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The expression levels of programmed cell death 5 (PDCD5) are down-regulated in many malignancies. SG611-pdcd5, a recombinant conditionally replicative adenovirus carrying pdcd5 gene expression cassette, can evidently kill the leukemic cells and protect selectively the normal cells. The purpose of this study was to investigate the synergistic killing effect of SG611-pdcd5 and low-dose etoposide (VP-16) on K562 cells. K562 cells were treated with different concentrations of VP-16 or different multiplicities of infection (MOI) of SG611-pdcd5. After 48 hours of incubation the cell viability was determined by using MTT assay. The results showed that the cell viability of SG611-pdcd5 (MOI = 40) plus VP-16 (0.5 µg/ml) group significantly decreased as compared with single SG611-pdcd5 (MOI = 40) treatment group or single VP-16(0.5 µg/ml) treatment group (42.00 ± 5.75% vs 59.45 ± 4.12%; 42.00 ± 5.75% vs 82.91 ± 3.41%, respectively, both p < 0.05). The synergistic killing effect of SG611-pdcd5 plus VP-16 was higher than that of PDCD5 protein plus VP-16 or that of non-replicating adenovirus carrying pdcd5 (Ad-pdcd5) plus VP-16 (both p < 0.05). The cell viability of VP-16 (4.0 µg/ml) plus SG611-pdcd5 (MOI = 40) group, VP-16 (4.0 µg/ml) plus proPDCD5 (40 µg/ml) group and VP-16 (4.0 µg/ml) plus Ad-pdcd5 (MOI = 80) group was 37.09 ± 1.89%, 52.36 ± 1.64% and 73.64 ± 4.33%, respectively. It is concluded that SG611-pdcd5 can promote K562 cell death induced by low-dose VP-16. The combination of SG611-pdcd5 and VP-16 can enhance the killing effect on leukemic cells.

PDCD5 (programmed cell death 5) accelerates apoptosis of certain tumor cells and is expressed at low levels in marrow-nucleated cells of AML and CML patients. In the present study, we evaluated the effects of PDCD5 overexpression on drug sensitivity of leukemia cells. K562 cells were treated with idarubicin (IDR) alone or in combination with adenoviral vectors expressing PDCD5 (Ad-PDCD5). As shown by annexin-V-FITC/PI dual labeling, apoptosis rates were markedly increased after combined treatment with Ad-PDCD5 compared to IDR treatment alone. We observed that PDCD5 overexpression significantly improves the antitumor effects of low dose IDR treatment in vivo. Tumor sizes were significantly decreased in combined Ad-PDCD5 and low dose IDR treatment groups compared with single IDR treatment groups. Similar results were obtained with combined systemic treatment of Ad-PDCD5 and low dose IDR, and combined treatment with Ad-PDCD5 local injection and low dose IDR i.p. injection. These results indicate that Ad-PDCD5 may be a promising agent for enhancing chemosensitivity.

This study was purposed to investigate the effect of adenovirus-mediated transfer of PDCD5 gene on apoptosis of K562 cells induced by etoposide. Recombinant adenovirus PDCD5 (Ad-PDCD5), control vectors Ad-null and Ad-eGFP were constructed by AdMax vector system respectively. After K562 cells were transfected by Ad-PDCD5, Ad-null or Ad-eGFP with different multiplicity of infection (MOI), the expression level of the PDCD5 gene was examined by RQ-RT-PCR assay. The effects of etoposide in combination with Ad-PDCD5 on the proliferation and apoptosis of K562 cells were measured by using MTT assay and flow cytometry with Annexin-V-FITC/PI dual labeling technique, respectively. The results showed that the transfection efficiencies of Ad-eGFP in K562, Jurkat and CEM cells ranged from 60% to 86%. Expression level of PDCD5 gene in K562 cells was evidently increased following transfection with Ad-PDCD5. The Ad-PDCD5 synergistically enhanced the apoptotic percentage of K562 cells induced by VP-16, as compared with that of Ad-null + VP16 and VP-16 alone respectively. It is concluded that Ad-PDCD5 may be a potential agent for enhancing the chemotherapy effect.

Apoptosis is reduced in the synovial tissue of patients with rheumatoid arthritis (RA), possibly due to decreased expression of pro-apoptotic genes. Programmed Cell Death 5 (PDCD5) has been recently identified as a protein that mediates apoptosis. Although PDCD5 is down-regulated in many human tumors, the role of PDCD5 in RA has not been investigated. Here we report that reduced levels of PDCD5 mRNA and protein are detected in RA synovial tissue (ST) and fibroblast-like synoviocytes (FLS) than in tissue and cells from patients with osteoarthritis (OA). We also report differences in the PDCD5 expression pattern in tissues from patients with these two types of arthritis. PDCD5 showed a scattered pattern in rheumatoid synovium compared with OA, in which the protein labeling was stronger in the synovial lining layer than in the sublining. We also observed increased expression and nuclear translocation of PDCD5 in RA patient-derived FLS undergoing apoptosis. Finally, overexpression of PDCD5 led to enhanced apoptosis and activation of caspase-3 in triptolide-treated FLS. We propose that PDCD5 may be involved in the pathogenesis of RA. These data also suggest that PDCD5 may serve as a therapeutic target to enhance sensitivity to antirheumatic drug-induced apoptosis in RA.

