Long-Term Remission in T315I+ Relapsed Philadelphia Chromosome-Positive Acute Lymphoblastic Leukemia with Blinatumomab and Allogeneic Stem Cell Transplantation: Two Case Studies.

BACKGROUND Philadelphia chromosome-positive acute lymphoblastic leukemia (Ph+ ALL) in adults has a poor prognosis with conventional chemotherapy. Treatment with tyrosine kinase inhibitors (TKIs) followed by allogeneic hematopoietic stem cell transplantation (allo-HSCT) has improved clinical outcomes; however, the relapse rate is still high. Therapeutic options for patients with relapsed/refractory (R/R) Ph+ ALL are scarce, with very few studies focusing on these patients. Blinatumomab is a novel bispecific T-cell engager antibody construct showing promising efficacy in R/R Ph+ ALL. CASE REPORT Here, we present 2 cases of relapsed Ph+ ALL with T315I mutation refractory to multiple TKIs and chemotherapy. Patient 1 was a 48-year-old woman who had increased leukocytes in her peripheral blood cells, with 90% abnormal cells and decreased platelets. Bone marrow (BM) smear showed 95% blasts. Patient 2 was a 20-year-old man who had leukocytosis with thrombocytopenia, while all other parameters were normal. BM aspirate showed 98% immature granulocytes/blasts. The immunophenotypic observations of both the patients on BM were consistent with the presence of ALL. Both patients were effectively treated with a combination of blinatumomab and allo-HSCT and achieved complete remission in 1 month with minimal residual disease negativity and remained in remission for a long period. CONCLUSIONS The findings suggest, that for patients with R/R Ph+ ALL with T315I mutation who respond poorly to TKIs, salvage therapy with blinatumomab is a potentially effective treatment for improving clinical outcomes. The treatment with blinatumomab can further act as a bridge to HSCT in these patients, helping them to attain deeper remission.

Identification and validation of a platelet-related signature for predicting survival and drug sensitivity in multiple myeloma.

Background: Significant progress has been achieved in the management of multiple myeloma (MM) by implementing high-dose therapy and stem cell transplantation. Moreover, the prognosis of patients has been enhanced due to the introduction of novel immunomodulatory drugs and the emergence of new targeted therapies. However, predicting the survival rates of patients with multiple myeloma is still tricky. According to recent researches, platelets have a significant impact in affecting the biological activity of tumors and are essential parts of the tumor microenvironment. Nonetheless, it is still unclear how platelet-related genes (PRGs) connect to the prognosis of multiple myeloma. Methods: We analyzed the expression of platelet-related genes and their prognostic value in multiple myeloma patients in this study. We also created a nomogram combining clinical metrics. Furthermore, we investigated disparities in the biological characteristics, immunological microenvironment, and reaction to immunotherapy, along with analyzing the drug susceptibility within diverse risk groups. Results: By using the platelet-related risk model, we were able to predict patients' prognosis more accurately. Subjects in the high-risk cohort exhibited inferior survival outcomes, both in the training and validation datasets, as compared to those in the low-risk cohort (p < 0.05). Moreover, there were differences in the immunological microenvironments, biological processes, clinical features, and chemotherapeutic drug sensitivity between the groups at high and low risk. Using multivariable Cox regression analyses, platelet-related risk score was shown to be an independent prognostic influence in MM (p < 0.001, hazard ratio (HR) = 2.001%, 95% confidence interval (CI): 1.467-2.730). Furthermore, the capacity to predict survival was further improved when a combined nomogram was utilized. In training cohort, this outperformed the predictive value of International staging system (ISS) alone from a 5-years area under curve (AUC) = 0.668 (95% CI: 0.611-0.725) to an AUC = 0.721 (95% CI: 0.665-0.778). Conclusion: Our study revealed the potential benefits of PRGs in terms of survival prognosis of MM patients. Furthermore, we verified its potential as a drug target for MM patients. These findings open up novel possibilities for prognostic evaluation and treatment choices for MM.

Prognostic significance of ß2-microglobulin decline index in multiple myeloma.

