1.Progress on RNA-based therapeutics for genetic diseases.
Ting LUO ; Chunxiao HUO ; Tianhua ZHOU ; Shanshan XIE
Journal of Zhejiang University. Medical sciences 2023;52(4):406-416
RNA therapeutics inhibit the expression of specific proteins/RNAs by targeting complementary sequences of corresponding genes or encode proteins for the synthesis desired genes to treat genetic diseases. RNA-based therapeutics are categorized as oligonucleotide drugs (antisense oligonucleotides, small interfering RNA, RNA aptamers), and mRNA drugs. The antisense oligonucleotides and small interfering RNA for treatment of genetic diseases have been approved by the FDA in the United States, while RNA aptamers and mRNA drugs are still in clinical trials. Chemical modifications can be applied to RNA drugs, such as pseudouridine modification of mRNA, to reduce immunogenicity and improve the efficacy. The secure and effective delivery systems such as lipid-based nanoparticles, extracellular vesicles, and virus-like particles are under development to address stability, specificity, and safety issues of RNA drugs. This article provides an overview of the specific molecular mechanisms of eleven RNA drugs currently used for treating genetic diseases, and discusses the research progress of chemical modifications and delivery systems of RNA drugs.
Aptamers, Nucleotide
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RNA, Small Interfering/therapeutic use*
;
RNA, Messenger
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Oligonucleotides, Antisense/therapeutic use*
2.MicroRNA expression profiles in enhancement of radiotherapy and the mechanisms.
Journal of Clinical Otorhinolaryngology Head and Neck Surgery 2014;28(21):1721-1726
MicroRNAs are a class of small (-22nt) non-coding RNAs that bind to the 3' untranslated regions (3'-UTRs) of their target mRNAs in a complementary or partially complementary manner via the seed sequence in their 5'-region to regulate biological effect. MicroRNAs also expressin malignant tumors and have close relations with occurrence, development and other biological characteristics of tumor. The effect of radiotherapy and the prognosis of cancer patients are limited and influenced by radioresistant all the time. In recent years, the application of microRNAs to improve the radiation sensitivity of tumor cells is a new field in tumor biotherapy. This paper mainly reviews the identification of related microRNAs participating in and regulating the formation of radiosensitivity/radioresistent, and the research progress of their possible mechanisms.
Humans
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MicroRNAs
;
metabolism
;
therapeutic use
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Neoplasms
;
metabolism
;
radiotherapy
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Prognosis
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RNA, Messenger
;
metabolism
;
Radiation Tolerance
3.Study of the clinical significance of ETAR mRNA expression in high-grade serous ovarian cancer and the inhibitory effect of ETAR derived fusion polypeptide on cancer progression.
Yan Ling ZHANG ; Xiao Kun XIA ; Meng ZHANG
Chinese Journal of Obstetrics and Gynecology 2023;58(12):930-938
Objective: To investigate the clinical significance of endothelin A receptor (ETAR) expression in high-grade serous ovarian carcinoma (HGSOC). To design ETAR carboxyl terminal (ETAR-C) amino acids derived polypeptide and to study the inhibitory effect on ovarian epithelial carcinoma cells in vitro. Methods: (1) A total of 126 patients who received surgical treatment and were diagnosed with HGSOC by postoperative pathological examination in Central Hospital of Xuzhou from January 1, 2007 to December 31, 2017 were selected. All patients had completed clinicopathological data and follow-up data. Cancer tissue samples were collected and ETAR mRNA expression in HGSOC tissues was detected by reverse transcript-PCR. The clinical significance was analyzed. (2) ETAR-C fusion polypeptide was designed based on the sequence of carboxyl terminal amino acids of ETAR, expressed and purified in vitro. The effects of ETAR-C fusion polypeptide on migration and invasion ability of ovarian cancer SKOV3 and CAOV3 cells were