Katsuyuki Kusuzaki

Soft-tissue tumors evaluated by line-scan diffusion-weighted imaging: influence of myxoid matrix on the apparent diffusion coefficient.

To compare the apparent diffusion coefficients (ADCs) of myxoid and nonmyxoid soft‐tissue tumors using line‐scan diffusion‐weighted imaging (LSDWI), and to investigate the myxoid matrix influence on ADCs of soft‐tissue tumors.

Successful treatment with Erwinia L-asparaginase for recurrent natural killer/T cell lymphoma.

We describe a patient with natural killer (NK)/T cell lymphoma who relapsed after autologous peripheral blood stem cell transplantation (auto-PBSCT) and was successfully treated with Escherichia coli (E. coli ) and Erwinia l -asparaginase. A 38-year-old male patient with ulcerated tumor at the left thigh was diagnosed as having nasal type NK/T cell lymphoma on the basis of histopathological and flowcytometric findings of tumor, revealing diffuse infiltration of atypical lymphoid cells into blood vessels and expression of CD7 and CD56 antigens, but not CD3. He had tumor infiltration in the bone marrow and at the right lower lung field. After five cycles of CHOP (cyclophosphamide, doxorubicin, vincristine and prednisolone) therapy, the patient achieved complete remission and received high-dose chemotherapy with auto-PBSCT, although the tumor recurred in the right leg 10 months later. Despite salvage chemotherapy, followed by local irradiation and surgical amputation, a tumor recurred at the left upper gingiva 10 days after. Using E. coli l -asparaginase (6000 U/m 2 /day), the tumor regressed, fever was alleviated and the serum lactate dehydrogenase decreased to normal range after several days. The asparagine synthetase expression in tumor cells was immunohistochemically negative on paraffin-embedded tissues. Because of the anaphylactoid reaction developing after E. coli l -asparaginase, alternative Erwinia l -asparaginase (6000 U/m 2 /day) was administered, resulting in regression of tumor and fever lysis. l -asparaginase is a promising agent for the treatment of NK/T cell lymphoma.

Decorin suppresses bone metastasis in a breast cancer cell line.

Decorin, the prototype of an expanding family of small leucine-rich proteoglycans, is involved in a number of cellular processes including matrix assembly, fibrillogenesis and the control of cell proliferation. In this study, we investigated the role of decorin in suppressing tumor aggressiveness and bone metastases. We used a metastatic breast cancer cell line, MDA-MB-231, to show that decorin causes marked growth suppression bothin vitro and in vivo. A cytomegaloviral vector containing the decorin transgene caused greatly reduced cell growth, motility and observed metastases. Bone metastases were decreased by >90% upon decorin transfection. These results demonstrate a novel role for decorin in the reduction or prevention of tumor metastases in this breast cancer model and could eventually lead to improved therapies for metastatic breast cancer.

Expression of hypoxia-inducible factor (HIF)-1α as a biomarker of outcome in soft-tissue sarcomas

Hypoxia-inducible factor (HIF)-1α is a transcription factor that supports the adaptation of human cancer cells to hypoxia and tumor growth and progression. The overexpression of HIF-1α protein has been reported to be associated with a worse prognosis in various cancers. However, the expression of HIF-1α in soft-tissue sarcomas has not yet been characterized. The expression of HIF-1α protein was immunohistochemically determined in 49 specimens of soft-tissue sarcomas including malignant fibrous histiocytoma (29 patients), synovial sarcoma (12 patients), leiomyosarcoma (four patients), and malignant peripheral nerve sheath tumors (four patients). The 49 samples consisted of 40 primary lesions and nine local recurrences. An immunohistochemical analysis showed the nuclear accumulation of HIF-1α protein in 35 (71.4%) samples. The expression of HIF-1α was negative in 14 (28.6%) cases, weak in nine (18.4%), moderate in 17 (35.4%), and strong in nine (18.4%). The patients with a strong or moderate expression of HIF-1α had a significantly shorter overall survival rate in comparison with those with a weak or negative expression in a univariate analysis (P = 0.029; log-rank test) and multivariate analysis (P = 0.018). This is the first report that demonstrated an overexpression of HIF-1α protein to be an independent prognostic factor for soft-tissue sarcomas.

