| 1. |
- Bermejo-Velasco, Daniel, et al.
(författare)
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Thiazolidine chemistry revisited : a fast, efficient and stable click-type reaction at physiological pH
- 2018
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Ingår i: Chemical Communications. - : Royal Society of Chemistry (RSC). - 1359-7345 .- 1364-548X. ; 54:88, s. 12507-12510
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Tidskriftsartikel (refereegranskat)abstract
- We describe the fast reaction kinetics between 1,2-aminothiols and aldehydes. Under physiological conditions such a click-type reaction afforded a thiazolidine product that remains stable and did not require any catalyst. This type of bioorthogonal reaction offers enormous potential for the coupling of biomolecules in an efficient and biocompatible manner.
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| 2. |
- Han, Yuanyuan, et al.
(författare)
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MicroRNA detection based on duplex-specific nuclease-assisted target recycling and gold nanoparticle/graphene oxide nanocomposite-mediated electrocatalytic amplification
- 2019
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Ingår i: Biosensors & bioelectronics. - : Elsevier BV. - 0956-5663 .- 1873-4235. ; 127, s. 188-193
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Tidskriftsartikel (refereegranskat)abstract
- DNA technology based bio-responsive nanomaterials have been widely studied as promising tools for biomedical applications. Gold nanoparticles (AuNPs) and graphene oxide (GO) sheets are representative zero- and two-dimensional nanomaterials that have long been combined with DNA technology for point-of-care diagnostics. Herein, a cascade amplification system based on duplex-specific nuclease (DSN)-assisted target recycling and electrocatalytic water-splitting is demonstrated for the detection of microRNA. Target microRNAs can form DNA: RNA heteroduplexes with DNA probes on the surface of AuNPs, which can be hydrolyzed by DSN. MicroRNAs are preserved during the reaction and released into the suspension for the digestion of multiple DNA probes. After the DSN-based reaction, AuNPs are collected and mixed with GO to form AuNP/GO nanocomposite on an electrode for the following electrocatalytic amplification. The utilization of AuNP/GO nanocomposite offers large surface area, exceptional affinity to water molecules, and facilitated mass diffusion for the water-splitting reaction. For let-7b detection, the proposed biosensor achieved a limit detection of 1.5 fM in 80 min with a linear detection range of approximately four orders of magnitude. Moreover, it has the capability of discriminating non-target microRNAs containing even single-nucleotide mismatches, thus holding considerable potential for clinical diagnostics.
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| 3. |
- Kadekar, Sandeep, et al.
(författare)
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Redox responsive Pluronic micelle mediated delivery of functional siRNA : a modular nano-assembly for targeted delivery
- 2021
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Ingår i: Biomaterials Science. - : Royal Society of Chemistry. - 2047-4830 .- 2047-4849. ; 9:11, s. 3939-3944
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Tidskriftsartikel (refereegranskat)abstract
- There is an unmet need to develop strategies that allow site-specific delivery of short interfering RNA (siRNA) without any associated toxicity. To address this challenge, we have developed a novel siRNA delivery platform using chemically modified pluronic F108 as an amphiphilic polymer with a releasable bioactive disulfide functionality. The micelles exhibited thermoresponsive properties and showed a hydrodynamic size of similar to 291 nm in DLS and similar to 200-250 nm in SEM at 37 degrees C. The grafting of free disulfide pyridyl groups enhanced the transfection efficiency and was successfully demonstrated in human colon carcinoma (HCT116; 88%) and glioma cell lines (U87; 90%), non-cancerous human dermal fibroblast (HDF; 90%) cells as well as in mouse embryonic stem (mES; 54%) cells. To demonstrate the versatility of our modular nanocarrier design, we conjugated the MDGI receptor targeting COOP peptide on the particle surface that allowed the targeted delivery of the cargo molecules to human patent-derived primary BT-13 gliospheres. Transfection experiments with this design resulted in similar to 65% silencing of STAT3 mRNA in BT-13 gliospheres, while only similar to 20% of gene silencing was observed in the absence of the peptide. We believe that our delivery method solves current problems related to the targeted delivery of RNAi drugs for potential in vivo applications.
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| 4. |
- Kadekar, Sandeep, et al.
