterial counts respectively after seven days of therapy and hence were chosen for <em>in vivo</em> efficacy studies in mice. </p> <p>In summary, we have successfully developed a series of biopharmaceutic, PK and PD experiments that help in an iterative PK/PD guided approach for development of spectinamides, a novel class of antituberculosis agents. The <em>in vivo</em> PK and <em>in vitro</em> PK/PD parameters obtained from these studies provide a basis for optimal compound as well as dose selection.</p>

Name thesis:degree_name

Doctor of Philosophy (PhD)

Level thesis:degree_level

Dissertation

Discipline thesis:degree_discipline

Pharmaceutical Sciences

Year dc:date.available

2011

Author dc:creator

• Vaddady, V. N. R. Pavan Kumar

Contributors dc:contributor

• Bernd Meibohm, Ph.D.

• In Vitro PK/PD Model Pharmacodynamics
• Pharmacokinetics
• Pharmacometrics
• Tuberculosis
• Bacterial Infections and Mycoses
• Diseases
• Medicinal and Pharmaceutical Chemistry
• Medicine and Health Sciences
• Pharmaceutics and Drug Design
• Pharmacy and Pharmaceutical Sciences

Repository record dc:identifier

https://dc.uthsc.edu/dissertations/278

OAI identifier oai:identifier

oai:dc.uthsc.edu:dissertations-1277