RNA-cleaving DNAzymes as a diagnostic and therapeutic agent against antimicrobial resistant bacteria.

The development of nucleic-acid-based antimicrobials such as RNA-cleaving DNAzyme (RCD), a short catalytically active nucleic acid, is a promising alternative to the current antibiotics. The current rapid spread of antimicrobial resistance (AMR) in bacteria renders some antibiotics useless against bacterial infection, thus creating the need for alternative antimicrobials such as DNAzymes. This review summarizes recent advances in the use of RCD as a diagnostic and therapeutic agent against AMR. Firstly, the recent diagnostic application of RCD for the detection of bacterial cells and the associated resistant gene(s) is discussed. The next section summarises the therapeutic application of RCD in AMR bacterial infections which includes direct targeting of the resistant genes and indirect targeting of AMR-associated genes. Finally, this review extends the discussion to challenges of utilizing RCD in real-life applications, and the potential of combining both diagnostic and therapeutic applications of RCD into a single agent as a theranostic agent.

Entropy-driven DNA logic circuits regulated by DNAzyme.

The catalytic DNA circuits play a critical role in engineered biological systems and molecular information processing. Actually, some of the natural or synthetic DNA circuits were triggered by covalent modifications, where conformational changes were induced to facilitate complex DNA engineering functions and signal transmissions. However, most of the reported artificial catalytic DNA circuits were regulated by the toehold-mediated reaction. Therefore, it is significant to propose a strategy to regulate the catalytic DNA circuit not only by the toehold-mediated mechanism, but also by involving the conformational changes induced by the covalent modification. In this study, we developed the catalytic DNA logic circuits regulated by DNAzyme. Here, a regulation strategy based on the covalent modification was proposed to control the DNA circuit, combing two reaction mechanisms: DNAzyme digestion and entropy-driven strand displacement. The DNAzyme and DNA catalyst can participate into the reactions alternatively, thus realizing the cascading catalytic circuits. Using the DNAzyme regulation, a series of logic gates (YES, OR and AND) were constructed. In addition, a two-layer cascading circuit and a feedback self-catalysis circuit were also established. The proposed DNAzyme-regulated strategy shows great potentials as a reliable and feasible method for constructing more complex catalytic DNA circuits.

Development and characterization of DNAzyme candidates demonstrating significant efficiency against human rhinoviruses.

BACKGROUND: Infections with human rhinoviruses (RVs) are responsible for millions of common cold episodes and the majority of asthma exacerbations, especially in childhood. No drugs specifically targeting RVs are available. OBJECTIVE: We sought to identify specific anti-RV molecules based on DNAzyme technology as candidates to a clinical study. METHODS: A total of 226 candidate DNAzymes were designed against 2 regions of RV RNA genome identified to be sufficiently highly conserved between virus strains (ie, the 5'-untranslated region and cis-acting replication element) by using 3 test strains: RVA1, RVA16, and RVA29. All DNAzymes were screened for their cleavage efficiency against in vitro-expressed viral RNA. Those showing any catalytic activity were subjected to bioinformatic analysis of their reverse complementarity to 322 published RV genomic sequences. Further molecular optimization was conducted for the most promising candidates. Cytotoxic and off-target effects were excluded in HEK293 cell-based systems. Antiviral efficiency was analyzed in infected human bronchial BEAS-2B cells and ex vivo-cultured human sinonasal tissue. RESULTS: Screening phase-generated DNAzymes characterized by either good catalytic activity or by high RV strain coverage but no single molecule represented a satisfactory combination of those 2 features. Modifications in length of the binding domains of 2 lead candidates, Dua-01(-L12R9) and Dua-02(-L10R11), improved their cleavage efficiency to an excellent level, with no loss in eminent strain coverage (about 98%). Both DNAzymes showed highly favorable cytotoxic/off-target profiles. Subsequent testing of Dua-01-L12R9 in BEAS-2B cells and sinonasal tissue demonstrated its significant antiviral efficiency. CONCLUSIONS: Effective and specific management of RV infections with Dua-01-L12R9 might be useful in preventing asthma exacerbations, which should be verified by clinical trials.

Selection and antitumor activity of anti-Bcl-2 DNAzymes.

