Targeting sodium/iodide symporter gene expression for estrogen-regulated imaging and therapy in breast cancer

Expression of the sodium iodide symporter (hNIS) has been detected in breast cancer tissue, but frequently, not at the levels necessary to mediate (131)I accumulation. Transducing the hNIS gene into breast cancer cells with adenovirus could be a tractable strategy to render breast cancer susceptible to radioiodide therapy. We constructed the replication-incompetent virus, AdSERE, in which an estrogen-responsive promoter directs the expression of hNIS. In vitro, we demonstrate that AdSERE mediates hNIS expression and iodide uptake in ER+ breast cancer cells. In vivo, we show that AdSERE-infected ER+ tumors can be imaged due to tracer accumulation; in addition, AdSERE in combination with therapeutic doses of (131)I suppresses tumor growth.     

Treatment of prostate cancer by radioiodine therapy after tissue-specific expression of the sodium iodide symporter

Causing prostate cancer cells to express functionally active sodium iodide symporter (NIS) by targeted NIS gene transfer might offer the possibility of radioiodine therapy of prostate cancer. Therefore, we investigated radioiodine accumulation and therapeutic effectiveness of 131I in NIS-transfected prostate cancer cells in vitro and in vivo. The human prostatic adenocarcinoma cell line LNCaP was stably transfected with NIS cDNA under the control of the prostate-specific antigen promoter. The stably transfected LNCaP cell line NP-1 showed perchlorate-sensitive, androgen-dependent iodide uptake in vitro that resulted in selective killing of these cells by 131I in an in vitro clonogenic assay. Xenografts were established in athymic nude mice and imaged using a gamma camera after i.p. injection of 500 microCi of 123I. In contrast to the NIS-negative control tumors (P-1) which showed no in vivo uptake of 123I, NP-1 tumors accumulated 25-30% of the total 123I administered with a biological half-life of 45 h. In addition, NIS protein expression in LNCaP cell xenografts was confirmed by Western blot analysis and immunohistochemistry. After a single i.p. application of a therapeutic 131I dose (3 mCi), significant tumor reduction was achieved in NP-1 tumors in the therapy group compared with P-1 tumors and tumors in the control group. In conclusion, a therapeutic effect of 131I has been demonstrated in prostate cancer cells after induction of tissue-specific iodide uptake activity by prostate-specific antigen promoter-directed NIS expression in vitro and in vivo. This study demonstrates the potential of NIS as a novel therapeutic gene for nonthyroidal cancers, in particular prostate cancer.     

Long-term radioiodine retention and regression of liver cancer after sodium iodide symporter gene transfer in wistar rats

Radioiodine therapy of nonthyroid cancers after sodium iodide symporter (NIS) gene delivery has been proposed as a potential application of gene therapy. However, it seems to be precluded by the rapid efflux of taken up iodine from most transduced xenografted tumors. We present an in vivo kinetic study of NIS-related hepatic iodine uptake in an aggressive model of hepatocarcinoma induced by diethylnitrosamine in immunocompetent Wistar rats. We followed the whole-body iodine distribution by repeated imaging of live animals. We constructed a rat NIS (rNIS) adenoviral vector, Ad-CMV-rNIS, using the cytomegalovirus (CMV) as a promoter. Injected in the portal vein in 5 healthy and 25 hepatocarcinoma-bearing rats and liver tumors in 9 hepatocarcinoma-bearing rats, Ad-CMV-rNIS drove expression of a functional NIS protein by hepatocytes and allowed marked (from 20 to 30% of the injected dose) and sustained (>11 days) iodine uptake. This contrasts with the massive iodine efflux found in vitro in human hepatic tumor cell lines. In vivo specific inhibition of NIS by sodium perchlorate led to a rapid iodine efflux from the liver, indicating that the sustained uptake was not attributable to an active retention mechanism but to permanent recycling of the effluent radioiodine via the high hepatic blood flow. Radioiodine therapy after Ad-CMV-rNIS administration achieved a strong inhibition of tumor growth, the complete regression of small nodules, and prolonged survival of hepatocarcinoma-bearing rats. This demonstrates for the first time the efficacy of NIS-based radiotherapy in a relevant preclinical model of nonthyroid human carcinogenesis.     

