Replenishable drug depot to combat post-resection cancer recurrence

Abstract

Local drug presentation made possible by drug-eluting depots has demonstrated benefits in a vast array of diseases, including in cancer, microbial infection and in wound healing. However, locally-eluting depots are single-use systems that cannot be refilled or reused after implantation at inaccessible sites, limiting their clinical utility. New strategies to noninvasively refill drug-eluting depots could dramatically enhance their clinical use. In this report we present a refillable hydrogel depot system based on bioorthogonal click chemistry. The click-modified hydrogel depots capture prodrug refills from the blood and subsequently release active drugs locally in a sustained manner. Capture of the systemically-administered refills serves as an efficient and non-toxic method to repeatedly refill depots. Refillable depots in combination with prodrug refills achieve sustained release at precancerous tumor sites to improve cancer therapy while eliminating systemic side effects. The ability to target tissues without enhanced permeability could allow the use of refillable depots in cancer and many other medical applications.

Keywords: Biomaterials; Cancer therapy; Drug delivery; Drug depot; Sustained release.

Figure 1

Refillable drug depots allow non-invasive targeting of tissues. Schematic for noninvasive refilling of therapeutic depots. Therapeutic depots (gray) carrying azide groups are implanted at tumor sites during surgery (left). Non-toxic prodrug consisting of cytotoxic drug conjugated to a targeting motif through a cleavable linker is administered systemically and is covalently captured at the depot (middle). Cleavage of the linker over a period of weeks allows release of drug (right).

Figure 2. Click chemistry-mediated targeting of a drug surrogate to subcutaneous space

Mice were injected with unconjugated control alginate hydrogels (left flank) or azide-conjugated hydrogels (right flank). Targeting to this gel was tested through i.v. injection of a fluorescently labeled DBCO. A) representative IVIS images, B) quantification of fluorescence at the gel site after 48 hours. Values represent the mean +/− SEM, n=3. p-value from Student’s two-tailed t-test (homoscedastic).

Figure 3. Click chemistry-mediated targeting to intratumoral hydrogels

Mice were injected with MDA-MB-231 tumor cells combined with hydrogels (azide-conjugated or control). Targeting to this gel was tested through i.v. injection of a fluorescently labeled DBCO at different time points. A) representative I VIS images, B: quantification of fluorescence at the gel site 24 hours after i.v. injection. Values represent the mean +/− SEM, n=6. ** = p <0.005 by Student’s two-tailed t-test.

Figure 4. A slow-cleaving, DBCO-conjugated doxorubicin prodrug

A) Synthesis of a chemotherapeutic prodrug linking doxorubicin (purple) to cyclooctyne (blue) through a hydrolyzable linker. B) LCMS quantitation of the hydrolysis of doxorubicin prodrug over time at different pH. C) In vitro toxicity of doxorubicin, doxorubicin prodrug and vehicle incubated with MDA-MB-231 mammalian cells. ** = p<01 determined by Holm-Sidak method for multiple comparison testing. n=3.

Figure 5. In vivo safety of click-enabled doxorubicin prodrugs

Mouse weight over time with administrations of doxorubicin (red), its cyclooctyne-linked prodrug (blue), or vehicle (green). Twice-weekly intraperitoneal administrations were evaluated for five weeks with doxorubicin or molar equivalent of prodrug. * = p<05 and ** = p<01 doxorubicin vs. dox prodrud for multiple comparison t-test with Holm-Sidak correction. n=5.

Figure 6. Efficacy of doxorubicin prodrugs in combination with refillable depots

A) Average tumor size over time in MDA-MB-231 bearing mice treated with azide-conjugated or control gels and twice-weekly adminsitration doxorubicin prodrug or vehicle. B) Mouse weight over time for the three experimental groups. * p<.05 by Wicoxon’s post-hoc comparisons testing. n=12. C) Repe- sentative tumor images.