Apoptosis pathway has become one of the important targets for therapeutic exploration for cancer therapy. The increased Bcl-2 protein level and phosphorylation is implicated in a decreased chemotherapeutic response in many cancers. BCL-2 inhibitors have been developed as direct inducers of apoptosis. However, resistance to BCL2 inhibitors has been emerging and thus considerable effort has been made to seek novel approaches to BCL2 suppression. In this report we describe an in vitro DNAzyme selection strategy resulting in molecules that are effective in suppressing expression of the target gene BCL-2 in vitro. A 3'-inverted modification was shown to significantly increase the DNAzyme stability in serum and the modified DNAzyme delivered by an osmotic pump chemosensitized human prostate cancer to Taxol in vivo. Thus this study provides an alternative strategy for potential BCL-2-targetd therapy.

Therapeutic potential of siRNA and DNAzymes in cancer.

Cancer is characterized by uncontrolled cell growth, invasion, and metastasis and possess threat to humans worldwide. The scientific community is facing numerous challenges despite several efforts to cure cancer. Though a number of studies were done earlier, the molecular mechanism of cancer progression is not completely understood. Currently available treatments like surgery resection, adjuvant chemotherapy, and radiotherapy are not completely effective in curing all the cancers. Recent advances in the antisense technology provide a powerful tool to investigate various cancer pathways and target them. Small interfering RNAs (siRNAs) could be effective in downregulating the cancer-associated genes, but their in vivo delivery is the main obstacle. DNA enzymes (DNAzymes) have great potential in the treatment of cancer due to high selectivity and significant catalytic efficiency. In this review, we are focusing on antisense molecules such as siRNA and DNAzymes in cancer therapeutics development. This review also describes the challenges and approaches to overcome obstacles involved in using siRNA and DNAzymes in the treatment of cancers.

Substantial Antiviral Potential of Deoxyribozymes Fixed on Anatase Nanoparticles Against Influenza A Viruses in vitro and in vivo.

Influenza A viruses (IAV) are a high threat to humanity because of a lack of proper effective antiviral drugs and resistance of viruses to existing vaccines. We describe the sufficient anti-IAV effect of Ans/PL-Dz nanocomposites that contain deoxyribozymes (Dz) immobilized on anatase TiO2 nanoparticles (Ans) through polylysine linker (PL). The Dz-containing nanocomposites appear to be more efficient than the Ans/PL-ODN nanocomposites that contain common oligodeoxyribonucleotides (ODN) targeted to the same RNA regions of the viral genome. The simultaneous use of nanocomposites that contain Dz and ODN, which are targeted to different sites of viral RNA provides a higher overall effect than the independent action of each of them (synergism). The inhibition of IAV with the proposed nanocomposites was shown to be effective, sequence-specific, and dose-dependent. The most efficient Ans/PL-Dz nanocomposite exhibited a high antiviral effect in vivo on mice models. The efficiency of IAV inhibition with this nanocomposite in vitro and in vivo is higher than that for the approved antiflu drug oseltamivir. The results open the prospect of creating a unique antiviral agent suitable for IAV suppression.

Inhibition of Transmural Inflammation in Crohn's Disease by Orally Administered Tumor Necrosis Factor-Alpha Deoxyribozymes-Loaded Pyroptosis Nanoinhibitors.

Crohn's disease (CD) is a refractory chronic inflammatory bowel disease (IBD) with unknown etiology. Transmural inflammation, involving the intestine and mesentery, represents a characteristic pathological feature of CD and serves as a critical contributor to its intractability. Here, this study describes an oral pyroptosis nanoinhibitor loaded with tumor necrosis factor-α (TNF-α) deoxyribozymes (DNAzymes) (DNAzymes@degradable silicon nanoparticles@Mannose, Dz@MDSN), which can target macrophages at the site of inflammation and respond to reactive oxygen species (ROS) to release drugs. Dz@MDSN can not only break the inflammatory cycle in macrophages by degrading TNF-α mRNA but also reduce the production of ROS mainly from macrophages. Moreover, Dz@MDSN inhibits excessive pyroptosis in epithelial cells through ROS clearance, thereby repairing the intestinal barrier and reducing the translocation of intestinal bacteria to the mesentery. Consequently, these combined actions synergistically contribute to the suppression of inflammation within both the intestine and the mesentery. This study likely represents the first successful attempt in the field of utilizing nanomaterials to achieve transmural healing for CD, which also provides a promising treatment strategy for CD patients.

