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Pleomorphic liposarcoma (PLPS) is a recalcitrant soft-tissue sarcoma (STS) subtype in need of transformative therapy. We have previously established a patient-derived orthotopic xenograft (PDOX) model, of PLPS with PDGFRA amplification, using surgical orthotopic implantation. In the current study, the PLPS PDOX model was randomized into 3 groups of 7 mice each: untreated control; doxorubicin (DOX)-treated; and treated with Salmonella typhimurium A1-R (S. typhimurium A1-R) expressing green fluorescent protein (GFP). Tumor volume and body weight were monitored during the treatment period. The PLPS PDOX was resistant to DOX. In contrast, the PLPS PDOX was highly sensitive to S. typhimurium A1-R. There was no significant body-weight loss among these 3 groups. Fluorescence imaging demonstrated that S. typhimurium A1-R-GFP was very effective to target the PLPS PDOX tumor. The current study demonstrates that a PLPS PDOX, resistant to first-line therapy DOX, was highly sensitive to tumor targeting S. typhimurium A1-R.

学位論文

The aim of the present study was to determine the usefulness of a patient-derived orthotopic xenograft (PDOX) nude-mouse model of a doxorubicin-resistant metastatic Ewing's sarcoma, with a unique combination of a FUS-ERG fusion and CDKN2A deletion, to identify effective drugs for third-line chemotherapy of the patient. Our previous study showed that cyclin-dependent kinase 4/6 (CDK4/6) and insulin-like growth factor-1 receptor (IGF-1R) inhibitors were effective on the Ewing's sarcoma PDOX, but not doxorubicin, similar to the patient's resistance to doxorubicin. The results of the previous PDOX study were successfully used for second-line therapy of the patiend. In the present study, the PDOX mice established with the Ewing's sarcoma in the right chest wall were randomized into 5 groups when the tumor volume reached 60 mm3: untreated control; gemcitabine combined with docetaxel (intraperitoneal [i.p.] injection, weekly, for 2 weeks); irinotecan combined with temozolomide (irinotecan: i.p. injection; temozolomide: oral administration, daily, for 2 weeks); pazopanib (oral administration, daily, for 2 weeks); yondelis (intravenous injection, weekly, for 2 weeks). All mice were sacrificed on day 15. Body weight and tumor volume were assessed 2 times per week. Tumor weight was measured after sacrifice. Irinotecan combined with temozolomide was the most effective regimen compared to the untreated control group (p=0.022). Gemcitabine combined with docetaxel was also effective (p=0.026). Pazopanib and yondelis did not have significant efficacy compared to the untreated control (p=0.130, p=0.818). These results could be obtained within two months after the physician's request and were used for third-line therapy of the patient.

The purpose of this study was to investigate whether splenectomy influences the tumor growth and metastatic pattern in an orthotopic syngeneic murine pancreatic cancer model. Murine pancreatic cancer cells (PAN02) were subcutaneously injected into the flanks of nude mice. A small tumor fragment (3 mm2), harvested from a subcutaneous tumor. was orthotopically implanted in the tail of the pancreas of C57/BL6 mice without splenectomy (control group, n=15) or with simultaneous splenectomy (splenectomy group, n=15). Tumor growth and metastatic patterns were analyzed by laparotomy at 21 days after surgery. No tumor growth was found in 5 mice (33.3%) of the control group and 1 mouse (6.7%) of the splenectomy group (p=0.169). Tumor volume was significantly larger in splenectomy group (p=0.013). Peritoneal seeding was more frequently observed in the splenectomy group (11 (73.3%) vs. 4 (26.7%), p=0.011). There were no differences in the number of liver and kidney metastasis between the two groups. The ratios of tumor-infiltrating CD4+ to FoxP3+ and CD8+ to FoxP3+ were significantly higher in the control group compared to the splenectomy group (8.2 ± 9.3 vs. 2.4 ± 1.5, p=0.046; 2.5 ± 1.4 vs. 1.5 ± 0.4, p=0.031, respectively). Splenectomy enhanced tumor growth and peritoneal seeding in an orthotopic syngeneic murine pancreatic cancer mouse model. The ramification of these results are discussed for pancreatic cancer treatment.

William B. Coley developed bacterial therapy of cancer more than 100years ago and had clinical success. James Ewing, a very famous cancer pathologist for whom the Ewing sarcoma is named, was Coley's boss at Memorial Hospital in New York and terminated Coley's bacterial therapy of cancer. A tumor from a patient with soft-tissue Ewing's sarcoma, who failed doxorubicin (DOX) therapy, was previously implanted in nude mice to establish a patient-derived orthotopic xenograft (PDOX) model. In the present study, the Ewing's sarcoma PDOX was treated with tumor-targeting S. typhimurium A1-R expressing green fluorescent (GFP), alone and in combination with DOX. S. typhimurium A1-R-GFP was detected in the tumors after intratumor (i.t.) or intravenous (i.v.) injection. The combination of S. typhimurium A1-R and DOX significantly reduced tumor weight (37.8 ± 15.6 mg) compared to the untreated control (73.8 ± 10.1mg, P < 0.01). S. typhimurium A1-R monotherapy-treated tumors tended to be smaller (50.9 ± 17.8mg, P = 0.051). DOX monotherapy did not show efficacy (66.3 ± 26.4mg, P = 0.82), as was the case with the patient. The PDOX model faithfully replicated the DOX resistance the Ewing's sarcoma had in the patient. S. typhimurium A1-R converted the Ewing's sarcoma from DOX resistant to sensitive. One can only wonder how bacterial therapy and immunotherapy of cancer would have developed over the past 80years if Ewing did not stop Coley.