(författare)
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Synthetic design of asymmetric miRNA with engineered 3′-overhang to improve strand selection
- 2019
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Ingår i: Molecular Therapy Nucleic Acids. - : Elsevier. - 2162-2531. ; 16, s. 597-604
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Tidskriftsartikel (refereegranskat)abstract
- We have developed a novel miRNA design that significantly improves strand selection within the RISC complex by engineering the 3′-end by adding extra nucleotides. Addition of seven nucleotides at the 3′-ends of the miR or miR* strand resulted in a thermodynamic asymmetry at either of the two-ends, which resulted in selective RISC recruitment as demonstrated by the stem-loop quantitative PCR experiment. Such selective recruitment was also corroborated at the protein level by Western blot analysis. In order to investigate the functional effect due to selective recruitment, we performed apoptosis and metastasis studies using human colon carcinoma cells (HCT116) and human osteosarcoma cells (MG63). These experiments indicated that the recruitment of miR strand is responsible for inducing apoptosis as well as to inhibit invasiveness of cancer cells. Recruitment of miR* strand, on the other hand, showed opposite effect. To the best of our knowledge, our strand engineering strategy is the first report of improved strand selection of desired miRNA strand by RISC without using any chemical modifications or mismatches. We believe such structural modifications of miR34a could mitigate some of the off-target effects of miRNA therapy and would also allow a better understanding of sequence-specific gene regulation. Such a design could also be adapted to other miRNA to enhance their therapeutic potential.
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| 5. |
- Mirajkar, Abhishek L., et al.
(författare)
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Synthetic green fluorescent protein (GFP) chromophore analog for rapid, selective and sensitive detection of cyanide in water and in living cells
- 2018
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Ingår i: Sensors and actuators. B, Chemical. - : ELSEVIER SCIENCE SA. - 0925-4005 .- 1873-3077. ; 265, s. 257-263
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Tidskriftsartikel (refereegranskat)abstract
- Here, we report Green Fluorescent Protein (GFP) chromophore analog as a turn-on fluorescent chemodosimeter (THBI) for selective detection of cyanide in water, on solid state and in living cells. The detection limit was found to be 0.17 mu M (4.5 ppb). The time dependent study revealed that there is a rapid enhancement in fluorescence intensity (in less than 5s) and was constant over the period of 1 h. Cell imaging data exhibited that THBI was successfully crossed cell membrane and visualized fluorescence response in live HCT cells.
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| 6. |
- Mittapelli, Lavanya L., et al.
(författare)
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A turn-on fluorescent GFP chromophore analog for highly selective and efficient detection of H2S in aqueous and in living cells
- 2019
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Ingår i: Sensors and actuators. B, Chemical. - : ELSEVIER SCIENCE SA. - 0925-4005 .- 1873-3077. ; 298
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Tidskriftsartikel (refereegranskat)abstract
- Hydrogen sulphide is a gaseous neurotransmitter responsible for neuronal function and controls vast range of physiological functions. Herein, we report the synthesis and evaluation of novel Green Fluorescent Protein (GFP) chromophore analog, acryloyl-4-(p-hydroxybenzylidene)-5-imidazolidinone (AHBI) for turn-on fluorescent detection of H2S over wide range of anions and various biologically important competitive thiols. AHBI probe exhibited high selectivity and sensitivity, high fluorescence stability, large stokes shift and lower detection limit (15.85 ppb) for H2S in complete water medium. Cell imaging studies in human colon cancer cells (HCT116) and normal human dermal fibroblasts (HDF) confirmed the compatibility and versatility of AHBI probe at micromolar level. Overall, we believe the AHBI, as an optical probe will be useful to investigate the role of H2S in various physiological processes, regulation of cancer cell growth, and in pathogenic events.
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| 7. |
- Nawale, Ganesh N., et al.
(författare)
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4 '-Guanidinium-modified siRNA : a molecular tool to control RNAi activity through RISC priming and selective antisense strand loading
- 2019
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Ingår i: Chemical Communications. - : ROYAL SOC CHEMISTRY. - 1359-7345 .- 1364-548X. ; 55:62, s. 9112-9115
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Tidskriftsartikel (refereegranskat)abstract
- We designed novel 4 '-C-guanidinocarbohydrazidomethyl-5-methyl uridine (GMU) modified small interfering RNA (siRNA) and evaluated its biophysical and biochemical properties. Incorporation of GMU units significantly increased the thermodynamic stability as well as the enzymatic stability against nucleases in human serum. A gene silencing experiment indicated that GMU modfied siRNA (siRNA6) resulted in approximate to 4.9-fold more efficient knockdown than unmodified siRNA.
