• Molecules and Cells
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• 제45권7호
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• pp.479-494
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• 2022

Human mesenchymal stem cells (MSCs) are multipotent stem cells that have been intensively studied as therapeutic tools for a variety of disorders. To enhance the efficacy of MSCs, therapeutic genes are introduced using retroviral and lentiviral vectors. However, serious adverse events (SAEs) such as tumorigenesis can be induced by insertional mutagenesis. We generated lentiviral vectors encoding the wild-type herpes simplex virus thymidine kinase (HSV-TK) gene and a gene containing a point mutation that results in an alanine to histidine substitution at residue 168 (TK(A168H)) and transduced expression in MSCs (MSC-TK and MSC-TK(A168H)). Transduction of lentiviral vectors encoding the TK(A168H) mutant did not alter the proliferation capacity, mesodermal differentiation potential, or surface antigenicity of MSCs. The MSC-TK(A168H) cells were genetically stable, as shown by karyotyping. MSC-TK(A168H) responded to ganciclovir (GCV) with an half maximal inhibitory concentration (IC₅₀) value 10-fold less than that of MSC-TK. Because MSC-TK(A168H) cells were found to be non-tumorigenic, a U87-TK(A168H) subcutaneous tumor was used as a SAE-like condition and we evaluated the effect of valganciclovir (vGCV), an oral prodrug for GCV. U87-TK(A168H) tumors were more efficiently ablated by 200 mg/kg vGCV than U87-TK tumors. These results indicate that MSC-TK(A168H) cells appear to be pre-clinically safe for therapeutic use. We propose that genetic modification with HSV-TK(A168H) makes allogeneic MSC-based ex vivo therapy safer by eliminating transplanted cells during SAEs such as uncontrolled cell proliferation.

• Asian Pacific Journal of Cancer Prevention
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• 제16권3호
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• pp.1241-1245
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• 2015

Background: Thyroid carcinoma is the most common malignancy of the endocrine organs. Although the majority of thyroid cancer patients experience positive outcomes, anaplastic thyroid carcinoma is considered one of the most aggressive malignancies. Current therapeutic regimens do not confer a significant survival benefit, and new therapies are urgently needed. Oncolytic herpes simplex virus (oHSV) may represent a promising therapy for cancer. In the present study, we investigated the therapeutic effects of a third-generation HSV vector, $G47{\Delta}$, on various human thyroid carcinoma cell lines in vitro. Two subcutaneous (s.c.) models of anaplastic thyroid carcinoma were also established to evaluate the in vivo anti-tumor efficacy of $G47{\Delta}$. Materials and Methods: The human thyroid carcinoma cell line ARO, FRO, WRO, and KAT-5, were infected with $G47{\Delta}$ at different multiplicities of infection (MOIs) in vitro. The survival rates of infected cells were calculated each day. Two s.c. tumor models were established using ARO and FRO cells in Balb/c nude mice, which were intratumorally (i.t.) treated with either $G47{\Delta}$ or mock. Tumor volumes and mouse survival times were documented. Results: $G47{\Delta}$ was highly cytotoxic to different types of thyroid carcinomas. For ARO, FRO, and KAT-5, greater than 30% and 80% of cells were killed at MOI=0.01 and MOI=0.1, respectively on day 5. WRO cells displayed modest sensitivity to $G47{\Delta}$, with only 21% and 38% of cells killed. In the s.c. tumor model, both of the anaplastic thyroid carcinoma cell lines (ARO and FRO) were highly sensitive to $G47{\Delta}$; $G47{\Delta}$ significantly inhibited tumor growth and prolonged the survival of mice bearing s.c. ARO and FRO tumors. Conclusions: The oHSV $G47{\Delta}$ can effectively kill different types of human thyroid carcinomas in vitro. $G47{\Delta}$ significantly inhibited growth of anaplastic thyroid carcinoma in vivo and prolonged animal survival. Therefore, $G47{\Delta}$ may hold great promise for thyroid cancer patients.

