NanoStringDiff: a novel statistical method for differential expression analysis based on NanoString nCounter data.

The advanced medium-throughput NanoString nCounter technology has been increasingly used for mRNA or miRNA differential expression (DE) studies due to its advantages including direct measurement of molecule expression levels without amplification, digital readout and superior applicability to formalin fixed paraffin embedded samples. However, the analysis of nCounter data is hampered because most methods developed are based on t-tests, which do not fit the count data generated by the NanoString nCounter system. Furthermore, data normalization procedures of current methods are either not suitable for counts or not specific for NanoString nCounter data. We develop a novel DE detection method based on NanoString nCounter data. The method, named NanoStringDiff, considers a generalized linear model of the negative binomial family to characterize count data and allows for multifactor design. Data normalization is incorporated in the model framework through data normalization parameters, which are estimated from positive controls, negative controls and housekeeping genes embedded in the nCounter system. We propose an empirical Bayes shrinkage approach to estimate the dispersion parameter in the model and a likelihood ratio test to identify differentially expressed genes. Simulations and real data analysis demonstrate that the proposed method performs better than existing methods.

Mutant IDH1 and thrombosis in gliomas.

Mutant isocitrate dehydrogenase 1 (IDH1) is common in gliomas, and produces D-2-hydroxyglutarate (D-2-HG). The full effects of IDH1 mutations on glioma biology and tumor microenvironment are unknown. We analyzed a discovery cohort of 169 World Health Organization (WHO) grade II-IV gliomas, followed by a validation cohort of 148 cases, for IDH1 mutations, intratumoral microthrombi, and venous thromboemboli (VTE). 430 gliomas from The Cancer Genome Atlas were analyzed for mRNAs associated with coagulation, and 95 gliomas in a tissue microarray were assessed for tissue factor (TF) protein. In vitro and in vivo assays evaluated platelet aggregation and clotting time in the presence of mutant IDH1 or D-2-HG. VTE occurred in 26-30 % of patients with wild-type IDH1 gliomas, but not in patients with mutant IDH1 gliomas (0 %). IDH1 mutation status was the most powerful predictive marker for VTE, independent of variables such as GBM diagnosis and prolonged hospital stay. Microthrombi were far less common within mutant IDH1 gliomas regardless of WHO grade (85-90 % in wild-type versus 2-6 % in mutant), and were an independent predictor of IDH1 wild-type status. Among all 35 coagulation-associated genes, F3 mRNA, encoding TF, showed the strongest inverse relationship with IDH1 mutations. Mutant IDH1 gliomas had F3 gene promoter hypermethylation, with lower TF protein expression. D-2-HG rapidly inhibited platelet aggregation and blood clotting via a novel calcium-dependent, methylation-independent mechanism. Mutant IDH1 glioma engraftment in mice significantly prolonged bleeding time. Our data suggest that mutant IDH1 has potent antithrombotic activity within gliomas and throughout the peripheral circulation. These findings have implications for the pathologic evaluation of gliomas, the effect of altered isocitrate metabolism on tumor microenvironment, and risk assessment of glioma patients for VTE.

The tumor suppressor prostate apoptosis response-4 (Par-4) is regulated by mutant IDH1 and kills glioma stem cells.

Prostate apoptosis response-4 (Par-4) is an endogenous tumor suppressor that selectively induces apoptosis in a variety of cancers. Although it has been the subject of intensive research in other cancers, less is known about its significance in gliomas, including whether it is regulated by key driver mutations, has therapeutic potential against glioma stem cells (GSCs), and/or is a prognostic marker. We found that patient-derived gliomas with mutant isocitrate dehydrogenase 1 have markedly lower Par-4 expression (P < 0.0001), which was validated by The Cancer Genome Atlas dataset (P = 2.0 E-13). The metabolic product of mutant IDH1, D-2-hydroxyglutarate (2-HG), can suppress Par-4 transcription in vitro via inhibition of promoter activity as well as enhanced mRNA degradation, but interestingly not by direct DNA promoter hypermethylation. The Selective for Apoptosis induction in Cancer cells (SAC) domain within Par-4 is highly active against glioma cells, including orthotopic xenografts of patient-derived primary GSCs (P < 0.0001). Among high-grade gliomas that are IDH1 wild type, those that express more Par-4 have significantly longer median survival (18.4 vs. 8.0 months, P = 0.002), a finding confirmed in two external GBM cohorts. Together, these data suggest that Par-4 is a significant component of the mutant IDH1 phenotype, that the activity of 2-HG is complex and can extend beyond direct DNA hypermethylation, and that Par-4 is a promising therapeutic strategy against GSCs. Furthermore, not every effect of mutant IDH1 necessarily contributes to the overall favorable prognosis seen in such tumors; inhibition of Par-4 may be one such effect.