Purpose: To assess the prognostic significance of ß2-microglobulin decline index (ß2M DI) in multiple myeloma (MM). Methods: 150 MM patients diagnosed with MM were enrolled in this study. Cox proportional hazards model was used to analyze the uni- and multivariate prognosis in training cohort (n=105). A new combined prognostic model containing ß2M DI was built up based on the data in training cohort. The validation group was used to verify the model. Results: ß2M DI showed significant correlation with prognosis in both uni- and multivariate analyses and had a good correlation with complete response (CR) rate and deep remission rate. The ROC and calibration curves in validation cohort (n=45) indicated a good predictive performance of the new model. Based on the median risk score of the training group, we classified patients into high- and low- risk groups. In both training and validation groups, patients in the low-risk group had longer overall survival (OS) time than that in the high-risk group (p<0.05). Conclusion: ß2M DI is a good predictive index for predicting treatment response and survival time in MM patients. The prognostic model added with ß2M DI showed a better correlation with OS.

Development and validation of a model for the early prediction of progression from essential thrombocythemia to post-essential thrombocythemia myelofibrosis: a multicentre retrospective study.

Background: Essential thrombocythemia (ET), a myeloproliferative neoplasm (MPN), has a substantial risk of evolving into post-essential thrombocythemia myelofibrosis (post-ET MF). This study aims to establish a prediction nomogram for early prediction of post-ET MF in ET patients. Methods: The training cohort comprised 558 patients from 8 haematology centres between January 1, 2010, and May 1, 2023, while the external validation cohort consisted of 165 patients from 6 additional haematology centres between January 1, 2010, and May 1, 2023. Univariable and multivariable Cox regression analysis was performed to identified independent risk factors and establish a nomogram to predict the post-ET MF free survival. Both bias-corrected area under the curve (AUC), calibration curves and concordance index (C-index) were employed to assess the predictive accuracy of the nomogram. Findings: Multivariate Cox regression demonstrated that elevated red blood cell distribution width (RDW), elevated levels of lactate dehydrogenase (LDH) and the level of haemoglobin (Hb), a history of smoking and the presence of splenomegaly were independent risk factors for post-ET MF. The C-index displayed of the training and validation cohorts were 0.877 and 0.853. The 5 years, 10 years AUC values in training and external validation cohorts were 0.948, 0.769 and 0.978, 0.804 respectively. Bias-corrected curve is close to the ideal curve and revealed a strong consistency between actual observation and prediction. Interpretation: We developed a nomogram capable of predicting the post-ET MF free survival probability at 5 years and 10 years in ET patients. This tool helps doctors identify patients who need close monitoring and appropriate counselling. Funding: This research was funded by the Key R&D Program of Zhejiang (No. 2022C03137); the Public Technology Application Research Program of Zhejiang, China (No. LGF21H080003); and the Zhejiang Medical Association Clinical Medical Research special fund project (No. 2022ZYC-D09).

Comprehensive analysis of ferroptosis-related genes in immune infiltration and prognosis in multiple myeloma.

Background: One particular type of cellular death that is known as ferroptosis is caused by the excessive lipid peroxidation. It is a regulated form of cell death that can affect the response of the tumor cells. Currently, it is not known if the presence of this condition can affect the prognosis of patients with multiple myeloma (MM). Methods: In this study, we studied the expression differences and prognostic value of ferroptosis-related genes (FRGs) in MM, and established a ferroptosis risk scoring model. In order to improve the prediction accuracy and clinical applicability, a nomogram was also established. Through gene enrichment analysis, pathways closely related to high-risk groups were identified. We then explored the differences in risk stratification in drug sensitivity and immune patterns, and evaluated their value in prognostic prediction and treatment response. Lastly, we gathered MM cell lines and samples from patients to confirm the expression of marker FRGs using quantitative real-time PCR (qRT-PCR). Results: The ability to predict the survival of MM patients is a challenging issue. Through the use of a risk model derived from ferroptosis, we were able to develop a more accurate prediction of the disease's prognosis. They were then validated by a statistical analysis, which showed that the model is an independent factor in the prognosis of MM. Patients of high ferroptosis risk scores had a much worse chance of survival than those in the low-risk groups. The calibration and power of the nomogram were also strong. We noted that the link between the ferroptosis risk score and the clinical treatment was suggested by the FRG's significant correlation with the immune checkpoint genes and the medication sensitivity. We validated the predictive model using qRT-PCR. Conclusion: We demonstrated the association between FRGs and MM, and developed a new risk model for prognosis in MM patients. Our study sheds light on the potential clinical relevance of ferroptosis in MM and highlights its potential as a therapeutic target for patients with this disease.

A novel glycolysis-related gene signature for predicting the prognosis of multiple myeloma.