Total Tumor Cell Elimination with Minimum Damage to Normal Tissues in Musculoskeletal Sarcomas following Photodynamic Therapy with Acridine Orange

Acridine orange (AO) has unique biological actions enabling tumor visualization (fluorovisualization) and a strong cytocidal effect (photodynamic therapy: AO-PDT) under illumination with blue light. Accordingly, in this study, we attempted to develop a new surgical technique for total tumor cell elimination using these photodynamic reactions with AO in a mouse osteosarcoma model. The results showed that local tumor recurrence was significantly inhibited (23%) in the group treated with curettage under fluorovisualization and AO-PDT, compared to that (80%) in the control group treated with curettage alone under ordinary light. Therefore, we concluded that the combination of curettage under fluorovisualization and AO-PDT may be useful for total tumor cell elimination with minimum damage to normal tissue in musculoskeletal sarcomas.

Clinical Outcome of a Novel Photodynamic Therapy Technique Using Acridine Orange for Synovial Sarcomas

Abstract Synovial sarcoma (SS) is one of common malignant soft-tissue tumors and is encountered most commonly in children and young adults. It frequently involves or invades major neurovascular structures and bones, and its local recurrence rate after simple resection has been reported to be as high as up to 80%. Because major nerves and vessels, as well as an adequate amount of bone, must be preserved to restore excellent limb function in cases of SS, a surgical technique entailing a low risk of local recurrence is needed. Based on the findings of recent experimental studies conducted by us using a mouse osteosarcoma model, we developed a novel therapeutic technique for SS, consisting of reduction surgery followed by photodynamic therapy using acridine orange (AO-PDT), with or without X-ray irradiation at 5 Gy. A preliminary study revealed that low-dose X-rays also excite AO like photons. After an initial study on cell cultures, this novel technique was applied to six cases of SS. A follow-up of the subjects to determine the clinical outcome revealed that none of the cases treated by AO-PDT, including the four cases treated by additional 5 Gy irradiation and the two cases not receiving any radiation, showed any evidence of recurrence or local/systemic complications during the follow-up period of 19–51 months after the surgery. Therefore, we believe that AO-PDT with 5 Gy irradiation may be an excellent novel therapeutic modality with reduction surgery to salvage excellent limb function in SS involving major nerves and vessels or bones.

Cytosolic chloride ion is a key factor in lysosomal acidification and function of autophagy in human gastric cancer cell

The purpose of the present study was to clarify roles of cytosolic chloride ion (Cl−) in regulation of lysosomal acidification [intra‐lysosomal pH (pHlys)] and autophagy function in human gastric cancer cell line (MKN28). The MKN28 cells cultured under a low Cl− condition elevated pHlys and reduced the intra‐lysosomal Cl− concentration ([Cl−]lys) via reduction of cytosolic Cl− concentration ([Cl−]c), showing abnormal accumulation of LC3II and p62 participating in autophagy function (dysfunction of autophagy) accompanied by inhibition of cell proliferation via G0/G1 arrest without induction of apoptosis. We also studied effects of direct modification of H+ transport on lysosomal acidification and autophagy. Application of bafilomycin A1 (an inhibitor of V‐type H+‐ATPase) or ethyl isopropyl amiloride [EIPA; an inhibitor of Na+/H+ exchanger (NHE)] elevated pHlys and decreased [Cl−]lys associated with inhibition of cell proliferation via induction of G0/G1 arrest similar to the culture under a low Cl− condition. However, unlike low Cl− condition, application of the compound, bafilomycin A1 or EIPA, induced apoptosis associated with increases in caspase 3 and 9 without large reduction in [Cl−]c compared with low Cl− condition. These observations suggest that the lowered [Cl−]c primarily causes dysfunction of autophagy without apoptosis via dysfunction of lysosome induced by disturbance of intra‐lysosomal acidification. This is the first study showing that cytosolic Cl− is a key factor of lysosome acidification and autophagy.