Targeting of tumor radioiodine therapy by expression of the sodium iodide symporter under control of the survivin promoter

To test the feasibility of using the survivin promoter to induce specific expression of sodium/iodide symporter (NIS) in cancer cell lines and tumors for targeted use of radionuclide therapy, a recombinant adenovirus, Ad-SUR-NIS, that expressed the NIS gene under control of the survivin promoter was constructed. Ad-SUR-NIS mediating iodide uptake and cytotoxicity was performed in vitro. Scintigraphic, biodistribution and radioiodine therapy studies were performed in vivo. PC-3 (prostate); HepG2 (hepatoma) and A375 (melanoma) cancer cells all exhibited perchlorate-sensitive iodide uptake after infection with Ad-SUR-NIS, approximately 50 times higher than that of negative control Ad-CMV-GFP-infected cells. No significant iodide uptake was observed in normal human dental pulp fibroblast (DPF) cells after infection with Ad-SUR-NIS. Clonogenic assays demonstrated that Ad-SUR-NIS-infected cancer cells were selectively killed by exposure to (131)I. Ad-SUR-NIS-infected tumors show significant radioiodine accumulation (13.3 ± 2.85% ID per g at 2 h post-injection), and the effective half-life was 3.1 h. Moreover, infection with Ad-SUR-NIS in combination with (131)I suppressed tumor growth. These results indicate that expression of NIS under control of the survivin promoter can likely be used to achieve cancer-specific expression of NIS in many types of cancers. In combination with radioiodine therapy, this strategy is a possible method of cancer gene therapy.     

Human sodium iodide symporter added to multidrug resistance 1 small hairpin RNA in a single gene construct enhances the therapeutic effects of radioiodine in a nude mouse model of multidrug resistant colon cancer

The objective of this study was to investigate the therapeutic potential of 131I added to doxorubicin therapy in multidrug resistance (MDR) mouse colon cancer coexpressing the MDR1 small hairpin RNA (shRNA) and human sodium iodide symporter (hNIS) gene in a single gene construct and to visualize the antitumor effects using molecular nuclear imaging. HCT-15 coexpressing shRNA for MDR1 gene (MDR1 shRNA) and hNIS gene with a single construct was established (referred to as MN61 cell). Inhibition of P-gp function by MDR1 shRNA and functional activity of hNIS gene was assessed using a ??(m)Tc sestamibi uptake and 12?I uptake, respectively. Cytotoxic effects by a combination of doxorubicin and 131I were determined in parental (HCT-15) or MN61 cells using an in vitro clonogenic assay. Therapeutic effect of either combination therapy (doxorubicin and 131I) or single therapy (doxorubicin or 131I alone) was evaluated by tumor volume measurement. ??(m)Tc-sestamibi, 123I, and ??(m)Tc-pertechnetate images of mice were acquired to evaluate functional assessment in vivo. Cellular uptake of ??(m)Tc-sestamibi and 12?I was approximately 2-fold and 100-fold higher in MN61 cells than in parental cells, respectively. Combination of 131I and doxorubicin resulted in higher cytotoxcity in MN61 cells as compared with parental cells. Scintigraphic imaging showed higher uptake of ??(m)Tc-sestamibi and 123I in MN61 tumor as compared with parental tumor. In mice treated with doxorubicin, there was a slight delay in tumor growth in the MN61 tumor but not in the parental tumor. Cancer treatment with 131I or doxorubicin induced a rapid reduction of tumor volume in the MN61 tumor but not in the parental tumor. Combination therapy further generated a rapid reduction of tumor volume as compared with 131I therapy alone (p?相似文献   

Kinetics of iodide uptake and efflux in various human thyroid cancer cells by expressing sodium iodide symporter gene via a recombinant adenovirus

