Daishiro Kato

Features of replicative senescence induced by direct addition of antennapedia-p16INK4A fusion protein to human diploid fibroblasts

The p16INK4A cyclin‐dependent kinase (Cdk) inhibitor is now recognized as a major tumor suppressor that is inactivated by a variety of mechanisms in a wide range of human cancers. It is also implicated in the mechanisms underlying replicative senescence since p16INK4A RNA and protein accumulate as cells approach their proscribed limit of population doublings in tissue culture. To obtain further evidence of its role in senescence, we have sought ways of overexpressing p16INK4A in primary human diploid fibroblasts (HDF). To circumvent the low transfection efficiency of primary cells we have exploited a recombinant form of the full‐length p16INK4A protein fused to a 16 amino acid peptide from the Drosophila antennapedia protein. This peptide has the capacity to cross both cytoplasmic and nuclear membranes allowing the direct introduction of the active protein to primary cells. Here, we show that antennapedia‐tagged wild‐type p16INK4A protein, but not a functionally compromised tumor‐specific variant, causes G1 arrest in early passage HDFs by inhibiting the phosphorylation of the retinoblastoma protein. Significantly, the arrested cells display several phenotypic features that are considered characteristic of senescent cells. These data support a role for p16INK4A in replicative senescence and raise the possibility of using the antennapedia‐tagged protein therapeutically.

Quadrilateral Space Syndrome: A Rare Complication of Thoracic Surgery

We report a case of quadrilateral space syndrome related to thoracic surgery. A 21-year-old man underwent video-assisted thoracic surgery for a left-sided pneumothorax. After the operation, he presented with difficulties in left arm abduction and paresthesia over the lateral aspect of the shoulder and upper arm. Deltoid muscle atrophy and tenderness over the quadrilateral space were also observed. On further examination, he was diagnosed with isolated paralysis of the axillary nerve, the so-called quadrilateral space syndrome. This is a rare complication, but it interferes with the activities of daily living, and thus one should pay attention to this syndrome.

Drug resistance to paclitaxel is not only associated with ABCB1 mRNA expression but also with drug accumulation in intracellular compartments in human lung cancer cell lines.

In order to clarify the mechanisms of resistance to paclitaxel in lung cancer, three human lung cancer cell lines which exhibit different sensitivity to paclitaxel were investigated from the following viewpoints: overexpression of ATP-binding cassette, sub-family B, member 1 (ABCB1), mutations on paclitaxel binding site of β-tubulin genes, quantity of polymerized tubulin and the intracellular localization of paclitaxel. ABCB1 expression was evaluated by real-time RT-PCR. No correlations were noted between the ABCB1 expression in the sensitive and resistant cell lines at the mRNA level. No mutations on the paclitaxel binding site of the β-tubulin genes were detected in either the resistant or sensitive cells. Live cell images obtained by confocal laser microscopy revealed that the resistant cell line, RERF-LC-KJ, had more accumulation of Oregon Green® 488 conjugated paclitaxel in the lysosomal and extra-lysosomal compartments of cytoplasm than other cell lines. The results obtained in this study indicated that the changes in the subcellular localization could contribute to the production of paclitaxel resistance in lung cancer cell lines. Further studies should be conducted to elucidate the molecular mechanisms that differentiate the intracellular localization of paclitaxel.

Growth Inhibition of A549 Human Lung Adenocarcinoma Cells by L‐Canavanine Is Associated with p21/WAF1 Induction

L‐Canavanine (CAV) is a higher plant nonprotein amino acid and a potent L‐arginine antimetabolite. CAV can inhibit the proliferation of tumor cells in vitro and in vivo, but little is known regarding the molecular mechanisms mediating these effects. We demonstrated that the treatment of human lung adenocarcinoma A549 cells with CAV caused growth inhibition; G1 phase arrest is accompanied by accumulation of an incompletely phosphorylated form of the retinoblastoma protein, whose phosphorylation is necessary for cell cycle progression from G1 to S phase. In addition, CAV induces the expression of p53 and subsequent expression of a cyclin‐dependent kinase inhibitor, p21/WAF1. The p53–dependent induction of p21/WAF1 and the following dephosphorylation of the retinoblastoma protein by CAV could account for the observed CAV‐mediated G1 phase arrest.

