Imaging the interaction of αv integrin-GFP in osteosarcoma cells with RFP-expressing host stromal cells and tumor-scaffold collagen in the primary and metastatic tumor microenvironment

Human osteosarcoma 143B cells were previously stably transfected with an α_v integrin green flourescent protein (GFP) vector. 143B cells expressing α_v integrin-GFP were transplanted orthotopically in the tibia of transgenic nude mice ubiquitously expressing red fluorescent protein (RFP). The primary tumors acquired RFP-expressing stroma and were passaged orthotopically in the tibia in noncolored nude mice, which maintained the RFP stroma. The interaction of α_v integrin-GFP expression in 143B cells with RFP-expressing host stromal cells was observed by confocal microscopy using the Olympus FV1000. Collagen fibers were imaged simultaneously in reflectance mode. The RFP-expressing stroma included cancer-associated fibroblasts (CAFs) and tumor-associated macrophages (TAMs) which persisted even 3 weeks after passage to nontransgenic nude mice. CAFs expressing RFP were aligned between collagen fibers and cancer cells expressing α_v integrin-GFP. Six weeks after transplantation, pulmonary metastases expressing α_v integrin-GFP could be identified. TAMs expressing RFP accompanied metastasized osteosarcoma cells expressing α_v integrin-GFP in the lung. The current study demonstrates the importance of α_v integrin interaction with stromal elements in osteosarcoma.     

The role of salicylic acid in the glutathione-mediated protection against photooxidative stress in rice

Salicylic acid (SA) is known to be an essential component responsible for disease resistance in dicotyledonous plants. In rice, however, tissue contains extremely high endogenous levels of SA that do not increase after pathogen infection, suggesting that the SA has other major functions in healthy leaves. Although involvement of SA in oxidative-stress response is known in some dicotyledonous plants, antioxidative role of SA in rice is obscure. In this study, we examined the involvement of SA in the protection against oxidative stress in rice, using transgenic plants expressing the bacterial nahG gene that encodes salicylate hydroxylase, an SA-degrading enzyme. In SA-deficient NahG rice, the glutathione pool size was constitutively diminished as compared with control plants. NahG seedlings showed a delayed development phenotype, an increased susceptibility to oxidative stress and they developed light-induced lesions in their leaves without pathogen infection. Conversely, treatment with an activator of the SA-mediated defense-signaling pathway, probenazole, increased the glutathione pool size and suppressed lesion formation. These results suggest that in rice, SA has an important role in the response to high-light-induced oxidative stress, through its regulatory effects on glutathione homeostasis.     

Dioxin interferes in chromosomal positioning through the aryl hydrocarbon receptor

Each chromosome occupies its own-specific space called a ‘territory’ within the interphase nucleus, and the arrangement of chromosome territories (CTs) is important in epigenetic mechanisms. The molecular mechanism to determine the positioning of CTs, however, remains unknown. On the other hand, dioxin is known to be the typical environmental pollutant that affects a wide variety of biological events in many species. Here, we show that dioxin enlarges the minimum distance between chromosome 12 and chromosome 16 territories in human preadipocyte cells, and the alteration of chromosome positioning is canceled by an aryl hydrocarbon receptor (AhR) antagonist α-naphthoflavone. Thus, AhR may be a key molecule to regulate chromosome positioning. Our results suggest a novel effect of dioxin toxicity, and demonstrate a clue to reveal the novel molecular mechanism for the arrangement of CTs.     

Eribulin regresses a doxorubicin‐resistant Ewing's sarcoma with a FUS‐ERG fusion and CDKN2A‐deletion in a patient‐derived orthotopic xenograft (PDOX) nude mouse model

Ewing's sarcoma is a recalcitrant tumor greatly in need of more effective therapy. The aim of this study was to determine the efficacy of eribulin on a doxorubicin (DOX)‐resistant Ewing's sarcoma patient derived orthotopic xenograft (PDOX) model. The Ewing's sarcoma PDOX model was previously established in the right chest wall of nude mice from tumor resected form the patient's right chest wall. In the previous study, the Ewing's sarcoma PDOX was resistant to doxorubicin (DOX) and sensitive to palbociclib and linsitinib. In the present study, the PDOX models were randomized into three groups when the tumor volume reached 60 mm³: G1, untreated control (n = 6); G2, DOX treated (n = 6), intraperitoneal (i.p.) injection, weekly, for 2 weeks); G3, Eribulin treated (n = 6, intravenous (i.v.) injection, weekly for 2 weeks). All mice were sacrificed on day 15. Changes in body weight and tumor volume were assessed two times per week. Tumor weight was measured after sacrifice. DOX did not suppress tumor growth compared to the control group (P = 0.589), consistent with the previous results in the patient and PDOX. Eribulin regressed tumor size significantly compared to G1 and G2 (P = 0.006, P = 0.017) respectively. No significant difference was observed in body weight among any group. Our results demonstrate that eribulin is a promising novel therapeutic agent for Ewing's sarcoma.     

