The combination of chemotherapy and gene therapy holds great promise for the treatment and eradication of tumors. However, due to significant differences in physicochemical properties between chemotherapeutic agents and functional nucleic acid drugs, direct integration into a single nano-agent is hindered, impeding the design and construction of an effective co-delivery nano-platform for synergistic anti-tumor treatments. In this study, we have developed an mRNA-responsive two-in-one nano-drug for effective anti-tumor therapy by the direct self-assembly of 2\'-fluoro-substituted antisense DNA against P-glycoprotein (2\'F-DNA) and chemo drug paclitaxel (PTX). The 2\'-fluoro modification of DNA could significantly increase the interaction between the therapeutic nucleic acid and the chemotherapeutic drug, promoting the successful formation of 2\'F-DNA/PTX nanospheres (2\'F-DNA/PTX NSs). Due to the one-step self-assembly process without additional carrier materials, the prepared 2\'F-DNA/PTX NSs exhibited considerable loading efficiency and bioavailability of PTX. In the presence of endogenous P-glycoprotein mRNA, the 2\'F-DNA/PTX NSs were disassembled. The released 2\'F-DNA could down-regulate the expression of P-glycoprotein, which decreased the multidrug resistance of tumor cells and enhanced the chemotherapy effect caused by PTX. In this way, the 2\'F-DNA/PTX NSs could synergistically induce the apoptosis of tumor cells and realize the combined anti-tumor therapy. This strategy might provide a new tool to explore functional intracellular co-delivery nano-systems with high bioavailability and exhibit potential promising in the applications of accurate diagnosis and treatment of tumors.

CXCL14 is one of the most evolutionarily conserved members of the chemokine family and is constitutionally expressed in multiple organs, suggesting that it is involved in the homeostasis maintenance of the system. CXCL14 is highly expressed in colon epithelial cells and shows obvious gene silencing in clinical colon cancer samples, suggesting that its silencing is related to the immune escape of cancer cells. In this paper, we analyzed the expression profiles of multiple human clinical colon cancer datasets and mouse colon cancer models to reveal the variation trend of CXCL14 expression during colitis, colon polyps, primary colon cancer, and liver metastases. The relationship between CXCL14 gene silencing and promoter hypermethylation was revealed through the colorectal carcinoma methylation database. The results suggest that CXCL14 is a tumor suppressor gene in colorectal carcinoma which is activated first and then silenced during the process of tumor occurrence and deterioration. Promoter hypermethylation is the main cause of CXCL14 silencing. The methylation level of CXCL14 is correlated with the anatomic site of tumor occurrence, positively correlated with patient age, and associated with prognosis. Reversing the hypermethylation of CXCL14 may be an epigenetic therapy for colon cancer.

Gene editing technologies (GETs) could induce gene knockdown or gene knockout for biomedical applications. The clinical success of gene silence by RNAi therapies pays attention to other GETs as therapeutic approaches. This review aims to highlight GETs, categories, mechanisms, challenges, current use, and prospective applications. The different academic search engines, electronic databases, and bibliographies of selected articles were used in the preparation of this review with a focus on the fundamental considerations. The present results revealed that, among GETs, CRISPR/Cas9 has higher editing efficiency and targeting specificity compared to other GETs to insert, delete, modify, or replace the gene at a specific location in the host genome. Therefore, CRISPR/Cas9 is talented in the production of molecular, tissue, cell, and organ therapies. Consequently, GETs could be used in the discovery of innovative therapeutics for genetic diseases, pandemics, cancer, hopeless diseases, and organ failure. Specifically, GETs have been used to produce gene-modified animals to spare human organ failure. Genetically modified pigs are used in clinical trials as a source of heart, liver, kidneys, and lungs for xenotransplantation (XT) in humans. Viral, non-viral, and hybrid vectors have been utilized for the delivery of GETs with some limitations. Therefore, extracellular vesicles (EVs) are proposed as intelligent and future cargoes for GETs delivery in clinical applications. This study concluded that GETs are promising for the production of molecular, cellular, and organ therapies. The use of GETs as XT is still in the early stage as well and they have ethical and biosafety issues.

