Pan-TRK positive uterine sarcoma in immunohistochemistry without neurotrophic tyrosine receptor kinase gene fusions:A case report

BACKGROUND The classification of uterine sarcomas is based on distinctive morphological and immunophenotypic characteristics,increasingly supported by molecular genetic diagnostics.Data on neurotrophic tyrosine receptor kinase(NTRK)gene fusionpositive uterine sarcoma,potentially aggressive and morphologically similar to fibrosarcoma,are limited due to its recent recognition.Pan-TRK immunohistochemistry(IHC)analysis serves as an effective screening tool with high sensitivity and specificity for NTRK-fusion malignancies.CASE SUMMARY We report a case of a malignant mesenchymal tumor originating from the uterine cervix,which was pan-TRK IHC-positive but lacked NTRK gene fusions,accompanied by a brief literature review.A 55-year-old woman presented to the emergency department with abdominal pain and distension,exhibiting significant ascites and multiple solid pelvic masses.Pelvic examination revealed a tumor encompassing the uterine cervix,extending to the vagina and uterine corpus.A punch biopsy of the cervix indicated NTRK sarcoma with positive immunochemical pan-TRK stain.However,subsequent next generation sequencing revealed no NTRK gene fusion,leading to a diagnosis of poorly differentiated,advanced-stage sarcoma.CONCLUSION The clinical significance of NTRK gene fusion lies in potential treatment with TRK inhibitors for positive sarcomas.Identifying such rare tumors is crucial due to the potential applicability of tropomyosin receptor kinase inhibitor treatment.

Detection of the SYT-SSX Fusion Gene in Synovial Sarcoma

Objective: To investigate the feasibility and significance of detecting SYT-SSX fusion gene in paraffin-embedded tissues of synovial sarcoma (SS) by reverse-transcriptase polymerase chain reaction(RT-PCR) methods. Methods: Twenty cases of SS tumors from archival materials were collected and all samples were formalin-fixed and paraffin-embedded (FFPE). SYT-SSX fusion transcript was detected by RT-PCR. Home-keeping gene Porphobilinogen Deaminase (PBGD) was regarded as internal control.Results: PBGD mRNA was detected in all 20 tumor cases (100%). SYT-SSX fusion transcript was detected in 18 tumor cases (90%). In 18 SYT-SSX positive SS cases, there are 12 present SYT-SSX1 fusion transcript and 6 present SYT-SSX2 fusion transcript. SYT-SSX1 fusion transcript can be seen in 9 monophasic SS and 3 biphasic SS. In 6 SYT-SSX2 positive SS cases, 4 were monophasic SS and 2 were biphasic. Conclusion: Detection of SYT-SSX fusion transcripts in FFPE tissues for diagnosis of SS is feasible and sensitive. Subtypes of SYT-SSX fusion gene may provide prognosis information.

貂源绿脓杆菌山东分离株exoA-fliC基因的原核表达及其免疫原性研究

采用重叠PCR方法扩增水貂绿脓杆菌山东分离株(PASD01株)的exoA与fli C基因,并将两段基因嵌合后,克隆到表达载体p ET-30a上,转化到宿主菌BL21(DE3)中诱导表达,并对表达产物进行免疫原性分析。结果表明,重组嵌合蛋白在37℃,0. 8 mmol/L IPTG诱导7 h时表达量最高;经SDS-PAGE和Western Blot分析,该蛋白在约68 k D处出现特异性条带,与预期结果一致;该重组嵌合蛋白免疫小鼠后,最高血清效价可达1∶640 000。结论:水貂绿脓杆菌exoA-fli C嵌合蛋白具有良好的免疫原性,为其进一步发展成为新型亚单位疫苗来抵抗水貂出血性肺炎奠定基础。

Detection of fusion gene in cell-free DNA of a gastric synovial sarcoma

Synovial sarcoma(SS) is genetically characterized by chromosomal translocation, which generates SYT-SSX fusion transcripts. Although SS can occur in any body part, primary gastric SS is substantially rare. Here we describe a detection of the fusion gene sequence of gastric SS in plasma cell-free DNA(cf DNA). A gastric submucosal tumor was detected in the stomach of a 27-year-old woman and diagnosed as SS. Candidate intronic primers were designed to detect the intronic fusion breakpoint and this fusion sequence was confirmed in intron 10 of SYT and intron 5 of SSX2 by genomic polymerase chain reaction(PCR) and direct sequencing. A locked nucleic acid(LNA) probe specificto the fusion sequence was designed for detecting the fusion sequence in plasma and the fusion sequence was detected in preoperative plasma cfD NA, while not detected in postoperative plasma cfD NA. This technique will be useful for monitoring translocation-derived diseases such as SS.

