Recent advances in the role of endogenous hydrogen sulphide in cancer cells.

Hydrogen sulphide (H2 S) is a gaseous neurotransmitter that can be self-synthesized by living organisms. With the deepening of research, the pathophysiological mechanisms of endogenous H2 S in cancer have been increasingly elucidated: (1) promote angiogenesis, (2) stimulate cell bioenergetics, (3) promote migration and proliferation thereby invasion, (4) inhibit apoptosis and (5) activate abnormal cell cycle. However, the increasing H2 S levels via exogenous sources show the opposite trend. This phenomenon can be explained by the bell-shaped pharmacological model of H2 S, that is, the production of endogenous (low concentration) H2 S promotes tumour growth while the exogenous (high concentration) H2 S inhibits tumour growth. Here, we review the impact of endogenous H2 S synthesis and metabolism on tumour progression, summarize the mechanism of action of H2 S in tumour growth, and discuss the possibility of H2 S as a potential target for tumour treatment.

Role of Nanomedicine-Based Therapeutics in the Treatment of CNS Disorders.

Central nervous system disorders, especially neurodegenerative diseases, are a public health priority and demand a strong scientific response. Various therapy procedures have been used in the past, but their therapeutic value has been insufficient. The blood-brain barrier (BBB) and the blood-cerebrospinal fluid barrier is two of the barriers that protect the central nervous system (CNS), but are the main barriers to medicine delivery into the CNS for treating CNS disorders, such as brain tumors, Parkinson's disease, Alzheimer's disease, and Huntington's disease. Nanotechnology-based medicinal approaches deliver valuable cargos targeting molecular and cellular processes with greater safety, efficacy, and specificity than traditional approaches. CNS diseases include a wide range of brain ailments connected to short- and long-term disability. They affect millions of people worldwide and are anticipated to become more common in the coming years. Nanotechnology-based brain therapy could solve the BBB problem. This review analyzes nanomedicine's role in medication delivery; immunotherapy, chemotherapy, and gene therapy are combined with nanomedicines to treat CNS disorders. We also evaluated nanotechnology-based approaches for CNS disease amelioration, with the intention of stimulating the immune system by delivering medications across the BBB.

Fluzoparib increases radiation sensitivity of non-small cell lung cancer (NSCLC) cells without BRCA1/2 mutation, a novel PARP1 inhibitor undergoing clinical trials.

PROPOSE: Poly (ADP-ribose) polymerase 1 inhibitors were originally investigated as anti-cancer therapeutics with BRCA1/2 genes mutation. Here, we investigate the effectiveness of a novel PARP1 inhibitor fluzoparib, for enhancing the radiation sensitivity of NSCLC cells lacking BRCA1/2 mutation. METHODS: We used MTS assays, western blotting, colony formation assays, immunofluorescence staining, and flow cytometry to evaluate the radiosensitization of NSCLC cells to fluzoparib and explore the underlying mechanisms in vitro. Through BRCA1 and RAD50 genes knockdown, we established dysfunctional homologous recombination (HR) DNA repair pathway models in NSCLC cells. We next investigated the radiosensitization effect of fluzoparib in vivo using human NSCLC xenograft models in mice. The expression of PARP1 and BRCA1 in human NSCLC tumor samples was measured by immunohistochemistry. Furthermore, we sequenced HR-related gene mutations and analyzed their frequencies in advanced NSCLC. RESULTS: In vitro experiments in NSCLC cell lines along with in vivo experiments using an NSCLC xenograft mouse model demonstrated the radiosensitization effect of fluzoparib. The underlying mechanisms involved increased apoptosis, cell-cycle arrest, enhanced irradiation-induced DNA damage, and delayed DNA-damage repair. Immunohistochemical staining showed no correlation between the expression of PARP1 and BRCA1. Moreover, our sequencing results revealed high mutation frequencies for the BRCA1/2, CHEK2, ATR, and RAD50 genes. CONCLUSION: The potential therapeutic value of fluzoparib for increasing the radiation sensitivity of NSCLC is well confirmed. Moreover, our findings of high mutation frequencies among HR genes suggest that PARP1 inhibition may be an effective treatment strategy for advanced non-small cell lung cancer patients.

Two validated liquid chromatography-mass spectrometry methods with different pretreatments for the quantification of an anti-CD47 monoclonal antibody in rat and cynomolgus monkey serum compared with an electrochemiluminescence method.