Previously, a patient-derived orthotopic xenograft (PDOX) model was established with a lung metastasis from an osteosarcoma patient which developed after adjuvant cisplatinum (CDDP) treatment. In this model, we previously demonstrated the efficacy of trabectedin (TRAB) and temozolomide (TEM) compared with CDDP. In the present report, osteosarcoma tissue was implanted orthotopically in the distal femur of mice which were randomized into the following groups when tumor volume reached approximately 100mm3; On day 14 after initiation of treatment, all but CDDP significantly inhibited tumor volume growth compared with untreated controls. Control (G1): 793.7 ± 215.0mm3; CDDP (G2): 588.1 ± 176.9mm3; Salmonella typhimurium A1-R (S. typhimurium A1-R) intravenous (i.v.) (G3): 269.7 ± 72.7mm3; S. typhimurium A1-R intra-arterial (i.a.) (G4): 70.2 ± 18.9mm3 (CDDP: p = 0.056; S. typhimurium A1-R i.v.: p = 0.0001; S. typhimurium A1-R i.a.: p = 0.00003, all vs. untreated controls). i.a. administration of S. typhimurium A1-R was significantly more effective than either CDDP (p = 0.00007), or i.v. administration of S. typhimurium A1-R (p = 0.00007) and significantly regressed the tumor volume compared with day 0 (p = 0.001). The new model of i.a. administration of S. typhimurium A1-R has great promise for the treatment of recalcitrant osteosarcoma.

Our laboratory pioneered patient-derived orthotopic xenograft (PDOX) mouse models using surgical orthotopic implantation (SOI). PDOX models are patient-like, in contrast to the ectopic subcutaneous-transplant cancer models. In the present study, we demonstrate that an undifferentiated pleomorphic soft-tissue sarcoma (UPS-STS) PDOX model acquired bright RFP-expressing stroma through one passage in red fluorescent protein (RFP) transgenic mice, which upon passage to non-colored nude mice was non-invasively imageable. A PDOX nude mouse model of UPS-STS was established in the biceps femoris of nude mice. After the tumors grew to a diameter of 10 mm, the tumors were subsequently passaged to RFP transgenic mice, and after tumor growth were then passaged to non-transgenic nude mice. Tumors were divided into small fragments and transplanted in the biceps femoris at each passage. The OV100 Small Animal Fluorescence Imaging System and FV1000 laser scanning confocal microscope were used to image RFP fluorescence in the UPS-STS PDOX models. UPS-STS PDOX tumors, previously grown in RFP transgenic nude mice for only one passage, had very bright fluorescence and after passage to non-transgenic nude mice maintained the bright fluorescence and were non-invasively imageable. FV1000 confocal imaging revealed diffusely distributed bright RFP stromal cells in the PDOX tumor, both in RFP transgenic mice and after passage to non-transgenic mice. These results demonstrate a powerful method to make the PDOX UPS-STS model brightly fluorescent for non-invasive imaging, as well as for confocal microscopy of individual stromal cells associated with the tumor. The RFP-labeled UPS PDOX has the potential to rapidly screen for novel effective agents for individual patients, including stroma-targeting drugs, whereby the stromal cells are a visual target. J. Cell. Biochem. 118: 361-365, 2017. © 2016 Wiley Periodicals, Inc.

大腿骨近位部骨折受傷患者の特徴を明らかにするために，受傷前の日常生活動作（以下ADL）のレベルと認知症の有無について調査を行った．対象は2009年9月から2010年1月までの期間に，沖縄県内の21施設で大腿骨近位部骨折の診断で入院加療を行った310人で，受傷前ADLをBarthel indexで，認知症の有無と程度を改訂版長谷川式認知症スケール（以下HDS-R）で評価した．受傷前ADLはBarthel indexで満点が31％，ある程度の自立が期待できる60点以上が64％で，年齢とBarthel indexには相関係数－0.369の負の相関が認められた．HDS-Rで14点以下を認知症有りと判断すると，52％に認知症が認められ，4点以下の高度認知症は27％であった．HDS-Rと年齢には相関係数－0.493の負の相関が認められた．Barthel indexとHDS-Rには相関係数0.681の正の相関関係が認められた．認知症の程度が高度になるにつれBarthel indexの点数が60点未満の者を多く含むようになり，逆に認知症を認めない者ではその多くがBarthel indexの点数は60点以上であった．