We evaluated the potential of radioiodide therapy in human sodium iodide symporter (hNIS)-defective thyroid cancer cells via exogenous hNIS expression. Three human thyroid cancer cells (ARO, FRO and NPA) of different origin were transduced by a recombinant adenovirus encoding hNIS expression cassette (Rad-hNIS). The cells were efficiently transduced by a recombinant adenovirus in a virus dose-dependent manner. Consequently, the hNIS protein could be readily detected by Western blot analysis 48-h post-infection at 10 infectious virus particles per cell. These hNIS-transduced cells actively transported iodide into the cytoplasm at the level of 11635.3, 61571.6, and 19367.5 pmoles/10(6) cells in ARO, FRO, and NPA, respectively. However, a significant amount of iodide was eluted to an iodide-free media within 60 min in all the cell lines. RT-PCR analysis revealed that the expression of genes related to iodide trapping (Tg, TSHR and TPO) was dramatically downregulated in these cells. The present study indicates that functional hNIS can be efficiently expressed and is responsible for active transport of iodide in hNIS-negative human thyroid cancer cells by a recombinant adenovirus. However, the human thyroid cancer cells, along with downregulation of iodide metabolism-related gene expression, lose the ability to maintain iodide. Therefore, these kinetic characteristics of iodide uptake and efflux may limit the therapeutic potential of hNIS/radioiodide-based treatment following exogenous hNIS expression in human thyroid cancer.     

重组腺病毒携带钠碘转运体基因介导放射性碘治疗小鼠胶质瘤

目的 验证转染人钠碘转运体基因(hNIS)介导放射性碘治疗肿瘤的有效性.方法 利用重组腺病毒将hNIS基因及人甲状腺过氧化物酶(hTPO)基因转染入人胶质瘤细胞系U251中,使肿瘤细胞获得hNIS和 hTPO基因,然后进行体外摄125I实验、过氯酸盐抑制实验、体外125I反流及内流实验.应用转染hNIS和hTPO基因及未转染hNIS和hTPO基因的细胞株,分别建立荷瘤裸鼠模型,并检测131I对肿瘤的抑制作用.结果 利用腺病毒可以将hNIS和hTPO基因成功转染到U251细胞系中,转染上述基因的肿瘤细胞系可以摄取碘,而且这种摄碘的功能是由hNIS基因所介导的,转染后的hNIS-U251细胞系摄碘能力是U251细胞的121.2倍,hNIS-hTPO-U251细胞系摄碘能力是U251细胞的172.0倍.131I对裸鼠移植瘤的治疗结果表明,在131I作用下,对照组肿瘤均继续迅速生长,而hNIS转染组及hNIS和hTPO共转染组移植瘤体积均有所减小.结论 在肿瘤细胞中转染hNIS和hTPO基因后,可以提高其摄取12I的活性.131I 可以有效杀伤荷瘤裸鼠体内的中瘤细胞.

Abstract：

Objective To explore the possibility of tranfecting hNIS and hTPO genes into gliomas cells by recombinant adenovirus for radioactive iodide treatment. Methods To tranfect hNIS gene into human glioma cell line U251 by recombinant adenovirus.The biological functions of the cells stably expressing hNIS and hTPO genes were assessed by 1251 uptake assay,125I influx-course and 12I-effluxcourse.A glioma model was established with inoculation of the U251 cells in nude mice,and the inhibiting effect of 131 I on the tumor growth was tested in the mouse models. Results The hNIS and hTPO genes were successfully transfected into human gliomas cell line U251 cells by recombinant adenovirus.The radioactive iodide could be intaken by the tumor cells mediated by hNIS gene.The uptake of 125I was higher in cell lines hNIS-U251 and hNIS-hTPO-U251 cells than in cell line U251 cells.The tumor volume of the mice after 131I treatment was significantly decreased in comparison with that before treatment.Conclusion Radioactive 131I treatment after HNIS-based gene transfer can be enhanced and effectively inhibite the tumor growth in nude mice.     

A novel oncolytic viral therapy and imaging technique for gastric cancer using a genetically engineered vaccinia virus carrying the human sodium iodide symporter

Background

Gastric cancers have poor overall survival despite recent advancements in early detection methods, endoscopic resection techniques, and chemotherapy treatments. Vaccinia viral therapy has had promising therapeutic potential for various cancers and has a great safety profile. We investigated the therapeutic efficacy of a novel genetically-engineered vaccinia virus carrying the human sodium iodide symporter (hNIS) gene, GLV-1 h153, on gastric cancers and its potential utility for imaging with ^99mTc pertechnetate scintigraphy and ¹²⁴I positron emission tomography (PET).

Methods

GLV-1 h153 was tested against five human gastric cancer cell lines using cytotoxicity and standard viral plaque assays. In vivo, subcutaneous flank tumors were generated in nude mice with human gastric cancer cells, MKN-74. Tumors were subsequently injected with either GLV-1 h153 or PBS and followed for tumor growth. ^99mTc pertechnetate scintigraphy and ¹²⁴I microPET imaging were performed.

