Neutrophil-Mediated Delivery of Dexamethasone Palmitate-Loaded Liposomes Decorated with a Sialic Acid Conjugate for Rheumatoid Arthritis Treatment.

PURPOSE: The aim of this research was to design dexamethasone palmitate (DP) loaded sialic acid modified liposomes, with the eventual goal of using peripheral blood neutrophils (PBNs) that carried drug-loaded liposomes to improve the therapeutic capacity for rheumatoid arthritis (RA). METHODS: A sialic acid - cholesterol conjugate (SA-CH) was synthesized and anchored on the surface of liposomal dexamethasone palmitate (DP-SAL). The physicochemical characteristics and in vitro cytotoxicity of liposomes were evaluated. Flow cytometry and confocal laser scanning microscopy were utilized to investigate the accumulation of liposomes in PBNs. The adjuvant-induced arthritis was adopted to investigate the targeting ability and anti-inflammatory effect of DP loaded liposomes. RESULTS: Both DP-CL and DP-SAL existed an average size less than 200 nm with remarkably high encapsulation efficiencies more than 90%. In vitro and in vivo experiments manifested SA-modified liposomes provided a reinforced accumulation of DP in PBNs. As well, DP-SAL displayed a greater degree of accumulation in the joints and a stronger anti-inflammatory effect in terms of RA suppression. CONCLUSIONS: SA-modified liposomal DP was a promising candidate for RA-targeting treatment through the neutrophil-mediated drug delivery system.

Dual-Ligand Modification of PEGylated Liposomes Used for Targeted Doxorubicin Delivery to Enhance Anticancer Efficacy.

Mannose receptor (CD206) and E-selectin are selectively expressed in M2-like tumor-associated macrophages (M2-TAMs) and activated endothelial cells of vessels surrounding tumor tissues. With the knowledge that D-mannose is the natural ligand of mannose receptors and L-fucose is the key calcium chelator for tumor-associated carbohydrate antigens (TACAs) binding to E-selectin, herein, we firstly reported D-mannose polyethylene glycol (PEG) conjugates (Man-PEG) and L-fucose PEG conjugates (Fuc-PEG) co-modified liposomal doxorubicin (DOX-MFPL) to improve tumor-targeting ability. The dual-ligand modified PEGylated liposomes (DOX-MFPL) were assessed by both in vitro and in vivo trials. Compared with the single-ligand D-mannose- or L-fucose-modified liposomes (DOX-MPL or DOX-FPL), DOX-MFPL achieved an increased distribution of DOX in tumor tissues. The antitumor study based on S180 tumor-bearing mice was conducted and the superior tumor inhibitory rate was shown with DOX-MFPL, probably owing to the superior tumor-targeting effect of DOX-MFPL and the modulation of the tumor microenvironment with the exhaustion of TAMs. In general, the dual-ligand drug delivery systems are expected to be promising in the development of specific and efficient methods for tumor treatment.

Use of Dual-Ligand Modification in Kupffer Cell-Targeted Liposomes To Examine the Contribution of Kupffer Cells to Accelerated Blood Clearance Phenomenon.

The "accelerated blood clearance (ABC) phenomenon" is known to be involved in the adaptive immune system. Regretfully, the relationship between the ABC phenomenon and innate immune system, especially with respect to Kupffer cells (KCs) has been largely unexplored. In this study, the contribution of KCs to ABC was examined using the 4-aminophenyl-α-d-mannopyranoside (APM) lipid derivative DSPE-PEG2000-APM (DPM) and the 4-aminophenyl-ß-l-fucopyranoside (APF) lipid derivative DSPE-PEG2000-APF (DPF) as ligands for mannose/fucose receptors on KCs, which were synthesized and modified on the surface of liposomes. The results of cellular liposome uptake in vitro and biodistribution in vivo indicated that DPM and DPF comodified liposomes (MFPL5-5) present the strongest capability of KC-targeting among all preparations tested. In rats pretreated with MFPL5-5 instead of PEGylated liposomes (PL), the ABC phenomenon was significantly enhanced and the distribution of liposomes in the liver was increased. Cellular uptake of the second injection of PL in vivo demonstrated that KCs was responsible for the uptake. Furthermore, compared to pretreatment with PL, the uptake of second injection of PL was more enhanced when pretreated with MFPL5-5. These findings suggest that KCs, which are considered traditional members of the innate immune system, play a crucial role in the ABC phenomenon and act as a supplement to the phenomenon.

Redox- and pH-Sensitive Glycan (Polysialic Acid) Derivatives and F127 Mixed Micelles for Tumor-Targeted Drug Delivery.

