Preparation of stimuli-responsive nanogels based on poly(<i>N,N</i>-diethylaminoethyl methacrylate) by a simple “surfactant-free” methodology

<p>The synthesis of poly(N,N-diethylaminoethyl methacrylate)-core-polyethyleneglycol-shell (PDEAEM-core-PEG-shell) nanogels was achieved by using a “surfactant-free” emulsion polymerization with the aid of commercially available polyethyleneglycol methyl ether methacrylates (PEGMAs) as polymerizable stabilizers. By adjusting the synthetic parameters like the choice of initiator, cross-linker, PEGMA:DEAEM ratio, and molecular weight of PEGMA, a series of core-shell nanogels varying in size (50–350 nm), PDEAEM content, and pH/temperature responsive behavior were obtained in reactions taking only 60 min. The nanogels were used as nanoreactors for the preparation of gold nanoparticles. The PEGylated nanogels have a great potential to be used for diagnosis and therapy.</p

Preparation of pH-sensitive nanogels bioconjugated with shark antibodies (VNAR) for targeted drug delivery with potential applications in colon cancer therapies.

Cancer is the second leading cause of death worldwide. To combat this disease, novel and specialized therapeutic systems are urgently needed. This is the first study to explore a system that combines shark variable domain (Fv) of new antigen receptor (VNAR) antibodies (hereinafter VNARs), PEGylated nanogels (pH-sensitive poly(N,N-diethylaminoethyl methacrylate, PDEAEM), and the anticancer drug 5-fluorouracil (5-FU) to explore its potential applications in colon cancer therapies. Nanogels were functionalized in a scalable reaction with an N-hydroxysuccinimide (NHS)-terminated polyethylene glycol derivative and bioconjugated with shark antibodies. Dynamic light scattering measurements indicated the presence of monodispersed nanogels (74 to 236 nm). All systems maintained the pH-sensitive capacity to increase in size as pH decreased. This has direct implications for the release kinetics of 5-FU, which was released faster at pH 5 than at pH 7.4. After bioconjugation, the ELISA results indicated VNAR presence and carcinoembryonic antigen (CEA) recognition. In vitro evaluations of HCT-116 colon cancer cells indicated that functionalized empty nanogels are not cytotoxic and when loaded with 5-FU, the cytotoxic effect of the drug is preserved. A 15% reduction in cell viability was observed after two hours of contact with bioconjugated nanogels when compared to what was observed with non-bioconjugated nanogels. The prepared nanogel system shows potential as an effective and site-specific nanocarrier with promising applications in in vivo studies of colon cancer therapies

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Cancer is the second leading cause of death worldwide. To combat this disease, novel and specialized therapeutic systems are urgently needed. This is the first study to explore a system that combines shark variable domain (Fv) of new antigen receptor (VNAR) antibodies (hereinafter VNARs), PEGylated nanogels (pH-sensitive poly(N,N-diethylaminoethyl methacrylate, PDEAEM), and the anticancer drug 5-fluorouracil (5-FU) to explore its potential applications in colon cancer therapies. Nanogels were functionalized in a scalable reaction with an N-hydroxysuccinimide (NHS)-terminated polyethylene glycol derivative and bioconjugated with shark antibodies. Dynamic light scattering measurements indicated the presence of monodispersed nanogels (74 to 236 nm). All systems maintained the pH-sensitive capacity to increase in size as pH decreased. This has direct implications for the release kinetics of 5-FU, which was released faster at pH 5 than at pH 7.4. After bioconjugation, the ELISA results indicated VNAR presence and carcinoembryonic antigen (CEA) recognition. In vitro evaluations of HCT-116 colon cancer cells indicated that functionalized empty nanogels are not cytotoxic and when loaded with 5-FU, the cytotoxic effect of the drug is preserved. A 15% reduction in cell viability was observed after two hours of contact with bioconjugated nanogels when compared to what was observed with non-bioconjugated nanogels. The prepared nanogel system shows potential as an effective and site-specific nanocarrier with promising applications in in vivo studies of colon cancer therapies.</div

Sizes of the PDEAEM:PEGMA:PEGA-NHS nanogels determined by dynamic light scattering (DLS).

a) Synthesis reproducibility and b) pH-responsive behavior of the nanogels.</p

<i>In vitro</i> analysis of VNAR-bioconjugated nanogels (N3).