Chemodynamic therapy (CDT) has been emerging as a promising strategy for cancer treatment. But the CDT efficiency is restricted by the insufficient intracellular hydrogen peroxide (H2O2) level. Herein, we present a method for H2O2 accumulation in tumor cells by silencing the catalase (CAT) gene with siRNA to achieve enhanced CDT. Cu-siRNA nanocomposites are fabricated by self-assembly of Cu2+ and CAT siRNA and then modified with hyaluronic acid (HA) for active tumor targeting. After tumor cell uptake, the released Cu2+ is reduced by highly expressed glutathione (GSH) to Cu+, which then catalyzes H2O2 to produce toxic hydroxyl radicals (•OH) to kill tumor cells. CAT siRNA can efficiently silence the CAT mRNA to inhibit the consumption of H2O2, resulting in H2O2 accumulation. The Cu2+-mediated GSH elimination and siRNA-induced endogenous H2O2 enrichment both potentiate CDT. Cu-siRNA@HA exhibits good biocompatibility and therapeutic efficiency. This work thus paves a new way to supply H2O2 in CDT and may hold potential for clinical application.

Neuropathic pain following spinal cord injury (SCI) remains a difficult problem that affects more than 80% of SCI patients. Growing evidence indicates that neuroinflammatory responses play a key role in neuropathic pain after SCI. Short hairpin RNA (shRNA) interference is an efficient tool for the knockdown of disease-related specific gene expression after SCI, yet insufficient data is available to establish guidelines. In this study, we have constructed the transient receptor potential ankyrin 1 (TRPA1) shRNA encoded-lentiviral vector (LV-shTRPA1) and P38 MAPK shRNA encoded-lentiviral vector (LV-shP38) to investigate the silencing effects of shRNAs and their ability to reprogram the neuroinflammatory responses, thereby enhancing somatosensory recovery after SCI. Our in vitro data employing HEK293-FT and activated macrophages demonstrated that delivered LV-shRNAs showed high transduction efficacy with no cytotoxicity. Furthermore, a combination of LV-shP38 and LV-shTRPA1 was found to be most effective at suppressing target genes, cutting the expression of pro-inflammatory and pro-nociceptive factors in the dorsal horn of the spinal cord and dorsal root ganglia, thus contributing to the alleviation of neuronal hypersensitivities after SCI. Overall, our data demonstrated that the combination LV-shP38/shTRPA1 produced a synergistic effect for immunomodulation and reduced neuropathic pain with a favorable risk-to-benefit ratio. Collectively, our LV-mediated shRNA delivery will provide an efficient tool for gene silencing therapeutic approaches to treat various incurable disorders.

Insulin-like growth factor 1 receptor (IGF1R), a cell surface receptor with tyrosine kinase (TK) activity, has ligands abnormally expressed in acute leukemia, multiple myeloma, breast, prostate, cervical, and nonsmall cell lung cancers, Ewing\'s sarcoma, and other malignant tumors. IGF1R mediates the malignant proliferation, invasion, and metastasis of tumor cells through a variety of signal transduction pathways, and it is also involved in tumor angiogenesis and tumor cell antiapoptosis. In this study, the neutral cytidinyl lipid DNCA and cystine skeleton cationic lipid CLD from our laboratory could be optimized to encapsulate antisense oligonucleotide (ASO) CT102 to form stable and uniform Mix/CT102 nanoparticles (NPs), which could specifically target tumor cells that highly expressed IGF1R in vivo by intravenous administration. Compared with naked CT102, the lipid complex could promote the uptake and late apoptosis levels of HepG2 and Huh-7 cells, inhibiting cell proliferation efficiently. We also found that Mix/CT102 could enter nucleus in about 2 h, effectively downregulating the mRNA level of IGF1R. The in vivo efficacy experiment demonstrated that in the group that received the optimal dose of Mix/CT102, tumor volume was reduced 8-fold compared with the naked dose group. Meanwhile, in vivo distribution studies showed that the nanoparticles had a predominant accumulation capacity in liver tissue. These results indicated that clinicians can expect the Mix/CT102 nanocomposite to be very effective in reducing the dose and frequency of clinically administered CT102, thereby reducing the side effects of ASOs.