Establishing reliable bioanalytical methods is essential to support pharmacokinetic (PK) studies in the preclinical and clinical evaluation of monoclonal antibody (mAb) drugs. Ligand binding assay (LBA) has always been the gold standard for protein quantification, whereas LC-MS has gradually become a promising alternative method for the study of pharmacokinetics of biotherapeutics with its advantages of accuracy and rapid method development. Here, we described for the first time two liquid chromatography-mass spectrometry (LC-MS) methods with different purification pretreatments, protein precipitation and immune affinity (IA) enrichment, along with one electrochemiluminescence (ECL) method for the quantification of an anti-CD47 monoclonal antibody (SHR-1603) in rat and cynomolgus monkey serum. An anti-adsorption reagent was added and digestion conditions were optimized to resolve the absorption issue of hydrophobic peptide in this study. These methods were all validated according to China Food and Drug Administration (CFDA) and European Medicines Agency (EMA) guidelines and were successfully applied to a preclinical study for the quantification of SHR-1603. The respective quantitative ranges of the three methods are respectively 250-500,000 ng/mL (protein precipitation), 100-100,000 ng/mL (IA) and 19.5-10,000 ng/mL (ECL). The two LC-MS methods were compared with ECL method respectively by the cross-validation using the Passing-Bablok regression and Bland-Altman plots. Systematic differences and proportional bias were observed between two LC-MS methods on the one hand and with the ECL method on the other hand. The drug concentrations obtained by the three methods showed good agreement in the low-dose group (ratios of drug exposure, 1.05-1.11), whereas the drug concentrations measured using the LC-MS methods were higher than those obtained by the ECL method in medium-dose and high-dose groups, which can be attributed to the forms of antibodies being determined (free and total). In conclusion, the established LC-MS methods exhibited superior accuracy, efficiency and cost-effectiveness for the PK assessment of SHR-1603 in the preclinical study. Thus, it provides a promising alternative to LBA in pre-clinical and clinical evaluation studies of mAb drugs in various matrices to facilitate the development of anti-tumor drugs.

SHR-A1403, a novel c-Met antibody-drug conjugate, exerts encouraging anti-tumor activity in c-Met-overexpressing models.

Emerging evidence demonstrates that a c-Met antibody-drug conjugate (ADC) has superior efficacy and safety profiles compared with those of currently available small molecules or antibody inhibitors for the treatment of c-Met-overexpressing cancers. Here we described both the in vitro and in vivo efficacies of SHR-A1403, a novel c-Met ADC composed of a humanized IgG2 monoclonal antibody against c-Met conjugated to a novel cytotoxic microtubule inhibitor. SHR-A1403 showed high affinity to c-Met proteins derived from human or monkey and potent inhibitory effects in cancer cell lines with high c-Met protein expression. In mice bearing tumors derived from cancer cell lines or patient HCC tissues with confirmed c-Met overexpression, SHR-A1403 showed excellent anti-tumor efficacy. Antibody binding with c-Met contributed to SHR-A1403 endocytosis; the subsequent translocation to lysosomes and cytotoxicity of the released toxin are speculated to be predominant mechanisms underlying the anti-tumor activity of SHR-A1403. In conclusion, SHR-A1403 showed significant anti-tumor activity in cancer cell lines, xenograft mouse models and an HCC PDX model, which all have high c-Met levels. These data provide references for SHR-A1403 as a potential therapy for the treatment of cancers with c-Met overexpression.

Pharmacologic characterization of fluzoparib, a novel poly(ADP-ribose) polymerase inhibitor undergoing clinical trials.

Poly(ADP-ribose) polymerase (PARP) enzymes play an important role in repairing DNA damage and maintaining genomic stability. Olaparib, the first-in-class PARP inhibitor, has shown remarkable clinical benefits in the treatment of BRCA-mutated ovarian or breast cancer. However, the undesirable hematological toxicity and pharmacokinetic properties of olaparib limit its clinical application. Here, we report the first preclinical characterization of fluzoparib (code name: SHR-3162), a novel, potent, and orally available inhibitor of PARP. Fluzoparib potently inhibited PARP1 enzyme activity and induced DNA double-strand breaks, G2 /M arrest, and apoptosis in homologous recombination repair (HR)-deficient cells. Fluzoparib preferentially inhibited the proliferation of HR-deficient cells and sensitized both HR-deficient and HR-proficient cells to cytotoxic drugs. Notably, fluzoparib showed good pharmacokinetic properties, favorable toxicity profile, and superior antitumor activity in HR-deficient xenografts models. Furthermore, fluzoparib in combination with apatinib or with apatinib plus paclitaxel elicited significantly improved antitumor responses without extra toxicity. Based on these findings, studies to evaluate the efficacy and safety of fluzoparib (phase II) and those two combinations (phase I) have been initiated. Taken together, our results implicate fluzoparib as a novel attractive PARP inhibitor.