Transgenic rice expressing a novel phytase-lactoferricin fusion gene to improve phosphorus availability and antibacterial activity

The developing trends of livestock production are efficiency availability of phytate phosphorus and abuse of antibiotics. safety and sustainability, which face two major challenges： low As a solution phytases and antimicrobial peptides are applied as feed additives. However, phytases and antimicrobial peptides are susceptible to proteases, costly by fermentation and potential toxic to production hosts. We transformed an optimized phytase-lactoferricin fusion gene PhyLfdriven by an en- dosperm-specific promoter Gt13aP and Bar （bialaphos resistance） gene as a selection maker into rice. The Bar and PhyLf genes were integrated into the rice genome, stably inherited and expressed. Their phosphinothricin acetyl transferase （PAT） protein content oftransgenic plants with glufosinate resistance varied between 50.45-93.39 IJg g-l. Fusion protein expressed especially in the seeds of transgenic rice had a summit phytase activity at 32.30 U g-l, which increased by 61.71-fold com- pared to the control/check group （CK） and 7.54-fold compared to un-optimized transgenic plant. The highest inorganic phosphorus （Pi） content of the transgenic seeds reached 13.15 mg g-~, increased by 12.77-fold compared to that of CK. Preliminary antibacterial experiments showed that the enterokinase hydrolysate product of fusion protein could inhibit the growth of Escherichia coil DH5a. These results indicated that the protein PhyLf has the potential to increase availability of feed phytate phosphorus, improve consumer＇s immunity and reduce the use of antibiotics.

PCA3 and TMPRSS2-ERG gene fusions as diagnostic biomarkers for prostate cancer

The incidence of prostate cancer （PCa） is rising steadily among males in many countries. Serum prostate-specific antigen （PSA） is widely applied to clinical diagnosis and screening of PCa. However, the so-called grey area of PSA levels 4.0-10.0 ng/mL has a low specificity of 25-40% resulting in a high rate of negative biopsy and overtreatment. So in order to treat PCa patients in early stage, there is an urgent need for new biomarkers in PCa diagnosis. The PCA3 gene, a non-coding RNA （ncRNA） that is highly expressed in prostate cancer （PCa） cells, has been identified as a molecular biomarkers to detect PCa, of which PCA3 has already under clinical application. PCA3 is strongly overexpressed in malignant prostate tissue compared to benign or normal adjacent one. Newly, PCA3 is considered to be a promising biomarker in clinical diagnosis and targeted therapy. The diagnostic significance of PCA3, however, is awaiting further researches. Moreover, it has been demonstrated recently that TMPRSS2-ERG gene fusion is identified as the predominant genetic change in patients diagnosed with PCa. Recent study revealed that combination of the PC43 and TMPRSS2-ERG gene fusion test optimizes PCa detection compared with that of single biomarker, which would lead to a considerable reduction of the number of prostate biopsies. In this review, we focused on the potential use of PCA3 and TMPRSS2-ERG gene fusion detection in the diagnosis of PCa.

Cloning of α-β fusion gene from Clostridium perfringens and its expression

AIM： To study the cloning of α-β fusion gene from Closindium perfringens and the immunogenidty of 0-6 fusion expression. METHODS： Cloning was accomplished after PCR amplification from strains NCTC64609 and C58-1 of the protective antigen genes of α-toxin and β-toxin. The fragment of the gene was cloned using plasmid pZCPAB. This fragment coded for the gene with the stable expression of α-β fusion gene binding. In order to verify the exact location of the α-β fusion gene, domain plasmids were constructed. The two genes were fused into expression vector pBV221. The expressed α-β fusion protein was identified by ELISA, SDS-PAGE, Western blotting and neutralization assay. RESULTS： The protective co-toxin gene （cpa906） and the β-toxin gene （cpb930） were obtained. The recombinant plasmid pZCPAB carrying α-β fusion gene was constructed and transformed into BL21（DE3）. The recombinant strain BL21（DE3）（pZCPAB） was obtained. After the recombinant strain BL21（DE3）（pZCPAB） was induced by 42℃, its expressed product was about 22.14% of total cellular protein at SDS-PAGE and thin-layer gel scanning analysis. Neutralization assay indicated that the antibody induced by immunization with α-βfusion protein could neutralize the toxicity of α-toxin and β-toxin. CONCLUSION： The obtained α-toxin and β-toxin genes are correct. The recombinant strain BL21（DE3）（pZCPAB） could produce α-β fusion protein. This protein can be used for immunization and is immunogenic. The antibody induced by immunization with α-β fusion protein could neutralize the toxicity of α-toxin and β-toxin.