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| 8. |
- Nawale, Ganesh N.
(författare)
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Incorporation of 4'-C-aminomethyl-2'-O-methylthymidine into DNA by thermophilic DNA polymerases
- 2012
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Ingår i: Chemical Communications. - : Royal Society of Chemistry (RSC). - 1359-7345 .- 1364-548X. ; 48:77, s. 9619-9621
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Tidskriftsartikel (refereegranskat)abstract
- The dual modified nucleotide 4'-C-aminomethyl-2'-O-methylthymidine 5'-triphosphate was synthesized and enzymatically incorporated into DNA by the thermophilic DNA polymerases Pfu and Therminator III. The dual ribose modification imparted increased exonuclease resistance to DNA compared to the well-known 2'-O-methyl modification.
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| 9. |
- Nawale, Ganesh N.
(författare)
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Synthesis, gene silencing, and molecular modeling studies of 4'-C-aminomethyl-2'-O-methyl modified small interfering RNAs.
- 2012
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Ingår i: Journal of Organic Chemistry. - : American Chemical Society (ACS). - 0022-3263 .- 1520-6904. ; 77:7, s. 3233-3245
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Tidskriftsartikel (refereegranskat)abstract
- The linear syntheses of 4'-C-aminomethyl-2'-O-methyl uridine and cytidine nucleoside phosphoramidites were achieved using glucose as the starting material. The modified RNA building blocks were incorporated into small interfering RNAs (siRNAs) by employing solid phase RNA synthesis. Thermal melting studies showed that the modified siRNA duplexes exhibited slightly lower T(m) (∼1 °C/modification) compared to the unmodified duplex. Molecular dynamics simulations revealed that the 4'-C-aminomethyl-2'-O-methyl modified nucleotides adopt South-type conformation in a siRNA duplex, thereby altering the stacking and hydrogen-bonding interactions. These modified siRNAs were also evaluated for their gene silencing efficiency in HeLa cells using a luciferase-based reporter assay. The results indicate that the modifications are well tolerated in various positions of the passenger strand and at the 3' end of the guide strand but are less tolerated in the seed region of the guide strand. The modified siRNAs exhibited prolonged stability in human serum compared to unmodified siRNA. This work has implications for the use of 4'-C-aminomethyl-2'-O-methyl modified nucleotides to overcome some of the challenges associated with the therapeutic utilities of siRNAs.
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| 10. |
- Paidikondala, Maruthibabu, 1985-, et al.
(författare)
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An Unexpected Role of Hyaluronic Acid in Trafficking siRNA Across the Cellular Barrier : The First Biomimetic, Anionic, Non-Viral Transfection Method
- 2019
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Ingår i: Angewandte Chemie International Edition. - : WILEY-V C H VERLAG GMBH. - 1433-7851 .- 1521-3773. ; 58:9, s. 2815-2819
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Tidskriftsartikel (refereegranskat)abstract
- Circulating nucleic acids, such as short interfering RNA (siRNA), regulate many biological processes; however, the mechanism by which these molecules enter the cell is poorly understood. The role of extracellular-matrix-derived polymers in binding siRNAs and trafficking them across the plasma membrane is reported. Thermal melting, dynamic light scattering, scanning electron microscopy, and computational analysis indicate that hyaluronic acid can stabilize siRNA via hydrogen bonding and Van der Waals interactions. This stabilization facilitated HA size- and concentration-dependent gene silencing in a CD44-positive human osteosarcoma cell line (MG-63) and in human mesenchymal stromal cells (hMSCs). This native HA-based siRNA transfection represents the first report on an anionic, non-viral delivery method that resulted in approximately 60% gene knockdown in both cell types tested, which correlated with a reduction in translation levels.
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| 11. |
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| 12. |
- Paidikondala, Maruthibabu, 1985-, et al.