Antiangiogenic and antitumoral effects mediated by a vascular endothelial growth factor receptor 1 (VEGFR-1)-targeted DNAzyme.

Antiangiogenesis is a promising antitumor strategy that inhibits tumor vascular formation to suppress tumor growth. DNAzymes are synthetic single-strand deoxyribonucleic acid (DNA) molecules that can cleave ribonucleic acids (RNAs). Here, we conducted a comprehensive in vitro selection of active DNAzymes for their activity to cleave the vascular endothelial growth factor receptor (VEGFR-1) mRNA and screened for their biological activity in a matrigel tube-formation assay. Among the selected DNAzymes, DT18 was defined as a lead molecule that was further investigated in several model systems. In a rat corneal vascularization model, DT18 demonstrated significant and specific antiangiogenic activity, as evidenced by the reduced area and vessel number in VEGF-induced corneal angiogenesis. In a mouse melanoma model, DT18 was shown to inhibit B16 tumor growth, whereas it did not affect B16 cell proliferation. We further assessed the DT18 effect in mice with established human nasopharyngeal carcinoma (NPC). A significant inhibition of tumor growth was observed, which accompanied downregulation of VEGFR-1 expression in NPC tumor tissues. To evaluate DT18 effect on vasculature, we performed dynamic contrast enhanced magnetic resonance imaging (DCE-MRI) on the human NPC xenograft mice treated with DT18 and showed a reduction of the parameter of K(trans) (volume constant for transfer of contrast agent), which reflects the condition of tumor microvascular permeability. When examining the safety and tolerability of DT18, intravenous administration of Dz18 to healthy mice caused no substantial toxicities, as shown by parameters such as body weight, liver/kidney function, and histological and biochemical analyses. Taken together, our data suggest that the anti-VEGFR-1 DNAzyme may be used as a therapeutic agent for the treatment of cancer, such as NPC.

[Design of deoxyribozymes for inhibition of influenza A virus].

Influenza A viruses take a significant place in human and animal pathology causing epidemics and epizootics. Therefore, the development of new antiflu drugs has become more and more urgent. Deoxyribozymes can be considered as promising antiviral agents due to their ability to efficiently and highly specifically cleave RNA molecules. In this study, a number ofgenomic sequences of the most relevant influenza A virus subtypes, H5N1, H3N2, and H1N1, were analyzed. Conservative regions were revealed in five the least variable segments of the fragmented viral RNA genome, and potential sites of their cleavage with "10-23" deoxyribozymes were determined. 46 virus-specific 33-mer deoxyribozymes with the general structure of 5'N8AGGCTAGCTACAACGAN9 were designed and synthesized. Screening of the antiviral activity of these agents in conjugation with lipofectin on the Madin-Darby Canine Kidney cells infected with highly pathogenic avian influenza virus A/chicken/Kurgan/05/2005 (H5N1) revealed 17 deoxyribozymes, which suppressed the titer of virus cytopathicity by more than 2.5 IgTCID50/mL (i.e. the virus neutralization index was more than 300), with five of them suppressing the virus titer by a factor of 1000 and more. The most active deoxyribozymes appeared to be specific to segment 5 of the influenza A virus genome, which encoded nucleoprotein (NP).

DNAzyme loaded nano-niosomes attenuate myocardial ischemia/reperfusion injury by targeting apoptosis, inflammation in a NF-κB dependent mechanism.

Although DNAzymes have been found to reduce injury after myocardial ischemia/reperfusion (MI/R), their efficiency have been limited due to rapid degradation in vivo. Thus, this study was conducted to extend their half-life by encapsulation into nano­niosomes and examine their cardioprotective effects in a rat model of myocardial infarction (MI). In order to synthesize nano­niosomes, surface active agent film hydration method was used. Characterization of nano­niosomes was performed using the atomic force microscopy (AFM). In order to establish MI/R model in rats, left anterior descending coronary artery (LAD) was ligated for 30 min. A single dose (150µL) of drug formulations was injected into the infarcted region. The cardiac function was evaluated using echocardiography. The expression of pro-inflammatory cytokines, apoptotic factors, and nuclear factor-κB (NF-κB) were evaluated using Western blot and immunohistochemistry, respectively. Particle size of only nano-niosomes was in the range of 60-90 nm, while a shift to 70-110 nm was seen after DNAzyme encapsulation. MI rats treated with DNAzyme­loaded nano­niosomes could markedly reduce Bax, caspase3, TNF-α, IL-1ß, and NF-κB as well as increase Bcl-2 compared to only MI/R group. Collectively, our finding show that nano­niosomes can be considered excellent drug delivery platforms to extend half-life and stability of DNAzyme, when it is used to reduce myocardial I/R injury.