A) ELISA of VNAR expression. On the horizontal axis: B (blank, Svelty milk 8%-PBS), N3 C- (nanogel without VNAR), C+ (CV043, 175 μg/mL), 1h (sample taken at hour 1 of the coupling reaction), 4h (sample taken at hour 4 of the coupling reaction), 8h (final sample taken at hour 8 of the coupling reaction), and PF (final product after dialysis [N3+VNAR]). B) ELISA assay of CEA recognition. On the horizontal axis: VNAR (CV043, 175 μg/mL), B (blank, Svelty milk 8%-PBS), c) cell viability by aqueous one solution cell proliferation assay (MTS) of empty selected nanogels at 24h, PBS (C-), DMSO 5% (C+), d) cell viability by MTS of 5-FU and nanogels containing 5-FU. On the horizontal axis: B (PBS 7.4), N3 (empty nanogels, 5-FU [20 μg/mL]), N3-VNAR-5FU (20 μg 5FU/mL), and N3-5-FU (20 μg 5FU/mL). ANOVA test, *p < 0.01, **p < 0.001, and ***p < 0.0001 versus N3 and PBS (C-). PDEAEM: poly(N,N-diethylaminoethyl methacrylate), PEGMA: poly(ethylene glycol) methyl ether methacrylate, PEGA-NHS: Acrylate-PEG3500-NHS, and CEA: carcinoembryonic antigen.</p

Model structures for bioconjugation of PDEAEM:PEGMA:PEGA-NHS nanogels.

A) UV-Vis of folic acid-bioconjugated nanogels. B) Fluorescence emission spectra of GFP-bioconjugated nanogels.</p

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Cancer is the second leading cause of death worldwide. To combat this disease, novel and specialized therapeutic systems are urgently needed. This is the first study to explore a system that combines shark variable domain (Fv) of new antigen receptor (VNAR) antibodies (hereinafter VNARs), PEGylated nanogels (pH-sensitive poly(N,N-diethylaminoethyl methacrylate, PDEAEM), and the anticancer drug 5-fluorouracil (5-FU) to explore its potential applications in colon cancer therapies. Nanogels were functionalized in a scalable reaction with an N-hydroxysuccinimide (NHS)-terminated polyethylene glycol derivative and bioconjugated with shark antibodies. Dynamic light scattering measurements indicated the presence of monodispersed nanogels (74 to 236 nm). All systems maintained the pH-sensitive capacity to increase in size as pH decreased. This has direct implications for the release kinetics of 5-FU, which was released faster at pH 5 than at pH 7.4. After bioconjugation, the ELISA results indicated VNAR presence and carcinoembryonic antigen (CEA) recognition. In vitro evaluations of HCT-116 colon cancer cells indicated that functionalized empty nanogels are not cytotoxic and when loaded with 5-FU, the cytotoxic effect of the drug is preserved. A 15% reduction in cell viability was observed after two hours of contact with bioconjugated nanogels when compared to what was observed with non-bioconjugated nanogels. The prepared nanogel system shows potential as an effective and site-specific nanocarrier with promising applications in in vivo studies of colon cancer therapies.</div

General characteristics of PDEAEM-based nanogels.

Cancer is the second leading cause of death worldwide. To combat this disease, novel and specialized therapeutic systems are urgently needed. This is the first study to explore a system that combines shark variable domain (Fv) of new antigen receptor (VNAR) antibodies (hereinafter VNARs), PEGylated nanogels (pH-sensitive poly(N,N-diethylaminoethyl methacrylate, PDEAEM), and the anticancer drug 5-fluorouracil (5-FU) to explore its potential applications in colon cancer therapies. Nanogels were functionalized in a scalable reaction with an N-hydroxysuccinimide (NHS)-terminated polyethylene glycol derivative and bioconjugated with shark antibodies. Dynamic light scattering measurements indicated the presence of monodispersed nanogels (74 to 236 nm). All systems maintained the pH-sensitive capacity to increase in size as pH decreased. This has direct implications for the release kinetics of 5-FU, which was released faster at pH 5 than at pH 7.4. After bioconjugation, the ELISA results indicated VNAR presence and carcinoembryonic antigen (CEA) recognition. In vitro evaluations of HCT-116 colon cancer cells indicated that functionalized empty nanogels are not cytotoxic and when loaded with 5-FU, the cytotoxic effect of the drug is preserved. A 15% reduction in cell viability was observed after two hours of contact with bioconjugated nanogels when compared to what was observed with non-bioconjugated nanogels. The prepared nanogel system shows potential as an effective and site-specific nanocarrier with promising applications in in vivo studies of colon cancer therapies.</div