Kazuma Masumoto

Small interfering RNA targeting epidermal growth factor receptor enhances chemosensitivity to cisplatin, 5-fluorouracil and docetaxel in head and neck squamous cell carcinoma.

Overexpression of epidermal growth factor receptor (EGFR) has been found in various epithelial malignancies, including head and neck squamous cell carcinoma (HNSCC), and is associated with increased tumor growth, metastasis, resistance to chemotherapeutic agents and poor prognosis. As such, EGFR is a potential target for antitumor therapy and several EGFR inhibitors have been investigated in preclinical or clinical settings. In the present study, we used small interfering RNA (siRNA) to downregulate EGFR expression while evaluating the effect of EGFR siRNA on cell proliferation, and the combined effects with cisplatin, 5‐fluorouracil (5‐FU) and docetaxel in HNSCC. Furthermore, HNSCC xenografts were treated with EGFR siRNA alone or in combination with cisplatin, and tumor growth was examined. EGFR expression, proliferation, angiogenesis and apoptosis index were evaluated by immunohistochemistry. The results showed that EGFR siRNA efficiently downregulated EGFR expression and inhibited cell growth of HNSCC. Treatment with EGFR siRNA in combination with cisplatin, 5‐FU and docetaxel enhanced chemosensitivity with a significant increase in apoptosis. EGFR siRNA delivered by atelocollagen enhanced the antitumor effect of cisplatin in the HNSCC xenograft model. These cumulative results suggest that EGFR siRNA combined with cisplatin, 5‐FU and docetaxel may be a feasible strategy to enhance the effects of chemotherapy in patients with HNSCC. (Cancer Sci 2006; 97: 1115–1124)

Detection of skin perforators by indocyanine green fluorescence nearly infrared angiography.

Perforator-based island flaps are widely used to reconstruct skin defects. For this procedure to succeed, a perforator with adequate blood flow must be selected, and precise preoperative prediction of the location of the perforators is required. To identify perforators, a variety of methods have bee

Interstitial photodynamic therapy with rotating and reciprocating optical fibers

Photodynamic therapy (PDT) is an effective treatment modality that allows selective destruction of malignant tumor cells. However, because of the difficulty in exposing deeper areas of tumors, the modality has strictly limited indications. In this study, the authors introduce a new method for delivering laser light to a three‐dimensional, wide area with the purpose of improving the therapeutic value of PDT.

Imaging mass spectrometry distinguished the cancer and stromal regions of oral squamous cell carcinoma by visualizing phosphatidylcholine (16:0/16:1) and phosphatidylcholine (18:1/20:4)

Most oral cancers are oral squamous cell carcinoma (OSCC). The anatomical features of OSCC have been histochemically evaluated with hematoxylin and eosin. However, the border between the cancer and stromal regions is unclear and large portions of the cancer and stromal regions are resected in surgery. To reduce the resected area and maintain oral function, a new method of diagnosis is needed. In this study, we tried to clearly distinguish the border on the basis of biomolecule distributions visualized by imaging mass spectrometry (IMS). In the IMS dataset, eleven signals were significantly different in intensity (p < 0.01) between the cancer and stromal regions. Two signals at m/z 770.5 and m/z 846.6 were distributed in each region, and a clear border was revealed. Tandem mass spectrometric (MS/MS) analysis identified these signals as phosphatidylcholine (PC) (16:0/16:1) at m/z 770.5 in the cancer region and PC (18:1/20:4) at m/z 846.6 in the stromal region. Moreover, the distribution of PC species containing arachidonic acid in the stromal region suggests that lymphocytes accumulated in response to the inflammation caused by cancer invasion. In conclusion, the cancer and stromal regions of OSCCs were clearly distinguished by use of these PC species and IMS analysis, and this molecular identification can provide important information to elucidate the mechanism of cancer invasion.