Contruction of the Genetic Engineering Strain Expressed Nontoxic ST_1-LT_B Fusion Protein Against Enterotoxigenic Eschenichia coli

Thermostable enterotoxinⅠ(ST1) mutant genes and thermolabile enterotoxin B subunit (LTB)genes were amplified by PCR from plasmids of Eschenichia coli C83902. The recombinantexpression plasmid pZST3LTB containing ST1-LTB fusion gene was constructed by recombinantDNA technique and then transformed into Escherichia coli BL21(DE3). The ST1-LTB fusionprotein was highly expressed in recombinant strain BL21(DE3)(pZST3LTB) and the fusionprotein was about 38.53% of total cellular protein by SDS-PAGE and thin-layer gelscanning analysis. More important, mice immunized with crude preparation containing thefusion protein inclusion bodies or inactivated recombinant strain produced antibodiesthat were able to recognize ST1 in vitro. These sera antibodies were able to neutralizethe biological activity of native ST1 in the suckling mouse assay. Hence the ST1-LTBfusion protein was nontoxic and immunogenic, the constructed recombinant strain BL21(DE3)(pZST3LTB) could be used as a candidate of vaccine strain.

Stable Expression of Hantavirus H8205 Strain G1/IL-2 Gene and Immune Protection of the Fusion Gene

To explore the feasibility of stable expression of Hantavirus H8205 strain G1 segment and human IL-2 fusion gene in Vero cells, and to examine the immune protection effects on mice vaccinated with this recombinant eukaryotic expression vector containing Hantavirus G1 gene and IL-2 gene. With the help of lipofectamine, the Vero cells were transfected with pcDNA3.1/HisB-IL-2-G1 and the positive cells were selected by G418. IFAT and SDS-PAGE elec- trophoresis were used to determine the stable transfection and expression of recombinant protein. Each mouse was inoculated with plasmids intramuscularly （i.m.） three times, 2 boosts were given at 2-week intervals, serum anti-hantavirus antibodies were detected by ELISA and neutralizing antibodies （NAb） were detected by Plaque Reduction Neutralization Test. The fusion protein expressed in Vero cells was 78 kD, corresponding to the estimated molecular size. The neutralizing antibody titers of mice with pcDNA3.1/HisB-IL-2-G1 were 1：20-1：80. IL-2/G1 fusion gene could be transferred in Vero cells and stably express the fusion protein. Specific humeral immune responses in mice can be induced with the recombinant eukaryotic expression vector containing the fusion gene, which lays the foundation for further development of therapeutic HTNV vaccine.

Bifunctional chimeric SuperCD suicide gene -YCD: YUPRT fusion is highly effective in a rat hepatoma model

AIM： To investigate the effects of catalytically superior gene-directed enzyme prodrug therapy systems on a rat hepatoma model. METHODS： To increase hepatoma cell chemosensitivity for the prodrug 5-fluorocytosine （5-FC）, we generated a chimeric bifunctional SuperCD suicide gene, a fusion of the yeast cytosine deaminase （YCD） and the yeast uracil phosphoribosyltransferase （YUPRT） gene. RESULTS： In vitro stably transduced Morris rat hepatoma cells （MH） expressing the bifunctional SuperCD suicide gene （MH SuperCD） showed a clearly marked enhancement in cell killing when incubated with 5-FC as compared with MH cells stably expressing YCD solely （MH YCD） or the cytosine deaminase gene of bacterial origin （MH BCD）, respectively. In vivo, MH SuperCD tumors implanted both subcutaneously as well as orthotopically into the livers of syngeneic ACI rats demonstrated significant tumor regressions （P〈0.01） under both high dose as well as low dose systemic 5-FC application, whereas MH tumors without transgene expression （MH naive） showed rapid progression. For the first time, an order of in vivo suicide gene effectiveness （SuperCD〉〉 YCD〉〉BCD〉〉〉negative control） was defined as a result of a direct in vivo comparison of all three suicide genes. CONCLUSION： Bifunctional SuperCD suicide gene expression is highly effective in a rat hepatoma model, thereby significantly improving both the therapeutic index and the efficacy of hepatocellular carcinoma killing by fluorocytosine.

