Activation of cyclin D1-kinase in murine fibroblasts lacking both p21(Cip1) and p27(Kip1)

Deregulation of D-type cyclin-dependent kinases (CDK4 and 6) is widely observed in various human cancers, illustrating their importance in cell cycle control. Like other cyclin-dependent kinases (CDKs), assembly with cyclins is the most critical step for activation of CDK4/6. As previously reported elsewhere, we observed that the level of cyclinD1-CDK4 complex and its associated kinase activity were significantly low in asynchronously proliferating mouse embryo fibroblasts lacking both p21(Cip1) and p27(Kip1) (p21/p27-null MEFs). These evidences imply that p21(Cip1) and p27(Kip1) CDK inhibitors are 'essential activators' of cyclin D-kinases. We, however, discovered here that both the assembly and activation of cyclin D1-CDK4 complex occur when quiescent p21/p27-null MEFs were stimulated to re-enter the cell cycle. This mitogen-induced cyclin D1-kinase activity was blocked by overexpression of p16(INK4a) and resulted in the inhibition of S phase entry in p21/p27-null MEFs. Furthermore, ectopic expression of p34(SEI-1), a mitogen-induced CDK4 binding protein, increased the levels of active cyclinD1-CDK4 complex in asynchronously proliferating p21/p27-null MEFs. Together, our results suggest that there are several independent ways to stimulate the assembly of cyclin D1-CDK4 kinases. Although p21(Cip1) and p27(Kip1) play a role in this process, our results demonstrate that additional mechanisms must occur in G0 to S phase transition.     

Survivin expression and its correlation with cell proliferation and prognosis in epithelial ovarian tumors

Survivin is a new member of the inhibitors of apoptosis proteins (IAP) family, selectively overexpressed in common human cancers but not in normal adult tissues, and associated with aggressiveness of the disease and unfavorable outcomes. Recent study also found that survivin expression is associated with cell proliferation. In order to gain insight into the role of survivin in ovarian tumors, we investigated the expression of survivin in a group of epithelial ovarian tumors, and examined the relationship of its expression with cell proliferation and clinical outcome. Immunohistochemical analysis was performed in 103 cases of epithelial ovarian tumors. Twenty-six of the 103 cases were evaluated by Western blot analysis. The results showed that survivin overexpression was detected in 21.2% (7 of 33) of benign tumors, 47.8% (11 of 23) of borderline tumors, and 51.1% (24 of 47) of ovarian carcinomas. The positive ratio was significantly higher in malignant or borderline tumors than in benign tumors, and the overexpression of survivin was significantly correlated with the size of residual disease. A positive correlation between survivin expression and proliferative activity of tumor cell measured by PCNA index was found. Kaplan-Meier analysis demonstrated that the patients with survivin overexpression have a short overall survival. These findings suggest that survivin overexpression may play a pivotal role in the progression of ovarian tumors and may provide an important prognostic implication for epithelial ovarian carcinomas.     

Metastatic urinary bladder tumor from extragonadal germ cell tumor: a case report

A 37-year old male patient complained of lower back pain. Investigations revealed a retroperitoneal tumor and left-sided cervical lymphadenopathy without abnormal findings in both testicles. The diagnosis of extragonadal germ cell tumor was made based on a lymph node biopsy. Following chemotherapy and retroperitoneal lymphoadenectomy, at outpatient follow-up, beta-HCG elevated again. He complained of gross hematuria, cystoscopy showed a bladder tumor, and abdominal MRI scan showed a right ureteral tumor. Total cystectomy and right nephroureterectomy were performed, which revealed that the bladder and ureteral tumors were metastatic germ cell tumors. Six months later, the patient developed hepatic and mesenteric lymph node metastases that failed to respond to treatment, and died. Germ cell tumors metastasizing to the urinary tract are extremely uncommon, and this is the first report of bladder metastases, other than through direct invasion, from an extratesticular germ cell tumor.     

Successful unrelated cord blood transplantation for relapse after autologous transplantation in non-Hodgkin's lymphoma

We report a 34-year-old male with relapsed non-Hodgkin's lymphoma (NHL) after autologous peripheral blood stem cell transplantation successfully treated with unrelated cord blood transplantation (CBT). The conditioning regimen included 12 Gy total body irradiation and cyclophosphamide. After the conditioning, a total of 3.14 x 10(7)/kg cord blood nucleated cells was infused on 14 February 2000. An absolute neutrophil count greater than 5 x 10(8)/l and a self-sustained platelet count greater than 50 x 10(9)/l were achieved on days 21 and 43, respectively. During the follow up period, grade I acute graft-versus-host disease (GVHD) and limited chronic GVHD occurred, but both were successfully treated with a dose modification of cyclosporine. After a follow-up period of 16 months, the patient is alive and free of disease. To our knowledge this is the first report of a successful unrelated CBT for an adult NHL patient who relapsed after autologous transplantation.     

Impaired infiltration of tumor-specific cytolytic T cells in the absence of interferon-gamma despite their normal maturation in lymphoid organs during CD137 monoclonal antibody therapy

Engagement of CD137 receptor by agonistic monoclonal antibodies (mAb) stimulates IFN-gamma production and eradicates established tumors in syngeneic mouse models. Using IFN-gamma-deficient mice or neutralizing mAb, we demonstrate that IFN-gamma is an absolute requirement for the antitumor effect of CD137 mAb. Despite progressive tumor growth in IFN-gamma-depleted mice, a fully competent CD8(+) cytolytic T cell (CTL) response developed in the lymph nodes. In addition, tumor cell sensitivity to IFN-gamma was not required because expression of a dominant-negative IFN-gamma receptor on the tumor did not affect the therapeutic effect of CD137 mAb. However, in the absence of IFN-gamma, the number of tumor-infiltrating CD8(+) CTLs was drastically decreased. Our results demonstrate that IFN-gamma is a critical factor regulating the infiltration of antigen-specific CTL into the tumor.     

