All-in-one glycol chitosan nanoparticles for co-delivery of doxorubicin and anti-PD-L1 peptide in cancer immunotherapy.

Synergistic immunotherapy of immune checkpoint blockade (ICB) and immunogenic cell death (ICD) has shown remarkable therapeutic efficacy in various cancers. However, patients show low response rates and undesirable outcomes to these combination therapies owing to the recycling mechanism of programmed death-ligand 1 (PD-L1) and the systemic toxicity of ICD-inducing chemotherapeutic drugs. Herein, we propose all-in-one glycol chitosan nanoparticles (CNPs) that can deliver anti-PD-L1 peptide (PP) and doxorubicin (DOX) to targeted tumor tissues for a safe and more effective synergistic immunotherapy. The PP-CNPs, which are prepared by conjugating ᴅ-form PP (NYSKPTDRQYHF) to CNPs, form stable nanoparticles that promote multivalent binding with PD-L1 proteins on the targeted tumor cell surface, resulting in effective lysosomal PD-L1 degradation in contrast with anti-PD-L1 antibody, which induces recycling of endocytosed PD-L1. Consequently, PP-CNPs prevent subcellular PD-L1 recycling and eventually destruct immune escape mechanism in CT26 colon tumor-bearing mice. Moreover, the ICD inducer, DOX is loaded into PP-CNPs (DOX-PP-CNPs) for synergistic ICD and ICB therapy, inducing a large number of damage-associated molecular patterns (DAMPs) in targeted tumor tissues with minimal toxicity in normal tissues. When the DOX-PP-CNPs are intravenously injected into CT26 colon tumor-bearing mice, PP and DOX are efficiently delivered to the tumor tissues via nanoparticle-derived passive and active targeting, which eventually induce both lysosomal PD-L1 degradation and substantial ICD, resulting in a high rate of complete tumor regression (CR: 60%) by a strong antitumor immune response. Collectively, this study demonstrates the superior efficacy of synergistic immunotherapy using all-in-one nanoparticles to deliver PP and DOX to targeted tumor tissues.

Sustained and Long-Term Release of Doxorubicin from PLGA Nanoparticles for Eliciting Anti-Tumor Immune Responses.

Immunogenic cell death (ICD) is a powerful trigger eliciting strong immune responses against tumors. However, traditional chemoimmunotherapy (CIT) does not last long enough to induce sufficient ICD, and also does not guarantee the safety of chemotherapeutics. To overcome the disadvantages of the conventional approach, we used doxorubicin (DOX) as an ICD inducer, and poly(lactic-co-glycolic acid) (PLGA)-based nanomedicine platform for controlled release of DOX. The diameter of 138.7 nm of DOX-loaded PLGA nanoparticles (DP-NPs) were stable for 14 days in phosphate-buffered saline (PBS, pH 7.4) at 37 °C. Furthermore, DOX was continuously released for 14 days, successfully inducing ICD and reducing cell viability in vitro. Directly injected DP-NPs enabled the remaining of DOX in the tumor site for 14 days. In addition, repeated local treatment of DP-NPs actually lasted long enough to maintain the enhanced antitumor immunity, leading to increased tumor growth inhibition with minimal toxicities. Notably, DP-NPs treated tumor tissues showed significantly increased maturated dendritic cells (DCs) and cytotoxic T lymphocytes (CTLs) population, showing enhanced antitumor immune responses. Finally, the therapeutic efficacy of DP-NPs was maximized in combination with an anti-programmed death-ligand 1 (PD-L1) antibody (Ab). Therefore, we expect therapeutic efficacies of cancer CIT can be maximized by the combination of DP-NPs with immune checkpoint blockade (ICB) by achieving proper therapeutic window and continuously inducing ICD, with minimal toxicities.

The safe and effective intraperitoneal chemotherapy with cathepsin B-specific doxorubicin prodrug nanoparticles in ovarian cancer with peritoneal carcinomatosis.