Tissue distribution of a new photosensitizer ATX-S10Na(II) and effect of a diode laser (670 nm) in photodynamic therapy

The aims of the present study were to analyse the quantitative tissue distribution of ATX-S10Na(II) and to investigate the maximal effect of a diode laser and the irradiation conditions required to obtain this effect in photodynamic therapy (PDT) with ATX-S10Na(II). Spectrofluorometry was used to obtain quantitative tissue distribution of ATX-S10Na(II) in Colon 26 carcinoma-bearing mice as a function of time following administration. Next, transplanted tumours of mice with or without ATX-S10Na(II) were treated with the diode laser under conditions in which power density and irradiation time were varied. Tumour tissue concentrations of ATX-S10Na(II) were higher than in all tissues at all intervals following administration. The uptake of ATX-S10Na(II) by most tissues was rapid, with maximal concentrations occurring 1 h after i.v. injection, and ATX-S10Na(II) was almost excreted within 24 h after administration. The maximal depth of necrosis induced by PDT in the treated tumour was 7.9 mm under conditions in which power density was 160 mW/cm2 and total dose was above 100 J/cm2. PDT with ATX-S10Na(II) and the diode laser is useful for the treatment of superficial cancers.

Photodynamic therapy with intradermal administration of 5-aminolevulinic acid for port-wine stains.

Objective: To investigate whether intradermal administration of 5-aminolevulinic acid (5-ALA) could achieve a sufficient amount of protoporphyrin IX (PpIX) to induce photochemical reaction in chicken comb, the animal model of port wine stains (PWSs). Methods: PpIX accumulation after 5-ALA administration through intradermal or intravenous injection was monitored for 24 hours. Localization of PpIX was observed under a confocal microscope. The comb was exposed to red light after intradermal 5-ALA injection, and the subsequent changes were observed grossly and microscopically. Results: In the comb, PpIX accumulation achieved the peak level at 5 and 4 hours after intravenous or intradermal injection of 5-ALA, respectively, and was almost completely eliminated within 24 hours. A similar amount of PpIX was observed in both groups. While in the body skin, a lower level of PpIX was observed after intradermal injection. A confocal microscope showed that PpIX distributed evenly in comb dermis without significant difference between the two groups. The vascular structure in comb was disrupted after laser irradiation based on intradermal administration of 5-ALA. Conclusions: Intradermal injection of 5-ALA is a safer administration route that could achieve the equivalent of PpIX accumulation and destroy vasculature after PDT. It might be applicable to the clinical treatment of PWSs.

Regulation of GATA-binding protein 2 levels via ubiquitin-dependent degradation by Fbw7: Involvement of cyclin B-cyclin-dependent kinase 1-mediated phosphorylation of Thr176 in GATA-binding protein 2

Background: SCF-Fbw7 participates in stability control of several Cdc4 phosphodegron-containing proteins phosphorylated by glycogen synthase kinase 3, an E3 ligase. Results: Ubiquitin-dependent degradation of GATA-binding protein 2 is promoted by Fbw7, is cyclin B-CDK1-mediated Thr176 phosphorylation-dependent, and influences hematopoietic cell differentiation. Conclusion: GATA-binding protein 2 is a novel target for Fbw7. Significance: The molecular mechanism of post-transcriptional control of GATA-binding protein 2 is clarified. A GATA family transcription factor, GATA-binding protein 2 (GATA2), participates in cell growth and differentiation of various cells, such as hematopoietic stem cells. Although its expression level is controlled by transcriptional induction and proteolytic degradation, the responsible E3 ligase has not been identified. Here, we demonstrate that F-box/WD repeat-containing protein 7 (Fbw7/Fbxw7), a component of Skp1, Cullin 1, F-box-containing complex (SCF)-type E3 ligase, is an E3 ligase for GATA2. GATA2 contains a cell division control protein 4 (Cdc4) phosphodegron (CPD), a consensus motif for ubiquitylation by Fbw7, which includes Thr176. Ectopic expression of Fbw7 destabilized GATA2 and promoted its proteasomal degradation. Substitution of threonine 176 to alanine in GATA2 inhibited binding with Fbw7, and the ubiquitylation and degradation of GATA2 by Fbw7 was suppressed. The CPD kinase, which mediates the phosphorylation of Thr176, was cyclin B-cyclin-dependent kinase 1 (CDK1). Moreover, depletion of endogenous Fbw7 stabilized endogenous GATA2 in K562 cells. Conditional Fbw7 depletion in mice increased GATA2 levels in hematopoietic stem cells and myeloid progenitors at the early stage. Increased GATA2 levels in Fbw7-conditional knock-out mice were correlated with a decrease in a c-Kit high expressing population of myeloid progenitor cells. Our results suggest that Fbw7 is a bona fide E3 ubiquitin ligase for GATA2 in vivo.