Results

GFP expression, a surrogate for viral infectivity, confirmed viral infection by 24 hours. At a multiplicity of infection (MOI) of 1, GLV-1 h153 achieved > 90% cytotoxicity in MNK-74, OCUM-2MD3, and AGS over 9 days, and >70% cytotoxicity in MNK- 45 and TMK-1. In vivo, GLV-1 h153 was effective in treating xenografts (p < 0.001) after 2 weeks of treatment. GLV-1 h153-infected tumors were readily imaged by ^99mTc pertechnetate scintigraphy and ¹²⁴I microPET imaging 2 days after treatment.

Conclusions

GLV-1 h153 is an effective oncolytic virus expressing the hNIS protein that can efficiently regress gastric tumors and allow deep-tissue imaging. These data encourages its continued investigation in clinical settings.     

Preclinical efficacy of the oncolytic measles virus expressing the sodium iodide symporter in iodine non-avid anaplastic thyroid cancer: a novel therapeutic agent allowing noninvasive imaging and radioiodine therapy

Anaplastic thyroid cancer is an extremely aggressive disease resistant to radioiodine treatment because of loss of sodium iodide symporter (NIS) expression. To enhance prognosis of this fatal cancer, we validated the preclinical efficacy of measles virus (MV)-NIS, the vaccine strain of the oncolytic MV (MV-Edm), modified to include the NIS gene. Western blotting analysis confirmed that a panel of eight anaplastic thyroid cancer (ATC)-derived cell lines do not express NIS protein, but do express CD46, the MV receptor. In vitro cell death assays and in vivo xenograft studies demonstrate the oncolytic efficacy of MV-NIS in BHT-101 and KTC-3, ATC-derived cell lines. Radioactive iodine uptake along with single-photon emission computed tomography (SPECT)-computed tomography imaging of KTC-3 xenografts after (99)Tc(m) administration confirmed NIS expression in vitro and in vivo, respectively, after virus treatment. Adjuvant administration of RAI, to MV-NIS-treated KTC-3 tumors showed a trend for increased tumor cell killing. As current treatment for ATC is only palliative, and MV-NIS is currently Food and Drug Administration approved for human clinical trials in myeloma, our data indicate that targeting ATC with MV-NIS could prove to be a novel therapeutic strategy for effective treatment of iodine-resistant ATC and will expedite its testing in clinical trials for this aggressive disease.     

Probasin promoter (ARR(2)PB)-driven, prostate-specific expression of the human sodium iodide symporter (h-NIS) for targeted radioiodine therapy of prostate cancer

Prostate cancer is one of the most promising candidates for sodium iodide symporter (NIS)-mediated gene therapy. Adenovirus-mediated expression of NIS that is driven by prostate-specific promoters induces generous radioiodine accumulation in prostate cancer cells that may be used for therapy with (131)I. We have recently developed a replication-deficient adenovirus carrying the human NIS cDNA linked to a composite probasin promoter, ARR(2)PB, aiming toward specific expression of the human NIS gene (h-NIS) in prostate tissue for targeted radioactive iodide therapy of prostate cancer (Ad-ARR(2)PB/hNIS). The ability of Ad-ARR(2)PB/hNIS to cause NIS expression in tumor cells was characterized by iodide uptake assay and compared with Ad-CMV/hNIS in which the h-NIS expression is driven by the cytomegalovirus (CMV) promoter. Androgen-dependent prostate cancer cell lines (LNCaP) and non-prostate origin tumor cell lines (SNU449, MCF-7, HCT116, OVCAR-3, and Panc-1) were infected with the viral constructs, and perchlorate-sensitive (125)I uptake and NIS protein expression were measured. Ad-ARR(2)PB/hNIS-infected LNCaP cells showed androgen-dependent and perchlorate-sensitive iodide uptake. Iodide accumulation in LNCaP cells infected with Ad-ARR(2)PB/hNIS, followed by incubation with synthetic androgen, was 5.3-fold increased compared with those coincubated with perchlorate (15,184 +/- 1,173 cpm versus 2,837 +/- 187 cpm). Ad-ARR(2)PB/hNIS-infected LNCaP cells revealed a 3.2-fold increase of iodide accumulation compared with those infected with Ad-CMV/hNIS (multiplicity of infection = 30). Iodide uptake in a panel of non-prostate tumor cell lines infected with Ad-ARR(2)PB/hNIS was no more than 2,500 cpm, demonstrating the tissue specificity of this construct. These results indicate that Ad-ARR(2)PB/hNIS can be used to achieve high-magnitude and tissue-specific expression of h-NIS in prostate tissue and is a promising candidate for cancer gene therapy of prostate cancer.     