Fusion genes in solid tumors: an emerging target for cancer diagnosis and treatment

Studies over the past decades have uncovered fusion genes, a class of oncogenes that provide immense diagnostic and therapeutic advantages because of their tumor-specific expression. Originally associated with hemotologic cancers, fusion genes have recently been discovered in a wide array of solid tumors, including sarcomas, carcinomas, and tumors of the central nervous system. Fusion genes are attractive as both therapeutic targets and diagnostic tools due to their inherent expression in tumor tissue alone. Therefore, the discovery and elucidation of fusion genes in various cancer types may provide more effective therapies in the future for cancer patients.

Fusion of EGFP and porcine α 1,3GT genes decrease GFP expression

Objective:To investigate the effect of fusion proteins expressed by the fused gene of porcineα1,3 galactosyltransierase(α1,3 GT) and enhanced green fluorescent protein(EGFP) on the green fluorescence intensity of EGFP.Methods:The fragment containingα1.3GT was firstly recovered after the pcDNA3.1-α1.3GT recombinant vector were digested with BamHl and EcoRI,and then,the resultant fragment was ligated to the pEGFP-N1 vector which was also digested with the same enzymes.The new recombinant eukaryotic expression pEGFP/a 1,3GT vector was obtained and sequenced.The pEGFP/α1,3GT was used to transfect human lung carcinoma cells A549 and HEKC 293FT,and the expression of EGFP was quantitatively analyzed by fluorescent microscope and flow cytometry.Results:The positive percentage of A549 was 80.5%,and that of 293 FT was 86.5%48 hours after the two cell lines both were transfected by pEGFP-N1.The positive percentage of A549 was 75.8%,and that of 293 FT was 81.2%48 hours after the two cell lines were transfected by pEGFP/α1.3GT.The mean fluorescence intensities of A549 transfected with pEGFP-N1 and pEGFP/α1.3GT were 1.21 and 0.956,respectively when compared with that of A549 without transfection.Meanwhile,the those of the 293FT that were transfected with pEGFP-N1 and pEGFP/αl,3GT were 7.66 and 1.00.respectively when compared with that of 293FT cells without transfection.Conclusions:These results suggested that the expression of EGFP gene fused with porcineα1,3GT gene was partly inhibited.

Cloned s-Lap Gene Coding Area, Expression and Localization of s-Lap/GFP Fusion Protein in Mammal Cells

s-Lap is a new gene sequence from pig retinal pigment epithelial(RPE) cells, which was found and cloned in the early period of apoptosis of RPE cells damaged with visible light. We cloned the coding area sequence of the novel gene of s-Lap and constructed its recombinant eukaryotic plasmid pcDNA3.1-GFP/s-lap with the recombinant DNA technique. The expression and localization of s-lap/GFP fusion protein in CHO and B_~16 cell lines were studied with the instantaneously transfected pcDNA3.1-GFP/s-lap recombinant plasmid. ~s-Lap/GFP fusion protein can be expressed in CHO and B_~16 cells with a high rate expression in the nuclei.

INHIBITION OF APOPTOSIS BY bcr-abl FUSION GENE IN K562 CELLS

Objective: To investigate the effect of bcr-abl fusion gene on CML cell apoptosis. Methods: Apoptosis of ex-vivo cultured K562 cells were observed after exposure to synthetic 18 mer antisense oligodeoxynucleotide complementary to the bcr-abl junction (b3a2). Results: Apoptosis of K562 cells was significantly increased associated with inhibition of bcr-abl expression. Conclusion: bcr-abl fusion gene formation due to chromosome translocation may be the major mechanism of CML via inhibition of apoptosis.

Detection of TMPRSS2：ERG fusion gene in circulating prostate cancer cells

Aim： To investigate the existence of TMPRSS2：ERG fusion gene in circulating tumor cells （CTC） from prostate cancer patients and its potential in monitoring tumor metastasis. Methods- We analyzed the frequency of TMPRSS2： ERG and TMPRSS2：ETV1 transcripts in 27 prostate cancer biopsies from prostatectomies, and TMPRSS2：ERG transcripts in CTC isolated from 15 patients with advanced androgen independent disease using reverse transcription polymerase chain reaction （RT-PCR）. Fluorescence in situ hybridization （FISH） was applied to analyze the genomic truncation of ERG, which is the result of TMPRSS2：ERG fusion in 10 of the 15 CTC samples. Results： TMPRSS2： ERG transcripts were found in 44% of our samples, but we did not detect expression of TMPRSS2：ETV1. Using FISH analysis we detected chromosomal rearrangements affecting the ERG gene in 6 of 10 CTC samples, including 1 case with associated TMPRSS2：ERG fusion at the primary site. However, TMPRSS2：ERG transcripts were not detected in any of the 15 CTC samples, including the 10 cases analyzed by FISH. Conclusion： Although further study is required to address the association between TMPRSS2：ERG fusion and prostate cancer metastasis, detection of genomic truncation of the ERG gene by FISH analysis could be useful for monitoring the appearance of CTC and the potential for prostate cancer metastasis.

