CCL18 from tumor-associated macrophages promotes angiogenesis in breast cancer.

The infiltration of tumor-associated macrophages (TAMs) is associated with extensive angiogenesis, which contributes to a poor prognosis in breast cancer. However, anti-angiogenic therapy with VEGF-specific monotherapy has been unsuccessful in treating breast cancer, and the molecular mechanisms associated with chemoresistance remain unclear. Here, we investigated whether CCL18, a chemokine produced by TAMs, can stimulate angiogenesis in breast cancer, as well as the underlying mechanisms. Double immunohistochemical staining for CCL18 and CD34/CD31/vWF was performed in 80 breast cancer samples to study the correlation between CCL18+ TAMs and microvascular density (MVD). Cocultures of TAMs with human umbilical vein endothelial cells (HUVECs) were used to model the inflammatory microenvironment, and CCL18-induced angiogenesis was evaluated both in vitro and in vivo. We demonstrated that CCL18+ TAM infiltration positively associated with MVD in breast cancer samples, which was correlated with tumor metastasis and poor prognosis. We confirmed, both in vitro and in vivo, that CCL18 and VEGF synergistically promoted endothelial cell migration and angiogenesis. Conversely, blocking CCL18 or VEGF with neutralizing antibodies synergistically inhibited the promigratory effects of TAMs. Silencing PITPNM3, a putative CCL18 receptor, on the surface of HUVECs abrogated CCL18-mediated promigration and the enhancement of HUVEC tube formation, independently of VEGFR signaling. Moreover, CCL18 exposure induced the endothelial-mesenchymal transformation and activated ERK and Akt/GSK-3ß/Snail signaling in HUVECs, thereby contributing to its pro-angiogenic effects. In conclusion, our findings suggest that CCL18 released from TAMs promotes angiogenesis and tumor progression in breast cancer; thus, CCL18 may serve as a novel target for anti-angiogenic therapies.

ScFv-decorated PEG-PLA-based nanoparticles for enhanced siRNA delivery to Her2⁺ breast cancer.

Patients with Her2-overexpressing (Her2(+)) breast cancers generally have a poorer prognosis due to the high aggressiveness and chemoresistance of the disease. Small interfering RNA (siRNA) targeting the gene encoding polo-like kinase 1 (Plk1; siPlk1) has emerged as an efficient therapeutic agent for Her2(+) breast cancers. Poly(ethylene glycol)-block-poly(D,L-lactide) (PEG-PLA)-based nanoparticles for siRNA delivery were previously developed and optimized. In this study, for targeted delivery of siPlk1 to Her2(+) breast cancer, anti-Her2 single-chain variable fragment antibody (ScFv(Her2))-decorated PEG-PLA-based nanoparticles with si Plk1 encapsulation (ScFv(Her2)-NP(si) Plk1) are developed. With the rationally designed conjugation site, ScFv(Her2)-NP(siRNA) can specifically bind to the Her2 antigen overexpressed on the surface of Her2(+) breast cancer cells. Therefore, ScFv(Her2)-NP(si) Plk1 exhibits improved cellular uptake, promoted Plk1 silencing efficiency, and induced enhanced tumor cell apoptosis in Her2(+) breast cancer cells, when compared with nontargeted NP(si) Plk1. More importantly, ScFv(Her2)-NP(siRNA) markedly enhances the accumulation of siRNA in Her2(+) breast tumor tissue, and remarkably improves the efficacy of tumor suppression. Dose-dependent anti-tumor efficacy further demonstrates that ScFvHer2 -decorated PEG-PLA-based nanoparticles with siPlk1 encapsulation can significantly enhance the inhibition of Her2(+) breast tumor growth and reduce the dose of injected siRNA. These results suggest that ScFvHer2 -decorated PEG-PLA-based nanoparticles show great potential for targeted RNA interference therapy of Her2(+) breast tumor.

Enhanced drug delivery to hepatocellular carcinoma with a galactosylated core-shell polyphosphoester nanogel.

Effective systemic therapy is often necessary to treat hepatocellular carcinoma (HCC). We synthesized a Gal-PPE nanogel consisting of a cross-linked polyphosphate core and galactosylated poly(ethylene glycol) arms for enhanced doxorubicin delivery to diethylnitrosamine-induced HCC in rats. The Gal-PPE nanogel exhibited high affinity to HepG2 cells in vitro, mediated by the asialoglycoprotein receptor. In vivo studies revealed that the Gal-PPE nanogel was taken up more efficiently by hepatocytes, in contrast to m-PPE nanogel. Consequently, doxorubicin delivery with Gal-PPE significantly inhibited the progress of HCC, reducing neoplastic liver nodules and prolonging the survival time of HCC rats more significantly. These results demonstrate the potential of Gal-PPE as a nanocarrier for improved HCC chemotherapy.