全反式维甲酸对甲状腺癌细胞NIS基因表达及吸碘能力影响的实验研究

目的：探讨全反式维甲酸（ATRA）对甲状腺癌细胞株钠/碘同向转运体（NIS）基因表达、吸碘能力的影响,为ATRA用于放射性碘治疗甲状腺癌提供理论依据。方法：分别以不同浓度（10^-7mol/L、10^-6mol/L、10^-5mol/L、10^-4mol/L）的ATRA处理体外培养的甲状腺癌细胞株（FTC-133）,48h后利用半定量RT-PCR检测细胞NISmRNA表达,γ-计数仪检测细胞吸碘能力。结果：ARTA浓度在（0～10%-5）mol/L范围内,细胞NIS基因表达及吸碘能力随ARTA剂量的增加而增加（P〈0.05）。当ARTA浓度达10%-4mol/L时,增加与前一浓度相比无统计学意义（P〉0.05）。结论：ATRA可上调甲状腺癌FTC-133细胞NIS基因表达,增强其吸碘能力,而且这种作用在一定浓度范围内具有剂量依赖性。     

Induction of uterine cervical neoplasias in mice by human papillomavirus type 16 E6/E7 genes.

We constructed a recombinant retrovirus containing the E6/E7 genes of human papillomavirus (HPV) 16 (ZE67) and examined the morphological changes in the cervicovaginal epithelium induced by inoculation of this virus into the vagina of mice. The ZNeo virus without HPV genes was used as a negative control. Moreover, cotreatment with phorbo-13-myristate-12-acetate or N-methyl-N'-nitro-N-nitrosoguanidine and these viruses was carried out. At the end of the observation period (9 months for CD-1 nu/nu and 15 months for C57BL/6J), of 31 CD-1 nude mice treated with ZE67, 7 and 19 had low-grade and high-grade dysplasia, respectively, while 5 of 22 mice treated with ZNeo had low-grade dysplasia (rank sum test, P less than 0.001). Similarly, 4 and 3 of 8 C57BL mice treated with ZE67 had low-grade and high-grade dysplasias, respectively, whereas 3 of the 8 control mice had low-grade dysplasias (P = 0.049). ZE67 plus phorbol-13-myristate-12-acetate and ZE67 plus N-methyl-N'-nitro-N-nitrosoguanidine cotreatments induced cervical cancers in 2 of 13 CD-1 nude mice and 6 of 15 C57BL mice, respectively. On the contrary, none of 6 CD-1 and 2 of 10 C57BL mice had cancer in the control groups (P = 0.0142 for phorbol-13-myristate-12-acetate treatment; P = 0.0173 for N-methyl-N'-nitro-N-nitrosoguanidine treatment). In addition, the existence and expression of HPV 16 E6/E7 genes were detected in the lesions induced by ZE67 but not in the lesions of the control mice by analysis by polymerase chain reaction and mRNA in situ hybridization. The present results suggest that HPV 16 E6/E7 genes induce dysplastic changes but require additional promoting or mutagenic stimulation for the development of cancer.     

Transfection of the Human Sodium/Iodide Symporter （NIS） Gene with Liposomes and the Expression of the NIS Protein in Human Lung A549 Cancer Cells

OBJECTIVE To examine the possibility of human sodium iodide symporter （hNIS） protein expression in lung cancer cells. METHODS Human lung A549 cancer cells were thawed and cultured in vitro. The cells were divided into an experimental group transfected with a recombinant pcDNA3-hNIS plasmid and a control group transfected only with a pcDNA3 plasmid. The recombinant plasmid vector encoding the hNIS gene （pcDNA3-hNIS） was amplified, purified and identified. The hNIS gene was followed by DNA sequencing. A Western blot and an immunohistochemical assay were applied to detect the hNIS protein expression in the transfected human lung A549 cancer cells. RESULTS Restriction enzyme digestion and DNA sequencing results showed the size and direction of the inserted gene in the recombinant pcD- NA3-hNIS plasmid was correct. The Western blot method and immunohistochemical analysis showed a positive NIS protein expression in the experimental group. The NIS protein was detected mainly in the cell membranes showing a positive rate up to 70.6% with no expression of the NIS protein in the control group. There was a significant difference between two groups （P=0.000）. CONCLUSION The hNIS gene was transfected effectively into human lung A549 cancer cells mediated by Lipofectamine 2000, and was expressed with its protein in vitro.     

