Browsing by Subject "Gene delivery"

Now showing 1 - 8 of 8
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    Enhancing Prednisone-Based Arthritis Therapy with Targeted IL-27 Gene Delivery
    (MDPI, 2022-06-09) Marin, Adriana A.; Decker, Richard E.; Kumar, Shreya; Lamantia, Zachary; Yokota, Hiroki; Emrick, Todd; Figueiredo, Marxa L.; Biomedical Engineering, School of Engineering and Technology
    Rheumatoid arthritis (RA) is a chronic autoimmune disease which is characterized primarily by synovial hyperplasia and accumulation of several types of immune infiltrates that promote progressive destruction of the articular structure. Glucocorticoids are often prescribed to treat RA because of their strong anti-inflammatory and immunosuppressive effects. However, their application must be limited to the short-term due to a risk of adverse events. In the present study, we examined the potential combination of low-dose prednisone with gene delivery of an agent of promising and complementary effectiveness in RA, interleukin (IL)-27. IL-27 has been shown to have anti-inflammatory potential, while also acting as an effective bone-normalization agent in prior reports. The present report examined a version of IL-27 targeted at the C-terminus with a short 'peptide L' (pepL, LSLITRL) that binds the interleukin 6 receptor α (IL-6Rα) upregulated during inflammation. By focusing on this targeted form, IL-27pepL or 27pL, we examined whether the anti-inflammatory potential of prednisone (at a relatively low dose and short duration) could be further enhanced in the presence of 27pL as a therapy adjuvant. Our results indicate that 27pL represents a novel tool for use as an adjuvant with current therapeutics, such as prednisone, against inflammatory conditions.
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    Hydrodynamic delivery for the study, treatment and prevention of acute kidney injury
    (2014-07-07) Corridon, Peter R.; Atkinson, Simon; Basile, David P.; Bacallao, Robert L.; Dunn, Kenneth William; Gattone II, Vincent H.
    Advancements in human genomics have simultaneously enhanced our basic understanding of the human body and ability to combat debilitating diseases. Historically, research has shown that there have been many hindrances to realizing this medicinal revolution. One hindrance, with particular regard to the kidney, has been our inability to effectively and routinely delivery genes to various loci, without inducing significant injury. However, we have recently developed a method using hydrodynamic fluid delivery that has shown substantial promise in addressing aforesaid issues. We optimized our approach and designed a method that utilizes retrograde renal vein injections to facilitate widespread and persistent plasmid and adenoviral based transgene expression in rat kidneys. Exogenous gene expression extended throughout the cortex and medulla, lasting over 1 month within comparable expression profiles, in various renal cell types without considerably impacting normal organ function. As a proof of its utility we by attempted to prevent ischemic acute kidney injury (AKI), which is a leading cause of morbidity and mortality across among global populations, by altering the mitochondrial proteome. Specifically, our hydrodynamic delivery process facilitated an upregulated expression of mitochondrial enzymes that have been suggested to provide mediation from renal ischemic injury. Remarkably, this protein upregulation significantly enhanced mitochondrial membrane potential activity, comparable to that observed from ischemic preconditioning, and provided protection against moderate ischemia-reperfusion injury, based on serum creatinine and histology analyses. Strikingly, we also determined that hydrodynamic delivery of isotonic fluid alone, given as long as 24 hours after AKI is induced, is similarly capable of blunting the extent of injury. Altogether, these results indicate the development of novel and exciting platform for the future study and management of renal injury.
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    Identification of a physiologic vasculogenic fibroblast state to achieve tissue repair
    (Springer Nature, 2023-02-28) Pal, Durba; Ghatak, Subhadip; Singh, Kanhaiya; Abouhashem, Ahmed Safwat; Kumar, Manishekhar; El Masry, Mohamed S.; Mohanty, Sujit K.; Palakurti, Ravichand; Rustagi, Yashika; Tabasum, Saba; Khona, Dolly K.; Khanna, Savita; Kacar, Sedat; Srivastava, Rajneesh; Bhasme, Pramod; Verma, Sumit S.; Hernandez, Edward; Sharma, Anu; Reese, Diamond; Verma, Priyanka; Ghosh, Nandini; Gorain, Mahadeo; Wan, Jun; Liu, Sheng; Liu, Yunlong; Castro, Natalia Higuita; Gnyawali, Surya C.; Lawrence, William; Moore, Jordan; Perez, Daniel Gallego; Roy, Sashwati; Yoder, Mervin C.; Sen, Chandan K.; Surgery, School of Medicine
    Tissue injury to skin diminishes miR-200b in dermal fibroblasts. Fibroblasts are widely reported to directly reprogram into endothelial-like cells and we hypothesized that miR-200b inhibition may cause such changes. We transfected human dermal fibroblasts with anti-miR-200b oligonucleotide, then using single cell RNA sequencing, identified emergence of a vasculogenic subset with a distinct fibroblast transcriptome and demonstrated blood vessel forming function in vivo. Anti-miR-200b delivery to murine injury sites likewise enhanced tissue perfusion, wound closure, and vasculogenic fibroblast contribution to perfused vessels in a FLI1 dependent manner. Vasculogenic fibroblast subset emergence was blunted in delayed healing wounds of diabetic animals but, topical tissue nanotransfection of a single anti-miR-200b oligonucleotide was sufficient to restore FLI1 expression, vasculogenic fibroblast emergence, tissue perfusion, and wound healing. Augmenting a physiologic tissue injury adaptive response mechanism that produces a vasculogenic fibroblast state change opens new avenues for therapeutic tissue vascularization of ischemic wounds.