Bcl-xL DNAzymes promote radiosensitivity and chemosensitivity in colorectal cancer cells via enhancing apoptosis.

BACKGROUND: RNA-cleaving deoxyribozymes (DNAzymes) are catalytic deoxyribonucleic acid molecules that have become a promising new class of gene suppressors by binding and cleaving target mRNA. This study investigated whether DNAzymes targeting Bcl-xL enhanced the effectiveness of radiotherapy and chemotherapy in colorectal cancer (CRC) cells. METHODS: Two types of CRC cells, SW480 and SW837, were transfected with five DNAzymes. Cell viability, Bcl-xL expression and apoptosis were examined. SW480 xenograft model was used to examine the combined effects of Bcl-xL DNAzymes and 5-FU (or X-rays) on tumor growth. RESULTS: Three Bcl-xL DNAzymes, DT882, DT883, and DT884 were identified to be effective in suppressing Bcl-xL expression and causing cell apoptosis. Furthermore, DT882 combined with 5-FU or radiotherapy addictively promoted cell apoptosis and significantly inhibited the growth of SW480 xenografts in vivo. CONCLUSIONS: These results suggest that Bcl-xL DNAzymes can enhance the radiosensitivity and chemosensitivity in CRC cells via inducing apoptosis.

Screening of deoxyribozyme with high reversal efficiency against multidrug resistance in breast carcinoma cells.

Specific inhibition of P-glycoprotein (Pgp) expression, which is encoded by multidrug resistance gene-1 (MDR1), is considered a well-respected strategy to overcome multidrug resistance (MDR). Deoxyribozymes (DRz) are catalytic nucleic acids that could cleave a target RNA in sequence-specific manner. However, it is difficult to select an effective target site for DRz in living cells. In this study, target sites of DRz were screened according to MDR1 mRNA secondary structure by RNA structure analysis software. Twelve target sites on the surface of MDR1 mRNA were selected. Accordingly, 12 DRzs were synthesized and their suppression effect on the MDR phenotype in breast cancer cells was confirmed. The results showed that 4 (DRz 2, 3, 4, 9) of the 12 DRzs could, in a dose-dependent response, significantly suppress MDR1 mRNA expression and restore chemosensitivity in breast cancer cells with MDR phenotype. This was especially true of DRz 3, which targets the 141 site purine-pyrimidine dinucleotide. Compared with antisense oligonucleotide or anti-miR-27a inhibitor, DRz 3 was more efficient in suppressing MDR1 mRNA and Pgp protein expression or inhibiting Pgp function. The chemosensitivity assay also proved DRz 3 to be the best one to reverse the MDR phenotype. The present study suggests that screening targets of DRzs according to MDR1 mRNA secondary structure could be a useful method to obtain workable ones. We provide evidence that DRzs (DRz 2, 3, 4, 9) are highly efficient at reversing the MDR phenotype in breast carcinoma cells and restoring chemosensitivity.

A therapeutic approach to nasopharyngeal carcinomas by DNAzymes targeting EBV LMP-1 gene.

Epstein-Barr virus (EBV)-encoded latent membrane protein 1 (LMP1) has been known to have oncogenic properties during latent infection in nasopharyngeal carcinoma (NPC). Genetic manipulation of LMP1 expression may provide a novel strategy for the treatment of NPC. DNAzymes are synthetic, single-stranded DNA catalysts that can be engineered to bind and cleave the target mRNA of a disease-causing gene. By targeting the LMP1 mRNA, we successfully obtained a phosphorothioate-modified ''10-23'' DNAzyme namely DZ1, through screening a series of DNAzymes. DZ1 could significantly down-regulate the expression of LMP1 in NPC cells, inhibit cell proliferation, metastasis, promote apoptosis and enhance radiosensitivity of NPC through interfering signal pathways which are abnormally activated by LMP1, including NF-κB, AP-1 and STAT3 signal pathways. Together, interfering LMP1 signaling pathway could be a promising strategy to target the malignant phenotypes of NPC.