detected by scratch test and invasion test, respectively. The effect of ETAR-C fusion polypeptide on chemosensitivity of cisplatin-resistant ovarian cancer SKOV3/cDDP and CAOV3/cDDP cells was determined by methyl thiazolyl tetrazolium (MTT) colorimetric assay. The effect of ETAR-C fusion polypeptide on β-arrestin-1 expression in ovarian cancer SKOV3 and CAOV3 cells was detected by western blot. Results: (1) The relative expression level of ETAR mRNA in HGSOC tissues was 18.6±5.1. Patients with HGSOC were divided into high ETAR mRNA expression (n=76) and low ETAR mRNA expression (n=50) with 61.7% as cut-off value analyzed by X-Tile software. High expression of ETAR mRNA was significantly correlated with abdominal water volume, platinum drug resistance, and cancer antigen 125 (CA125) value in HGSOC patients (all P<0.05), but was not related to the age of patients with HGSOC and the size of postoperative residual lesions (all P>0.05). The 5-year progression free survival rates were 18.4% and 28.0%, and the 5-year overall survival rates were 38.2% and 52.0% in HGSOC patients with high and low ETAR mRNA expression respectively, there were statistically significant differences (P=0.046, P=0.034). (2) The results of scratch test and invasion test showed that the scratch healing rate and cell invasion rate of SKOV3 or CAOV3 cells treated with endothelin-1 (ET-1) and ET-1+ETAR-C were respectively compared, and the differences were statistically significant (all P<0.05). MTT assay showed that the inhibition rates of ETAR-C fusion polypeptide treated in SKOV3/cDDP and CAOV3/cDDP cells were significantly higher than those of control cells after the addition of 4, 6, 8, 10, 12, and 24 μg/ml cisplatin (all P<0.05). Western blot analysis showed that the relative expression levels of β-arrestin-1 in SKOV3 or CAOV3 cells treated with ET-1 and ET-1+ETAR-C were 1.85±0.09 and 1.13±0.09 (SKOV3 cells), 2.14±0.15 and 1.66±0.12 (CAOV3 cells), respectively. The differences were statistically significant (all P<0.05). Conclusions: The prognosis of HGSOC patients with high expression of ETAR mRNA is significantly worse than those with low expression of ETAR mRNA. ETAR might be a new target for HGSOC treatment. The ETAR-C fusion polypeptide that interferes with the interaction of ETAR and β-arrestin-1 has good inhibitory effect on ovarian cancer cells in vitro, and might have clinical application potential.
Female
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Humans
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Amino Acids/therapeutic use*
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beta-Arrestins/therapeutic use*
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Cell Line, Tumor
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Cisplatin/pharmacology*
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Clinical Relevance
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Ovarian Neoplasms/pathology*
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Receptor, Endothelin A/therapeutic use*
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RNA, Messenger/metabolism*
4.Correlation between Serum miR-34a Level and Thrombocytopenia after Chemotherapy in Patients with Diffuse Large B-Cell Lymphoma.
Hong-Li ZHANG ; Adina UTICUL ; Xiao-Wei XU ; Yu-Wei SHI
Journal of Experimental Hematology 2022;30(3):784-789
OBJECTIVE:
To analyze the relationship between serum miR-34a level and thrombocytopenia after chemotherapy in patients with diffuse large B-cell lymphoma (DLBCL).
METHODS:
A total of 69 eligible DLBCL patients who received chemotherapy in our hospital from January 2018 to January 2020 were prospectively included as the research subjects, all patients received R-CHOP 21 regimen (rituximab + cyclophosphamide + adriamycin + vincristine + prednisone) for chemotherapy, 3 weeks was 1 cycle, and 2 cycles of chemotherapy were used. The patients were divided into a reduction group and a non reduction group according to whether there was thrombocytopenia after chemotherapy, the general data and laboratory indexes of the two groups were investigated and compared, the relationship between serum miR-34a before chemotherapy and thrombocytopenia after chemotherapy in patients was analyzed.