Background: Metabolic reprogramming is an important hallmark of cancer. Glycolysis provides the conditions on which multiple myeloma (MM) thrives. Due to MM's great heterogeneity and incurability, risk assessment and treatment choices are still difficult. Method: We constructed a glycolysis-related prognostic model by Least absolute shrinkage and selection operator (LASSO) Cox regression analysis. It was validated in two independent external cohorts, cell lines, and our clinical specimens. The model was also explored for its biological properties, immune microenvironment, and therapeutic response including immunotherapy. Finally, multiple metrics were combined to construct a nomogram to assist in personalized prediction of survival outcomes. Results: A wide range of variants and heterogeneous expression profiles of glycolysis-related genes were observed in MM. The prognostic model behaved well in differentiating between populations with various prognoses and proved to be an independent prognostic factor. This prognostic signature closely coordinated with multiple malignant features such as high-risk clinical features, immune dysfunction, stem cell-like features, cancer-related pathways, which was associated with the survival outcomes of MM. In terms of treatment, the high-risk group showed resistance to conventional drugs such as bortezomib, doxorubicin and immunotherapy. The joint scores generated by the nomogram showed higher clinical benefit than other clinical indicators. The in vitro experiments with cell lines and clinical subjects further provided convincing evidence for our study. Conclusion: We developed and validated the utility of the MM glycolysis-related prognostic model, which provides a new direction for prognosis assessment, treatment options for MM patients.

Identification and validation of a novel cuproptosis-related gene signature in multiple myeloma.

Background: Cuproptosis is a newly identified unique copper-triggered modality of mitochondrial cell death, distinct from known death mechanisms such as necroptosis, pyroptosis, and ferroptosis. Multiple myeloma (MM) is a hematologic neoplasm characterized by the malignant proliferation of plasma cells. In the development of MM, almost all patients undergo a relatively benign course from monoclonal gammopathy of undetermined significance (MGUS) to smoldering myeloma (SMM), which further progresses to active myeloma. However, the prognostic value of cuproptosis in MM remains unknown. Method: In this study, we systematically investigated the genetic variants, expression patterns, and prognostic value of cuproptosis-related genes (CRGs) in MM. CRG scores derived from the prognostic model were used to perform the risk stratification of MM patients. We then explored their differences in clinical characteristics and immune patterns and assessed their value in prognosis prediction and treatment response. Nomograms were also developed to improve predictive accuracy and clinical applicability. Finally, we collected MM cell lines and patient samples to validate marker gene expression by quantitative real-time PCR (qRT-PCR). Results: The evolution from MGUS and SMM to MM was also accompanied by differences in the CRG expression profile. Then, a well-performing cuproptosis-related risk model was developed to predict prognosis in MM and was validated in two external cohorts. The high-risk group exhibited higher clinical risk indicators. Cox regression analyses showed that the model was an independent prognostic predictor in MM. Patients in the high-risk group had significantly lower survival rates than those in the low-risk group (p < 0.001). Meanwhile, CRG scores were significantly correlated with immune infiltration, stemness index and immunotherapy sensitivity. We further revealed the close association between CRG scores and mitochondrial metabolism. Subsequently, the prediction nomogram showed good predictive power and calibration. Finally, the prognostic CRGs were further validated by qRT-PCR in vitro. Conclusion: CRGs were closely related to the immune pattern and self-renewal biology of cancer cells in MM. This prognostic model provided a new perspective for the risk stratification and treatment response prediction of MM patients.

A review of the therapeutic role of the new third-generation TKI olverembatinib in chronic myeloid leukemia.

Several tyrosine kinase inhibitors (TKIs) have been developed as targeted therapies to inhibit the oncogenic activity of several tyrosine kinases in chronic myeloid leukemia (CML), acute lymphoid leukemia (ALL), gastrointestinal stromal tumor (GIST), and other diseases. TKIs have significantly improved the overall survival of these patients and changed the treatment strategy in the clinic. However, approximately 50% of patients develop resistance or intolerance to imatinib. For second-generation TKIs, approximately 30%-40% of patients need to change therapy by 5 years when they are used as first-line treatment. Clinical study analysis showed that the T315I mutation is highly associated with TKI resistance. Developing new drugs that target the T315I mutation will address the dilemma of treatment failure. Olverembatinib, as a third-generation TKI designed for the T315I mutation, is being researched in China. Preliminary clinical data show the safety and efficacy in treating CML patients harboring the T315I mutation or who are resistant to first- or second-line TKI treatment. Herein, we review the characteristics and clinical trials of olverembatinib. We also discuss its role in the management of CML patients.

