Drug-induced amplification of nanoparticle targeting to tumors

Abstract

Nanomedicines have the potential to significantly impact cancer therapy by improving drug efficacy and decreasing off-target effects, yet our ability to efficiently home nanoparticles to disease sites remains limited. One frequently overlooked constraint of current active targeting schemes is the relative dearth of targetable antigens within tumors, which restricts the amount of cargo that can be delivered in a tumor-specific manner. To address this limitation, we exploit tumor-specific responses to drugs to construct a cooperative targeting system where a small molecule therapeutic modulates the disease microenvironment to amplify nanoparticle recruitment in vivo. We first administer a vascular disrupting agent, ombrabulin, which selectively affects tumors and leads to locally elevated presentation of the stress-related protein, p32. This increase in p32 levels provides more binding sites for circulating p32-targeted nanoparticles, enhancing their delivery of diagnostic or therapeutic cargos to tumors. We show that this cooperative targeting system recruits over five times higher doses of nanoparticles to tumors and decreases tumor burden when compared with non-cooperative controls. These results suggest that using nanomedicine in conjunction with drugs that enhance the presentation of target antigens in the tumor environment may be an effective strategy for improving the diagnosis and treatment of cancer.

Keywords: Nanomedicine; active targeting; cancer therapy; liposomes; magnetic nanoworms; vascular disrupting agent.

Figure 1. Schematic of cooperative targeting approach

Inducing agent, ombrabulin, disrupts the tumor vasculature, which initiates a cascade of intratumoral effects that lead to upregulated presentation of the p32 protein. LyP-1 coated nanoparticles, which target the p32 protein, are then able to home to the tumor.

Figure 2. Characterization of ombrabulin effect on tumor microenvironment

(A) Schematic of ombrabulin-induced enhancement in p32 presentation. (B) Tumors at 24 hr post injection (p.i.) revealing ombrabulin mediated hemorrhaging. (C) Hematoxylin and eosin staining of tumors harvested from mice injected with ombrabulin at different timepoints p.i. (Con., control (0 mg/kg); scale bar = 100 µm). (D) Immunofluorescent staining of tumors without (0 mg/kg) and with (60 mg/kg) ombrabulin (red = p32 staining, blue = nuclear stain; scale bar = 1 mm). (E) Quantification of percentage p32 positive area of immunofluorescent staining of tumors receiving different dosages of ombrabulin at different timepoints p.i. (n = 3 mice, s.e.). (F) Quantification of percentage p32 positive human cells from tumors receiving different dosages of ombrabulin at different timepoints as determined by flow cytometry (*** P < 0.005, one-way ANOVA with Tukey post test; n = 3 mice, s.e.).

Figure 3. Ombrabulin mediated amplification of NW delivery

(A) Schematic of ombrabulin signaling to NWs. Ombrabulin upregulates the presentation of p32 in tumors, which is then targeted by NW-LyP1. (B) Experimental timeline for testing the signaling system. (C) Quantification of NW homing to tumors as a function of ombrabulin dosage (** P < 0.01, Student’s t-test; n = 3–4 mice, s.e.). (D) Representative near-infrared fluorescent scans of NW homing to tumors in response to increasing doses of ombrabulin. Tumors were excised and imaged at 24 hrs post-NW injection. (E) Immunofluorescent staining of NWs in tumors without (0 mg/kg) and with (60 mg/kg) ombrabulin (green = NW, red = p32 staining, blue = nuclear stain; scale bar = 100 µm).

Figure 4. Ombrabulin mediated amplification of LP delivery

(A) Schematic of ombrabulin signaling to LPs. Ombrabulin upregulates the presentation of p32 in tumors, which is then targeted by LP-LyP1. (B) Quantification of doxorubicin-loaded LP homing to tumors as a function of ombrabulin dosage (*** P < 0.005, Student’s t-test; n = 3 mice, s.e.). (C) Immunofluorescent staining of LPs in tumors without (0 mg/kg) and with (60 mg/kg) ombrabulin (green = LP, red = p32 staining, blue = nuclear stain; scale bar = 100 µm). (D) Quantification of doxorubicin-loaded LP biodistribution in organs without (0 mg/kg) and with (60 mg/kg) ombrabulin (No significance, one-way ANOVA with Tukey post test; n = 3–6 mice, s.d.).

Figure 5. Therapeutic efficacy of cooperative targeting system

(A) Tumor volumes of different groups following three weeks of treatment. Black arrow head denotes time of ombrabulin (60 mg/kg) administration; orange arrow head denotes time of LP (1 mg/kg by dox) administration (* P < 0.05, ** P < 0.01, *** P < 0.005, two-way ANOVA with Bonferroni post test, n = 7 mice, s.e.). (B) Change in body weight of different groups following three weeks of treatment (n = 7 mice, s.e.). (C) Survival rate of different groups in the therapeutic efficacy study (** P < 0.01, log rank test; n = 7 mice).