Preclinical pharmacokinetics of a novel anti-c-Met antibody-drug conjugate, SHR-A1403, in rodents and non-human primates.

1. The in vivo pharmacokinetics (PK) profiles of a novel c-Met antibody-drug conjugate (ADC), SHR-A1403, were investigated and characterized in mice, rats and monkeys. 2. Serum concentrations of ADC and total antibody were detected using validated ELISA methods. The results showed low systemic clearance of both ADC and total antibody in all three species as reflected by gradual decrease in serum concentrations. Half-life (t1/2) of ADC ranged from 4.6 to 11.3 days in the three species. 3. Tissue distribution study in tumor-bearing mice showed high accumulation of 125I-SHR-A1403 in tumor tissues over the other organs/tissues, indicating the favorable safety of SHR-A1403 and characteristics of an ADC drug. 4. Relatively low grade of anti-drug antibody (ADA) in monkeys had no impact on PK profile of the ADC. 5. During discovery stage, undesirable exposure and/or ADA incidence were observed for SHR-A1403 with high or low drug-antibody ratio (DAR), which was DAR = 5 to 6 and DAR = 1, respectively, and therefore prompted selection of an appropriate DAR value (DAR = 2) for SHR-A1403 used in preclinical development and clinical trials. 6. In conclusion, our work demonstrated favorable PK characterization of SHR-A1403, and supported for investigational new drug application (IND) and the ongoing first-in-human trial in the US.

BMSCs pre-treatment ameliorates inflammation-related tissue destruction in LPS-induced rat DIC model.

This study aimed to investigate the effect of bone marrow-derived mesenchymal stem cells (BMSCs) on disseminated intravascular coagulation (DIC) model rats and to further explore the underlying mechanism. A rat model of lipopolysaccharide (LPS)-induced DIC was successfully established, as indicated by impaired plasma hemostatic parameters and damaged organ functions in rats. Importantly, pre-treatment with rat allogeneic BMSCs before LPS injection significantly alleviated systemic intravascular coagulation, reduced plasma levels of organ dysfunction indicators and pro-inflammatory cytokines, suppressed fibrin microthrombi formation, ameliorated liver, heart, and renal injuries, and increased 24-hour survival rates in LPS-induced DIC rats. The protection of BMSCs against DIC was in a moderately dose-dependent manner. Further investigation revealed that BMSCs co-cultured with peripheral blood mononuclear cells (PBMCs) significantly inhibited the LPS-stimulated PBMCs proliferation and the release of pro-inflammatory cytokines from PBMCs. Of note, upregulation of immunosuppressive factors including indoleamine 2,3-dioxygenase and interleukin-10, which was induced by interferon-γ, contributed to BMSCs-mediated inhibition of LPS-stimulated PBMCs proliferation. These effects do not depend on the direct cell-cell contact. In conclusion, BMSCs pre-treatment ameliorates inflammation-related tissue destruction in LPS-induced DIC model rats. The protection of BMSCs may be attributed to their anti-inflammatory and immunomodulatory properties, which render BMSCs a promising source for stem cell-based therapeutic approaches in inflammation-related DIC.

Discovery and development of pyrotinib: A novel irreversible EGFR/HER2 dual tyrosine kinase inhibitor with favorable safety profiles for the treatment of breast cancer.

The discovery and development of a novel irreversible EGFR/HER2 dual tyrosine kinase inhibitor SHR1258 (pyrotinib) for the treatment of HER2-postive breast cancer is presented. The structure-activity relationship of lead series and their pharmacokinetic properties were evaluated to identify the potential candidates for further in vivo efficacy studies and preclinical safety assessments. Metabolic pathway and drug-drug interaction were also investigated in preclinical settings. In particular, major metabolites in human and animal species were assessed with regard to potential toxicity or off-target side effects. Overall, the potent and selective EGFR/HER2 dual inhibitor, pyrotinib, displayed robust anti-tumor effects on HER2-overexpressing xenograft models and sufficiently safety windows in animals as well as favorable pharmacokinetic properties in human, which substantially ensures current clinical development. Finally, recent advances of pyrotinib in clinical studies are highlighted with very encouraging outcomes in patients with HER2-postive advanced breast cancer.