Pneumocystis jirovecii pneumonia in non-Hodgkin's lymphoma after rituximab-based chemotherapy: a case series.

Background: The incidence of Pneumocystis jirovecii pneumonia (PCP) has been increasing in patients with hematologic malignancies due to the use of glucocorticoid therapy and immunosuppressive medication. The reports of PCP in non-Hodgkin's lymphoma (NHL) after rituximab-based chemotherapy are still rare. We reported a case series of PCP in NHL to show the clinical features and prognosis in those patients. Methods: We conducted a retrospective review of 15 NHL patients who developed PCP after rituximab-based chemotherapy during June 30, 2014 to June 1, 2020. We analyzed the laboratory and radiographic findings for those patients through descriptive statistics analysis. Results: The study revealed that PCP in NHL patients was complicated by chemotherapy after about 4 courses (range, 2 to 6 courses). Most patients had a standard lymphocyte count before treatment, and 14 of 15 patients (93.3%) had lymphopenia at the time of diagnosis of PCP. In addition to typical symptoms such as fever and dyspnea at the diagnosis of PCP, most patients had abnormal laboratory indexes such as marked elevations of C-reactive protein (CRP) and lactic dehydrogenase (LDH) both before and at the time of diagnosis. The (1,3)-ß-D-glucan test was also revealed as a sensitive index for PCP. Bilateral ground-glass opacity was detected in 14 cases through computed tomography (CT) scans. Positive results of microbiological testing were observed in 7 cases; sputum culture was positive in 3 and next-generation sequencing (NGS) was positive in 3 of these 7 patients, and the other case was positive in both sputum culture and NGS. Patients received high-dose trimethoprim/sulfamethoxazole (TMP/SMZ), caspofungin, and steroids as the treatment for PCP. Ventilatory support was required by 3 patients, so they were admitted to the intensive care unit (ICU), and 1 patient died from PCP. Conclusions: Dynamic monitoring of CRP, LDH, and (1,3)-ß-D-glucan test during the treatment of NHL may have a predictive value for the diagnosis of PCP. Additionally, we should use NGS as a rapid and sensitive method for the early diagnosis of PCP. When patients are classified as 'probable PCP', early and effective treatment has obvious significance to improve the prognosis.

[Toxicity Management and Efficacy Evaluation of BCMA-CART in the Treatment of Relapsed and Refractory Multiple Myeloma].

OBJECTIVE: To investigate the toxicity management and efficacy evaluation of BCMA-chimeric antigen receptor T cells(CART) in the treatment of relapsed and refractory multiple myeloma (MM). METHODS: The efficacy and adverse reactions of 21 patients with MM who received BCMA-CART treatment at the First Affiliated Hospital of Wenzhou Medical University from December 2017 to September 2020 were evaluated, and the efficacy assessment and survival analysis for high-risk patients and non-high-risk patients were evaluated. RESULTS: After infusion of BCMA-CART cells in 21 MM patients, the number of effective cases was 17, of which the complete remission (sCR/CR) was 10, and the partial remission (VGPR/PR) was 7. The median OS time for all patients was 19.4 months, and the median PFS time was 7.9 months. The number of patients with extramedullary disease(EMD), high-risk genetics, and ISS stage Ⅲ were 5, 15 and 8, and the effective number was 3, 11 and 6, respectively. The treatment of 3 patients without high-risk factors was effective. The median OS and median PFS of patients with EMD were 14.2 and 2.5 months, respectively, which were shorter than those of patients without EMD (19.4 months and 8.9 months, respectively). The median OS and median PFS of patients with high-risk cytogenetic factors and ISS Ⅲ were not significantly different from those of non-high-risk patients. Cytokine release syndrane (CRS) occurred in 20 patients, of which 14 cases were Grade 1 CRS, while 6 were Grade 2, no CRS of Grade 3 or above occurred. IL-6 receptor inhibitors were used in 9 patients. All CRS were controlled effectively, and no patients had neurological toxicity. CONCLUSION: BCMA-CART is a certain curative effect in the treatment of relapsed and refractory multiple myeloma, and the adverse reactions can be well controlled through close monitoring and timely treatment.