The objective of this study was to estimate a novel apoptosis-promoting molecule PDCD5 expression in the bone marrow cells from adult acute myeloid leukemia (AML) for investigation of its significance in the pathogenesis of AML. Flow cytometry assay was used for detection of PDCD5 expression in the different groups of cells from bone marrow of AML patients and normal controls by using 21 monoclonal antibodies with different fluorescent markers. The PDCD5 expressions in bone marrow cells from some AML patients and normal controls were also detected by Western blot. The results showed that the mean PDCD5 fluorescence intensity in bone marrow nucleated cells (MNC) from the bone marrow of 36 untreated AML patients was significantly lower than that from the bone marrow of 30 normal controls (3059 +/- 1392) vs (7432 +/- 1261) (P < 0.01). The mean PDCD5 fluorescence intensity was lower in the marrow granulocytes, monocytes, blast cells, and lymphocytes from untreated AML patients than that from normal (3939 +/- 2121) vs (8367 +/- 1045); (3156 +/- 1635) vs (5917 +/- 2329); (2824 +/- 1592) vs (3998 +/- 2106); (1474 +/- 816) vs (3355 +/- 2042) respectively, (all P < 0.01). Western blot analysis demonstrated that PDCD5 expression was significantly decreased in the AML cells, as compared with normal cells. It is concluded that PDCD5 expression in MNC in untreated AML patients is lower than that in the normal. PDCD5 expression in the marrow granulocytes, monocytes, blast cells, and lymphocytes of untreated AML patients is significantly lower than that in the normal. It suggests that the abnormally low expression of PDCD5 may be involved in the pathogenesis of AML.

The objectives of the present study were to define whether amlodipine induces apoptosis and what mechanism is involved in the process in human resistant and non-resistant leukemia cells following co-administration of stealth liposomal topotecan with amlodipine, a novel antiresistant liposomes developed by our institution. In three leukemias, K562, HL-60, and multidrug resistant (MDR) HL-60, cytotoxicity of topotecan was potentiated by amlodipine, while topotecan alone was resistant to MDR HL-60 cells. In two selected K562 or MDR HL-60 cells, the apoptotic effects were increased by addition of amlodipine, showing a dose-dependent manner. The activities of caspase 3 and 7 (marked as caspase 3/7), and caspase 8 were significantly activated by topotecan with amlodipine co-treated as the stealth liposomes. The deletions of intracellular Ca2+ stores induced by amlodipine correlated with the activated activities of caspase 3/7, or 8, respectively. In xenograft model with MDR HL-60 in nude mice, antitumor activity of stealth liposomal topotecan with amlodipine was significantly enhanced as compared to that of stealth liposomal topotecan or topotecan alone. In conclusion, apoptotic effect is associated with deletion of intracellular Ca2+ by amlodipine through activation of caspase 8 and then 3/7 activities. The enhanced antitumor activities by stealth liposomal topotecan with amlodipine are mainly due to the potentiating apoptotic effect and reversing the resistance by amlodipine. Stealth liposomal encapsulation of anticancer agent with a modulator may provide a novel strategy for improving the chemotherapeutic effects.

Abstract: To clarify whether expression of the programmed cell death 5 (PDCD5) gene in leukemic cells is abnormal, real-time quantitative reverse transcription polymerase chain reaction (RQ-RT-PCR) was used to examine its expression in marrow cells from leukemia patients. We found lower PDCD5 in both AML and CML marrow cells than in normal donor marrow cells. A negative correlation was found between relative levels of PDCD5 and BCR/ABL expression in all CML patients and in CML patients in the advanced phase. Treatment with the ABL tyrosine kinase inhibitor Imatinib mesylate increased PDCD5 expression in K562 and MEG-01 cells. These findings suggest that abnormal expression of PDCD5 in leukemia may be involved in the pathomechanism of AML and CML. [Copyright &y& Elsevier]

To further elucidate the role of vascular endothelial growth factor (VEGF) in the pathogenesis of chronic myeloid leukemia (CML), we transfected K562 cells with a VEGF121cDNA sense vector (S), an antisense (AS) vector or vector (V) alone. The growth of transfected cells was investigated by MTT and colony-formation assays, and apoptosis was measured by flow cytometry (FCM) of Annexin-V-FITC/PI dual labeled cells. Transfected cells were subcutaneously transplanted into nude mice and the microvessel density (MVD) of tumor masses was determined by vWF immunohistochemistry staining. We tested the supernatant of different transfected K562 cells against human bone marrow endothelial cells (BMECs), and examined the synergic effects of antisense VEGF121cDNA and IFNα or STI571 on the proliferation and apoptosis of K562 cells. We found that K562/AS transfectants exhibited a 49% reduction in VEGF secretion, whereas K562/S transfectants exhibited a 3-fold increase in VEGF secretion, all in comparison to the vector controls. K562 cells transfected with antisense VEGF121cDNA showed growth retardation in vitro. In transplanted nude mice in vivo, transfection of implanted cells with antisense VEGF121cDNA resulted in decreased tumor MVD, and increased apoptosis in the presence of IFNα. Taken together, these results suggest that VEGF may be involved in the pathogenesis of CML through autocrine and paracrine mechanisms, and that anti-VEGF therapy alone or in combination with conventional treatment may be beneficial for CML patients. [Copyright &y& Elsevier]