Improved technique for evaluating oral free flaps by pinprick testing assisted by indocyanine green near-infrared fluorescence angiography

In head and neck surgery, free-flap reconstruction using a microvascular anastomosis is an indispensable option after tumor ablation. Because the success of free-flap reconstruction is enhanced by rapid identification and salvage of failing flaps, postoperative monitoring of free flaps is essential. We describe a new technique using indocyanine green (ICG) near-infrared angiography and pinprick testing to monitor intraoral free flaps. A solution of ICG (Diagnogreen, 5 ml) was intravenously injected, and scanning was performed with a near-infrared video camera system. Thirty seconds after ICG injection, a pinprick test was performed by placing a 24-gage needle through the dermis to the subcutaneous fat of the flap. Pinprick testing during ICG fluorescence imaging was performed in 30 patients. Flap perfusion was confirmed in all patients, and all flaps survived postoperatively. ICG fluorescence imaging demonstrated that flap perfusion was maintained.

E3 Ubiquitin Ligases as Molecular Targets in Human Oral Cancers

The ubiquitin-proteasome pathway is involved in various biological processes. Several oncogenic E3 ligases target tumor suppressor proteins for ubiquitin-mediated degradation. Alternatively, some other E3 ligases play as a tumor suppressor specifically targeting oncogene products. Deregulation of these E3 ligases induces unbalance between oncogenic signal and tumor suppressor pathway and leads to cellular transformation, tumor growth and metastasis in various human malignancies including oral, and head and neck cancers. Facilitated degradation of the cyclin-dependent kinase (CDK) inhibitor p27(Kip1) has been observed in oral, and head and neck cancers, and is correlated with their poor prognosis. SCF(Skp2), KPC complex, Pirh2 and CRL4(DDB2-Artemis) have been reported as E3 ligases targeting p27(Kip1) for degradation. In oral cancers, it is reported that overexpression of Skp2 and Pirh2 is associated with poor prognosis. Thus, chemical inhibitors against these E3 ligases are applicable for oral cancer therapy. Some potential compounds that inhibit E3 ligase activity of SCF(Skp2) have been reported. Moreover, the HECT-type E3 ligase WWP family and Smurf1 are also involved in the development and growth of human oral cancers. Therefore, small molecule inhibitors against HECT-type E3 ligases are discussed as anti-oral cancer drugs.

Matrix-assisted laser desorption/ionization imaging mass spectrometry revealed traces of dental problem associated with dental structure

Periodontal disease is a serious dental problem because it does not heal naturally and leads to tooth loss. In periodontal disease, inflammation at periodontal tissue is thought as predominant, and its effect against tooth itself remains unclear. In this study, we applied matrix-assisted laser desorption/ionization imaging mass spectrometry (IMS) to teeth for the first time. By comparing anatomical structure of tooth affected with periodontal disease with normal ones, we analyzed traces of the disease on tooth. We found signals characteristic of enamel, dentin, and dental pulp, respectively, in mass spectra obtained from normal teeth. Ion images reconstructed using these signals showed anatomical structures of the tooth clearly. Next, we performed IMS upon teeth of periodontal disease. Overall characteristic of the mass spectrum appeared similar to normal ones. However, ion images reconstructed using signals from the tooth of periodontal disease revealed loss of periodontal ligament visualized together with dental pulp in normal teeth. Moreover, ion image clearly depicted an accumulation of signal at m/z 496.3 at root surface. Such an accumulation that cannot be examined only from mass spectrum was revealed by utilization of IMS. Recent studies about inflammation revealed that the signal at m/z 496.3 reflects lyso-phosphatidylcholine (LPC). Infiltration of the signal is statistically significant, and its intensity profile exhibited the influence has reached deeply into the tooth. This suggests that influence of periodontal disease is not only inflammation of periodontal tissue but also infiltration of LPC to root surface, and therefore, anti-inflammatory treatment is required besides conventional treatments.