Masafumi Kataoka

Activation of caspase-3 and cleavage of Rb are associated with p16-mediated apoptosis in human non-small cell lung cancer cells

The p16 tumor suppressor gene is frequently inactivated in human cancer tissues and cell lines. We previously reported that wild-type p16 expression from an adenovirus vector (Adv/p16) induced p53-dependent apoptotic cell death in non-small cell lung cancer (NSCLC) cell lines. Here we show the potential mechanism of apoptosis induced by Adv/p16 infection. Infection of human NSCLC cell line A549, which carries the wild-type p53 gene, with Adv/p16 resulted in activation of caspase-3, accompanied by the cleavage of its substrate poly (ADP-ribose) polymerase (PARP), on day 3 of infection. The retinoblastoma (Rb) cell cycle regulator protein was also cleaved after activation of caspase-3; when the levels of Rb significantly diminished, apoptosis began. When A549 cells were pretreated with the caspase-inhibitory peptide N-acetyl-asp-Glu-Val-Asp-CHO (aldehyde) (Ac-DEVD-CHO), Adv/p16-mediated apoptosis and Rb cleavage were greatly inhibited. Furthermore, MDM2, a negative regulator of p53 expression was upregulated 3 days after Adv/p16 infection, and MDM2 was subsequently cleaved by caspase-3; MDM2 cleavage was inhibited by Ac-DEVD-CHO treatment. These data implied that cleavage of Rb, in addition to activation of caspase-3, represented a mechanism by which Adv/p16 induced apoptotic cell death in human NSCLC cells. Our results support the clinical relevance of Adv/p16 as a treatment for p16-null human NSCLC that express wild-type p53.

Contribution of CD95 Ligand-Induced Neutrophil Infiltration to the Bystander Effect in p53 Gene Therapy for Human Cancer

Clinical trials of adenoviral p53 gene therapy provide the evidence that the bystander effect induced by the wild-type p53 gene transfer on adjacent tumor cells contributes to tumor progression; its mechanism, however, remains uncharacterized. We report in this work that injection of adenovirus expressing the human wild-type p53 gene (Ad5CMVp53) into established human colorectal tumors in nu/nu mice resulted in CD95 ligand (CD95L) overexpression, followed by a massive neutrophil infiltration. Culture supernatants of human colorectal cancer cells infected with Ad5CMVp53 exhibited a potent chemotactic activity against murine polymorphonuclear neutrophils, which could be abolished by the anti-CD95L mAb (NOK-1). In vivo cell depletion experiments indicated that neutrophils were in part responsible for the antitumor effect of the Ad5CMVp53 infection. Our data directly suggest that overexpression of CD95L by the wild-type p53 gene transfer induces neutrophil infiltration into human colorectal tumors, which may play a critical role in the bystander effect of p53 gene therapy.

Adenovirus-mediated p53 gene therapy for human cancer.

Recent advances in molecular biology have fostered remarkable insights into the molecular basis of neoplasms. Considerable evidence has accumulated that among the mechanisms of human cancer development are overexpression of dominant oncogenes, expression of mutant oncogenes, or specific chromosomal deletions or mutations that induce inactivation of tumor-suppressor genes. This understanding of cancer pathogenesis suggests that restoration of the function of critical gene products could halt or reverse these abnormalities, thus having a therapeutic effect. The p53 tumor suppressor gene has been implicated in many inherited and sporadic forms of malignancies in humans. Preclinical experiments have demonstrated that restoration of wildtype p53 function in the cancer cell by gene transfer is sufficient to cause antitumor effects such as cell-cycle arrest and induction of apoptosis. This approach has entered clinical testing and provided intriguing information about the intratumoral administration of an adenovirus vector expressing the wildtype p53 gene in non-small-cell lung cancer. The clinical study has also provided evidence of the bystander phenomenon, which is important for potential clinical efficacy. This article reviews recent highlights in this rapidly evolving field: p53 gene therapy for human cancer.