With increasing application of PEGylated products, drawbacks are beginning to emerge such as the "PEG dilemma". Other promising materials may need to be found in the current situation. Endogenous polysialic acid (PSA), which is highly expressed on mammalian, bacterial, and malignant surface, may be a promising material in oncology. In this study, a dual-responsive amphiphilic PSA cholesterol derivative (PSA-CS-CH) was synthesized to explore the opportunity of PSA in targeted drug delivery systems. PSA-CS-CH, F127 mixed micelles (PF-M), and pure F127 micelles (F-M) were prepared for comparative antitumor experiments. The in vitro experiments showed that modification of PSA-CS-CH significantly increased cytotoxicity and cellular uptake. PF-M had excellent tumor microenvironment response release behavior on acidic media with high GSH levels. The in vivo fluorescence imaging and antitumor experiments showed that PF-M had excellent tumor targeting ability and great tumor suppression ability. In summary, biodegradable PSA may contribute to cancer therapy.

Persistent immune response: Twice tumor exfoliation induced by sialic acid-modified vincristine sulfate liposomes.

Studies have shown that tumor-associated macrophages (TAMs) are crucial for the establishment and maintenance in immunosuppressive tumor immune microenvironment (TIME), which can help tumor cells to achieve immune escape and attenuate antitumor therapy. Siglecs, the receptors of sialic acid (SA), widely exist in TAMs, which could be targeted to disrupt TIME and inhibit tumor growth at the root. Therefore, a SA-modified VCR liposome was reported (VCR-SSAL). Cellular and pharmacodynamic experiments showed that VCR-SSAL exhibited strong TAMs targeting and tumor-killing ability. Interestingly, VCR-SSAL treatment induced a phenomenon in which the cancerous tissues were "fell off" from the growth site, after which the wound gradually healed. Three months after the wound healed, the mice whose tumors fell off were re-inoculated, and the tumor fell off again without treatment, with an exfoliation rate of 100%. We speculated that this special efficacy might be due to that VCR loaded in VCR-SSAL could activate adaptive immunity by inducing DNA damage, promoting cytotoxic T lymphocytes (CTLs) infiltration into tumor sites, and enhancing the antitumor immune response. Thus, this study might provide new insights into the application of traditional chemotherapeutic drugs.

Discovery in polyethylene glycol immunogenicity: The characteristic of intergenerational inheritance of anti-polyethylene glycol IgG.

Several studies have reported the prevalence of anti-polyethylene glycol (PEG) antibodies (APAs) in healthy people, highlighting the widespread existence of APAs. The prevalence of anti-PEG immunoglobulin (Ig)G is significantly negatively correlated with age. Here, we used Wistar rats as model organism to examine whether APAs in parental rats can affect the production of antibodies in their offspring. After being pre-stimulated with blank PEGylated nanoemulsions (PE) to induce APAs production, parental rats were paired in cages. The presence of antibodies in the parents and offspring was detected using enzyme-linked immunosorbent assay. The presence of antibodies in the parental rats led to significant anti-PEG IgG positivity in their offspring, indicating that anti-PEG IgG exhibits intergenerational inheritance. Moreover, anti-PEG IgG in the offspring rats could bind to PE and accelerate its blood clearance. To the best of our knowledge, this is the first report on the intergenerational properties of APAs.

Sialic acid conjugate-modified liposomal platform modulates immunosuppressive tumor microenvironment in multiple ways for improved immune checkpoint blockade therapy.

Immune checkpoint blockade (ICB) treatment is promising for the clinical therapy of numerous malignancies. However, most cancer patients rarely benefit from such single-agent immunotherapies because of the complexity of both the tumor and tumor microenvironment. A tumor-specific liposomal vehicle (DOX-SAL) modified with a sialic acid-cholesterol conjugate (SA-CH) and remotely loaded with doxorubicin (DOX) is herein reported for improving chemoimmunotherapy. The intravenous administration of DOX-SAL dramatically downregulates tumor-associated macrophage (TAM)-mediated immunosuppression, inhibits immunoregulatory functions, and promotes intratumoral infiltration of CD8+ T cells. Compared to conventional liposomes, DOX-SAL-mediated combination therapy with a PD-1-blocking monoclonal antibody (aPD-1 mAb) almost completely eliminates B16F10 tumors and efficiently inhibits 4T1 tumors. Moreover, cancer stem cells exhibit efficient tumor-initiating, tumor-propagating, and immunosuppressive tumor microenvironment-shaping capabilities. To further improve the treatment efficacy of an immunologically "cold" tumor, metformin (MET), which selectively eradicates breast cancer tumor stem cells, is co-encapsulated with DOX into liposomes to develop DOX/MET-SAL. The combination therapy with DOX/MET-SAL and aPD-1 mAb in a 4T1 orthotopic mouse model indicates their synergetic benefit on primary tumor inhibition, metastasis suppression, and survival rate improvement. Thus, the multifunctional liposomal platform has potential value for ICB combination immunotherapy.

Sialic acid-conjugate modified liposomes targeting neutrophils for improved tumour therapy.