Strategies for antigen loading of dendritic cells to enhance the antitumor immune response

Dendritic cells (DCs) primed with tumor antigens can effectively mediate the regression of a variety of established solid malignancies in both murine and human models. Despite such clinical efficacy, the optimal means of DC priming is unknown. The goal of this study was to compare three methods of tumor preparation: irradiation, boiling, or freeze thaw lysis for DC priming. Mouse bone marrow-derived DCs were loaded with defined ratios of E.G7 tumor cells expressing a model tumor antigen, OVA. Sensitized DCs were used for stimulation of OVA-specific CTLs derived from OT-1 T-cell receptor transgenic mice. IFN-gamma release, determined by ELISA at 24 and 48 h, was used to assess the expression of antigens by DCs. DCs loaded with irradiated tumors were effective stimulators for OT-1 CTLs, whereas DCs stimulated with freeze-thawed or boiled tumors did not stimulate IFN-gamma production. Freeze-thaw lysis appeared to inhibit CTL activity in vitro and in two of three cases, this effect was not overcome by the addition of OVA. The ability to load irradiated tumor cells was reproduced in two analogous human melanoma models using melanoma cell lines expressing gp100 and CTL clones specific for a gp100 melanoma antigen. Consistent with the in vitro data, only DC/irradiated tumor vaccines were effective in preventing or delaying outgrowth of E.G7 and a poorly immunogenic murine squamous cell carcinoma (SCCVII), on local tumor challenge. These data demonstrate that the method of tumor cell preparation clearly influences the ability of DCs to present antigen to T cells. Correlation of in vitro data with the generation of protective immunity in vivo suggests the utility of irradiated tumor-primed DCs as a means to generate protective immunity in patients with solid malignancies.     

Cell therapy for renal cell carcinoma

Renal cell carcinoma (RCC) is an immunogenic tumor that has shown some response to cytokine-therapy and other types of immune-based treatment. Since the advent of lymphocyte culture techniques in the 1980s, clinical trials of lymphokine-activatedkiller cells and tumor infiltrating lymphocytes have been conducted. Although these approaches have not shown apparent benefit compared to the standard cytokine therapy, further trials are ongoing using dendritic cell or gene-modified tumor vaccines in order to induce a tumor-specific cytotoxic T cell response in vivo. Recently, several investigators have indicated that RCC is susceptible to a graft-versus-tumor effect promoted by allogeneic stem-cell transplantation. This article reviews the clinical results of these cell therapies for metastaic RCC.     

A case of locally extended rectal cancer responding to intra-arterial infusion therapy via the internal iliac artery

Chemotherapy with combined administration of 5-FU and l-Leucovorin has been reported to be effective for advanced rectal cancer. A 61-year-old woman with a huge, locally extended advanced rectal cancer was treated with intra-arterial infusion therapy via internal iliac artery. Catheters were inserted from the bilateral femoral arteries to the opposite internal iliac arteries and the bilateral upper and lower gluteal arteries and lateral sacral artery were embolized with a metallic coil. The port was positioned under the skin of her lower abdominal wall. The chemotherapy regimen was 5-FU (500 mg/body) and l-Leucovorin (250 mg/m2), administered over 5 hours once weekly to bilateral reservoirs through an infuser pump. After 5 sessions of the chemotherapy, the perineal pain decreased and the patient no longer needed morphine. Following 34 administrations (at 10 months) of this regimen, reductions of tumor size on pelvic CT and CEA level were confirmed. Side effects of the intra-arterial infusion chemotherapy were pygal dermatitis, leg desensitization, infection of circumferential reservoir and obstruction of the catheter due to flection were observed. Local intra-arterial infusion chemotherapy via the internal iliac artery appears to be effective for local advanced rectal cancer.     

A case of postoperative multiple hepatic metastasis from gastric cancer successfully treated by percutaneous microwave coagulation therapy and hepatic arterial infusion chemotherapy

A 61-year-old male who underwent radical resection for gastric cancer was diagnosed with multiple hepatic metastasis 2 years and 2 months after the surgery. He first underwent percutaneous microwave hepatic coagulation therapy with segmental hepatic blood flow occlusion and obtained complete coagulation of the main tumor. Consecutively, he received hepatic arterial infusion chemotherapy (FAP: 5-FU, cisplatin, adriamycin) against residual multiple hepatic tumors. These hepatic recurrent lesions disappeared completely after 3 sessions of arterial infusion chemotherapy. At present, this patient is alive with no recurrent lesions, 1 year and 6 months from the beginning of treatment for hepatic metastasis. Recently, we tried hepatic arterial infusion chemotherapy (FAP) in four cases in which the recurrence from gastric cancer was not only in the liver but elsewhere. The response rate (CR and PR) was 75% and no major side effects were observed. In conclusion, some cases can obtain longer survival if the multimoderate therapy including hepatic arterial infusion chemotherapy (FAP) and microwave coagulation therapy are effective.