Expression of GST-IL-1 fusion gene

Two GST-IL-1 fusion genes were constructed by inserting different cDNA fragments of human interleukin1 (IL-1) into the 3'-terminus of GST gene in the fusion protein expression vector pGEX-4T. After IPTG induction ,SDS-PAGE was employed to detect the gene expression. No corresponding protein encoded by GST gene fused with the whole-length 816 bp IL-1 cDNA was observed, nor was free GST protein. However, the fusion protein of GST and IL-1 cDNA without the 189 bp at the 5'- terminus was detected, amounting to 30% of the total bacterial protein expressed. This might suggest that the sequence of 1-189 bp of IL-1 cDNA affected the expression of the fusion gene. That is to say, the downstream sequence distant from the translation start codon AUG in the target gene could significantly affect the expression of the fusion gene.

Profiling of gene fusion involving targetable genes in Chinese gastric cancer

BACKGROUND Approximately half of all new cases of gastric cancer(GC)and related deaths occur in China.More than 80%of patients with GC are diagnosed at an advanced stage,which results in poor prognosis.Although HER2-directed therapy and immune checkpoint inhibitors have been somewhat successful,new drugs are still needed for the treatment of GC.Notably,several gene fusion-targeted drugs have been approved by the United States Food and Drug Administration for solid tumors,including GC,such as larotrectinib for NTRK fusion-positive cancers and zenocutuzumab for NRG1 fusion-positive cancers.However,gene fusions involving targetable genes have not been well characterized in Chinese patients with GC.AIM To identify the profile of fusions involving targetable genes in Chinese patients with GC using clinical specimens and determine the distribution of patients with gene fusion variants among the molecular subtypes of GC.METHODS We retrospectively analyzed gene fusion events in tumor tissue samples from 954 Chinese patients with GC.Clinicopathological characteristics were obtained from their medical records.Genetic alterations,such as single nucleotide variants,indels,amplifications,and gene fusions,were identified using a targeted sequencing panel containing 825 genes.Fusions were validated by fluorescence in situ hybridization(FISH)using break-apart probes.The microsatellite instability(MSI)status was evaluated using MSIsensor from the targeted sequencing panel data.Tumor mutational burden(TMB)was calculated using the total number of nonsynonymous mutations divided by the total genomic targeted region.Chi-square analysis was used to determine the enrichment of gene fusions associated with the molecular subtypes of GC.RESULTS We found that 1.68%(16/954)of patients harbored 20 fusion events involving targetable genes.RARA fusions(n=5)were the most common,followed by FGFR2,BRAF,MET,FGFR3,RET,ALK,EGFR,NTRK2,and NRG1 fusions.Two of the RARA fusions,EML4-ALK(E6:E20)and EGFRSEPTIN14(E7:E10),have been identified in other tumors but not in GC.Surprisingly,18 gene fusion events were previously not reported in any cancer types.Twelve of the eighteen novel gene fusions included complete exons encoding functional domains of targetable genes,such as the tyrosine kinase domain of receptor tyrosine kinases and the DNA-and ligand-binding domains of RARA.Consistent with the results of detection using the targeted sequencing fusion panel,the results of FISH(fluorescence in situ hybridization)confirmed the rearrangement of FGFR2 and BRAF in tumors from patients 04 and 09,respectively.Genetic analysis indicated that the fusion genes were significantly enriched in patients with ERBB2 amplification(P=0.02);however,there were no significant differences between fusion-positive and fusion-negative patients in age,sex,MSI status,and TMB.CONCLUSION We characterized the landscape of fusions involving targetable genes in a Chinese GC cohort and found that 1.68%of patients with GC harbor potential targetable gene fusions,which were enriched in patients with ERBB2 amplification.Gene fusion detection may provide a potential treatment strategy for patients with GC with disease progression following standard therapy.