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    Interleukin-27 Gene Delivery Targeting IL-6R -Expressing Cells as a Stress Response Therapy
    (MDPI, 2020-02) Neto, Manoel Figueiredo; Liu, Shengzhi; Wes Salameh, Janelle; Yokota, Hiroki; Figueiredo, Marxa Leão; Biomedical Engineering, School of Engineering and Technology
    Interleukin-27 (IL-27) has shown promise in halting tumor growth and mediating tumor regression in several models, including prostate cancer. We describe our findings on the effects of IL-27 on the gene expression changes of TC2R prostate adenocarcinoma cells. We utilized RNAseq to assess profile differences between empty vector control, vector delivering IL-27 modified at its C-terminus with a non-specific peptide, and IL-27 modified at the C-terminus with a peptide targeting the IL-6-Rα. The targeted IL-27 had higher bioactivity and activity in vivo in a recent study by our group, but the mechanisms underlying this effect had not been characterized in detail at the gene expression level on tumor cells. In the present work, we sought to examine potential mechanisms for targeted IL-27 enhanced activity directly on tumor cells. The targeted IL-27 appeared to modulate several changes that would be consistent with an anti-tumor effect, including upregulation in the Interferon (IFN) and Interferon regulatory factor (IRF), oxidative phosphorylation, Janus kinase/Signal transducers and activators of transcription (JAK/STAT), and eukaryotic initiation factor 2 (EIF2) signaling. Of these signaling changes predicted by ingenuity pathway analyses (IPA), the novel form also with the highest significance (-log(Benjamini-Hochberg (B-H)) p-value) was the EIF2 signaling upregulation. We validated this predicted change by assaying for eukaryotic initiation factor 2 alpha (eIF2α), or phosphorylated eIF2α (p-eIF2α), and caspase-3 levels. We detected an increase in the phosphorylated form of eIF2α and in the cleaved caspase-3 fraction, indicating that the EIF2 signaling pathway was upregulated in these prostate tumor cells following targeted IL-27 gene delivery. This approach of targeting cytokines to enhance their activity against cancer cells is a novel approach to help augment IL-27's bioactivity and efficacy against prostate tumors and could be extended to other conditions where it could help interfere with the EIF2α pathway and promote caspase-3 activation.
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    Metformin Bicarbonate-Mediated Efficient RNAi for Precise Targeting of TP53 Deficiency in Colon and Rectal Cancers
    (Elsevier, 2022) Xu, Jiangsheng; Liu, Yunhua; Liu, Sheng; Ou, Wenquan; White, Alisa; Stewart, Samantha; Tkaczuk, Katherine H.R.; Ellis, Lee M.; Wan, Jun; Lu, Xiongbin; He, Xiaoming; Medical and Molecular Genetics, School of Medicine
    Colon and rectal cancers are the leading causes of cancer-related deaths in the United States and effective targeted therapies are in need for treating them. Our genomic analyses show hemizygous deletion of TP53, an important tumor suppressor gene, is highly frequent in both cancers, and the 5-year survival of patients with the more prevalent colon cancer is significantly reduced in the patients with the cancer harboring such deletion, although such reduction is not observed for rectal cancer. Unfortunately, direct targeting TP53 has been unsuccessful for cancer therapy. Interestingly, POLR2A, a gene essential for cell survival and proliferation, is almost always deleted together with TP53 in colon and rectal cancers. Therefore, RNA interference (RNAi) with small interfering RNAs (siRNAs) to precisely target/inhibit POLR2A may be an effective strategy for selectively killing cancer cells with TP53 deficiency. However, the difficulty of delivering siRNAs specifically into the cytosol where they perform their function, is a major barrier for siRNA-based therapies. Here, metformin bicarbonate (MetC) is synthesized to develop pH-responsive MetC-nanoparticles with a unique “bomb” for effective cytosolic delivery of POLR2A siRNA, which greatly facilitates its endo/lysosomal escape into the cytosol and augments its therapeutic efficacy of cancer harboring TP53 deficiency. Moreover, the MetC-based nanoparticles without functional siRNA show notable therapeutic effect with no evident toxicity or immunogenicity.