Autophagy and oxidative stress in gliomas with IDH1 mutations.

IDH1 mutations in gliomas associate with longer survival. Prooxidant and antiproliferative effects of IDH1 mutations and its D-2-hydroxyglutarate (2-HG) product have been described in vitro, but inconsistently observed. It is also unclear whether overexpression of mutant IDH1 in wild-type cells accurately phenocopies the effects of endogenous IDH1-mutations on tumor apoptosis and autophagy. Herein we investigated the effects of 2-HG and mutant IDH1 overexpression on proliferation, apoptosis, oxidative stress, and autophagy in IDH1 wild-type glioma cells, and compared those results with patient-derived tumors. 2-HG reduced viability and proliferation of U87MG and LN18 cells, triggered apoptosis in LN18 cells, and autophagy in U87MG cells. In vitro studies and flank xenografts of U87MG cells overexpressing R132H IDH1 exhibited increased oxidative stress, including increases of both manganese superoxide dismutase (MnSOD) and p62. Patient-derived IDH1-mutant tumors showed no significant differences in apoptosis or autophagy, but showed p62 accumulation and actually trended toward reduced MnSOD expression. These data indicate that mutant IDH1 and 2-HG can induce oxidative stress, autophagy, and apoptosis, but these effects vary greatly according to cell type.

Association between serum albumin concentration change trajectory and risk of hypertension: a cohort study in China.

Background: We sought to assess the risk of hypertension based on the trajectory of changes in serum albumin concentrations. Methods: A total of 11,946 nonhypertension adults aged 30-60 years who underwent at least 3 medical examinations between 2009 and 2016 were included in this study. Group-based trajectory models were obtained for 4 category groups, and logistic regression models were used to estimate odds ratios (ORs) and 95% confidence intervals (CIs) for each category group of serum albumin concentration and the risk of hypertension. Results: During a mean follow-up period of 4.30 years, 1,537 hypertension events occurred in 11,946 subjects without hypertension. A high stable trajectory of serum albumin concentrations (OR, 0.70, 95% CI, 0.51-0.96) was associated with a significantly lower risk of developing hypertension. The results of the sensitivity analysis of the high stable trajectory (OR, 0.64, 95% CI, 0.43-0.96) remained statistically significant. Subjects with normal weight and those ≥45 years of age had a significantly lower risk of hypertension at moderate increase (P = 0.053 or 0.026) and high stable trajectories (P = 0.011 or 0.016). In males and overweight subjects, the risk of hypertension was significantly lower in the high stable trajectory (P = 0.038 or 0.044). Conclusion: In this study, we found that moderate increase in serum albumin concentrations and a high stable trajectory were significantly associated with a reduced risk of hypertension in subjects aged ≥45 years and those with normal weight and that high stable serum albumin concentrations were significantly associated with a reduced risk of hypertension in males and overweight subjects.

BACE2 expression increases in human neurodegenerative disease.

ß-Secretase, the rate-limiting enzymatic activity in the production of the amyloid-ß (Aß) peptide, is a major target of Alzheimer's disease (AD) therapeutics. There are two forms of the enzyme: ß-site Aß precursor protein cleaving enzyme (BACE) 1 and BACE2. Although BACE1 increases in late-stage AD, little is known about BACE2. We conducted a detailed examination of BACE2 in patients with preclinical to late-stage AD, including amnestic mild cognitive impairment, and age-matched controls, cases of frontotemporal dementia, and Down's syndrome. BACE2 protein and enzymatic activity increased as early as preclinical AD and were found in neurons and astrocytes. Although the levels of total BACE2 mRNA were unchanged, the mRNA for BACE2 splice form C (missing exon 7) increased in parallel with BACE2 protein and activity. BACE1 and BACE2 were strongly correlated with each other at all levels, suggesting that their regulatory mechanisms may be largely shared. BACE2 was also elevated in frontotemporal dementia but not in Down's syndrome, even in patients with substantial Aß deposition. Thus, expression of both forms of ß-secretase are linked and may play a combined role in human neurologic disease. A better understanding of the normal functions of BACE1 and BACE2, and how these change in different disease states, is essential for the future development of AD therapeutics.