Intraperitoneal (IP) chemotherapy has shown promising efficacy in ovarian cancer with peritoneal carcinomatosis (PC), but in vivo rapid clearance and severe toxicity of free anticancer drugs hinder the effective treatment. Herein, we propose the safe and effective IP chemotherapy with cathepsin B-specific doxorubicin prodrug nanoparticles (PNPs) in ovarian cancer with PC. The PNPs are prepared by self-assembling cathepsin B-specific cleavable peptide (FRRG) and doxorubicin (DOX) conjugates, which are further formulated with pluronic F68. The PNPs exhibit stable spherical structure and cytotoxic DOX is specifically released from PNPs via sequential enzymatic degradation by cathepsin B and intracellular proteases. The PNPs induce cytotoxicity in cathepsin B-overexpressing ovarian (SKOV-3 and HeyA8) and colon (MC38 and CT26) cancer cells, but not in cathepsin B-deficient normal cells in cultured condition. With enhanced cancer-specificity and in vivo residence time, IP injected PNPs efficiently accumulate within PC through two targeting mechanisms of direct penetration (circulation independent) and systemic blood vessel-associated accumulation (circulation dependent). As a result, IP chemotherapy with PNPs efficiently inhibit tumor progression with minimal side effects in peritoneal human ovarian tumor-bearing xenograft (POX) and patient derived xenograft (PDX) models. These results demonstrate that PNPs effectively inhibit progression of ovarian cancer with peritoneal carcinomatosis with minimal local and systemic toxicities by high cancer-specificity and favorable in vivo PK/PD profiles enhancing PC accumulation.

Doxorubicin-Loaded PLGA Nanoparticles for Cancer Therapy: Molecular Weight Effect of PLGA in Doxorubicin Release for Controlling Immunogenic Cell Death.

Direct local delivery of immunogenic cell death (ICD) inducers to a tumor site is an attractive approach for leading ICD effectively, due to enabling the concentrated delivery of ICD inducers to the tumor site. Herein, we prepared doxorubicin (DOX)-loaded poly(lactic-co-glycolic acid) (PLGA) nanoparticles (NPs) using different molecular weight PLGA (7000 g/mol and 12,000 g/mol), showing different drug release kinetics. The different release kinetics of DOX might differently stimulate a tumor cell-specific immune response by releasing damage-associated molecular patterns (DAMPs), resulting in showing a different antitumor response in the living body. DOX-PLGA7K NPs showed faster DOX release kinetics than DOX-PLGA12K NPs in the physiological condition. DOX-PLGA7K NPs and DOX-PLGA12K NPs were successfully taken up by the CT-26 tumor cells, subsequently showing different DOX localization times at the nucleus. Released DOX successfully lead to cytotoxicity and HMGB1 release in vitro. Although the DOX-PLGA7K NPs and DOX-PLGA12K NPs showed different sustained DOX release kinetics in vitro, tumor growth of the CT-26 tumor was similarly inhibited for 28 days post-direct tumor injection. Furthermore, the immunological memory effect was successfully established by the ICD-based tumor-specific immune responses, including DC maturation and tumor infiltration of cytotoxic T lymphocytes (CTLs). We expect that the controlled release of ICD-inducible chemotherapeutic agents, using different types of nanomedicines, can provide potential in precision cancer immunotherapy by controlling the tumor-specific immune responses, thus improving the therapeutic efficacy.

Inhibitory effect of tramadol on vasorelaxation mediated by ATP-sensitive K+ channels in rat aorta.

PURPOSE: Tramadol produces a conduction block similar to lidocaine by exerting a local anesthetic-like effect. The aims of this in vitro study were to determine the effects of tramadol on the vasorelaxant response induced by the adenosine triphosphate-sensitive K(+) (K(ATP)) channel opener, levcromakalim, in an endothelium-denuded rat aorta, and to determine whether this effect of tramadol is stereoselective. METHODS: The effects of tramadol (racemic, R(-) and S(+): 10(-6), 10(-5), 5 x 10(-5) M), and glibenclamide on the levcromakalim dose-response curve were assessed in aortic rings that had been pre-contracted with phenylephrine. In the rings pretreated independently with naloxone, and glibenclamide, the levcromakalim dose-response curves were generated in the presence or absence of tramadol. The effect of tramadol on the dose-response curve of diltiazem was assessed. RESULTS: Racemic, R(-) and S(+) tramadol (10(-5), 5 x 10(-5) M) attenuated (P < 0.0001) levcromakalim-induced relaxation in the ring with or without naloxone in a dose-dependent manner. The magnitude of the R(-)-tramadol-induced attenuation of vasorelaxant response induced by levcromakalim was greater (P < 0.05) than that induced by S(+)-tramadol. Glibenclamide almost abolished the levcromakalim-induced relaxation. Tramadol, 5 x 10(-5) M, did not significantly alter the diltiazem-induced relaxation. CONCLUSION: These results suggest that a supraclinical dose (10(-5) M) of tramadol [racemic, R(-) and S(+)] attenuates the vasorelaxation mediated by the K(ATP) channels in the rat aorta. The R(-) tramadol-induced attenuation of vasorelaxation induced by levcromaklim was more potent than that induced by S(+) tramadol. This attenuation is independent of opioid receptor activation.