Anti-Her2 single-chain antibody mediated DNMTs-siRNA delivery for targeted breast cancer therapy.

The targeted delivery of small interfering RNA (siRNA) to specific tumor tissues and tumor cells remains as one of the key challenges in the development of RNA interference as a therapeutic application. To target breast cancer, we developed a therapeutic delivery system using a fusion protein of an anti-Her2 single-chain antibody fragment with a positively charged protamine, namely F5-P, as the carrier to specifically deliver siRNA-targeting DNA methyltransferases 1 and/or 3b genes (siDNMTs) into Her2-expressing breast tumor cells. The carrier F5-P, expressed by the Escherichia coli system, was able to bind siRNA molecules and specifically deliver the siRNA to Her2-expressing BT474 breast cancer cells but not Her2-nonexpressing MDA-MB-231 breast cancer cells, while delivery of siDNMTs to BT474 cells successfully silenced the expression of targeted DNA methyltransferases (DNMTs) and facilitated the de-methylation of the RASSF1A tumor suppressor gene promoter, leading to the suppression of tumor cell proliferation. Moreover, as demonstrated in the BT474 xenograft murine model, F5-P successfully delivered siRNA into a Her2-expressing breast tumor, and tumor growth inhibition was mediated by an intravenous injection of F5-P/siDNMTs complex by down-regulating the expression of DNMTs and restoring tumor suppressor gene expression. These data suggest that the delivery of siDNMTs by F5-P could be used to treat Her2-expressing breast cancer.

Single-step assembly of cationic lipid-polymer hybrid nanoparticles for systemic delivery of siRNA.

The clinical success of therapeutics of small interfering RNA (siRNA) is still hindered by its delivery systems. Cationic polymer or lipid-based vehicles as the major delivery systems of siRNA cannot sufficiently satisfy siRNA therapeutic applications. It is hypothesized that cationic lipid-polymer hybrid nanoparticles may take advantage of both polymeric and lipid-based nanoparticles for siRNA delivery, while diminishing the shortcomings of both. In this study, cationic lipid-polymer hybrid nanoparticles were prepared by a single-step nanoprecipitation of a cationic lipid (N,N-bis(2-hydroxyethyl)-N-methyl-N-(2-cholesteryloxycarbonyl aminoethyl) ammonium bromide, BHEM-Chol) and amphiphilic polymers for systemic delivery of siRNA. The formed hybrid nanoparticles comprised a hydrophobic polylactide core, a hydrophilic poly(ethylene glycol) shell, and a cationic lipid monolayer at the interface of the core and the shell. Such hybrid nanoparticles exhibited excellent stability in serum and showed significantly improved biocompatibility compared to that of pure BHEM-Chol particles. The hybrid nanoparticles were capable of delivering siRNA into BT474 cells and facilitated the escape of loaded siRNA from the endosome into the cytoplasm. The hybrid nanoparticles carrying polo-like kinase 1 (Plk1)-specific siRNA (siPlk1) remarkably and specifically downregulated expression of the oncogene Plk1 and induced cancer cell apoptosis both in vitro and in vivo and significantly suppressed tumor growth following systemic administration. We demonstrate that this system is stable, nontoxic, highly efficient, and easy to scale up, bringing the clinical application of siRNA therapy one important step closer to reality.

Targeted delivery of PLK1-siRNA by ScFv suppresses Her2+ breast cancer growth and metastasis.

A major obstacle to developing small interfering RNAs (siRNAs) as cancer drugs is their intracellular delivery to disseminated cancer cells. Fusion proteins of single-chain fragmented antibodies (ScFvs) and positively charged peptides deliver siRNAs into specific target cells. However, the therapeutic potential of ScFv-mediated siRNA delivery has not been evaluated in cancer. Here, we tested whether Polo-like kinase 1 (PLK1) siRNAs complexed with a Her2-ScFv-protamine peptide fusion protein (F5-P) could suppress Her2(+) breast cancer cell lines and primary human cancers in orthotopic breast cancer models. PLK1-siRNAs transferred by F5-P inhibited target gene expression, reduced proliferation, and induced apoptosis of Her2(+) breast cancer cell lines and primary human cancer cells in vitro without triggering an interferon response. Intravenously injected F5-P/PLK1-siRNA complexes concentrated in orthotopic Her2(+) breast cancer xenografts and persisted for at least 72 hours, leading to suppressed PLK1 gene expression and tumor cell apoptosis. The intravenously injected siRNA complexes retarded Her2(+) breast tumor growth, reduced metastasis, and prolonged survival without evident toxicity. F5-P-mediated delivery of a cocktail of PLK1, CCND1, and AKT siRNAs was more effective than an equivalent dose of PLK1-siRNAs alone. These data suggest that F5-P could be used to deliver siRNAs to treat Her2(+) breast cancer.