Metabolic analysis of the effect of rheum on a taurocholate-induced acute pancreatitis rat model

ABSTRACT The effects of rheum on serum parameters in a taurocholate-induced acute pancreatitis (AP) rat model were investigated using pathological and biochemical tests, and a proton nuclear magnetic resonance (1H NMR)-based metabonomic strategy. Healthy rats and rats with AP were either treated with rheum (7.5% at a dose of 1.5 g/kg) or left untreated. Serum samples were collected from the AP and rheum-treated groups at 6, 12, and 24 h after treatment. The effect of rheum on pathological changes in the pancreatic was investigated to validate the AP model. We obtained 1H NMR spectra and analyzed the results using the partial least squares discriminant method. The results of the pathological and metabolic analyses revealed an amelioration of multiple metabolic abnormalities and an increase in the aerobic respiration ratio after treatment, compared with the AP groups. These results were attributed to improvements in energy supply and the elimination of metabolic products. The study also promoted NMR-based metabonomic analysis as a feasible method of assessing traditional Chinese drugs.

Metabolic characterization of pyrotinib in humans by ultra-performance liquid chromatography/quadrupole time-of-flight mass spectrometry.

Pyrotinib is a novel irreversible tyrosine kinase inhibitor developed for the treatment of human epidermal growth factor receptor 2 (HER2)-positive breast cancer. The results of phase I clinical trial demonstrated that pyrotinib was well tolerated and exhibited potent antitumor activity. As a promising therapeutic agent for HER2-positive breast cancer, it is of great importance to investigate the biotransformation of pyrotinib in humans and identify the major enzymes involved in its metabolism during its early stage of development for safety consideration. For this purpose, a robust analytical method based on ultra-performance liquid chromatography/quadrupole time-of-flight mass spectrometry (UPLC/Q-TOF MS) was established to characterize the metabolites of pyrotinib in human plasma, feces, and urine, and identify the primary enzymes responsible for its metabolism. As a result, a total of 24 metabolites were identified, including 16 phase I metabolites resulting from dealkylation, oxidation, dehydrogenation, and carbonylation, and 8 phase II metabolites originating from cysteine and N-acetylcysteine conjugation. Pyrotinib was absorbed into blood by 1h, reached its peak level at 4h, and afterwards underwent slow elimination. The principal metabolites detected in humans (M1, M2, and M5) were products resulting from O-depicoline and pyrrolidine lactam formation, whose structures have been confirmed by the synthetic references. In addition, fecal clearance was the major route of excretion for pyrotinib. Further phenotyping experiment proved that CYP3A4 was the most active enzyme responsible for the biotransformation of pyrotinib, implying the vital necessity of the assessment of the potential CYP3A-mediated drug-drug interactions in humans. Taken together, this study provided valuable metabolic data to explicate the dynamic process of pyrotinib in humans, and important reference basis for its safety evaluation and rational clinical application. The results will also benefit the assessment of the contributions to the overall activity or toxicity from the key metabolites.

Totally thoracoscopic closure of ventricular septal defect without a robotically assisted surgical system: a summary of 119 cases.