Neutrophils are the most abundant white blood cells in humans. Many tumor-treatment methods that are related to tissue infiltration and the activation of neutrophils have been developed. In particular, one strategy, which aims to improve tumor treatment, involves the exploitation or targeting of activated neutrophils. Peripheral blood neutrophils (PBNs) from tumor-bearing mice display high expression of l-selectin, which is well known to be targeted by the sialic acid (SA) ligand. Hence, in this research, we developed a drug delivery platform involving liposomes modified with an SA conjugate that targets activated PBNs. The uptake of doxorubicin (DOX)-loaded liposomes by PBNs did not alter their activation and transmigration. Furthermore, in tumor-bearing mice, SA-modified liposomes displayed a greater tumor-targeting ability and stronger tumor treatment efficacy, which were mediated by the neutrophil infiltration induced by inflammatory factors released from the tumor microenvironment. In conclusion, SA-modified liposomal DOX was shown to be an effective neutrophil-mediated drug delivery system for tumor therapy.

Exhausting tumor associated macrophages with sialic acid-polyethyleneimine-cholesterol modified liposomal doxorubicin for enhancing sarcoma chemotherapy.

To overstep the dilemma of chemical drug degradation within powerful lysosomes of tumor associated macrophages (TAMs), a sialic acid-polyethylenimine-cholesterol (SA-PEI-CH) modified liposomal doxorubicin (DOX-SPCL) was designed with both TAMs targeting and smart lysosomal trafficking. The modified liposome DOX-SPCL performed particle size as 103.2 ±â€¯3.1 nm and zeta potential as -4.5 ±â€¯0.9 mV with encapsulation efficiency as 95.8 ±â€¯0.5%. In in vitro cell experiments, compared with conventional liposomal doxorubicin (DOX-CL) and PEGylated liposomal doxorubicin (DOX-PL), DOX-SPCL showed a selective binding on TAMs and a mere lysosomal concentration. In pharmacokinetic study, DOX-SPCL effectively impeded/delayed the disposition of mononuclear phagocyte system (MPS) with a value of AUC0-t as 796.03 ±â€¯66.93 mg L-1 h. In S180 sarcomas bearing mice, DOX-SPCL showed the greatest tumor inhibition rate (92.7% ±â€¯3.6%) compared with DOX-CL (46.4% ±â€¯2.0%) or DOX-PL (58.8% ±â€¯7.6%). The <0.5% positive region of TAMs in tumor section indicated a super TAMs exhaustion for DOX-SPCL treatment. Conclusively, DOX-SPCL was supposed as a safe and effective liposomal preparation for clinical sarcoma treatment via TAMs targeting/deletion delivery strategy.

Targeted delivery of pixantrone to neutrophils by poly(sialic acid)-p-octadecylamine conjugate modified liposomes with improved antitumor activity.

Based on the knowledge that poly(sialic acid) is a critical element for tumour development and that the receptors for its monomer are expressed on neutrophils, which play important roles in the progression and invasion of tumours, a poly(sialic acid)-p-octadecylamine conjugate (PSA-p-ODA) was synthesised and used to modify the surface of liposomal pixantrone (Pix-PSL) to improve the delivery of Pix to peripheral blood neutrophils (PBNs). The liposomes were fabricated using a remote loading technology via a pH gradient, and were then assessed for particle size, encapsulation efficiency, in vitro release, in vitro cytotoxicity, and pharmacokinetics. Simultaneously, in vitro and in vivo cellular uptake studies demonstrated that Pix-PSL provided an enhanced accumulation of Pix in PBNs. An in vivo study showed that the anti-tumour activity of Pix-PSL was superior to that of other formulations, probably owing to the efficient targeting of PBNs by Pix-PSL, after which PBNs containing Pix-PSL (Pix-PSL/PBNs) in the circulatory system are recruited by the tumour microenvironment. These findings suggest that PSA-p-ODA-decorated liposomal Pix may provide a neutrophil-mediated drug delivery system (DDS) for the eradication of tumours, and thus represents a promising approach for the tumour targeting of chemotherapeutic treatments.

Neutrophil-mediated delivery of pixantrone-loaded liposomes decorated with poly(sialic acid)-octadecylamine conjugate for lung cancer treatment.

Poly(sialic acid) (PSA) is a natural hydrophilic biodegradable and non-immunogenic biopolymer, receptors for its monomer are expressed on peripheral blood neutrophils (PBNs), which plays important roles in the progression and invasion of tumors. A poly(sialic acid)-octadecylamine conjugate (PSA-ODA) was synthesized and then anchor it on the surface of liposomal pixantrone (Pix-PSL), to achieve an improved anticancer effect. The liposomes were prepared using a remote loading method via a pH gradient, and then assessed for particle size, zeta potential encapsulation efficiency, in vitro release, and in vitro cytotoxicity. Simultaneously, in vitro and in vivo cellular uptake studies confirmed that PSA-decorated liposomes provided an enhanced accumulation of liposomes in PBNs. An in vivo study presented that the anti-tumor activity of Pix-PSL was superior to that of other Pix formulations, probably due to the efficient targeting of PBNs by Pix-PSL, after which PBN containing Pix-PSL (Pix-PSL/PBNs) in the blood circulation are recruited by the tumor microenvironment. These findings suggest that PSA-decorated liposomal Pix may provide a neutrophil-mediated drug delivery system (DDS) for the eradication of tumors, which represents a promising approach for the tumor targeting of chemotherapeutic treatments.