Decreased expression of DCC mRNA in gastric and colorectal cancer

The deleted-in-colorectal-cancer (DCC) gene, located on chromosome 18q 21.3, is considered to be a tumor suppressor gene related to cellular adhesion receptors. A loss of heterozygosity (LOH) on chromosome 18q is frequently observed in adenomatous polyposis coli, as well as in sporadic colon carcinoma and its liver metastatic loci. In this study, we investigated the expression of DCC mRNA in the resected specimens of 38 gastric cancers and 28 colorectal cancers by a reverse transcription-polymerase chain reaction method. In the gastric cancer patients, the mean expression level of DCC mRNA in the tumors was significantly lower than that in normal tissues (P=0.009), but no difference was observed in the colorectal cancer patients. DCC mRNA expression was decreased in 15 gastric cancers (40%) and 10 colorectal cancers (36%), and there was a significant correlation between the decreased expression of DCC mRNA and nodal metastasis in colorectal cancer (x2=7.049, DF=1, P=0.0079). Two of four gastric cancer patients and none of seven colorectal cancer patients whose cancers were confined to the muscularis propria without metastasis showed decreased expression of DCC mRNA. These findings demonstrate that decreased expression of DCC mRNA may occur at an early stage in gastric cancer and at a late stage in colorectal cancer and that this decreased expression correlates with the potential to develop nodal metastasis.

Randomized feasibility study of S-1 for adjuvant chemotherapy in completely resected Stage IA non–small-cell lung cancer: results of the Setouchi Lung Cancer Group Study 0701

The feasibility of the S-1 administration schedules (the 4-week versus the 2-week) showed no significant difference for adjuvant chemotherapy among pathological-Stage IA non–small-cell lung cancer patients.

A case of a pancreatic tumor that was diagnosed as metastasis from lung cancer by endoscopic ultrasound-guided fine needle aspiration.

Metastatic tumors are rare in the pancreas, and some cases are difficult to distinguish from pancreatic cancer. However, distinguishing between them is very important to formulate a treatment plan. A case of a rare disease, called overlap cancer, involving metastatic tumors to the pancreas and right kidney from lung cancer, and duodenal papilla cancer, is described. Endoscopic ultrasound-guided fine needle aspiration (EUS-FNA) is useful for diagnosing metastatic pancreatic tumors, particularly in patients with multiple cancers.

Abstract 4976: Telomerase-specific replication-selective virotherapy combined with radiation therapy for oral squamous cell carcinoma cells

Proceedings: AACR 103rd Annual Meeting 2012‐‐ Mar 31‐Apr 4, 2012; Chicago, IL Replication-selective tumor-specific viruses present a novel approach for treating neoplastic disease. These vectors are designed to induce virus-mediated lysis of tumor cells after selective viral propagation within the tumor. Telomerase activation is considered to be a critical step in carcinogenesis, and its activity is closely correlated with human telomerase reverse transcriptase (hTERT) expression. We previously, investigated the antitumor effect of the hTERT-specific replication-competent adenovirus (Telomelysin: OBP-301) on oral squamous cell carcinoma (OSCC) cells in vitro and in vivo orthotopic graft model. OBP-301 replicated efficiently and induced more over 50% cell killing in a panel of human OSCC cell lines, but not in normal human fibroblasts, which were lacking telomerase activity. In orthotopic graft model, intratumoral injection of OBP-301 resulted in a significant inhibition of tumor growth and prolongation of survival. Moreover, OBP-301 in use has limited potency when administered alone; however, combination therapy using these agents and conventional anticancer agents, such as radiation exhibits encouraging levels of efficacy. Here, we describe the mechanistic basis for synergy of irradiation and OBP-301. This combination therapy was more effective compared with OBP-301 alone in OSCC cells. These results illustrate the potential of combining oncolytic virotherapy and ionizing radiation as a promising strategy in the management of human oral cancer. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 4976. doi:1538-7445.AM2012-4976

Die Kombination aus systemisch verabreichtem adenoviralen p53 Gentransfer und 2-Methoxyestradiol führt zu additiver Tumorhemmung in vivo

Inspite of high transduction efficiencies in vitro after adenoviral gene transfer, the number of viral particles remains low in the tumor after systemic administration, which results in low efficiency of this therapeutic approach. We investigated the combination of adenoviral gene transfer using wt-p53 (Ad-p53) with 2-methoxyestradiol (2-ME2), which has been shown to stabilize the p53 protein, increasing the rate of apoptosis. A small cell lung cancer cell line A549 has been injected in mice to induce lung metastases. Intravenous injection of Ad-p53 alone did not show any tumor inhibition. 2-ME2 alone resulted in a reduction of the lung colonies of 42.3% (p < 0.0001). The combination of 2-ME2 and Ad-p53 resulted in a more than additive reduction of lung colonies to 72.6% (p < 0.05 against 2-ME2 alone). The average diameter of the single tumor nodules was also reduced. In non-treated control mice, we found an average tumor size of 245 µm. After 2-ME2 alone, the size was reduced to 108 µm, and in the combination group it was further reduced to 54 µm (p = 0.0001). The total tumor burden was reduced by 336 fold compared to control non-treated mice. This is the first report, showing a significant tumor inhibition by systemically administered adenoviral gene transfer.