Treatment of Melanoma by Nano-conjugate-Delivered Wee1 siRNA.

Small interfering RNA (siRNA)-based drugs have shown tremendous potential to date in cancer gene therapy. Despite the considerable efforts in siRNA design and manufacturing, unsatisfactory delivery systems persist as a limitation for the application of siRNA-based drugs. In this work, the cholesterol, cell-penetrating peptide conjugate cRGD (R8-cRGD), and polyethylene glycol (PEG) were introduced into low-molecular-weight polyethyleneimine (LMW PEI) to form cRGD-R9-cholesterol-PEI-PEG (RRCPP) nanoparticles with specific targeting and highly penetrating abilities. The enhanced siRNA uptake efficiency of the RRCPP delivery system benefited from R8-cRGD modification. Wee1 is an oncogenic nuclear kinase that can regulate the cell cycle as a crucial G2/M checkpoint. Overexpression of Wee1 in melanoma may lead to a poor prognosis. In the present study, RRCPP nanoparticles were designed for Wee1 siRNA delivery to form an RRCPP/siWee1 complex, which significantly silenced the expression of the WEE1 gene (>60% inhibition) and induced B16 tumor cell apoptosis by abrogating the G2M checkpoint and DNA damage in vitro. Furthermore, the RRCPP/siWee1 complex suppressed B16 tumor growth in a subcutaneous xenograft model (nearly 85% inhibition rate) and lung metastasis (nearly 66% inhibition rate) with ideal in vivo safety. Briefly, our results support the validity of RRCPP as a potential Wee1 siRNA carrier for melanoma gene therapy.

Local and systemic delivery of mRNA encoding survivin-T34A by lipoplex for efficient colon cancer gene therapy.

Background: In vitro transcribed (IVT) mRNA has been applied as an alternative therapeutic molecule to plasmid DNA in the field of cancer therapy and biomedical research studies. mRNA-based therapy has demonstrated several advantages over its DNA counterparts. However, its further therapeutic application is largely restricted by delivery method. Methods: In this work, a liposome-protamine lipoplex (CLPP) was prepared to deliver IVT mRNA encoding survivin-T34A gene, forming a novel core-shell structured nanoparticle formulation (CLPP/mSur-T34A). Results: The prepared CLPP/mSur-T34A particle had an average size of 186.1±3.1 nm, displaying high mRNA transfecting and expression efficiency on C26 tumor cells through lipid rafts-mediated endocytosis. CLPP/mSur-T34A mRNA formulation demonstrated obvious therapeutic effects on various models of C26 colon cancer both in vitro and in vivo. Particularly, local and systemic administration of CLPP/mSur-T34A particle exhibited superior antitumor effect regarding its DNA plasmid counterpart with high safety. Conclusion: Our results indicated the high delivery capacity of liposome-protamine lipoplex and further suggested CLPP/mSur-T34A mRNA formulation to be a potential candidate for colon cancer therapy.

Cationic micelle-based siRNA delivery for efficient colon cancer gene therapy.

Small interfering RNA (siRNA)-based gene therapy has provided an alternative strategy for cancer therapy. One of the key components within gene therapy process is the delivery system. As a novel non-viral gene vector, DMP, prepared by modifying mPEG-PCL micelle with cationic DOTAP lipid, has been prepared and successfully applied in plasmid DNA-based colon cancer gene therapy study. However, its potential in siRNA delivery is unknown. In this study, the preparation process of DMP was optimized and the anti-cancer efficacies of the DMP/siMcl1 and DMP/siBcl-xl complexes were studied on a mouse colon cancer model. Our results demonstrated that DMP cationic micelle-delivered siRNAs could effectively inhibit the growth of C26 colon cancer cells in vitro. Meanwhile, intratumoral administration of DMP/siMcl1 and DMP/siBcl-xl complexes obviously suppressed subcutaneous tumor model in vivo. These results suggest the DMP/siRNA complex to be a potential candidate for cancer gene therapy.