Mycobiota of the Takamatsuzuka and Kitora Tumuli in Japan,focusing on the molecular phylogenetic diversity of <Emphasis Type="Italic">Fusarium</Emphasis> and <Emphasis Type="Italic">Trichoderma</Emphasis>

In an effort to clarify the cause of the deterioration of the colorfully painted murals that adorn the inner walls of the small stone chambers in the Takamatsuzuka and Kitora Tumuli in Japan, we enumerated the fungi that were isolated from moldy spots on the plaster walls collected between May 2004 and April 2005. The 262 fungal isolates from 79 samples of both tumuli were identified as approximately 100 species based on their phenotypic characters. Fusarium, Trichoderma, and Penicillium species were the predominant colonizers in the stone chamber interior and adjacent areas of both tumuli. In addition to the 28S phylogeny, neighbor-joining and Bayesian phylogenies of partial EF-1-alpha gene sequences revealed 24 genetically diverse fusaria in the Takamatsuzuka and Kitora Tumuli. Most of the fusaria were nested in clade 3 of the Fusarium solani species complex (FSSC); however, a few isolates were members of the F. oxysporum species complex (FOSC) clade or the F. avenaceum/F. tricinctum species complex clade. The FSSC isolates were compared with those detected in the Lascaux cave in France. In addition, a partial EF-1α gene phylogeny indicated that 13 Trichoderma isolates clustered in the Harzianum-Virens clade and 5 isolates in the Viride clade or Trichoderma sect. Longibrachiatum. Our analyses suggest that most of the fungi recovered from both tumuli are typically soil dwellers. First two authors contributed equally to this work     

Epstein-Barr Virus and Human Papillomavirus Infections and Genotype Distribution in Head and Neck Cancers

ObjectiveTo investigate the prevalence, genotypes, and prognostic values of Epstein-Barr virus (EBV) and human papillomavirus (HPV) infections in Japanese patients with different types of head and neck cancer (HNC).ResultsOf the 209 HNC patients, 63 (30.1%) had HPV infection, and HPV-16 was the most common subtype (86.9%). HPV E6/E7 mRNA expression was found in 23 of 60 (38.3%) HPV DNA-positive cases detected. The site of highest prevalence of HPV was the oropharynx (45.9%). Among 146 (69.9%) HNCs in which EBV DNA was identified, 107 (73.3%) and 27 (18.5%) contained types A and B, respectively, and 124 (84.9%) showed the existence of del-LMP-1. However, only 13 (6.2%) HNCs were positive for EBER, 12 (92.3%) of which derived from the nasopharynx. Co-infection of HPV and EBER was found in only 1.0% of HNCs and 10.0% of NPCs. Kaplan-Meier survival analysis showed significantly better disease-specific and overall survival in the HPV DNA+/mRNA+ oropharyngeal squamous cell carcinoma (OPC) patients than in the other OPC patients (P = 0.027 and 0.017, respectively). Multivariate analysis showed that stage T1–3 (P = 0.002) and HPV mRNA-positive status (P = 0.061) independently predicted better disease-specific survival. No significant difference in disease-specific survival was found between the EBER-positive and -negative NPC patients (P = 0.155).ConclusionsOur findings indicate that co-infection with HPV and EBV is rare in HNC. Oropharyngeal SCC with active HPV infection was related to a highly favorable outcome, while EBV status was not prognostic in the NPC cohort.     

The irony of highly-effective bacterial therapy of a patient-derived orthotopic xenograft (PDOX) model of Ewing's sarcoma,which was blocked by Ewing himself 80 years ago

William B. Coley developed bacterial therapy of cancer more than 100 years 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 ± 10.1 mg, P < 0.01). S. typhimurium A1-R monotherapy-treated tumors tended to be smaller (50.9 ± 17.8 mg, P = 0.051). DOX monotherapy did not show efficacy (66.3 ± 26.4 mg, 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 80 years if Ewing did not stop Coley.     

Patient-derived orthotopic xenograft (PDOX) mouse model of adult rhabdomyosarcoma invades and recurs after resection in contrast to the subcutaneous ectopic model

Rhabdomyosarcoma (RMS) is a rare mesenchymal tumor. The aim of the present study was to develop a patient-derived orthotopic xenograft (PDOX) mouse model of RMS and compare the PDOX model to a subcutaneous (s.c.)-transplant model. A patient RMS from a striated muscle was grown orthotopically in the right biceps femoris muscle and right quadriceps muscle of nude mice to establish a PDOX model, as well as under the skin to establish an s.c. model. PDOX tumors grew at a statistically-significant faster rate compared to the s.c. tumors. Recurrence after surgical resection occurred only in PDOX tumors, not in the s.c. model. Histologically, only the PDOX model was shown to be invasive. In conclusion, these results indicate that the PDOX model of adult RMS is malignant and the subcutaneous model is benign. These results emphasize that a proper tumor microenvironment is necessary for patient-like behavior of a tumor in a mouse model.     

Regorafenib regresses an imatinib-resistant recurrent gastrointestinal stromal tumor (GIST) with a mutation in exons 11 and 17 of c-kit in a patient-derived orthotopic xenograft (PDOX) nude mouse model

Gastrointestinal stromal tumor (GIST) with a mutation in exons 11 and 17 of c-kit is a rare type of sarcoma. The aim of this study was to determine drug sensitivity for a regionally-recurrent case of GIST using a patient-derived orthotopic xenograft (PDOX) model. The PDOX model was established in the anterior wall of the stomach. GIST PDOX models were randomized into 5 groups of 6 mice each when the tumor volume reached 60 mm³: G1, control group; G2, imatinib group (oral administration (p.o.), daily, for 3 weeks); G3, sunitinib group (p.o., daily, for 3 weeks); G4, regorafenib (p.o., daily, for 3 weeks); G5, pazopanib (p.o., daily, for 3 weeks). All mice were sacrificed on day 22. Tumor volume was evaluated on day 0 and day 22 by laparotomy. Body weight were measured 2 times per week. Though regorafenib is third-line therapy for GIST, it was the most effective drug and regressed the tumor significantly (p < 0.001). Sunitinib suppressed tumor growth compared to the control group (p = 0.002). Imatinib, first-line therapy for GIST, and pazopanib did not have significant efficacy compared to the control group (p = 0.886, p = 0.766). The implications of this result is discussed for GIST patients.