CT findings after lipiodol marking performed before video-assisted thoracoscopic surgery for small pulmonary nodules

Background In preoperative lipiodol marking for small pulmonary nodules, lipiodol has a potential risk of distribution in the surrounding lung structure. There are no reports about the detailed accumulation and distribution of lipiodol. Purpose To evaluate computed tomography (CT) findings after lipiodol marking before thoracoscopic surgery for pulmonary nodules. Material and Methods Sixty-four consecutive CT-guided lipiodol markings for 103 nodules were performed in 55 patients. Lipiodol (0.2–0.4 mL) was injected using a 21-gauge needle near the nodule. The appearance of lipiodol spots was classified into the following three types on CT: type 1, dense; type 2, punctate; and type 3, unclear. The distribution of lipiodol was also investigated. Statistical analyses were performed on the accumulation and distribution related to nodule factors. Incidences of complications were also investigated. Results A total of 110 markings were performed because of seven additional procedures due to insufficient marking. All nodules were successfully resected on the same day. The appearances of the lipiodol spots were type 1 (82%), type 2 (11%), and type 3 (7.3%). The areas of distribution were lung parenchyma (54%), central bronchus (39%), peripheral bronchovascular bundle (24%), needle tract (20%), pleural space (19%), another segment of ipsilateral lung (5.5%), and contralateral lung (0.9%). Distribution into pleural space and central bronchus was frequently seen in the shallow nodules (P < 0.05). Complications were pneumothorax (61%) and pulmonary hemorrhage (35%). There were no serious symptoms. Conclusion The appearance of the lipiodol spot was dense in most cases, despite frequent distribution in the surrounding lung structures without serious complications.

Lung metastasis of thymoma manifesting as myasthenia gravis 12 years after thymomectomy: report of a case.

A 51-year-old woman, who had undergone thymomectomy for asymptomatic noninvasive thymoma 12 years before, was admitted to our hospital with blepharoptosis caused by myasthenia gravis. A chest computed tomogram (CT) showed an abnormal shadow in the right lower lung field and CT-guided needle biopsy revealed findings of a thymoma. We performed extended thymectomy and partial resection of the right lung using three different approaches, via a cervical incision and bilateral video-assisted thoracoscopic surgery. The resected specimen was a lung metastasis of thymoma, and the residual mediastinal tissue showed no sign of malignancy. Because thymoma and post-thymomectomy myasthenia gravis can both recur, we recommend performing extended thymectomy or thymothymectomy, even for patients who are asymptomatic.

Pneumonia occurring after injection of Lipiodol to localize pulmonary nodules before fluoroscopy-aided thoracoscopic resection:

We report the case of a 33-year-old man with a testicular tumor and multiple lung metastases. After high orchiectomy, chemotherapy was administered. Subsequently, fluoroscopy-assisted thoracoscopic resection was performed for pulmonary lesions that had decreased in size after the chemotherapy but had persisted. Because four pulmonary lesions were too small to detect by fluoroscopy during the operation, Lipiodol marking was done before thoracoscopic resection. Sixteen days after the surgery, pneumonia after Lipiodol injection occurred on the lung contralateral to that injected with Lipiodol. Pneumonia improved rapidly after intravenous infusion of antibiotics.