Local and Systemic Delivery of Interleukin-12 Gene by Cationic Micelles for Cancer Immunogene Therapy.

Immunogene therapy is an alternative strategy for cancer gene therapy. By stimulating the activities of immune cells in the tumor microenvironment, the genetic materials boost the immune response to attack cancer cells resulting in therapeutic effects. Interleukin-12 is an important regulator with great potential in modulating both innate and adaptive immunities. Here, a cancer immunogene therapy strategy was established and evaluated by delivering the IL-12 gene with a novel non-viral gene vector DMP. The DMP cationic micelles were prepared by modifying monomethoxy poly(ethylene glycol)-poly(ε-caprolactone) with the DOTAP lipid via self-assembly. The anti-cancer efficacy of the DMP/IL-12 complex was studied on multiple murine cancer models via both local and systemic administration. Our results demonstrated that the secretory expressed IL-12 cytokine effectively enhanced lymphocytes activities resulting in strong inhibition of cancer cell growth in vitro. Meanwhile, there were obvious tumor regressions achieved in tumor models of C26 colon carcinoma and LL/2 lung cancer in vivo. Multiple anti-cancer mechanisms including T cell infiltration, TNF-α secretion, apoptosis induction and angiogenesis inhibition are involved; no pathology changes were observed in healthy tissues. These results suggest that the DMP/IL-12 complex is a potential candidate for cancer immunogene therapy.

Delivery of interleukin-22 binding protein (IL-22BP) gene by cationic micelle for colon cancer gene therapy.

Gene therapy has provided an alternative strategy for cancer therapy. As an important cytokine, interleukin-22 (IL-22) is not only critical in reinforcing innate immune defenses and tissue regeneration, but also involved in the initial establishment of tumors. A soluble-secreted receptor of the cytokine IL-22, IL-22 binding protein (IL-22BP), binds IL-22 and prevents its binding to the functional transmembrane receptor IL-22R1 complex, inhibiting IL-22-based intracellular cancer proliferation signal. In this work, a novel IL-22BP-based cancer gene therapy strategy was reported for the first time. It was established by delivering IL-22BP gene with a newly developed non-viral gene vector DMP. The DMP cationic micelles were prepared by modifying monomethoxy poly(ethylene glycol)-poly(ε-caprolactone) with DOTAP lipid through self-assembling. The anti-cancer efficacy of the DMP/IL-22BP complex was studied on a colon cancer model by intraperitoneal administration. Our results demonstrated that the secretory expressed IL-22BP cytokine effectively inhibited cancer growth both in vitro and in vivo. Multiple anti-cancer mechanisms including IL-22 blocking, apoptosis inducing, lymphocyte infiltration and angiogenesis inhibition were indicated to be involved while no pathology changes were observed in healthy tissues. These results suggest the DMP/IL-22BP complex to be a potential candidate for cancer gene therapy.

Synthesis of Cyclic Ethers with Fluorinated Side Chains.