Thrombin Inhibitor, Argatroban, Prevents Tumor Cell Migration and Bone Metastasis

It is well known that malignant cells show procoagulant activity, which is associated with their metastatic potential. Thrombin, the key enzyme of the blood coagulation system, is generated around tumor cells, promoting the migration and metastasis of tumor cells. In this study, we evaluated the effect of argatroban, a specific thrombin inhibitor, on the migration and metastasis of B16BL6 melanoma cells. In vitro argatroban dose-dependently inhibited cell migration, the maximum inhibition being observed in the presence of 10 µM argatroban (p < 0.0001). In order to investigate the antimetastatic effect of the thrombin inhibitor, we used an animal model that we have reported previously. C57BL6 mice which had received a bone (femur or tibia) transplanted into the dorsal subcutis were injected with B16 melanoma cells into the left heart ventricle. Intraperitoneal injection of argatroban (9 mg/kg/day for 4 weeks) significantly reduced the number of limbs with metastatic lesions as compared to a placebo (p < 0.05). These results suggest that argatroban was associated with reduced melanoma metastases by inhibiting cell migration. Our results showed that argatroban is effective for treatment of bone metastasis.

Transfection of NF-κB decoy oligodeoxynucleotide suppresses pulmonary metastasis by murine osteosarcoma.

Nuclear factor-kappa B (NF-κB) has a pivotal role in the progression and distant metastasis of cancers, including malignant bone tumors. To inhibit NF-κB activation, a new molecular therapy using synthetic double-stranded oligodeoxynucleotide (ODN) as a ‘decoy’ cis element against NF-κB has been developed. To determine whether pulmonary metastasis of osteosarcoma is reduced by inhibiting the action of NF-κB, NF-κB decoy ODN was transfected into the nuclei of murine osteosarcoma cells with high pulmonary metastatic potential, the LM8 cell line, using a three-dimensional alginate spheroid culture model. An in vitro study demonstrated the successful transfection of LM8 cells cultured in alginate beads by ‘naked’ NF-κB decoy ODN and that the activation of NF-κB signaling was significantly suppressed. Tumor growth was not affected by transfection of NF-κB decoy ODN, however, the expression of vascular endothelial growth factor (VEGF) and intercellular adhesion molecule 1 (ICAM-1) mRNA was markedly decreased. Furthermore, the transfection of ‘naked’ NF-κB decoy ODN effectively suppressed pulmonary metastasis in an in vivo alginate bead transplantation model. Our results suggest that NF-κB has a central and specific role in the regulation of tumor metastasis and could be a molecular target for development of anti-metastatic treatments for osteosarcoma.

Calcaneal osteosarcoma associated with Werner syndrome. A case report with mutation analysis.

In 1934, Oppenheimer and Kugel established the eponym of Werner syndrome13. This disease, which occurs with equal frequency in males and females, usually becomes manifest in the decade after adolescence, when previously normal development is altered by graying of the hair, impairment of normal growth, and loss of subcutaneous tissue and muscle mass in the extremities. The limbs become slender, and the trunk becomes disproportionately stocky. The shiny, atrophic, hyperkeratotic skin, unpadded by subcutaneous fat, is pulled tightly over the osseous prominences. Painful, circumscribed callosities occur, especially on the soles of the feet, and indolent ulcers appear in the regions of the malleoli of the ankles, the Achilles tendons, and the heels and toes. Baldness appears, and the hair thins in the regions of the eyebrows, face, axillae, and pubis. The facial appearance is often characteristically altered as the taut skin of the cheeks causes beaking of the nose, as shallow orbits and loss of periorbital connective tissue produce the appearance of proptosis, and as artificial lenses are required after the extraction of rapidly progressing cataracts. A peculiar thickening and vascularity of the vocal cords is associated with a weak, high-pitched voice. Arteriosclerosis is strikingly premature, and sexual underdevelopment results in sterility. Diabetes mellitus is common, and the skeleton is frequently affected by osteoporosis and hypertrophic arthritis. The fully developed syndrome creates a remarkably constant picture of premature senility, short stature, slender extremities, a stocky trunk, a beak-nosed face, and scleroderma-like changes14. Patients who have Werner syndrome have been reported to be at high risk for the development of malignant lesions2,4,7. In addition, it has also been emphasized that neoplasms originating from mesenchymal tissue compose more than half of all malignant processes (such as leukemia, melanoma, and myelodysplastic syndrome) in …