OBJECTIVES: To summarize the clinical outcomes of totally thoracoscopic closure of a ventricular septal defect (VSD). METHODS: Totally thoracoscopic VSD closure was performed in 119 patients (66 boys; mean age, 7.1 ± 3.6 years). An additional 35 patients undergoing open-chest VSD closure were selected as a control group. Using 3 port incisions in the right chest, pericardiotomy, bicaval occlusion, atriotomy, and VSD closure were performed by thoracoscopy without the aid of a robotically assisted surgical system. RESULTS: Cardiopulmonary bypass and aortic crossclamp times were 42.2 ± 9.8 and 32.5 ± 7.3 minutes, respectively. There were no deaths but 1 patient required insertion of a permanent pacemaker as a result of postoperative atrioventricular conduction block. The length of stay in the intensive care unit (11.0 ± 2.6 vs 22.9 ± 4.9 hours, P < .01) or postoperative hospital stay (4.2 ± 1.1 vs 6.6 ± 2.1 days, P < .03) in the thoracoscopic group were shorter than in the control group. The percentage of patients who required postoperative opioid analgesics in the thoracoscopic group was lower than in the control group (31.9% vs 74.2%, P < .001). Rate of blood transfusion during the operation (17.6% vs 65.7%, P = .001) and the postoperative use of opioid analgesics (31.9% vs 74.3%, P = .003) in the thoracoscopic group was lower than in the control group. Transesophageal echocardiographic analysis 4.6 ± 2.3 months after the operation showed complete closure of the defect. CONCLUSIONS: Totally thoracoscopic closure of VSD through a 3-port entry was safe and effective.

The impact of pulmonary hypertension on outcomes of patients with low left ventricular ejection fraction: a propensity analysis.

BACKGROUND AND AIM OF THE STUDY: Pulmonary hypertension (PH) is commonly described as a risk factor in cardiac surgery; however, the effect of a low left ventricular ejection fraction (LVEF) on PH has not been assessed. Hence, the study aim was to determine whether PH increases operative mortality and survival outcome in patients with a low LVEF. METHODS: Between January 2001 and September 2009, a total of 845 consecutive patients with LVEF < 40% was enrolled into the study. Among these patients, 444 had a pulmonary pressure < 40 mmHg (NPH group), while in 401 patients the pulmonary pressure was > or = 40 mmHg. RESULTS: Preoperatively, the PH patients were older (p < 0.001), had a lower LVEF (p = 0.001), and had a higher logistic EuroSCORE (p < 0.001) and serum creatinine level (p < 0.026) when compared to NPH patients. The PH patients showed a greater tendency to develop postoperative complications (p < 0.001). After adjusting by propensity score, the in-hospital mortality was significantly higher among PH patients (p < 0.001), while multivariate logistic regressions revealed PH as an independent predictor for in-hospital mortality (p = 0.036). The 12-, 36-, and 60-month follow up mortality rates were significantly higher in the PH group. By using a Cox logistic regression model, PH was shown to be an independent predictor for follow up mortality (p = 0.035). CONCLUSION: Pulmonary hypertension increased the morbidity and mortality in patients with a low LVEF who were undergoing cardiac surgery. Future studies may identify subgroups that may benefit from a preoperative optimization of PH and/or intra- and postoperative therapies directed at minimizing the effects of the condition.

[Experimental studies of therapeutic effect of Rheum officinale on acute pancreatitis].

OBJECTIVE: To investigate the therapeutic effect of Rheum officinale on acute pancreatitis. METHODS: Buffered sodium taurocholate (3% m/V) was injected into the pancreatico-biliary duct to induce acute pancreatitis. Death rate,coefficient of pancreas, serum amylyse (AMY), hemocuprein (SOD), TNF-alpha and IL-1beta level were examined at 6, 12 and 24 hours after operations. Pathology analysis were also obtained. RESULTS: Compared with corresponding pancreatitis groups,death rate, coefficient of pancreas, serum TNF-alpha and IL-1beta level of drug groups decreased remarkably (P < 0.05), while serum SOD level significantly increased (P < 0.01). Serum AMY level of drug groups increased at 6 h (P < 0.01), decreased at 12 h (P < 0.01) and had no statistics disparity at 24 h (P > 0.05) compared with respective pancreatitis group. Although score points of all drug groups were lower than corresponding pancreatitis groups, the growth tendency of both were similar. CONCLUSION: Rheum officinale Baill has the effect of prevention to pancreas pathological changes in the animal pattern, but not able to reverse the tendency.

[In vitro study on two kind of recombinant adeno-associated virus-mediated transfection efficiency with enhanced green fluorescent protein into rats osteoblasts].