Association between hyperuricemia and hypertension and the mediatory role of obesity: a large cohort study in China.

OBJECTIVE: This study aimed to investigate the sex-specific association between hyperuricemia and the risk of hypertension and whether obesity mediates this association. METHODS: This study included 31,395 (47.0% women) adults without hypertension, cardiovascular disease, or cancer at baseline who completed at least one follow-up annual examination between 2009 and 2016. Cox regression models were performed to calculate hazard ratios and 95% confidence intervals. Mediation analysis was conducted to estimate the effect of body mass index on the association between hyperuricemia and hypertension. RESULTS: During a median 2.9-year follow-up, hyperuricemia was significantly associated with a higher risk of hypertension (HR 1.15, 95%CI 1.07-1.24 for all participants; HR 1.12, 95%CI 1.03-1.22 for men; and HR 1.23, 95%CI 1.02-1.48 for women) after adjustment for potential confounders. Additional adjustment for body mass index attenuated this association (HR 1.09, 95%CI 1.08-1.10 for all participants; HR 1.07; 95%CI 0.98-1.16 for men; HR 1.18; 95%CI 0.96-1.44 for women). Mediation analysis showed that BMI partially mediated the relationship between hyperuricemia and incident hypertension (indirect effect HR 1.09, 95%CI 1.08-1.10; direct effect: HR 1.08, 95%CI 1.02-1.15). The percentage of the mediation effect was 53.2% (95%CI 37.9-84.5). CONCLUSION: Hyperuricemia is associated with a risk of hypertension in both sexes, and BMI partially mediates hyperuricemia-related incident hypertension.

Submesoscale inverse energy cascade enhances Southern Ocean eddy heat transport.

Oceanic eddy-induced meridional heat transport (EHT) is an important process in the Southern Ocean heat budget, the variability of which significantly modulates global meridional overturning circulation (MOC) and Antarctic sea-ice extent. Although it is recognized that mesoscale eddies with scales of ~40-300 km greatly contribute to the EHT, the role of submesoscale eddies with scales of ~1-40 km remains unclear. Here, using two state-of-the-art high-resolution simulations (resolutions of 1/48° and 1/24°), we find that submesoscale eddies significantly enhance the total poleward EHT in the Southern Ocean with an enhancement percentage reaching 19-48% in the Antarctic Circumpolar Current band. By comparing the eddy energy budgets between the two simulations, we detect that the primary role of submesoscale eddies is to strengthen mesoscale eddies (and thus their heat transport capability) through inverse energy cascade rather than directly through submesoscale heat fluxes. Due to the submesoscale-mediated enhancement of mesoscale eddies in the 1/48° simulation, the clockwise upper cell and anti-clockwise lower cell of the residual-mean MOC in the Southern Ocean are weakened and strengthened, respectively. This finding identifies a potential route to improve the mesoscale parameterization in climate models for more accurate simulations of the MOC and sea ice variability in the Southern Ocean.

Circulating neprilysin clears brain amyloid.

The use of the peptidase neprilysin (NEP) as a therapeutic for lowering brain amyloid burden is receiving increasing attention. We have previously demonstrated that peripheral expression of NEP on the surface of hindlimb muscle lowers brain amyloid burden in a transgenic mouse model of Alzheimer's disease. In this study we now show that using adeno-associated virus expressing a soluble secreted form of NEP (secNEP-AAV8), NEP secreted into plasma is effective in clearing brain Abeta. Soluble NEP expression in plasma was sustained over the 3-month time period it was measured. Secreted NEP decreased plasma Abeta by 30%, soluble brain Abeta by approximately 28%, insoluble brain Abeta by approximately 55%, and Abeta oligomersby 12%. This secNEP did not change plasma levels of substance P or bradykinin, nor did it alter blood pressure. No NEP was detected in CSF, nor did the AAV virus produce brain expression of NEP. Thus the lowering of brain Abeta was due to plasma NEP which altered blood-brain Abeta transport dynamics. Expressing NEP in plasma provides a convenient way to monitor enzyme activity during the course of its therapeutic testing.

Expression of neprilysin in skeletal muscle reduces amyloid burden in a transgenic mouse model of Alzheimer disease.