A biodegradable amphiphilic and cationic triblock copolymer for the delivery of siRNA targeting the acid ceramidase gene for cancer therapy.

One of the key challenges in the development of RNA interference-based cancer therapy is the lack of an efficient delivery system for synthetic small interfering RNAs (siRNAs) that would enable efficient uptake by tumor cells and allow for significant knockdown of a target transcript in vivo. Here, we describe a micelleplex system based on an amphiphilic and cationic triblock copolymer, which can systemically deliver siRNA targeting the acid ceramidase (AC) gene for cancer therapy. This triblock copolymer, consisting of monomethoxy poly(ethylene glycol), poly(ε-caprolactone) and poly(2-aminoethyl ethylene phosphate), self-assembles into micellar nanoparticles (MNPs) in aqueous solution with an average diameter of 60 nm and a zeta potential of approximately 48 mV. The resulting micelleplex, formed by the interaction of MNPs and siRNA, was effectively internalized by BT474 breast cancer cells and siRNA was subsequently released, resulting in significant gene knockdown. This effect was demonstrated by significant down-regulation of luciferase expression in BT474-luciferase cells which stably express luciferase, and suppression of AC expression in BT474 cells at both the transcriptional and protein level, following delivery of specific siRNAs by the micelleplex. Furthermore, a micelleplex carrying siRNA targeting the AC (micelleplex(siAC)) gene was found to induce remarkable apoptosis and reduce the proliferation of cancer cells. Systemic delivery of micelleplex(siAC) significantly inhibited tumor growth in a BT474 xenograft murine model, with depressed expression of AC and no positive activation of the innate immune response, suggesting therapeutic promise for micelleplex siRNA delivery in cancer therapy.

Simultaneous delivery of siRNA and paclitaxel via a "two-in-one" micelleplex promotes synergistic tumor suppression.

Combination of two or more therapeutic strategies with different mechanisms can cooperatively prohibit cancer development. Combination of chemotherapy and small interfering RNA (siRNA)-based therapy represents an example of this approach. Hypothesizing that the chemotherapeutic drug and the siRNA should be simultaneously delivered to the same tumoral cell to exert their synergistic effect, the development of delivery systems that can efficiently encapsulate two drugs and successfully deliver payloads to targeted sites via systemic administration has proven to be challenging. Here, we demonstrate an innovative "two-in-one" micelleplex approach based on micellar nanoparticles of a biodegradable triblock copolymer poly(ethylene glycol)-b-poly(ε-caprolactone)-b-poly(2-aminoethyl ethylene phosphate) to systemically deliver the siRNA and chemotherapeutic drug. We show clear evidence that the micelleplex is capable of delivering siRNA and paclitaxel simultaneously to the same tumoral cells both in vitro and in vivo. We further demonstrate that systemic administration of the micelleplex carrying polo-like kinase 1 (Plk1) specific siRNA and paclitaxel can induce a synergistic tumor suppression effect in the MDA-MB-435s xenograft murine model, requiring a thousand-fold less paclitaxel than needed for paclitaxel monotherapy delivered by the micelleplex and without activation of the innate immune response or generation of carrier-associated toxicity.

[Experience of endovenous radiofrequency combined with TriVex in treatment of chronic venous insufficiency in lower extremity].

OBJECTIVE: To evaluate therapeutic results of endovenous radiofrequency in combination with TriVex in treatment of venous insufficiency in lower extremities. METHODS: One hundred and fifty patients with chronic venous insufficiency (150 limbs) were randomly assigned to Group A (75 limbs) and Group B (75 limbs). Patients in Group A were treated with long saphenous veins radiofrequency ablation procedures in combination with TriVex. Patients in Group B were treated with long saphenous veins traditional stripping operation in combination with TriVex. The postoperative pain, average hospital stay and short-term results in hospital were compared between the two groups. Self-assessment of the operation 4 weeks after, changes of CEAP classification, venous clinical severity score (VCSS) and chronic venous insufficiency questionnaire (CIVIQ) score were compared between the two groups. RESULTS: The operation time in Group A was (67 +/- 11) min, compared with (59 +/- 9) min in Group B (P > 0.05). Postoperative pain and average hospital stay in Group A were significantly lower than those in Group B (P < 0.05). The scores of self-assessment of the operation in Group A was higher than that in Group B 4 weeks after operation (P < 0.05). The change of CEAP classification, VCSS and quality of life were significant after operation in both groups. The VCSS of Group A decreased by 4.6 +/- 2.5 compared with 4.3 +/- 2.7 in Group B (P > 0.05). CONCLUSIONS: Endovenous radiofrequency combined with TriVex for treatment of venous insufficiency in lower extremity is available, effective and with less trauma and faster recovery. CEAP classification, VCSS and CIVIQ are useful tools for assessing outcomes after radiofrequency in these patients.