Nucleic acid-cleaving catalytic DNA for sensing and therapeutics.

DNAzymes with nucleic acid-cleaving catalytic activity are increasing in versatility through concerted efforts to discover new sequences with unique functions, and they are generating excitement in the sensing community as cheap, stable, amplifiable detection elements. This review provides a comprehensive list and detailed descriptions of the DNAzymes identified to date, classified by their associated small molecule or ion needed for catalysis; of note, this classification clarifies conserved regions of various DNAzymes that are not obvious in the literature. Furthermore, we detail the breadth of functionality of these DNA sequences as well as the range of reaction conditions under which they are useful. In addition, the utility of the DNAzymes in a variety of sensing and therapeutic applications is presented, detailing both their advantages and disadvantages.

Development of Novel Chemically-Modified Nucleic Acid Molecules for Efficient Inhibition of Human MAPT Gene Expression.

The hyperphosphorylation of the microtubule-associated protein tau (MAPT) has been implicated in various neurological diseases, including Alzheimer's disease. It has been hypothesized that the reduction of MAPT would result in depolymerizing neurofibrillary tangles and could be a potential strategy for the treatment of Alzheimer's disease and other tauopathies. In this study, we report the development of novel DNAzymes and splice-modulating antisense oligonucleotides (AOs) for the efficient inhibition of MAPT. We designed and synthesized a range of DNAzymes and 2'-O-methyl (2'-OMe)-modified AOs on a phosphorothioate (PS) backbone targeting various exons across the MAPT gene transcript. Our results demonstrated that RNV563, an arm-loop-arm-type DNAzyme targeting exon 13, and an AO candidate AO4, targeting exon 4, efficiently downregulated MAPT RNA expression by 58% and 96%, respectively. In addition, AO4 also reduced the MAPT protein level by 74%. In line with our results, we believe that AO4 could be used as a potential therapeutic molecule for Alzheimer's disease and other tauopathies.

Metal organic framework coated MnO2 nanosheets delivering doxorubicin and self-activated DNAzyme for chemo-gene combinatorial treatment of cancer.

The RNA-cleaving DNAzyme (DZ) holds promising potential for RNA interference (RNAi) applications and is favored over siRNA owing to its high chemical stability, biocompatibility, predictable activity, and substrate versatility. However, its pharmaceutical applications for disease treatment are limited by the requirement of metal cofactor for activation, as well as the lack of effective co-delivery systems to combine with other therapeutic modalities. Herein, we designed and constructed metal organic framework (MOF) coated MnO2 nanosheets to realize the co-delivery of a survivin inhibiting DZ and doxorubicin (DOX) for chemo-gene combinatorial treatment of cancer. In our design, the DOX was adsorbed on MnO2 planar surface, and the DZ was loaded into the MOF shell layer through the coordination between Mn2+ and tannic acid. The nano-system could stably encapsulate the payloads under physiological condition, but rapidly degraded after endocytose into tumor cells in response to intracellular stimuli, resulting in triggered drugs release. Notably, the coreleased Mn2+ could act as metal cofactor for effective DZ activation. Both in vitro and in vivo studies have demonstrated the enhanced anti-tumor efficacy of the nanosystem, with co-contributions from anti-neoplastic DOX, survivin silencing effect of DZ, and to some extent, ROS generation by Mn2+. This work provides an ingenious strategy to address the key limitation of DZ for RNAi applications and realize the combination of DZ with other therapeutic modalities, in which the DZ can be in-situ activated for target gene silencing.

EBV-encoded LMP1 regulates Op18/stathmin signaling pathway by cdc2 mediation in nasopharyngeal carcinoma cells.