Maintenance therapy with all-trans retinoic acid and arsenic trioxide improves relapse-free survival in adults with low- to intermediate-risk acute promyelocytic leukemia who have achieved complete remission after consolidation therapy.

BACKGROUND: Currently, the optimal maintenance therapy for patients with acute promyelocytic leukemia (APL) who have achieved complete remission (CR) after completing consolidation chemotherapy remains controversial. The comparative effectiveness of the all-trans retinoic acid (ATRA) plus arsenic trioxide (As2O3) maintenance strategy with classic ATRA plus chemotherapy has not been evaluated. In this study, we compared the efficacy and toxicity of maintenance therapy with ATRA plus As2O3 and classic ATRA plus chemotherapy in low- to intermediate-risk APL patients reaching the first CR after induction and consolidation therapy. METHODS: A retrospective review of 58 adult patients diagnosed with APL was conducted. After receiving consolidation therapy and achieving CR, 30 patients were administered maintenance therapy with an ATRA plus As2O3 regimen (ATRA+As2O3 group), whereas 28 patients were administered 3-monthly cycles of an ATRA plus chemotherapy regimen (ATRA+chemotherapy group). RESULTS: Grade 3-4 neutropenia was significantly more frequent in the ATRA+chemotherapy group (N=9, 32.1%) than in the ATRA+As2O3 group (N=0) (P=0.001). At a median follow-up of 49.1 months (range: 9.7-97.4 months) from the completion of consolidation, no relapses were observed in the ATRA+As2O3 group, whereas seven relapses occurred in the ATRA+chemotherapy group. The risk of relapse in the patients administered ATRA+As2O3 maintenance was significantly lower than that in those administered ATRA+chemotherapy maintenance (P=0.004). Based on log-rank analysis, only maintenance therapy with ATRA and As2O3 was associated with a significantly higher relapse-free survival (P=0.0159). CONCLUSION: Maintenance therapy with ATRA and As2O3 was beneficial in low- to intermediate-risk APL patients who were effectively treated to achieve CR. Further clinical trials with reliable designs are needed to confirm these observations.

[Effect of hydroquinone on expression of topoisomerase enzyme IIα in human bone marrow mononuclear cells].

OBJECTIVE: To investigate the effects of hydroquinone (HQ) on expression of topoisomerase IIα (TOPOIIα) in human bone marrow mononuclear cells, and to explore the role and possible regulatory mechanism of TOPOIIα involved in toxicity of HQ to hematopoietic cells. METHODS: After human bone marrow mononuclear cells were exposed to 50 µmol/L HQ (used the cells which were exposed to sterile distilled water as control); the activity of TOPOII was measured by TOPOII assay kit; the expression levels of TOPOIIα mRNA and protein were detected by RT-PCR technique and Western blotting method respectively; the chromatin immunoprecipitation (ChIP) assay was carried out to study the possible mechanism of TOPOIIα expression changes. RESULTS: (1) The activity of TOPOII was inhibited obviously; the protein and mRNA expression of TOPOIIα were 0.017 ± 0.029 and 0.610 ± 0.128, significantly lower than that in the control with the significant difference (P < 0.01) after treated with HQ for 10 h; (2) The decreased content of TOPOIIα was associated with descended level of histone H4 acetylation than in the control, from 1.198 ± 0.056 to 0.324 ± 0.229, with the significant difference (P < 0.01), without accompanied descended level of histone H3 acetylation, from 1.253 ± 0.045 to 1.177 ± 0.025 (P > 0.05); (3) TOPOIIα mRNA expression decreased gradually after HQ processing, and the chemical modification (histone H4 acetylation) of TOPOIIα promoter happened prior to the mRNA expression. CONCLUSION: HQ could repress the expression of TOPOIIα in human bone marrow mononuclear cells; the change of histone chemical modification plays an important role in the benzene's hematopoietic toxicity.

[Anti-cD20scFv/CD80/CD28/zeta specific T lymphocytes eradicate primary chronic lymphocytic leukemia cells in vitro].