Neprilysin (NEP) is a zinc metallopeptidase that efficiently degrades the amyloid beta (Abeta) peptides believed to be involved in the etiology of Alzheimer disease (AD). The focus of this study was to develop a new and tractable therapeutic approach for treating AD using NEP gene therapy. We have introduced adeno-associated virus (AAV) expressing the mouse NEP gene into the hindlimb muscle of 6-month-old human amyloid precursor protein (hAPP) (3X-Tg-AD) mice, an age which correlates with early stage AD. Overexpression of NEP in muscle decreased brain soluble Abeta peptide levels by approximately 60% and decreased amyloid deposits by approximately 50%, with no apparent adverse effects. Expression of NEP on muscle did not affect the levels of a number of other physiological peptides known to be in vitro substrates. These findings demonstrate that peripheral expression of NEP and likely other peptidases represents an alternative to direct administration into brain and illustrates the potential for using NEP expression in muscle for the prevention and treatment of AD.

Peripherally expressed neprilysin reduces brain amyloid burden: a novel approach for treating Alzheimer's disease.

A number of therapeutic strategies for treating Alzheimer's disease have focused on reducing amyloid burden in the brain. Among these approaches, the expression of amyloid beta peptide (Abeta)-degrading enzymes in the brain has been shown to be effective but to date not practical for treating patients. We report here a novel strategy for lowering amyloid burden in the brain by peripherally expressing the Abeta-degrading enzyme neprilysin on leukocytes in the 3xTg-AD mouse model of Alzheimer's disease. Through transplantation of lentivirus-transduced bone marrow cells, the Abeta-degrading protease neprilysin was expressed on the surface of leukocytes. This peripheral neprilysin reduced soluble brain Abeta peptide levels by approximately 30% and lowered the accumulation of amyloid beta peptides by 50-60% when transplantation was performed at both young and early adult age. In addition, peripheral neprilysin expression reduced amyloid-dependent performance deficits as measured by the Morris water maze. Unlike other methods designed to lower Abeta levels in blood, which cause a net increase in peptide, neprilysin expression results in the catabolism of Abeta to small, innocuous peptide fragments. These findings demonstrate that peripherally expressed neprilysin, and likely other Abeta-degrading enzymes, has the potential to be utilized as a therapeutic approach to prevent and treat Alzheimer's disease and suggest that this approach should be explored further.

Association between hyperuricemia and hypertension and the mediatory role of obesity: a large cohort study in China

SUMMARY OBJECTIVE: This study aimed to investigate the sex-specific association between hyperuricemia and the risk of hypertension and whether obesity mediates this association. METHODS: This study included 31,395 (47.0% women) adults without hypertension, cardiovascular disease, or cancer at baseline who completed at least one follow-up annual examination between 2009 and 2016. Cox regression models were performed to calculate hazard ratios and 95% confidence intervals. Mediation analysis was conducted to estimate the effect of body mass index on the association between hyperuricemia and hypertension. RESULTS: During a median 2.9-year follow-up, hyperuricemia was significantly associated with a higher risk of hypertension (HR 1.15, 95%CI 1.07-1.24 for all participants; HR 1.12, 95%CI 1.03-1.22 for men; and HR 1.23, 95%CI 1.02-1.48 for women) after adjustment for potential confounders. Additional adjustment for body mass index attenuated this association (HR 1.09, 95%CI 1.08-1.10 for all participants; HR 1.07; 95%CI 0.98-1.16 for men; HR 1.18; 95%CI 0.96-1.44 for women). Mediation analysis showed that BMI partially mediated the relationship between hyperuricemia and incident hypertension (indirect effect HR 1.09, 95%CI 1.08-1.10; direct effect: HR 1.08, 95%CI 1.02-1.15). The percentage of the mediation effect was 53.2% (95%CI 37.9-84.5). CONCLUSION: Hyperuricemia is associated with a risk of hypertension in both sexes, and BMI partially mediates hyperuricemia-related incident hypertension.

In vitro and in vivo degradation of Abeta peptide by peptidases coupled to erythrocytes.

It is generally believed that amyloid beta peptides (Abeta) are the key mediators of Alzheimer's disease. Therapeutic interventions have been directed toward impairing the synthesis or accelerating the clearance of Abeta. An equilibrium between blood and brain Abeta exists mediated by carriers that transport Abeta across the blood-brain barrier. Passive immunotherapy has been shown to be effective in mouse models of AD, where the plasma borne antibody binds plasma Abeta causing an efflux of Abeta from the brain. As an alternative to passive immunotherapy we have considered the use of Abeta-degrading peptidases to lower plasma Abeta levels. Here we compare the ability of three Abeta-degrading peptidases to degrade Abeta. Biotinylated peptidases were coupled to the surface of biotinylated erythrocytes via streptavidin. These erythrocyte-bound peptidases degrade Abeta peptide in plasma. Thus, peptidases bound to or expressed on the surface of erythroid cells represent an alternative to passive immunotherapy.