RESULTS:
Among the 69 DLBCL patients, 36 patients developed thrombocytopenia after 2 cycles of R-CHOP 21 regimen for chemotherapy, the incidence was 52.17%; the level of serum IL-11 and the relative expression of miR-34a mRNA in the reduction group were significantly lower than the non reduction group (P<0.05), compared other data between groups, there was no statistical significant difference (P>0.05); after Logistic regression analysis, the results showed that the level of serum IL-11 and the relative expression of miR-34a mRNA were related to thrombocytopenia after chemotherapy in DLBCL patients, low expression of each index may be a risk factor of thrombocytopenia after chemotherapy in DLBCL patients (OR>1, P<0.05); ROC curve was drawn, and the results showed that the AUC of serum IL-11 level and miR-34a mRNA relative expression before chemotherapy alone and in combination predicted the risk of thrombocytopenia after chemotherapy in DLBCL patients were all >0.80, and the predictive value was ideal, when the cut-off value of serum IL-11 level before chemotherapy was 42.094 pg/ml, and the cut-off value of miR-34a mRNA relative expression was 3.894, the combined prediction value was the best.
CONCLUSION
The relative expression of miR-34a mRNA is associated with thrombocytopenia after chemotherapy in DLBCL patients, which may be a risk factor for thrombocytopenia in patients after chemotherapy, has certain value in predicting the risk of thrombocytopenia of patients after chemotherapy.
Antineoplastic Combined Chemotherapy Protocols/therapeutic use*
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Cyclophosphamide
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Doxorubicin
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Humans
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Interleukin-11/therapeutic use*
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Lymphoma, Large B-Cell, Diffuse/genetics*
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MicroRNAs/genetics*
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Prednisone/therapeutic use*
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Prognosis
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RNA, Messenger
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Rituximab/therapeutic use*
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Thrombocytopenia
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Vincristine
5.The effect of recombinant HSP47siRNA on the pathological scar in nude mice: in vivo study.
Pei-Sheng JIN ; Jun-Jie CHEN ; Ying CEN ; Ai-Jun ZHANG ; Chang-Bo TAO ; Xue-Yang LI
Chinese Journal of Plastic Surgery 2009;25(5):377-380
OBJECTIVETo study the significance of HSP47 gene in the development of pathological scar.
METHODSThe nude mice were used to reconstruct animal model of pathological scar. 16 days later, the mixture of recombinant HSP47siRNA and liposome was injected into the pathological scar in experimental group. In the control group, 0.25 ml PBS was injected intraperitoneally. 7 days after injection, the specimens were collected for detection of mRNA of HSP47, the collagen and for immunohistochemical study.
RESULTSIn the control and experimental group, the collagen content was (91.71 +/- 1.24)% and (82.12 +/- 4.79)%, respectively; the expression of HSP47mRNA was 1 042 862.01 +/- 604 194.36 and 306 123.68 +/- 105 857.08, respectively; the expression of collagen I mRNA was 10 228 614.70 +/- 2 532 879.04 and 6 011 841.97 +/- 2 886 897.17, respectively;the scar volume was (255.60 +/- 21.34) mm3 and (132.99 +/- 24.06) mm3, respectively. All the above results showed significant difference between the two groups (P < 0.05).
CONCLUSIONSThe collagen production can be reduced through suppression of the expression of HSP47 gene. It indicates that HSP47 gene enhance the development of keloid and could be used as the target of treatment.
Animals ; Cicatrix ; genetics ; pathology ; therapy ; Collagen ; biosynthesis ; Genetic Therapy ; Genetic Vectors ; HSP47 Heat-Shock Proteins ; genetics ; therapeutic use ; Liposomes ; therapeutic use ; Mice ; Mice, Nude ; RNA, Messenger ; genetics ; RNA, Small Interfering ; therapeutic use
6.Targeting glycyrrhetinic acid to hepatic stellate cells in treating rat liver fibrosis.
