Drug retention after intradiscal administration

Abstract

Intradiscal drug delivery is a promising strategy for treating intervertebral disk degeneration (IVDD). Local degenerative processes and intrinsically low fluid exchange are likely to influence drug retention. Understanding their connection will enable the optimization of IVDD therapeutics. Release and retention of an inactive hydrophilic fluorine-19 labeled peptide (¹⁹F-P) as model for regenerative peptides was studied in a whole IVD culture model by measuring the ¹⁹F-NMR (nuclear magnetic resonance) signal in culture media and IVD tissue extracts. In another set-up, noninvasive near-infrared imaging was used to visualize IR-780, as hydrophobic small molecular drug model, retention upon injection into healthy and degenerative caudal IVDs in a rat model of disk degeneration. Furthermore, IR-780-loaded degradable polyester amide microspheres (PEAM) were injected into healthy and needle pricked degenerative IVDs, subcutaneously, and in knee joints with and without surgically-induced osteoarthritis (OA). Most ¹⁹F-P was released from the IVD after 7 days. IR-780 signal intensity declined over a 14-week period after bolus injection, without a difference between healthy and degenerative disks. IR-780 signal declined faster in the skin and knee joints compared to the IVDs. IR-780 delivery by PEAMs enhanced disk retention beyond 16 weeks. Moreover, in degenerated IVDs the IR-780 signal was higher over time than in healthy IVDs while no difference between OA and healthy joints was noted. We conclude that the clearance of peptides and hydrophobic small molecules from the IVD is relatively fast. These results illustrate that development of controlled release formulations should take into account the target anatomical location and local (patho)biology.

Keywords: Drug retention; intervertebral disk degeneration; peptide; polyester amide microsphere; small molecule.

Graphical abstract

Figure 1.

Schematic overview of the study designs including ex vivo bovine IVD explants and in vivo rat study. IVD: intervertebral disk; IVDD: intervertebral disk degeneration; NIR: near-infrared; PEA: polyesteramide.

Figure 2.

19F-P release and retention in cultured bovine caudal intervertebral disk (IVD) after intradiscal injection evaluated by nuclear magnetic resonance. (A) Evaluation and (B) quantification of ¹⁹F-P in bovine AF and NP tissue extracts, 3 h after intradiscal injection using ¹⁹F-NMR spectroscopy. (C) The release profile and (D) retention of ¹⁹F-P in bovine caudal IVDs cultured with daily dynamic loading for 7 days after intradiscal injection. AF: annulus fibrosus; NP: nucleus pulposus.

Figure 3.

Retention of free IR-780 after intradiscal injection in healthy or degenerated intervertebral disks (IVDs). (A) Decreased disk height index (DHI), 4 weeks after degeneration induction by NP needle puncture. *p < 0.05. (B) Histologic overview of a healthy and degenerated IVD, stained with picrosirius red/alcian blue. The dotted circle indicates a distinct NP (left), and black arrows indicate disrupted endplates (right). Scale bar = 500 μm. (C) Quantification of IR-780 intensity signal after IR-780 injection in healthy (n = 4) or degenerated (n = 4) rat IVDs. The IR-780 intensity was normalized to 2 weeks after injection. Data represent mean ± SEM. (D) IR-780 signal after 4 and 8 weeks in tail IVDs, detected by fluorescence optical imaging. AF: annulus fibrosus; NP: nucleus pulposus.

Figure 4.

Tissue-dependent retention of IR-780 released from polyesteramide (PEA) microspheres over 16 weeks. (A) Quantification of IR-780 intensity signal after IR-780 injection in healthy skin (n = 5), knee joint (n = 5), and IVD (n = 4). The IR-780 intensity was normalized to the signal 2 weeks after injection. Data represent mean ± SEM. A significant increase in IR-780 intensity was found in IVD compared to skin and knee joints, indicated by *p < 0.05. (B) Fluorescence optical images 4 and 8 weeks after IR-780 loaded PEA microsphere injections in rats. The yellow dotted box indicates healthy IVD. IVD: intervertebral disk.

Figure 5.

Disease-dependent joint retention of IR-780 over 16 weeks, released from polyesteramide (PEA) microspheres in rats. (A) Quantification of IR-780 signal after IR-780 loaded PEA microsphere (PEA-IR780) injection in healthy IVDs (n = 4) versus degenerated IVDs (n = 4) in rats. NIR intensity was normalized to 2 weeks after injection. Data represent mean ± SEM. A significant increase of IR-780 intensity released from PEA microspheres was found in degenerated IVDs compared to healthy IVDs (*p < 0.05). (B) Fluorescence optical images 4 and 8 weeks after PEA-IR780 injections in healthy and degenerated tail IVDs in rats. The blue arrows indicate healthy IVD, the red arrows indicate degenerated IVD. (C) Quantification of IR-780 intensity signal after PEA-IR780 injection in healthy knee joints (n = 5) versus degenerated knee joints (n = 5). The IR-780 intensity was normalized to 2 weeks after injection. Data represent mean ± SEM, and no significant differences were found. (D) Fluorescence optical images 4 and 8 weeks after PEA-IR780 injections in healthy and degenerated knee joints in rats. IVDs: intervertebral disks.