Enhanced antitumor effects by combination gene therapy using MDR1 gene shRNA and HSV1-tk in a xenograft mouse model

The use of a novel therapeutic vector containing HSV1-thymidine kinase (HSV1-tk) and a short hairpin RNA for the MDR1 gene (shMDR) was proposed previously. We investigated the antitumor effects in an in vivo mouse model of colon cancer and assessed treatment response by serial non-invasive imaging. shMDR-TK expressing (MTKG) tumors for the dual therapy group mice with ganciclovir and doxorubicin showed a decrease in size, while tumors in the single therapy group mice showed a moderate increase (p < 0.05). The ¹³¹I-5-iodo-2′-fluoro-2′deoxy-1-β-d-arabinofuranosyluracil (FIAU) uptake ratio of MTKG-to-parent HCT-15 tumors decreased as treatment progressed for single or dual therapy group mice, while that of the control group mice increased gradually. This study demonstrates the enhanced antitumor effects with combination gene therapy compared with a single therapeutic approach, and provides the potential of therapeutic response monitoring.     

Effective cancer therapy with the alpha-particle emitter [211At]astatine in a mouse model of genetically modified sodium/iodide symporter-expressing tumors.

PURPOSE: The sodium/iodide symporter (NIS) gene is currently explored in several trials to eradicate experimental cancer with radiodine ((131)I) by its beta-emission. We recently characterized NIS-specific cellular uptake of an alternative halide, radioastatine ((211)At), which emits high-energy alpha-particles. The aim of this study was to investigate in vivo effects of the high linear energy transfer (LET) emitter (211)At on tumor growth and outcome in nude mice. EXPERIMENTAL DESIGN: We administered radioastatide in a fractionated therapy scheme to NMRI nude mice harboring rapidly growing solid tumors established from a papillary thyroid carcinoma cell line genetically modified to express NIS (K1-NIS). Animals were observed over 1 year. Tumor growth, body weight, blood counts, survival, and side effects were measured compared with control groups without therapy and/or lack of NIS expression. RESULTS: Within 3 months, radioastatide caused complete primary tumor eradication in all cases of K1-NIS tumor-bearing nude mice (n = 25) with no tumor recurrence during 1 year follow-up. Survival rates of the K1-NIS/(211)At group were 96% after 6 months and 60% after 1 year, in contrast to those of control groups (maximum survival 40 days). CONCLUSION: Our study indicates that (211)At represents a promising substrate for NIS-mediated therapy of various cancers either with endogenous or gene transfer-mediated NIS expression.     

COMBINED IL-2/IL-3 GENE THERAPY FOR G422 MOUSE GLIOBLASTOMA BY INTRATUMORAL INJECTION OF RECOMBINANT ADENOVIRUSES

ＣＯＭＢＩＮＥＤＩＬ２／ＩＬ３ＧＥＮＥＴＨＥＲＡＰＹＦＯＲＧ４２２ＭＯＵＳＥＧＬＩＯＢＬＡＳＴＯＭＡＢＹＩＮＴＲＡＴＵＭＯＲＡＬＩＮＪＥＣＴＩＯＮＯＦＲＥＣＯＭＢＩＮＡＮＴＡＤＥＮＯＶＩＲＵＳＥＳ１ＨｏｎｇＢｏ２洪波ＣａｏＸｕｅｔａｏ３曹雪涛Ｙｕ...     