OBJECTIVE: To compare the transfection efficiency of two kind of recombinant adeno-associated virus-mediated transfection to rats osteoblasts with enhanced green fluorescent protein and assess the feasibility of it as a vector for gene therapy of osteoblast lesions. METHODS: The osteoblasts of rats were isolated, cultured and identified with type I collagen staged digestion method. According to different multiplicity of infection (MOI) (MOI = 1 x 10(3), 1 x 10(4), 1 X 10(4), 5 x 10(5)), rAAV-EGFP was transfected into osteoblasts with rAAV only and rAAV-ADV co-transfection respectively. The expression of EGFP along with the transfection time was observed under inverted fluorescence microscope. The transfection efficiency and fluorescence intensity was evaluated by flow cytometry. The best MOI value was analysed and the cell growth curves were obtained according to the best MOI value to evaluate the toxic effects of rAAV-EGFP. RESULTS: The cultured cells possessed the biological behaviors of osteoblasts. The transfection efficiency of the rAAV was increased with the increasing of MOI. The EGFP expression reached the maximum on day 5 in ADV(-) group, the transfection efficiency of rAAV2/6-EGFP and rAAV2/9-EGFP was 90.2% and 66.1% respectively when MOI was 1 x 10(5) and no significant increase was observed when MOI was 5 x 10(5). In ADV(+) group, EGFP expression reached its maximum on day 3, the transfection efficiency of rAAV2/6-EGFP and rAAV2/9-EGFP was 47.6% and 30.5% respectively when MOI was 5 x 10(5). And no significant biologic effects on the cyto-activity was observed. CONCLUSION: The transfection efficiency of two kind of virus vectors was both very high and rAAV2/6's is higher than that of rAAV2/9. This suggested the potential of rAAV-EGFP as a safe and efficient vector for gene therapy.

[Relationship of apoptotic receptor expression in normal human peripheral blood lymphocytes to cell apoptosis].

The aim of this study was to detect the expression and cell cycle specificity of Fas, TNFRI and TNFRII in human peripheral blood lymphocytes (PBL), and to study the potential role of Fas, TNFRIand TNFRII in cell cycle specific apoptosis. The improved double-parameter flow cytometry was used to detect the expressions of Fas, TNFRI and TNFRII and cell cycle specificity in PBL which were incubated for 24 hours in the presence or absence of phytohaematoagglutinin (PHA) respectively. Apoptosis induced by IgM type anti-Fas and TNF-alpha was detected by API method. The results showed that compared with PBL treated in the absence of PHA in G(0) phase, the ratio of Fas, TNFRI and TNFRII expressions in PHA-stimulated PBL entering cell cycle increased (35.55 +/- 6.63)%, (30.63 +/- 2.66)%, (26.62 +/- 5.14)% respectively (P < 0.01), and mainly appeared at G(1)-phase; no apoptosis was induced by anti-Fas and TNF-alpha in G(0)-phase PBL cultured in the absence of PHA. On the contrary, the apoptosis was induced by anti-Fas and TNF-alpha in PBL which entered cell cycle after stimulation with PHA and mainly initiated at G(1)-Phase. It is concluded that there is evident dose-effect relationship between apoptotic receptor and receptor-mediated apoptosis. Moreover, the cell cycle specificity of receptor-mediated apoptosis is correlated with the cell cycle specific expressions of apoptotic receptor. The induction of apoptosis by apoptotic factors (anti-Fas and TNF-alpha) depends on whether cell entering cell cycle or not.

[Correlation of receptor-mediated apoptosis to cell cycle].

BACKGROUND & OBJECTIVE: Chondriosome-mediated apoptosis is closely related to cell cycle, however, the correlation of receptor-mediated apoptosis to cell cycle progression is unclear yet. This study was to observe the receptor-mediated apoptosis and cell cycle specificity in cultured normal and tumor lymphocytes, and investigate their correlation. METHODS: Exponentially growing human leukemia cell lines Molt-4 and Jurkat were treated with tumor necrosis factor-alpha (TNF-alpha) or Anti-Fas. Peripheral blood lymphocytes (PBLs) from healthy donors were stimulated by phytohemagglutinin (PHA), and further incubated with the presence of TNF-alpha or anti-Fas. Annexin V/PI was used to detect the apoptosis, and API method was used to illustrate the cell cycle specificity of apoptotic cells. RESULTS: Unstimulated PBLs kept blunt to stimulation with TNF-alpha or anti-Fas, and the apoptotic rate was 6%-8%. Molt-4 cells, Jurkat cells, and stimulated PBLs which were treated with TNF-alpha or anti-Fas went to apoptosis, and the apoptosis rates were 15%-28%. Most receptor-mediated apoptosis happened in early G1 phase. CONCLUSION: Receptor-mediated apoptosis is closely related to cell cycle and presents cell cycle specificity.