Qi-sheng ZHANG ; John M LUK ; Jian ZHANG ; Geng-yuan TIAN
Chinese Journal of Hepatology 2005;13(9):664-667
OBJECTIVESWe synthesized M6P26-HSA as a carrier for hepatic stellate cells (HSC) and coupled it with glycyrrhetinic acid (GA) to get a new conjugate GA-HSA-M6P26. Its organ distribution, specific combination with HSC and anti-fibrotic effect on livers were studied.
METHODSThe GA-HSA-M6P26 was labeled with 125I and its organ distribution was detected radiologically. Selective combination of GA-HSA-M6P26 was observed with double immunocytochemic staining and collagen staining of the liver preparations was carried out using Sirius red staining method. The effect of the conjugate on mRNA expression of type I procollagen was studied with real-time PCR in vivo. RT-PCR was used for the effect on mRNA expression of alpha-SMA, MMP-9 and TIMP-1.
RESULTS10 minutes after GA-HSA-M6P26 i.v. injection, 55%+/-5% of it was distributed in the livers. Double immunocytochemic staining showed that most of GA-HSA-M6P26 was taken up by HSC. With GA- HSA- M6P26 treatment, the collagen deposition in the liver decreased significantly compared with GA and M6P26-HSA treated rats. Similarly, the mRNA expression of type I procollagen and alpha-SMA dropped significantly. As to MMP-9 and TIMP-1, no significant change was shown.
CONCLUSIONGA-HSA-M6P26 was selectively delivered to HSC and it showed a significant anti-fibrotic effect on rat liver fibrosis.
Animals ; Drug Delivery Systems ; Glycyrrhetinic Acid ; therapeutic use ; Hepatocytes ; cytology ; metabolism ; Liver Cirrhosis, Experimental ; drug therapy ; Male ; RNA, Messenger ; biosynthesis ; genetics ; therapeutic use ; Rats ; Rats, Sprague-Dawley
8.Research progress on the expression and effect of glucocorticoid receptors in developing brain.
Chinese Journal of Contemporary Pediatrics 2007;9(3):278-280
Animals
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Brain
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growth & development
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metabolism
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Emotions
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Glucocorticoids
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therapeutic use
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Humans
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Memory
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RNA, Messenger
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analysis
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Receptors, Glucocorticoid
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analysis
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genetics
;
physiology
9.Pro-pigmentary action of 5-fluorouracil through the stimulated secretion of CXCL12 by dermal fibroblasts.
Zhi-Kai LIAO ; Shuang-Hai HU ; Bin-Yu HAN ; Xie QIU ; Shan JIANG ; Tie-Chi LEI
Chinese Medical Journal 2021;134(20):2475-2482
BACKGROUND:
There is growing evidence that 5-fluorouracil (5-FU) combined with therapeutic trauma can effectively induce skin repigmentation in vitiligo patients who are unresponsive to conventional treatments. Previous studies have mainly focused on identifying the antimitotic activity of 5-FU for the treatment of skin cancer, but few studies have investigated its extra-genotoxic actions favoring melanocyte recruitment.
METHODS:
We utilized the full thickness excisional skin wound model in Dct-LacZ transgenic mice to dynamically assess the migration of melanocytes in the margins of wounds treated with or without 5-FU. The in-situ expression of CXCL12 was examined in the wound beds using immunofluorescence staining. Quantitative real-time polymerase chain reaction and Western blotting analyses were performed to detect the expression levels of CXCL12 mRNA and protein in primary mouse dermal fibroblasts treated with or without 5-FU. Transwell assays and fluorescein isothiocyanate (FITC)-phalloidin staining were used to observe cell migration and filamentous actin (F-actin) changes of melan-a murine melanocytes.