siRNA下调EIF4E基因对人乳腺癌MDA-MB-231细胞增殖和周期的影响

目的:应用小干扰RNA(small interfering RNA, siRNA)乳腺癌MDA-MB-231细胞中真核细胞翻译起始因子4E (eukaryotic translation initiation factor 4E, EIF 4E )基因的表达,探讨其对乳腺癌细胞增殖和生长周期的影响。方法:构建针对EIF 4E 基因的siRNA表达质粒,采用脂质体法将表达质粒pGPU6/GFP/Neo-EIF4E转染人乳腺癌MDA-MB-231细胞;分别采用RT-PCR、Western 印迹法和免疫细胞化学法检测转染EIF4E siRNA后, 对MDA-MB-231细胞中EIF4E及cyclinD1 mRNA和蛋白的表达水平的影响;MTT法、平板克隆形成实验和FCM检测MDA-MB-231细胞增殖活性、克隆形成率及细胞周期。结果:成功构建EIF4E siRNA表达质粒。RT-PCR、Western 印迹法和免疫细胞化学法检测结果显示,MDA-MB-231细胞中EIF4E及cyclinD1 mRNA和蛋白表达均明显下降(P <0.05)。转染EIF4E siRNA组细胞生长缓慢,集落形成数明显减少,与空白对照组和转染空载体组比较,差异有统计学意义(P<0.05)。FCM检测结果显示,EIF4E siRNA转染组G1期细胞所占百分比为(71.30±0.47)%, 较空白对照组的( 53.10±0.43)%明显增加,而S期细胞所占百分比为(12.53±0.13)%,较空白对照组的(26.47±0.38)%明显减少(P<0.05)。结论:EIF4E siRNA可显著下调EIF4E基因在乳腺癌MDA-MB-231细胞中的表达水平,在一定程度上抑制乳腺癌细胞的增殖,有可能成为临床治疗乳腺癌的靶点之一。     

The antitumor activity of the human FOLR1-specific monoclonal antibody,farletuzumab, in an ovarian cancer mouse model is mediated by antibody-dependent cellular cytotoxicity

Because of its high mortality rate, ovarian cancer is a leading cause of death among women and a highly unmet medical need. New therapeutic agents that are effective and well tolerated are needed and cancer antigen-specific monoclonal antibodies that have direct pharmacologic effects or can stimulate immunological responses represent a promising class of agents for the treatment of this disease. The human folate receptor α (FOLR1), which is overexpressed in ovarian cancer but largely absent in normal tissues, appears to play a role in the transformed phenotype in ovarian cancer, cisplatin sensitivity, and growth in depleted folate conditions and therefore has potential as a target for passive immunotherapy. The anti-FOLR1 monoclonal antibody MORAb-003 (farletuzumab) was previously shown to elicit antibody dependent cellular cytotoxicity (ADCC) and inhibit tumor growth of human tumor xenografts in nude mice. Because of its promising preclinical profile, farletuzumab has been evaluated in clinical trials as a potential therapeutic agent for ovarian cancer. In this report, we demonstrated that farletuzumab’s antitumor effect against an experimental model of ovarian cancer is mediated by its ADCC activity.     

Growth and angiogenesis of human breast cancer in a nude mouse tumour model is reduced by NK4, a HGF/SF antagonist

Hepatocyte growth factor/scatter factor (HGF/SF) is a cytokine primarily produced by stromal fibroblasts and is a known angiogenic and invasion-inducing factor. It is increased in patients with breast cancer. This study examined the effect of NK4, a newly described HGF/SF antagonist, on HGF/SF-promoted growth of a human breast cancer. Both in vitro (invasion and migration assays) and in vivo (murine tumour model) methods were used to ascertain the effect of NK4 on HGF/SF from two sources: human fibroblast-derived HGF/SF and recombinant HGF/SF. In the in vitro invasion assay and migration assay, both HGF/SF and human fibroblasts, which secrete bioactive HGF/SF, increased the invasiveness and migration of the breast cancer cells (MDA MB 231). NK4 significantly reduced this invasiveness and motility. In the animal model, tumour volume and weight was significantly reduced with addition of NK4. It also suppressed HGF/SF-induced growth and markedly retarded tumour growth induced by fibroblasts (MRC5), secreting bioactive HGF/SF. Tumour angiogenesis was assessed by immunohistochemical staining of primary tissue sections using VE-cadherin (an endothelial cell specific cell-cell adhesion molecule). Again, NK4 reduced the effects of both HGF/SF and fibroblasts. We conclude that NK4 has a significant effect on the growth of human breast tumours in nude mice, particularly when stimulated by HGF/SF or fibroblasts. This may occur by decreasing angiogenesis. This gives a clear indication of the therapeutic worth of NK4.