Oncoprotein 18/stathmin (Op18/stathmin) plays a crucial role in maintaining cell biological characteristics by regulating microtubule dynamics, especially entry into mitosis; phosphorylated Op18/stathmin promotes microtubule polymerization to form the mitotic spindle, which is essential for chromosome segregation and cell division. Cdc2 is a critical kinase in starting M phase events in cell-cycle progression and is a positive regulator of the cell cycle. Latent membrane protein 1 (LMP1) is an Epstein-Barr virus (EBV)-encoded oncogenic protein that is able to induce carcinogenesis via various signaling pathways. This study focused on regulation by LMP1 of Op18/stathmin signaling in nasopharyngeal carcinoma (NPC) cells and showed that LMP1 regulates Op18/stathmin signaling through cdc2 mediation, LMP1 upregulates cdc2 kinase activity, and Op18/stathmin phosphorylation promotes the interaction of cdc2 with Op18/stathmin and microtubule polymerization during mitosis, and inhibition of LMP1 expression attenuates the interaction of cdc2 and Op18/stathmin and promotes microtubule depolymerization. These results reveal a new pathway via which LMP1 regulates Op18/stathmin signaling by cdc2 mediation; this new signaling pathway not only perfects the LMP1 regulation network but also elucidates the molecular mechanism of LMP1 that leads to carcinogenesis.

c-Jun Is critical for the progression of osteosarcoma: proof in an orthotopic spontaneously metastasizing model.

The oncogene c-Jun has been found to be up-regulated in a variety of cancers including osteosarcoma. DNA enzymes (DNAzymes) are oligonucleotides capable of specific catalysis of target mRNA. A c-Jun DNAzyme inhibited the growth and metastasis of osteosarcoma in an orthotopic spontaneously metastasizing model of the disease. c-Jun down-regulation-mediated apoptosis in osteosarcoma cells involved caspase-1, caspase-2, and caspase-8, but not the Fas/FasL pathway. Clinically, knockdown of c-Jun with DNAzymes may proffer an improved treatment outcome for these tumors originating in bone.

Specific repression of mutant K-RAS by 10-23 DNAzyme: sensitizing cancer cell to anti-cancer therapies.

Point mutations of the Ras family are frequently found in human cancers at a prevalence rate of 30%. The most common mutation K-Ras(G12V), required for tumor proliferation, survival, and metastasis due to its constitutively active GTPase activity, has provided an ideal target for cancer therapy. 10-23 DNAzyme, an oligodeoxyribonucleotide-based ribonuclease consisting of a 15-nucleotide catalytical domain flanked by two target-specific complementary arms, has been shown to effectively cleave the target mRNA at purine-pyrimidine dinucleotide. Taking advantage of this specific property, 10-23 DNAzyme was designed to cleave mRNA of K-Ras(G12V)(GGU-->GUU) at the GU dinucleotide while left the wild-type (WT) K-Ras mRNA intact. The K-Ras(G12V)-specific 10-23 DNAzyme was able to reduce K-Ras(G12V) at both mRNA and protein levels in SW480 cell carrying homozygous K-Ras(G12V). No effect was observed on the WT K-Ras in HEK cells. Although K-Ras(G12V)-specific DNAzymes alone did not inhibit proliferation of SW480 or HEK cells, pre-treatment of this DNAzyme sensitized the K-Ras(G12V) mutant cells to anti-cancer agents such as doxorubicin and radiation. These results offer a potential of using allele-specific 10-23 DNAzyme in combination with other cancer therapies to achieve better effectiveness on cancer treatment.

Deoxyribozyme-mediated knockdown of xylosyltransferase-1 mRNA promotes axon growth in the adult rat spinal cord.

In the injured spinal cord, proteoglycans (PGs) within scar tissue obstruct axon growth through their glycosaminoglycan (GAG)-side chains. The formation of GAG-side chains (glycosylation) is catalysed by xylosyltransferase-1 (XT-1). Here, we knocked down XT-1 mRNA using a tailored deoxyribozyme (DNAXTas) and hypothesized that this would decrease the amount of glycosylated PGs and, consequently, promote axon growth in the adult rat spinal cord. A continuous 2-week delivery of DNAXTas near the rostral border of a peripheral nerve graft bridging the transected dorsal columns in the thoracic spinal cord resulted in an 81% decrease in XT-1 mRNA, an average of 1.4-fold reduction in GAG-side chains of chondroitin sulphate or heparan sulphate-PGs and 2.2-fold reduction in neurocan and brevican core proteins in scar tissue. Additionally, compared to control deoxyribozyme, the DNAXTas treatment resulted in a 9-fold increase in length and a 4-fold increase in density of ascending axons growing through the nerve graft and scar tissue present at the rostral spinal cord. Together our data showed that treatment with a deoxyribozyme against XT-1 mRNA decreased the amount of glycosylated PGs and promoted axon growth through scar tissue in the injured spinal cord. The deoxyribozyme approach may become a contributing factor in spinal cord repair strategies.