RESULTS:
Whole mount and cryosection X-gal staining showed that the cell numbers of LacZ-positive melanocytes were much higher in the margins of dorsal and tail skin wounds treated with 5-FU compared with the controls. Meanwhile, CXCL12 immunostaining was significantly increased in the dermal compartment of wounds treated with 5-FU (control vs. 5-FU, 22.47 ± 8.85 vs. 44.69 ± 5.97, P < 0.05). Moreover, 5-FU significantly upregulated the expression levels of CXCL12 mRNA (control vs. 5-FU, 1.00 ± 0.08 vs. 1.54 ± 0.06, P < 0.05) and protein (control vs. 5-FU, 1.00 ± 0.06 vs. 2.93 ± 0.10, P < 0.05) in cultured fibroblasts. Inhibition of the CXCL12/CXCR4 axis suppressed melanocyte migration in vitro using a CXCL12 small interfering RNA (siRNA) or a CXCR4 antagonist (AMD3100).
CONCLUSION
5-FU possesses a pro-pigmentary activity through activation of the CXCL12/CXCR4 axis to drive the chemotactic migration of melanocytes.
Animals
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Cell Movement
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Cell Proliferation
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Chemokine CXCL12/genetics*
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Fibroblasts
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Fluorouracil/therapeutic use*
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Humans
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Mice
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RNA, Messenger
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Receptors, CXCR4
10.Effect of Recombinant Human Thrombopoietin (rhTPO) on Long-term Hematopoietic Recovery in Mice with Acute Radiation Sickness and Relative Mechanism.
Hao LUAN ; Shuang XING ; Jing-Kun YANG ; Ye-Mei WANG ; Xue-Wen ZHANG ; Zi-Zhi QIAO ; Xing SHEN ; Zu-Yin YU
Journal of Experimental Hematology 2023;31(2):546-552
OBJECTIVE:
To investigate the effect and relative mechanism of Recombinant Human Thrombopoietin (rhTPO) on long-term hematopoietic recovery in mice with acute radiation sickness.
METHODS:
Mice were intramuscularly injected with rhTPO (100 μg/kg) 2 hours after total body irradiation with 60Co γ-rays (6.5 Gy). Moreover, six months after irradiation, peripheral blood, hematopoietic stem cells (HSC) ratio, competitive transplantation survival rate and chimerization rate, senescence rate of c-kit+ HSC, and p16 and p38 mRNA expression of c-kit+ HSC were detected.
RESULTS:
Six months after 6.5 Gy γ-ray irradiation, there were no differences in peripheral blood white blood cells, red blood cells, platelets, neutrophils and bone marrow nucleated cells in normal group, irradiated group and rhTPO group (P>0.05). The proportion of hematopoietic stem cells and multipotent progenitor cells in mice of irradiated group was significantly decreased after irradiation (P<0.05), but there was no significant changes in rhTPO group (P>0.05). The counts of CFU-MK and BFU-E in irradiated group were significantly lower than that in normal group, and rhTPO group was higher than that of the irradiated group(P<0.05). The 70 day survival rate of recipient mice in normal group and rhTPO group was 100%, and all mice died in irradiation group. The senescence positive rates of c-kit+ HSC in normal group, irradiation group and rhTPO group were 6.11%, 9.54% and 6.01%, respectively (P<0.01). Compared with the normal group, the p16 and p38 mRNA expression of c-kit+ HSC in the irradiated mice were significantly increased (P<0.01), and it was markedly decreased after rhTPO administration (P<0.01).
CONCLUSION
The hematopoietic function of mice is still decreased 6 months after 6.5 Gy γ-ray irradiation, suggesting that there may be long-term damage. High-dose administration of rhTPO in the treatment of acute radiation sickness can reduce the senescence of HSC through p38-p16 pathway and improve the long-term damage of hematopoietic function in mice with acute radiation sickness.
Humans
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Mice
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Animals
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Thrombopoietin/metabolism*
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Hematopoietic Stem Cells
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Blood Platelets
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Recombinant Proteins/therapeutic use*
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Radiation Injuries
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RNA, Messenger/metabolism*