Inhibition of human B-cell lymphoma by an anti-CD20 antibody and its chimeric F(ab')2 fragment via induction of apoptosis.

Monoclonal antibodies (mAb) directed against CD20, either unmodified or in radiolabeled forms, have been successfully used in clinic as effective therapeutic agents in the management of non-Hodgkin's B-cell lymphoma. Despite all clinical success the exact mechanisms of action of various anti-CD20 antibodies remains mostly unclear. Several mechanisms have been proposed to be responsible for the therapeutic activity of anti-CD20 antibodies, including antibody-dependent cell-mediated cytotoxicity, complement-mediated cytotoxicity, and direct inhibition of tumor growth via induction of apoptosis. We previously produced an anti-CD20 mAb, HI47, and showed that the antibody effectively blocked human B-cell proliferation in vitro and inhibited xenografted B-cell lymphoma in nude mice. In this study, we engineered the chimeric versions of both the Fab and F(ab)'2 fragments of HI47 and produced the fragments in E. coli. Both fragments competed efficiently with HI47 for binding to CD20+ B cells, and inhibited proliferation of B-lymphoma cells in a dose-dependent manner. Mechanistic studies revealed that both antibody fragments induced significant degree of B-cell apoptosis that is independent of any cross-linking agents. Further, both the F(ab)'2 and Fab fragments when administered in vivo significantly inhibited the growth of human B-cell lymphoma xenografts in nude mice. The bivalent F(ab)'2 fragment showed consistently better efficacy compared to its monovalent Fab counterpart in inducing apoptosis and inhibiting B-cell lymphoma growth both in vitro and in vivo. Taken together, these observations suggest that HI47 and its fragments most likely exert their antitumor activity through induction of cell apoptosis, and cross-linking/dimerization of CD20 molecules on B- cell surface is an important, but not essential, process for therapeutic efficacy of HI47 and its fragments.

[High level expression of chimeric antibody fragment F(ab')2 directed against CD20 in Escherichia coli].

The use of tumor antigen specific antibody for the delivery of therapeutic agents offers the possibility of targeting therapy with reduced toxicity to normal tissues compared to conventional treatments. In previous work, the human-mouse chimeric antibody fragment Fab' directed against CD20 was constructed from the new anti-CD20 antibody HI47 (a mouse IgG3, K). The chimeric antibody fragment Fab' could reduce its antigenicity, but the yield, quality and affinity of chimeric antibody fragment Fab' restrict its use. To improve affinity of chimeric antibody fragment Fab', a new phasmid pYZcpp3, which expresses chimeric antibody fragment F(ab')2, was constructed by adding a sequence encoding a small peptide, (CPP)3, to C-terminus of heavy chain constant region of chimeric antibody fragment Fab'. Using the pYZcpp3 to transform E. coli. 16c9, the genetically engineered bacteria 10916# was obtained. 10916# can secret the soluble chimeric antibody fragment Fab' and F(ab')2 into periplasmic. The yield was up to 360 mg/L with the percent of F(ab')2 up to 45% in 19L fermentor by the high density fermentation technology. Without denaturation and renaturation, the F(ab')2 has possessed the native three-dimensional structure. The purity of F(ab')2 was more than 90% after the purification of protein G affinity chromatography and S200 size exclusion chromatography. The F(ab')2 could distinguish and bind to Raji cells (CD20+) by FACS. F(ab')2 could inhibit the proliferation of Raji cells in vitro by MTT, IC50 was 22.8 microg/mL. HI47 and its chimeric fragments F(ab')2 induced a significant level of apoptosis (23.5%, 20.8%, respectively), independent of any cross-linking agents, in Raji cells after 24 h incubation. The chimeric antibody fragment F(ab')2 directed against CD20 is possible to apply to tumor therapy in clinic in the future.

[Expression of chimeric anti-CD3 IgG antibody in mammalian cells and analysis of its biological activity].

