Catch and release: photocleavable cationic diblock copolymers as a potential platform for nucleic acid delivery

Green, Matthew D.; Foster, Abbygail A.; Greco, Chad T.; Roy, Raghunath; Lehr, Rachel; Epps, Thomas H.; Sullivan, Millicent O.

doi:10.1039/c4py00638k

Cited by 26 publications

(42 citation statements)

References 57 publications

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“…This behavior was consistent with the typical cleavage behavior of o-NB-containing polymers, which disassemble in response to irradiation with UV light to form a carboxylic acid and nitrosobenzaldehyde. [29] The spectral shifts also were similar to those previously noted for mPEG-b-P(APNBMA) n in pDNA polyplexes, [19] and the decrease in absorbance as a function of irradiation time followed an exponential decay for both the polymer and siRNA polyplex preparations. In particular, both the free polymer and polyplex spectra suggested complete cleavage of the o-NB group, based on the change in absorbance at 316 nm.…”

Section: Uv-induced Unpackaging Of Sirna/mpeg-b-p(apnbma)236 Polyplexessupporting

confidence: 69%

“…[34] However, as previously demonstrated, mPEG-b-P(APNBMA) n copolymers exhibited significantly reduced cytotoxicity as compared to PEI, and pDNA/mPEG-b-P(APNBMA) 7.9 polyplexes exhibited minimal cytotoxicity both with and without UV irradiation. [19] Thus, we verified the expected lack of cytotoxicity in cell populations transfected by siRNA polyplexes under similar conditions using the Alamar Blue (AB) cell survival assay. AB-based quantification revealed greater than 90% cell survival in all samples, and no additive/synergistic effects were found in cells that were treated with UV light in combination with siRNA/mPEG-b-P(APNBMA) 23.6 polyplexes vs. UV light alone ( Figure 8).…”

Section: Nih/3t3 Cell Viabilitymentioning

confidence: 99%

“…Free mPEG-b-P(APNBMA) 23.6 block copolymer as well as siRNA/mPEG-b-P(APNBMA) 23.6 polyplexes were irradiated with 365 nm light, and UV/Vis spectroscopy was used to monitor changes in the characteristic absorbance of the o-NB ester at 316 nm to determine the extent of photocleavage ( Figure 5). [19] The free polymer absorbance spectra displayed dramatic decreases in absorbance at 316 nm for UV exposure times up to 10 min, and smaller changes at longer UV exposure times ( Figure 5A). Irradiation of siRNA/mPEG-b-P(APNBMA) 23.6 polyplexes yielded similar results under the same conditions, with a large decrease in absorbance after 10 min of irradiation ( Figure 5B).…”

Section: Uv-induced Unpackaging Of Sirna/mpeg-b-p(apnbma)236 Polyplexesmentioning

confidence: 99%

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Light‐Mediated Activation of siRNA Release in Diblock Copolymer Assemblies for Controlled Gene Silencing

Foster¹,

Greco²,

Green³

et al. 2015

Adv Healthcare Materials

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Controllable release is particularly important for the delivery of small interfering RNA (siRNA), as siRNAs have a high susceptibility to enzymatic degradation if release is premature, yet lack silencing activity if they remain inaccessible within the cytoplasm. To overcome these hurdles, novel and tailorable mPEG-b-poly(5-(3-(amino)propoxy)-2-nitrobenzyl methacrylate) (mPEG-b-P(APNBMA) n ) diblock copolymers containing light-sensitive o-nitrobenzyl moieties and pendant amines are employed to provide both efficient siRNA binding, via electrostatic and hydrophobic interactions, as well as triggered charge reversal and nucleic acid release. In particular, siRNA/ mPEG-b-P(APNBMA) 23.6 polyplexes show minimal aggregation in physiological salt and serum, and enhanced resistance to polyanion-induced unpackaging compared to polyethylenimine preparations. Cellular delivery of siRNA/mPEG-b-P(APNBMA) 23.6 polyplexes reveal greater than 80% cellular transfection, as well as rapid and widespread cytoplasmic distribution. Additionally, UV irradiation indicates ~70% reduction in targeted gene expression following siRNA/mPEG-b-P(APNBMA) 23.6 polyplex treatment, as compared to 0% reduction in polyplex treated cells without UV irradiation, and only ~30% reduction for Lipofectamine treated cells. The results herein highlight the potential of these light-sensitive copolymers with a well-defined on/off switch for applications including cellular patterning for guided cell growth and extension, and cellular microarrays for exploring protein and drug interactions that require enhanced spatiotemporal control of gene activation.