• Journal of Microbiology and Biotechnology
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• 제28권6호
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• pp.849-859
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• 2018

Herpes simplex virus type 2 (HSV-2) infection has been a public health concern worldwide. It is the leading cause of genital herpes and a contributing factor to cervical cancer and human immunodeficiency virus (HIV) infection. No vaccine is available yet for the treatment of HSV-2 infection, and routinely used synthetic nucleoside analogs have led to the emergence of drug resistance. The small molecule $Retro-2^{cycl}$ has been reported to be active against several pathogens by acting on intracellular vesicle transport, which also participates in the HSV-2 lifecycle. Here, we showed that Retro-2.1, which is an optimized, more potent derivative of $Retro-2^{cycl}$, could inhibit HSV-2 infection, with 50% inhibitory concentrations of $5.58{\mu}M$ and $6.35{\mu}M$ in cytopathic effect inhibition and plaque reduction assays, respectively. The cytotoxicity of Retro-2.1 was relatively low, with a 50% cytotoxicity concentration of $116.5{\mu}M$. We also preliminarily identified that Retro-2.1 exerted the antiviral effect against HSV-2 by a dual mechanism of action on virus entry and late stages of infection. Therefore, our study for the first time demonstrated Retro-2.1 as an effective antiviral agent against HSV-2 in vitro with targets distinct from those of nucleoside analogs.

• 대한미생물학회지
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• 제35권2호
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• pp.191-201
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• 2000

The viruses of Herpes Simplex Virus 1 (HSV-1), Herpes Simplex Virus 2 (HSV-2) and Varicella-Zoster virus (VZV) which belong to the alpha herpes subfamily are important human pathogens. When eruptions were not fully developed from these viral infections, clinical diagnosis was not always easy and required virological confirmation test. The above viruses were reactivated in individuals who were compromised in immune competence for one reason or another. Polymerase chain reaction (PCR) enables rapid and sensitive detection of HSV and VZV DNAs. Its sensitivity was largely influenced by choice of primers. Authors conducted a study to detect of those three viruses in human specimens including vesicle fluid and joint fluid and serum using PCR methods. Primers used for this study were the general primer pair GPHV-RU which was known to amplify within the genes enjoying the highest degree of homology between UL15 of HSV and UL42 of VZV. PCR with primers hybridized pair GPHV-RU amplifies a 396 bp with THP-1 and HSV-2 standard strain DNA and 405 bp with VZV standard strain DNA. Restriction enzyme cleavage with HpaII and DdeI were used to detect and distinguish DNAs of THP-1 and HSV-2 and VZV. The purpose of this study was a rapid and easy detection of VZV and THP-1 or HSV-2 from various clinical specimens (vesicle fluid, serum and joint fluid) by PCR method. Used methods were: HSV PCR with primer 1, 2 and HpaII RE digestion; VZV nested PCR; HSV PCR with primer A, Band BssHII RE digestion. 1) In 33 cases (33/42, 78.6%) VZV was detected single or mixed infection from 42 clinical specimens which included vesicle fluid (5), serum form respiratory infected children (10), serum from immune suppressed adult cancer patients (7) and joint fluid from arthritis patients (20). 2) In 20 cases (20/42, 47.6%) HSV was detected singly or mixed infection and 19 of those cases were HSV-2 and 1 case was THP-1. 3) In 19 cases (19/42, 45.2%) VZV was singly detected which included serum from respiratory infected children (6 cases), joint fluid from arthritis patients (9 cases), vesicle fluid (2 cases) and serum form immunosuppressed cancer patients (2 cases). 4) HSV was singly detected in 6 cases (6/42, 14.3%) which included joint fluid from arthritis patients (5 cases) and serum form respiratory infected children (1 cases). 5) 14 cases of VZV and HSV mixed infection (14/42, 33.3%) were detected. They included vesicle fluid (3 cases), serum form immunosuppressed cancer patients (4 cases), serum from respiratory infected children (2 cases) and joint fluid from arthritis patients (5 cases). 6) HSV-1 and HSV-2 detection and typing by HSV PCR with primer A, Band BssHII RE digestion method was more sensitive and the results were easier to detect than on other method.