OBJECTIVE: To construct anti-CD20scFv/CD80/CD28/zeta recombinant gene modified T cells, test its effectiveness of eradicating CD20 positive primary chronic lymphocytic leukemia (CLL) cells and provide a promising tool for tumor adoptive immunotherapy. METHODS: The recombinant vectors were transduced into PA 317 cells and high titer retroviruses were obtained to infect human peripheral blood T lymphocytes. Resistant T cells were obtained by G418 selection for one week. Then transduced T lymphocytes and primary CLL cells were co-cultured. The status of primary chronic lymphocytic leukemia cells were observed by microscope. The level of IL-2 and IFN-gamma in the culture medium were measured. RESULTS: Primary T cells expressing anti-CD20scFv/IgGFc/CD80/CD28/zeta could be constructed successfully. These T cells were able to lyse CD20+ targets and secrete high levels of IL-2 (1301.00 pg/ml) and IFN-gamma (602.18 pg/ml) in vitro. CONCLUSION: (1) Recombinant gene modified T cells can be constructed successfully. (2) Recombinant gene modified T cells can specially kill CD20 positive primary CLL cells in vitro.

Immunotherapy of lymphomas with T cells modified by anti-CD20 scFv/CD28/CD3zeta recombinant gene.

One of the approaches to make anti-CD20 antibody more efficient is to express this antibody on the surface of T cells. scFv from anti-CD20 antibody has been expressed on T cell surface to bind to CD20 positive cells and CD3zeta has been expressed as a fusion partner to transduct signals. T cells grafted with this chimeric scFv/CD3zeta were able to redirect grafted T cells to an MHC/Ag-independent antitumor response. To test the effects of CD28 signal on the cellular activation and antitumor effectiveness of chimeric scFv/CD3zeta modified T cells, we constructed a recombinant anti-CD20 scFv/CD28/CD3zeta gene in a retroviral vector. T cells expressing anti-CD20 scFv/CD28/CD3zeta specifically lysed CD20 positive target tumor cells and secreted not only IFN-gamma but also IL-2 after binding to their target cells. Our data indicate that CD3 and CD28 signalling can be delivered in one molecule, which is sufficient for complete T cell activation without exogenous B7/CD28 costimulation.

[Construction of recombinant retroviruses expressing anti-CD20 scFv/CD80 /CD28/zeta gene and expression in Jurkat cell line].

AIM: To construct recombinant retroviruses expressing anti-CD20 scFv/CD80/CD28/zeta gene and detect its expression in Jurkat cells. METHOD: CD28-zetacDNA were amplified from plasmids pBULLET and inserted into pLNCX vector that contained anti-CD20 scFv/CD80 gene. The recombinant plasmids were transfected into PA 317 cells. Retroviruses were harvested from culture medium of PA 317 cells. Then NIH 3T3 were transfected with retroviruses. Objective gene expression was determined by PCR and FACS. Jurkat cells were transfected with high titer of retroviruses and resistant clones were obtained by G418 selection. Objective mRNA was determined by RT- PCR. RESULTS: The recombinant eukaryotic vector was constructed successfully by PCR and enzyme digestion analysis and objective gene was amplified from NIH 3T3 cells transfected with retroviruses by PCR; FACS showed that objective protein could be expressed in NIH 3T3 cells. Objective gene was amplified from Jurkat cells transfected with retroviruses by RT-PCR. CONCLUSION: Recombinant retrovirus expressing anti-CD20 scFv/CD80/CD28/zeta gene was successfully constructed and objective protein could be expressed in Jurkat cells.

[Construction of anti-cD20scFv/CD80/CD28/zeta recombinant gene modified T cell and research on its targeting cytotoxicity].

OBJECTIVE: To construct anti-CD20scFv/CD80/CD28/zeta recombinant gene modified T cells, test its effectiveness of eradicating CD20+ lymphoma cells and provide a probably new approach to tumor adoptive immunotherapy. METHODS: CD28-zeta cDNA were amplified from vector pBULLET and inserted into pLNCX vector that contained anti-CD20scFv/CD80 gene. The recombinant vectors were transduced into PA317 cells and high titer retroviruses were obtained to infect human peripheral blood T lymphocytes. Resistant T cells were obtained by G418 selection at one week. Then transduced T lymphocytes and lymphoma cell lines Daudi Raji were cocultured. The cytotoxicity and cytokine production of transduced T cells were determined by non-radio-activation cytotoxicity assay and ELISA respectively. RESULTS: The recombinant eukaryotic vector was constructed successfully as proved by enzyme digestion analysis and sequencing. These T cells were able to lyse CD20+ target cells and secrete high levels of IL-2 and IFN-gamma in vitro. CONCLUSION: Recombinant gene modified T cells can be constructed successfully. It can specially kill CD20 positive lymphoma cells in vitro.