Intuitive Touch Panel Navigation System through Kyoto Digital Sosui Network

We have developed multi-touch panel robotic arm control system for thoracoscopic surgery. Our multi-touch panel navigation system consists of green laser guide maker, robotic arms with four spindles, multi-touch panel monitor, and PCs for control regulation. The multi-touch panel monitor recognized the finger-touch; thereby the tip of the robot arm is controlled just on the green maker. For estimation in remote control use, the PC for the robot control was placed in Kyoto Prefectural University of Medicine, which was connected through the Kyoto Digital Sosui Network, a gigabit Ethernet with layer-2 security. Another computer for the remote control of the PC for the robot control was placed in Yamashiro Public Hospital, 35 kilometers south of Kyoto city, through the same gigabit Ethernet. Using virtual network computing (VNC), which is a graphical desktop sharing system to control another computer, we demonstrated a remote control of the robot and collected the packets to examine the traffic bandwidth. The robot was successfully controlled in remote conditions. The throughput of the Ethernet was 16.4 ±3.6 Mbps in the remote control of the robot using VNC. The round-trip time was 19.9 ± 0.39 msec, while 8000 bytes data was sent by using ping command. We demonstrated an intuitive touch panel navigation system, which a surgical robot stopped an arterial bleeding in an animal model. The robot system was successfully controlled through a gigabit Ethernet between two distant hospitals.

Significance of PD-L1 expression in pulmonary metastases from head and neck squamous cell carcinoma

BACKGROUND The mechanism by which tumors escape the immune system has been actively investigated and is partly explained by the programmed death-1 (PD-1) and its ligand (PD-L1) pathway. This study is aimed at clarifying the prognostic significance of PD-L1 expression in patients with surgically resected pulmonary metastases of head and neck squamous cell carcinoma (HNSCC). PATIENTS AND METHODS A retrospective review was conducted of 26 HNSCC patients who underwent complete resection of pulmonary metastases. PD-L1 expression in both the primary and metastatic tumors was evaluated using immunohistochemistry (anti-PD-L1 antibody, clone SP263). High PD-L1 expression was defined as ≥50% of tumor cells with positive staining. Survival and prognostic impacts following pulmonary metastasectomy were analyzed based on the PD-L1 expression level. RESULTS The patients included 23 men and 3 women, with a median age of 65 years. Six (23%) of the pulmonary metastatic cases showed high PD-L1 expression, while their corresponding primary lesions had low PD-L1 expression. The 5-year overall survival rate after pulmonary metastasectomy was 57.6% in all cases. The 5-year overall survival rates were 72.5% and 16.7% in the low and high PD-L1 groups, respectively (p < 0.001). Multivariate analysis demonstrated that high PD-L1 expression and older age (>65 years) correlated independently with a shorter overall survival (p < 0.001). CONCLUSIONS High PD-L1 expression in pulmonary metastases could be an independent predictor of poor outcome in HNSCC patients undergoing pulmonary metastasectomy. This is the first report evaluating the prognostic implication of PD-L1 expression in metastatic HNSCC.

Successful treatment with afatinib for pancreatic metastasis of lung adenocarcinoma: a case report

Metastatic lung cancer of the pancreas is rare and optimal treatment protocols have not been determined. An asymptomatic 53-year-old man with primary lung cancer underwent a right lower lobectomy and was diagnosed with acinar adenocarcinoma, pT2bN2M0 stage IIIA. An epidermal growth factor receptor (EGFR) mutation (exon 19 deletion L747-A750insP) was detected in the primary tumor. Adjuvant platinum-based chemotherapy was administered. The patients serum carcinoembryonic antigen (CEA) level had increased to 38.6 ng/mL 32 months after surgery, and positron emission tomography-computed tomography (PET-CT) revealed a 2.5-cm, hypermetabolic nodule in the pancreatic body. Using endoscopic ultrasound-guided fine-needle aspiration (EUS-FNA), the nodule was pathologically diagnosed as a pancreatic metastasis of lung adenocarcinoma. The EGFR status of the pancreatic metastasis was confirmed to be the same as that of the primary lung tumor. The patient started afatinib therapy and his serum CEA level immediately decreased and remained at approximately the reference limit. On a follow-up PET-CT scan after 10 months of treatment with afatinib, the metastatic nodule in the pancreas had disappeared and no accumulation of fluorine-18-deoxyglucose (FDG) was detected. To the best of our knowledge, this is the first reported case of a complete response of pancreatic metastatic lung adenocarcinoma to EGFR-tyrosine kinase inhibitors (TKIs).