The anti-CD3 antibody can improve success rate of organs transplant. HIT3a, a mouse anti-CD3 antibody, was chimerized by using gene engineering methods to decrease its immunogenity. The anti-CD3 genes, heavy chain and light chain, were cloned using PCR from the vector pCANTAB 5E containing anti-CD3 scFv gene fragment, and two PCR fragments were recombined into the expression vector pKN100 with human antibody light constant domain and pG1D105 with human antibody heavy constant domain, respectively. The two vectors were co-transfected into CHO cells using liposome. The anti-CD3 antibody was detected by ELISA and Western blot assay in supernatant of transfected CHO cells culture. The primary results of competitive assays by FACS showed that anti-CD3 antibody could partially block the sites through which parent antibody (HIT3a) bind to CD3+ Jurkat cells. The result of 3H-TdR incorporation showed that the chimeric anti-CD3 antibody could stimulated proliferation of peripheral blood mononuclear cells (PBMC) as the parent antibody. In this thesis, the results of some experiments indicated that the chimeric anti-CD3 antibody expressed in CHO cells was an antibody with native biological activity, and it is possible to apply to in clinic in the future.

[One amino acid mutation in an anti-CD20 antibody fragment that affects the yield bacterial secretion and the affinity].

Monoclonal antibodies (mAb) directed against CD20, either unmodified or in radiolabeled forms, have been successfully exploited in clinic as effective therapeutic agents in the management of non-Hodgkin's B-cell lymphoma. The antibody fragment is a potential agent in image and therapy of tumor. To further improve the soluble expression of anti-CD20 antibody Fab' fragment, PCR was used to mutate the anti-CD20 VL and VH genes and its biological activity was identified. The expression vector of chimeric antibody Fab' was constructed and expressed in E. coli. The data of mutant clone DNA sequence showed that the amino acid of light chain gene of the parent anti-CD20 antibody (H47) was successful mutated as Ser (GAG)-Asn (CAG). The soluble expression of mutated anti-CD20 Fab' (CD20-7) was 3.8 mg/g dry cell weight, while the parent (CD20-2) was 1.3 mg/g dry cell weight. The affinity constant Ka of CD20-7 was 2.2 x 10(9) L/mol. The primary results of competitive assays by FACS showed that CD20-7 could partially block the sites through which parent antibody (HI47) bind to Raji cells. There was difference in the Raji cells (CD20+)-binding activity between the mutant CD20-7 and parent CD20-2. The site mutation of anti-CD20 Fab' gene make it possible that the anti-CD20 antibody fragment was succeeded to obtain higher expression. In this thesis, we succeeded in completing mutation and expression of anti-CD20 Fab' genes, distinguishing its biological activity, and obtaining its highly expression. These period results will lay a foundation for development of other kind of anti-CD20 engineering antibody (for instance: Fab' Diabody and miniantibody), and make it possible for anti-CD20 antibody to be applied to tumor therapy in civil in the future.

[Apoptosis of human B cell lymphoma Raji cells induced by monovalent anti-CD20 antibody].

BACKGROUND & OBJECTIVE: The anti-CD20 antibodies and fragments have been applied for treatment of non-Hodgkin's lymphomas (NHL) in clinic. The new anti-CD20 antibodies and their fragments (unmodified or radiolabeled) yet have been exploited for those patients with incomplete response to rituximab. The chimeric antibody fragments Fab and F(ab) '(2) derived from HI(47)(a mouse anti-CD20 antibody) have been constructed. The present study was designed to determine the effect of HI(47) and chimeric anti-CD20 antibody fragments on growth inhibition and apoptosis of lymphoma cells. METHODS: The binding of anti-CD20 antibodies to CD20 positive human B cell lymphoma Raji cells was examined using immunofluorescence assay. MTT method was used to evaluate the effect of chimeric antibody fragments on Raji cells growth. Annexin V staining and DNA ladder were used to examine the apoptosis of Raji cells induced by chimeric antibody fragments. RESULTS: HI(47) and its chimeric antibody fragments were capable of binding to CD20 positive Raji cells with the binding rate of above 90%. HI(47) did not compete with rituximab in binding with Raji cells. The growth of Raji cells was inhibited by HI(47) and its fragments with the inhibition rates of (57+/-1.5)%, (65.2+/-2.5)%,and (77.2+/-3.2)%, respectively, at the concentration of 100 microg/ml. The monovalent antibody fragment Fab (20 microg/ml) induced the apoptosis of Raji cells with early apoptosis rate of 17%. CONCLUSION: The chimeric anti-CD20 antibody fragments derived from HI(47) have inhibitory effect on Raji cells and can induce apoptosis of Raji cells.