• Asian Pacific Journal of Cancer Prevention
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• 제13권1호
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• pp.339-342
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• 2012

Background: About one third of the human population suffer cancer during their lifetime and more than 20% of total morbidity is related to neoplasia. Cervical cancer is generally the most common cancer in developing countries and the second most common in women globally. The role of human papilloma viruses viruses in its induction is clear. However, the involvement of hepres simplex virus type 2 (HSV-2) is controversial. Therefore a survey was conducted of the prevalence of HSV-2 in patients with cervical cancer and also healthy people with sensitive and quantitative Taq Man real-time PCR assay. Materials and methods: Seventy six formaldehyde fixed paraffin embedded tissue specimens from patients with histologically proven history of cervical cancer as well as 150 control blocks were sectioned for deparaffinization and DNA extraction. Results: There was no HSV-2 DNA in our patient specimens but four control samples were positive, all with a history of hysterectomy. Conclusion: Considering the absence of any positive viral HSV-2 DNA in our patients and also the presence of four positive specimens among our controls, we did not find any relationship between the presence of HSV-2 DNA and cervical cancer.

• 대한세포병리학회지
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• 제1권1호
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• pp.74-84
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• 1990

Herpes simplex virus type 1 and 2(HSV-1, HSV-2) are the ubiquitous human pathogens responsible for a variety of afflictions. HSV-2 is one of the viruses that were suspected of promoting carcinogenesis in the uterine cervix. Certainly, there is a need for the more sensitive and accurate laboratory techniques for HSV detection. We examined total 80 cases of smears including 17 Tzanck smears of skin and 63 cases of Papanicolaou smears from total 77 patients with clinical impression of herpetic infections, from September, 1985 through August, 1989. Immunohistochemical typings for HSV-1 and HSV-2 were performed together with routine cytologic findings and compared. The results are as follows : 1) Patients were 9 males and 33 females, and age distribution was between 5 and 71 years. 2) Subjective symptoms such as ulceration, vesicle, vaginal discharge, pruritus, and pain were complained in 36 patients and 38 cases were genital herpes. Recurrence was noted in 11 cases. 3) Positive results were obtained in 42 among 80 cases. 4) Both routine cytology and immunohistochemical staining were positive in 13 cases and in 24 cases only immunohistochemical staining were positive. 5 cases were positive only in routine cytologic smears. 5) The cases that immunocytochemical stain had been performed were 37 cases, which were all positive in type 2. Among the above 37 cases, type 1 also were positive in 5 cases. The results show that the immunoperoxidase technique is one of the rapid and reliable method to confirm the herpetic infection when suspected and that it is particularly useful when the Papanicolaou smear findings are equivocal.

• Journal of Pharmaceutical Investigation
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• 제33권4호
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• pp.267-272
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• 2003

Herpes simplex virus (HSV) thymidine kinase (TK) has been widely used for suicidal gene therapy in combination with nucleoside analogs such as ganciclovir (GCV). The use of HSV-TK is limited due to the side effect of GCV at high concentrations. Previous studies showed that stable transfectants of mutant HSV-TK with enhanced affinity to GCV were killed at lower GCV concentrations. In this study, we tested whether mutant HSV-TK can provide enhanced suicidal effect when transiently transfected with Epstein-Barr virus (EBV)-based plasmid. EBV-based plasmid which contains OriP and EBNA-1 sequence is well known for a stable episomal maintenance in human cells. Optimal transfection condition was assessed for SNU-638 gastric cancer cell line using polyetylnimine (PEI). Maximum transfection efficiency was achieved when DNA:PEI was 1:3 (w/v). Cytotoxicities of mutant and wild type HSV-TK were compared before and after partially selecting transfected cells. The cells were sensitive to $100\;{\mu}g/ml$ hygromycin. Following GCV treatment, more cells were killed after hygromycin selection than before selection. The mutant HSV-TK showed enhanced cytotoxicity compared with the wild type HSV-TK. Our results suggest that the EBV-based plasmid encoding mutant HSV-TK may be useful to treat the diseases caused by uncontrolled cell proliferation such as cancer and rheumatoid arthritis.