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    Modeling the gene delivery process of the needle array-based tissue nanotransfection
    (Springer, 2022) Li, Zhigang; Xuan, Yi; Ghatak, Subhadip; Guda, Poornachander R.; Roy, Sashwati; Sen, Chandan K.; Surgery, School of Medicine
    Hollow needle array-based tissue nanotransfection (TNT) presents an in vivo transfection approach that directly translocate exogeneous genes to target tissues by using electric pulses. In this work, the gene delivery process of TNT was simulated and experimentally validated. We adopted the asymptotic method and cell-array-based model to investigate the electroporation behaviors of cells within the skin structure. The distribution of nonuniform electric field across the skin results in various electroporation behavior for each cell. Cells underneath the hollow microchannels of the needle exhibited the highest total pore numbers compared to others due to the stronger localized electric field. The percentage of electroporated cells within the skin structure, with pore radius over 10 nm, increases from 25% to 82% as the applied voltage increases from 100 to 150 V/mm. Furthermore, the gene delivery behavior across the skin tissue was investigated through the multilayer-stack-based model. The delivery distance increased nonlinearly as the applied voltage and pulse number increased, which mainly depends on the diffusion characteristics and electric conductivity of each layer. It was also found that the skin is required to be exfoliated prior to the TNT procedure to enhance the delivery depth. This work provides the foundation for transition from the study of murine skin to translation use in large animals and human settings.
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    Optimization of human papillomavirus-based pseudovirus techniques for efficient gene transfer
    (Nature Publishing group, 2020-09-23) Gilson, Timra D.; Gibson, Ryan T.; Androphy, Elliot J.; Dermatology, School of Medicine
    Human papillomavirus (HPV) L1 and L2 capsid proteins self-assemble into virions capable of efficiently packaging either its 8 kilobase genome or non-viral DNA. The ability of HPV capsids to package non-viral DNA makes these a useful tool for delivering plasmids to study proteins of interest in a variety of cell types. We describe optimization of current methods and present new protocols for using HPV capsids to deliver non-viral DNA thereby providing an alternative to DNA transfection. Using keratinocyte generated extracellular matrices can enhance infection efficiency in keratinocytes, hepatocytes and neuronal cells. Furthermore, we describe a suspension-based efficient technique for infecting different cell types.
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    Ultrasound-mediated gene delivery of factor VIII plasmids for hemophilia A gene therapy in mice
    (Elsevier, 2022-01-10) Song, Shuxian; Lyle, Meghan J.; Noble-Vranish, Misty L.; Min-Tran, Dominic M.; Harrang, James; Xiao, Weidong; Unger, Evan C.; Miao, Carol H.; Pediatrics, School of Medicine
    Gene therapy offers great promises for a cure of hemophilia A resulting from factor VIII (FVIII) gene deficiency. We have developed and optimized a non-viral ultrasound-mediated gene delivery (UMGD) strategy. UMGD of reporter plasmids targeting mice livers achieved high levels of transgene expression predominantly in hepatocytes. Following UMGD of a plasmid encoding human FVIII driven by a hepatocyte-specific promoter/enhancer (pHP-hF8/N6) into the livers of hemophilia A mice, a partial phenotypic correction was achieved in treated mice. In order to achieve persistent and therapeutic FVIII gene expression, we adopted a plasmid (pHP-hF8-X10) encoding an FVIII variant with significantly increased FVIII secretion. By employing an optimized pulse-train ultrasound condition and immunomodulation, the treated hemophilia A mice achieved 25%–150% of FVIII gene expression on days 1–7 with very mild transient liver damage, as indicated by a small increase of transaminase levels that returned to normal within 3 days. Therapeutic levels of FVIII can be maintained persistently without the generation of inhibitors in mice. These results indicate that UMGD can significantly enhance the efficiency of plasmid DNA transfer into the liver. They also demonstrate the potential of this novel technology to safely and effectively treat hemophilia A.