VISUALIZATION OF HEAD AND NECK CANCER MODELS WITH A TRIPLE FUSION REPORTER GENE

The development of experimental animal models for head and neck tumors generally rely on the biol uminescence imaging to achieve the dynamic monitoring of the tumor growth and metastasis due to the complicated anatomical structures.Since the bioluminescence imaging is largely affected by the intracellular luciferase expression level and external D-luciferin concentrations,its imaging accuracy requires further confirmation.Here,a new triple fusion reportelr gene,which consists of a herpes simplex virus type 1 thymidine kinase(TK)gene for radioactive imaging,a far-red fuorescent protein(mLumin)gene for fuorescent imaging,and a firefly luciferase gene for bioluminescence imaging,was introduced for in vrivo observation of the head and neck tumors through multi-modality imaging.Results show that fuorescence and bioluminescence signals from mLumin and luciferase,respectively,were clearly observed in tumor cells,and TK could activate suicide pathway of the cells in the presence of nucleotide analog-ganciclovir(GCV),demonstrating the effecti veness of individual functions of each gene.Moreover,subcutaneous and metastasis animal models for head and neck tumors using the fusion reporter gene-expressing cell lines were established,allowing multi-modality imaging in vio.Together,the established tumor models of head and neck cancer based on the newly developed triple fusion reporter gene are ideal for monitoring tumor growth,assessing the drug therapeutic efficacy and verifying the effec-tiveness of new treatments.

Secretory carcinoma——impact of translocation and gene fusions on salivary gland tumor

Secretory carcinoma（SC）, previously described as mammary analogue secretory carcinoma（MASC）, is a recently described salivary gland tumor which morphologically resembles mammary secretory carcinoma. The first description of SC/MASC, reported by Skálová et al. in 2010, was as a rare salivary carcinoma imitating secretory carcinoma of the breast. SC/MASC is a unique salivary gland tumor with morphological overlap with acinic cell carcinoma（Aci CC）, mucoepidermoid carcinoma（MEC）, and adenocarcinoma not otherwise specified（ADCNOS）. SC/MASC shares similar clinicopathological features with Aci CC. As a critical difference between SC/MASC and Aci CC, SC/MASC characteristically has the chromosomal translocation t（12;15）（p13;q25） which leads to a fusion gene between the ETV6 gene on chromosome 12 and the NTRK3 gene on chromosome 15. This genetic background is an important differential diagnostic finding for excluding other salivary gland tumors and may be a critical factor determining the prognosis for patients with SC/MASC. Research in recent years has provided a large body of new data on SC/MASC and suggests the possibility that the ETV6-NTRK3 translocation could be a therapeutic target. Here, we review the morphological and clinicopathological features of SC/MASC and discuss new directions for therapy.

Construction of the recombinant expression vector for CD80-IgG fusion gene and its expression in Chinese hamster ovary cells

To construct the recombinant expression functionally in Chinese hamster ovary cells in order vector for CD80-IgG fusion gene and to express it to be used as an effective method to eliminate the immune escape of leukemic cells, the cDNA encoding the signal and extracellular domains of murine CD80 was generated by PCR amplification from plasmid pcDNMB7 containing the full length cDNA of murine CD80 and those of murine IgG1, in which the Fc fragment was obtained through RT-PCR amplification from murine spleen cells. These two cDNAs were then cloned in tandem into eukaryotic expression vector pcDNA3.0 and the resultant recombinant plasmid pcDNA/CD80-IgG was then transfected to Chinese hamster ovary cells with liposome transfection reagent. The cell clones constitutively expressing CD80-IgG fusion protein were obtained by G418 screening. Western blotting and dot ELISA assay were used to detect the expression of the fusion protein in the supernatants of these cells. Meanwhile, the fusion protein expressed was then purified with affinity chromatography, and its biological activity was demonstrated by flow cytometry, MTr colorimetry and ELISA assay. The experimental resuits showed that these two inserts were successfully cloned into plasmid pcDNA3.0, and the highly purified fusion protein was obtained. This fusion protein was proved to be able to upregulate the density of CD80 on leukemic cells, deliberately promote the proliferative reactions of mouse allogenic lymphocytes and increase the killing activity against WEHI-3 cells from 49.7 % up to 84.6 %. In addition, this fusion protein could also enhance the IL-2 secretion from allogenic lymphocytes activated by tumorspecific antigens. It is concluded that the recombinant vector constructed can be functionally expressed in the mammalian cells, thus providing a solid foundation for the further investigation on the mechanism to eliminate the immune escape of leukemic cells in vivo.