• Bulletin of the Korean Chemical Society
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• 제25권7호
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• pp.991-996
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• 2004

New series of 6-(arylthio)uracils, 6-(4-substituted-1-piperazinyl)uracils, 2,4,5-trioxo-1H,3H-benzothiopyrano[2,3-d]pyrimidine and 5-aryl-2,4-dioxo-1H,3H-pyrimido[5,4-f]benzo[1,4]thiazepines have been prepared and screened for their in vitro activity against herpes simplex-1 virus (HSV-1) and human immunodeficiency virus-1 (HIV-1). The in vitro cytotoxic activity was also evaluated. The results of biological testing revealed that compound 5b showed marginal activity against HSV-1, while compounds 5b and 5f exhibited marginal activity against HIV-1. The rest of the tested compounds were found devoid of antiviral activity against both HSV-1 and HIV-1.

• 한국응용약물학회:학술대회논문집
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• 한국응용약물학회 1993년도 제2회 신약개발 연구발표회 초록집
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• pp.112-112
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• 1993

바이러스 치료제 개발을 위하여 합성된 핵산유도체 11개에 대한 in vitro 항바이러스 약효검색을 수행하였다. 검색대상 바이러스로서 외피보유 DNA 바이러스인 human herpesvirus에 속하는 herpes simplex virus type 1과 type 2에 대해서는 Vero 세포체계에서 3일 후 CPE 저해정도를 MTT 검색법으로 cytomegalovirus에 대해서는 HEL 세포체계에서 7일 후 Giemsa 염색법으로 약효를 측정하였다. 외피비보유 RNA 바이러스인 picornavirus에 속하는 poliovirus type 1과 type 3과 coxsackie B virus type 3에 대한 약효를 HeLa 세포체계에서 2일 후 CPE 저해정도를 MTT 검색법으로 측정하였다. 아울러 selectivity index를 구하기 위하여 Vero와 HeLa 세포에 대한 약물자체의 독성인 cytocidal effect를 MIT 검색법으로 측정하였다. 그 결과 항 herpesvirus 약효는 어떤 물질에서도 발견되지 않았으나 한 물질이 poliovirus type 1과 3에 대하여 selectivity index 10정도 (CC$_{60}$ 38 ug/ml, EC$_{50}$ 1-4 ug/ml)를 나타내었고 자세한 기작은 좀 더 조사할 필요가 있다.

• BMB Reports
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• 제55권12호
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• pp.615-620
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• 2022

The murine leukemia virus-based semi-retroviral replicating vectors (MuLV-based sRRV) had been developed to improve safety and transgene capacity for cancer gene therapy. However, despite the apparent advantages of the sRRV, improvements in the in vivo transduction efficiency are still required to deliver therapeutic genes efficiently for clinical use. In this study, we established a gibbon ape leukemia virus (GaLV) envelope-pseudotyped semi-replication-competent retrovirus vector system (spRRV) which is composed of two transcomplementing replication-defective retroviral vectors termed MuLV-Gag-Pol and GaLV-Env. We found that the spRRV shows considerable improvement in efficiencies of gene transfer and spreading in both human glioblastoma cells and pre-established human glioblastoma mouse model compared with an sRRV system. When treated with ganciclovir after intratumoral injection of each vector system into pre-established U-87 MG glioblastomas, the group of mice injected with spRRV expressing the herpes simplex virus type 1-thymidine kinase (HSV1-tk) gene showed a survival rate of 100% for more than 150 days, but all control groups of mice (HSV1-tk/PBS-treated and GFP/GCV-treated groups) died within 45 days after tumor injection. In conclusion, these findings sug-gest that intratumoral delivery of the HSV1-tk gene by the spRRV system is worthy of development in clinical trials for the treatment of malignant solid tumors.