(författare)
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Insights into siRNA Transfection in Suspension : Efficient Gene Silencing in Human Mesenchymal Stem Cells Encapsulated in Hyaluronic Acid Hydrogel
- 2019
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Ingår i: Biomacromolecules. - : American Chemical Society (ACS). - 1525-7797 .- 1526-4602. ; 20:3, s. 1317-1324
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Tidskriftsartikel (refereegranskat)abstract
- Small interfering RNAs (siRNAs) are powerful toolsfor post-transcriptional gene silencing, which offers enormousopportunities for tissue engineering applications. However, poorserum stability, inefficient intracellular delivery, and inevitabletoxicity of transfection reagents are the key barriers for their clinicaltranslation. Thus, innovative strategies that allow safe and efficientintracellular delivery of the nucleic acid drugs at the desired site isurgently needed for a smooth clinical translation of therapeuticallyappealing siRNA-based technology. In this regard, we havedeveloped an innovative siRNA transfection protocol that employsa short incubation time of just 5 min. This allows easy transfection insuspension followed by transplantation of the cells in a hyaluronicacid (HA) hydrogel system. We also report here the unique ability ofsiRNA to bind HA that was quantified by siRNA release andrheological characterization of the HA-hydrogel. Such interactions also showed promising results to deliver functional siRNA insuspension transfection conditions within 30 min using native HA, although removal of excess HA by centrifugation seem to beessential. In the 2D experiments, suspension transfection of hMSCs with RNAiMAX resulted in ≈90% gene silencing (with orwithout removal of the excess reagent by centrifugation), while HA demonstrated a modest ≈40% gene silencing after removalof excess reagent after 30 min. Transplantation of such transfected cells in the HA-hydrogel system demonstrated an improvedknockdown (≈90% and ≈60% with RNAiMAX and HA respectively after 48 h), with lower cytotoxicity (up to 5-days) asdetermined by PrestoBlue assay. The gene silencing efficiency in the 2D and 3D conditions were also confirmed at the proteinlevels by Western blot analysis. We postulate this novel transfection method could be applied for in vivo applications as it allowsminimal manipulation of cells that are to be transplanted and reduce toxicity.
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| 13. |
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| 14. |
- Rangasami, Vignesh K., et al.
(författare)
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Pluronic Micelle-Mediated Tissue Factor Silencing Enhances Hemocompatibility, Stemness, Differentiation Potential, and Paracrine Signaling of Mesenchymal Stem Cells
- 2021
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Ingår i: Biomacromolecules. - : American Chemical Society (ACS). - 1525-7797 .- 1526-4602. ; 22:5, s. 1980-1989
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Tidskriftsartikel (refereegranskat)abstract
- Mesenchymal stem/stromal cells (MSCs) evoke great excitement for treating different human diseases due to their ability to home inflamed tissues, suppress inflammation, and promote tissue regeneration. Despite great promises, clinical trial results are disappointing as allotransplantation of MSCs trigger thrombotic activity and are damaged by the complement system, compromising their survival and function. To overcome this, a new strategy is presented by the silencing of tissue factor (TF), a transmembrane protein that mediates procoagulant activity. Novel Pluronic-based micelles are designed with the pendant pyridyl disulfide group, which are used to conjugate TF-targeting siRNA by the thiol-exchange reaction. This nanocarrier design effectively delivered the payload to MSCs resulting in similar to 72% TF knockdown (KD) without significant cytotoxicity. Hematological evaluation of MSCs and TF-KD MSCs in an ex vivo human whole blood model revealed a significant reduction in an instant-blood-mediated-inflammatory reaction as evidenced by reduced platelet aggregation (93% of free platelets in the TF-KD group, compared to 22% in untreated bone marrow-derived MSCs) and thrombin- antithrombin complex formation. Effective TF silencing induced higher MSC differentiation in osteogenic and adipogenic media and showed stronger paracrine suppression of proinflammatory cytokines in macrophages and higher stimulation in the presence of endotoxins. Thus, TF silencing can produce functional cells with higher fidelity, efficacy, and functions.
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| 15. |
- Wang, Shujiang, et al.
(författare)
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Dynamic covalent crosslinked hyaluronic acid hydrogels and nanomaterials for biomedical applications
- 2022
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Ingår i: Biomaterials Science. - : Royal Society of Chemistry (RSC). - 2047-4830 .- 2047-4849. ; 10:22, s. 6399-6412
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Forskningsöversikt (refereegranskat)abstract
- Hyaluronic acid (HA), one of the main components of the extracellular matrix (ECM), is extensively used in the design of hydrogels and nanoparticles for different biomedical applications due to its critical role in vivo, degradability by endogenous enzymes, and absence of immunogenicity. HA-based hydrogels and nanoparticles have been developed by utilizing different crosslinking chemistries. The development of such crosslinking chemistries indicates that even subtle differences in the structure of reactive groups or the procedure of crosslinking may have a profound impact on the intended mechanical, physical and biological outcomes. There are widespread examples of modified HA polymers that can form either covalently or physically crosslinked biomaterials. More recently, studies have been focused on dynamic covalent crosslinked HA-based biomaterials since these types of crosslinking allow the preparation of dynamic structures with the ability to form in situ, be injectable, and have self-healing properties. In this review, HA-based hydrogels and nanomaterials that are crosslinked by dynamic-covalent coupling (DCC) chemistry have been critically assessed.