Di-tert-butyl peroxide initiated free radical addition of THF to various fluorinated alkenes (CF(2)=CH(2), CF(2)=CFH, CH(2)=CHCF(3), CF(2)=CFCF(3), CF(2)=CFC(5)F(11), CF(2)=CFOCF(2)CF(CF(3))OCF(2)CF(2)SO(2)F) gives either bidirectional addition products [CH(2)CH(2)CH(2)OCH(CF(2)CH(3)) (1), CH(2)CH(2)CH(2)OCH(CH(2)CHF(2)) (2), CH(2)CH(2)CH(2)OCH(CF(2)CH(2)F) (3), and CH(2)CH(2)CH(2)OCH(CFHCHF(2)) (4)] or unidirectional products [CH(2)CH(2)CH(2)OCH(CH(2)CH(2)CF(3)) (5), CH(2)CH(2)CH(2)OCH(CF(2)CHFCF(3)) (6), CH(2)CH(2)CH(2)OCH(CF(2)CHFC(5)F(11)) (7), and CH(2)CH(2)CH(2)OCH(CF(2)CHFOCF(2)CF(CF(3))OCF(2)CF(2)SO(2)F) (8)] depending on the structure of the alkene. Reaction of dioxane with CF(2)=CFOCF(2)CF(CF(3))OCF(2)CF(2)SO(2)F gives a single product, CH(2)OCH(2)CH(2)OCH(CF(2)CHFOCF(2)CF(CF(3))OCF(2)CF(2)SO(2)F) (9). In the case of hexafluoropropene or perfluoroallylbenzene, reaction with an excess of tetrahydrofuran gives only the monosubstituted products CH(2)CH(2)CH(2)OCH(CF(2)CHFCF(3)) (6) and CH(2)CH(2)CH(2)OCH(CF(2)CFHCF(2)C(6)F(5)) (11) respectively. When tetrahydrofuran is reacted with a 3:1 molar excess of the same two perfluoroalkenes, the disubstituted products CH(2)CH(2)CH(CF(2)CHFCF(3))OCH(CF(2)CHFCF(3)) (10) and CH(2)CH(2)CH(CF(2)CFHCF(2)C(6)F(5))OCH(CF(2)CFHCF(2)C(6)F(5)) (12) are formed respectively. When 18-crown-6 is reacted in the same way with fluoroalkenes in a 1:1 molar ratio, the monosubstituted products 18-crown-6-CH(2)CH(2)CF(3) (13), 18-crown-6-CF(2)CHFCF(3) (14), 18-crown-6-CF(2)CFHCF(2)C(6)F(5) (15), and 18-crown-6-CF(2)CHFOCF(2)CF(CF(3))OCF(2)CF(2)SO(2)F (16) are obtained. Polyfluorinated 18-crown-6 products containing three and two polyfluroalkyl/aryl groups are prepared when 18-crown-6 is reacted with a 3:1 molar excess of perfluoropropene and perfluoroallylbenzene to give 18-crown-6-(CF(2)CHFCF(3))(3) (17) and 18-crown-6-(CF(2)CFHCF(2)C(6)F(5))(2) (18), respectively. (Pentafluorophenoxy)trimethylsilane reacts with 11 in the presence of a catalytic amount of cesium fluoride to give compound 19, CH(2)CH(2)CH(2)OCH(CF(2)CFHCF(2)C(6)F(4)OC(6)F(5)). Two molecules of 11 are bridged by reaction with Me(3)SiOCH(2)CF(2)CF(2)CF(2)CH(2)OSiMe(3) to give CH(2)CH(2)CH(2)OCHCF(2)CFHCF(2)C(6)F(4)OCH(2)CF(2)CF(2)CF(2)CH(2)OC(6)F(4)CF(2)CHFCF(2)CHOCH(2)CH(2)CH(2) (20), while 12 forms the macroheterocycle OCHCH(2)CH(2)CHCF(2)CFHCF(2)C(6)F(4)OCH(2)CF(2)CF(2)CF(2)CH(2)OC(6)F(4)CF(2)CFHCF(2) (21) under similar reaction conditions. The lanthanum triflate complexes of 18-crown-6 (22) and 18-crown-6(CF(2)CFHCF(3)) (23) were prepared and the structures were obtained via single-crystal X-ray analysis. Although crystals suitable for single-crystal X-ray analysis could not be formed, lanthanum triflate complexes were formed with polyfluorinated ethers 15 and 16 to give the fluorinated complexes La(OSO(2)CF(3))(3)(18-crown-6-CF(2)CFHCF(2)C(6)F(5))(H(2)O) (24) and [La(OCH(2)CH(2))(5)OCH(2)CHCF(2)CFHOCF(2)C(CF(3))FOCF(2)CF(2)SO(2)F](3+)[CF(3)SO(3)(-)](3) (25) respectively. The acid salt La[N(SO(2)CF(3))(2)](3) (26) was also prepared and characterized, and reacted with dibenzo-18-crown-6 to give the complex dibenzo-18-crown-6-La[N(SO(2)CF(3))(2)](3) (27).