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Section: Uv-induced Unpackaging Of Sirna/mpeg-b-p(apnbma)236 Polyplexessupporting

confidence: 69%

Section: Nih/3t3 Cell Viabilitymentioning

confidence: 99%

Section: Uv-induced Unpackaging Of Sirna/mpeg-b-p(apnbma)236 Polyplexesmentioning

confidence: 99%

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Light‐Mediated Activation of siRNA Release in Diblock Copolymer Assemblies for Controlled Gene Silencing

Foster¹,

Greco²,

Green³

et al. 2015

Adv Healthcare Materials

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“…The mPEG- b -P(APNBMA) n polymers (M n = 7,900 g mol −1 , n = 7.9; M n = 13,100 g mol −1 , n = 23.6) were synthesized via atom-transfer radical polymerization as described previously. [46] Dulbecco's Modified Eagle Medium (DMEM) and Dulbecco's phosphate-buffered saline (DPBS, 150 mM NaCl, pH of 7.4) were obtained from Corning Life Sciences – Mediatech Inc. (Manassas, VA). Opti-MEM medium, SuperSignal West Dura Chemiluminescent Substrate, Phalloidin-660, Hoescht 33258, TRIzol Reagent, and AlamarBlue were purchased from Life Technologies (Carlsbad, CA).…”

Section: Methodsmentioning

confidence: 99%

Attenuation of Maladaptive Responses in Aortic Adventitial Fibroblasts through Stimuli‐Triggered siRNA Release from Lipid–Polymer Nanocomplexes

et al. 2017

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Lipid-siRNA assemblies are modified with photo-responsive polymers to enable spatiotemporally-controlled silencing of interleukin 1 beta (IL1β) and cadherin 11 (CDH11), two genes that are essential drivers of maladaptive responses in human aortic adventitial fibroblasts (AoAFs). These hybrid nanocomplexes address the critical challenge of locally mitigating fibrotic actions that lead to the high rates of vascular graft failures. In particular, the lipid-polymer formulations provide potent silencing of IL1β and CDH11 that is precisely modulated by a photo-release stimulus. Moreover, a dynamic modeling framework is used to design a multi-dose siRNA regimen that sustains knockdown of both genes over clinically-relevant timescales. Multi-dose suppression illuminates a cooperative role for IL1β and CDH11 in pathogenic adventitial remodeling and is directly linked to desirable functional outcomes. Specifically, myofibroblast differentiation and cellular proliferation, two of the primary hallmarks of fibrosis, are significantly attenuated by IL1β silencing. Meanwhile, the effects of CDH11 siRNA treatment on differentiation become more pronounced at higher cell densities characteristic of constrictive adventitial remodeling in vivo. Thus, this work offers a unique formulation design for photo-responsive gene suppression in human primary cells and establishes a new dosing method to satisfy the critical need for local attenuation of fibrotic responses in the adventitium surrounding vascular grafts.

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“…The polymers have tunable molecular weights, low dispersities, and photocleavable moieties that permit light-induced charge reversal to initiate nucleic acid release. [43] A nonfouling PEG block was incorporated to provide stability in physiological environments and resistance to opsonization. These BCPs have proven biocompatibility and protect siRNA in salt, serum, and nuclease solutions, while simultaneously stimulating siRNA release and gene-specific knockdown upon application of a cytocompatible photo-stimulus.…”

Section: Introductionmentioning

confidence: 99%

Efficient tuning of siRNA dose response by combining mixed polymer nanocarriers with simple kinetic modeling

et al. 2017

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Two of the most prominent challenges that limit the clinical success of siRNA therapies are a lack of control over cargo release from the delivery vehicle and an incomplete understanding of the link between gene silencing dynamics and siRNA dosing. Herein, we address these challenges through the formulation of siRNA polyplexes containing light-responsive polymer mixtures, whose varied compositions and triggered release behavior provide enhanced gene silencing and controlled dose responses that can be predicted by simple kinetic models. Through the straightforward mixing of two block copolymers, the level of gene knockdown was easily optimized to achieve the maximum level of GAPDH protein silencing in NIH/3T3 cells (70%) using a single siRNA dose. The kinetic model was used to describe the dynamic changes in mRNA and protein concentrations in response to siRNA treatment. These predictions enabled the application of a second dose of siRNA to maximally suppress gene expression over multiple days, leading to a further 50% reduction in protein levels relative to those measured following a single dose. Furthermore, polyplexes remained dormant in cells until exposed to the photo-stimulus, demonstrating the complete control over siRNA activity as well as the stability of the nanocarriers. Thus, this work demonstrates that pairing advances in biomaterials design with simple kinetic modeling provides new insight into gene silencing dynamics and presents a powerful strategy to control gene expression through siRNA delivery.

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Catch and release: photocleavable cationic diblock copolymers as a potential platform for nucleic acid delivery

Cited by 26 publications

References 57 publications

Light‐Mediated Activation of siRNA Release in Diblock Copolymer Assemblies for Controlled Gene Silencing

Light‐Mediated Activation of siRNA Release in Diblock Copolymer Assemblies for Controlled Gene Silencing

Attenuation of Maladaptive Responses in Aortic Adventitial Fibroblasts through Stimuli‐Triggered siRNA Release from Lipid–Polymer Nanocomplexes

Efficient tuning of siRNA dose response by combining mixed polymer nanocarriers with simple kinetic modeling

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