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| 16. |
- Wang, Shujiang, et al.
(författare)
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Influence of ions to modulate hydrazone and oxime reaction kinetics to obtain dynamically cross-linked hyaluronic acid hydrogels
- 2019
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Ingår i: Polymer Chemistry. - : ROYAL SOC CHEMISTRY. - 1759-9954 .- 1759-9962. ; 10:31, s. 4322-4327
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Tidskriftsartikel (refereegranskat)abstract
- Dynamic covalent chemistry forming hydrazone and oxime linkages is attractive due to its simplicity, selectivity and compatibility under aqueous conditions. However, the low reaction rate at physiological pH hampers its use in biomedical applications. Herein, we present different monovalent and bivalent aqueous salt solutions as bio-friendly, non-toxic catalysts which can drive the hydrazone and oxime reactions with excellent efficacy at physiological pH. Direct comparison of hydrazone and oxime reactions using a small molecule model, without any salt catalysis, indicated that oxime formation is 6-times faster than hydrazone formation. Addition of different salts (NaCl, NaBr, KCl, LiCl, LiClO4, Na2SO4, MgCl2 and CaCl2) accelerated the pseudo-first-order reaction kinetics by similar to 1.2-4.9-fold for acylhydrazone formation and by similar to 1.5-6.9-fold for oxime formation, in a concentration-dependent manner. We further explored the potential of such catalysts to develop acylhydrazone and oxime cross-linked hyaluronic acid (HA) hydrogels with different physicochemical properties without changing the degree of chemical modification. Analogous to the small molecule model system, the addition of monovalent and divalent salts as catalysts significantly reduced the gelling time. The gelling time for the acylhydrazone cross-linked HA-hydrogel (1.6 wt%) could be reduced from 300 min to 1.2 min by adding 100 mM CaCl2, while that for the oxime cross-linked HA-hydrogel (1.2 wt%) could be reduced from 68 min to 1.1 min by adding 50 mM CaCl2. This difference in the gelling time also resulted in hydrogels with differential swelling properties as measured after 24 h. Our results are the first to demonstrate the use of salts, for catalyzing hydrogel formation under physiologically relevant conditions.
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| 17. |
- Wang, Shujiang, et al.
(författare)
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Saline Accelerates Oxime Reaction with Aldehyde and Keto Substrates at Physiological pH
- 2018
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Ingår i: Scientific Reports. - : NATURE PUBLISHING GROUP. - 2045-2322. ; 8
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Tidskriftsartikel (refereegranskat)abstract
- We have discovered a simple and versatile reaction condition for oxime mediated bioconjugation reaction that could be adapted for both aldehyde and keto substrates. We found that saline accelerated the oxime kinetics in a concentration-dependent manner under physiological conditions. The reaction mechanism is validated by computational studies, and the versatility of the reaction is demonstrated by cell-surface labeling experiments. Saline offers an efficient and non-toxic catalytic option for performing the bioorthogonal-coupling reaction of biomolecules at the physiological pH. This saline mediated bioconjugation reaction represents the most biofriendly, mild and versatile approach for conjugating sensitive biomolecules and does not require any extensive purification step.
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| 18. |
- Wang, Shujiang, et al.
(författare)
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Saline catalyse oxime reaction at physiological pH : overcoming a major limitation of bioorthogonal reaction
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Tidskriftsartikel (refereegranskat)abstract
- We have discovered a simple and versatile reaction condition for oxime mediated bioconjugation reaction that could be adapted for both aldehyde and keto substrates. We found that saline accelerated the oxime kinetics in a concentration dependent manner under physiological conditions. The reaction mechanism is validated by computational studies, and the versatility of the reaction is demonstrated by cell-surface labeling experiments. Saline offers an efficient and non-toxic catalytic option for performing the bioorthogonal-coupling reaction of biomolecules at the physiological pH. This saline mediated bioconjugation reaction represents the most bio-friendly, mild and versatile approach for conjugating sensitive biomolecules and does not require any extensive purification step.
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