Using small interfering RNA (siRNA) for the specific gene-silencing has been a novel therapeutic method for the treatment of incurable diseases such as malignancies. However, it remains a challenge whether siRNA can be safely and effectively delivered into target cells. Therefore, we synthesized fluorescent carbon dots (CDs) as a gene vector at the siRNA delivery system that induced efficient gene knockdown in vitro while binding aptamer AS1411 to resolve the difficulty in cell targeting. We found that CDs with adequate biocompatibility can improve the efficiency of cellular uptake of siRNA. CLSM and FCM results showed that CDs were mainly localized in the cytoplasm and emitted bright green fluorescence. In addition, the CD/siRNA delivery system mediated by the aptamer AS1411 effectively silenced the expression of Fragile X mental retardation protein (FMRP) and successfully inhibited the migration and invasive propensity of hepatocellular carcinoma (HCC) cells. In summary, we have synthesized a valuable siRNA delivery vector enabling not only bioimaging but also effective downregulation of gene expression, which is indicative of an efficient potential for gene delivery and therapy.

The inflammatory dysfunction of microglia from excess amyloid-β peptide (Aβ) disposal is an overlooked but pathogenic event in Alzheimer\'s disease (AD). Here, we exploit a native high-density lipoprotein (HDL)-inspired nanoscavenger (pHDL/Cur-siBACE1) that combines the trinity of phosphatidic acid-functionalized HDL (pHDL), curcumin (Cur), and β-site APP cleavage enzyme 1 targeted siRNA (siBACE1) to modulate microglial dysfunction. By mimicking the natural lipoprotein transport route, pHDL can penetrate the blood-brain barrier and sequentially target Aβ plaque, where Aβ catabolism is accelerated without microglial dysfunction. The benefit results are from a three-pronged modulation strategy, including promoted Aβ clearance with an antibody-like Aβ binding affinity, normalized microglial dysfunction by blocking the NF-κB pathway, and reduced Aβ production by gene silence (44%). After treatment, the memory deficit and neuroinflammation of APPswe/PSEN 1dE9 mice are reversed. Collectively, this study highlights the double-edged sword role of microglia and provides a promising tactic for modulating microglial dysfunction in AD treatment.

The functionality of DNA biomacromolecules has been widely excavated, as therapeutic drugs, carriers, and functionalized modification derivatives. In this study, we developed a series of DNA tetrahedron nanocages (Td), via synchronous conjugating different numbers of i-(X) and therapeutic siRNA on four vertexes of tetrahedral DNA nanocage (aX-Td@bsiRNA, a+b = 4). This i-motif-conjugated Td exhibited good endosomal escape behaviours in A549 tumor cells, and the escape efficiency was affected by the number of i-motif. Furthermore, the downregulating mRNA and protein expression level of epidermal growth factor receptor (EGFR) caused by this siRNA embedded Td were verified in A549 cells. The tumor growth inhibition efficiency of the 2X-Td@2siRNA treated group in tumor-bearing mice was significantly higher than that of non-i-motif-conjugated Td@2siRNA (3.14-fold) and free siRNA (3.63-fold). These results demonstrate a general strategy for endowing DNA nanostructures with endosomal escape behaviours to achieve effective in vivo gene delivery and therapy.

Pancreatic carcinoma (PC) is greatly induced by the KRAS gene mutation, but effective targeted delivery for gene therapy has not existed. Small interfering ribonucleic acid (siRNA) serves as an advanced therapeutic modality and holds great promise for cancer treatment. However, the development of a non-toxic and high-efficiency carrier system to accurately deliver siRNA into cells for siRNA-targeted gene silencing is still a prodigious challenge. Herein, polyethylenimine (PEI)-modified hydroxyapatite (HAp) nanoparticles (HAp-PEI) were fabricated. The siRNA of the KRAS gene (siKras) was loaded onto the surface of HAp-PEI via electrostatic interaction between siRNA and PEI to design the functionalized HAp-PEI nanoparticle (HAp-PEI/siKras). The HAp-PEI/siKras was internalized into the human PC cells PANC-1 to achieve the maximum transfection efficiency for active tumor targeting. HAp-PEI/siKras effectively knocked down the expression of the KRAS gene and downregulated the expression of the Kras protein in vitro. Furthermore, the treatment with HAp-PEI/siKras resulted in greater anti-PC cells\' (PANC-1, BXPC-3, and CFPAC-1) efficacy in vitro. Additionally, the HAp-PEI exhibited no obvious in vitro cytotoxicity in normal pancreatic HPDE6-C7 cells. These findings provided a promising alternative for the therapeutic route of siRNA-targeted gene engineering for anti-pancreatic cancer therapy.