Denosumab administration for bone metastases from solid tumors: a retrospective cross-sectional study.

BACKGROUND: Little is known on how denosumab reduces skeletal-related events (SREs) by bone metastases from solid tumors. We sought to evaluate the effect of denosumab administration in patients with bone metastases from solid tumors. METHODS: Data of patients treated with denosumab were collected from electronic medical charts (n = 496). Eligible participants in this study were adult patients (age ≥ 18 years) with metastatic bone lesions from solid tumors treated with denosumab. SREs, surgical interventions, the spinal instability neoplastic score (SINS) for spinal region, and Mirels' score for the appendicular region were evaluated. To assess whether denosumab could prevent SREs and associated surgery, the SINS and Mirels' score were compared between patients with and without SREs. RESULTS: A total of 247 patients (median age, 65.5 years old; median follow-up period, 13 months) treated with denosumab for metastatic bone lesions from solid tumors were enrolled in this study. SREs occurred in 19 patients (7.7%). SREs occurred in 2 patients (0.8%) who took denosumab administration before SREs. Surgical interventions were undertaken in 14 patients (5.7%) (spinal and intradural lesions in five patients and appendicular lesions in nine patients). The mean SINS of patients without SREs compared to those with SREs were 7.5 points and 10.2 points, respectively. The mean Mirels' scores of non-SREs patients and those with SREs were 8.07 points and 10.7 points, respectively. Patients with SREs had significantly higher Mirels' score than non-SREs patients (p < 0.01). Patients with SREs had higher SINS than non-SREs patients (p = 0.09). CONCLUSIONS: SREs occurred in patients with higher SINS or Mirels' scores. Two patients suffered from SREs though they took denosumab administration before SREs. Appropriate management of denosumab for patients with bone metastasis is significant. Surgical interventions may be needed for patients who with higher SINS or Mirel's scores.

What Are the Complications, Function, and Survival of Tumor-devitalized Autografts Used in Patients With Limb-sparing Surgery for Bone and Soft Tissue Tumors? A Japanese Musculoskeletal Oncology Group Multi-institutional Study.

BACKGROUND: Tumor-devitalized autografts treated with deep freezing, pasteurization, and irradiation are biological reconstruction methods after tumor excision for aggressive or malignant bone or soft tissue tumors that involve a major long bone. Tumor-devitalized autografts do not require a bone bank, they carry no risk of viral or bacterial disease transmission, they are associated with a smaller immunologic response, and they have a better shape and size match to the site in which they are implanted. However, they are associated with disadvantages as well; it is not possible to assess margins and tumor necrosis, the devitalized bone is not normal and has limited healing potential, and the biomechanical strength is decreased owing to processing and tumor-related bone loss. Because this technique is not used in many countries, there are few reports on the results of this procedure such as complications, graft survival, and limb function. QUESTIONS/PURPOSES: (1) What was the rate of complications such as fracture, nonunion, infection, or recurrence in a tumor-devitalized autograft treated with deep freezing, pasteurization, and irradiation, and what factors were associated with the complication? (2) What were the 5-year and 10-year grafted bone survival (free from graft bone removal) of the three methods used to devitalize a tumor-containing autograft, and what factors were associated with grafted bone survival? (3) What was the proportion of patients with union of the tumor-devitalized autograft and what factors were associated with union of the graft-host bone junction? (4) What was the limb function after the tumor-devitalized autograft, and what factors were related to favorable limb function? METHODS: This was a retrospective, multicenter, observational study that included data from 26 tertiary sarcoma centers affiliated with the Japanese Musculoskeletal Oncology Group. From January 1993 to December 2018, 494 patients with benign or malignant tumors of the long bones were treated with tumor-devitalized autografts (using deep freezing, pasteurization, or irradiation techniques). Patients who were treated with intercalary or composite (an osteoarticular autograft with a total joint arthroplasty) tumor-devitalized autografts and followed for at least 2 years were considered eligible for inclusion. Accordingly, 7% (37 of 494) of the patients were excluded because they died within 2 years; in 19% (96), an osteoarticular graft was used, and another 10% (51) were lost to follow-up or had incomplete datasets. We did not collect information on those who died or were lost to follow-up. Considering this, 63% of the patients (310 of 494) were included in the analysis. The median follow-up was 92 months (range 24 to 348 months), the median age was 27 years (range 4 to 84), and 48% (148 of 310) were female; freezing was performed for 47% (147) of patients, pasteurization for 29% (89), and irradiation for 24% (74). The primary endpoints of this study were the cumulative incidence rate of complications and the cumulative survival of grafted bone, assessed by the Kaplan-Meier method. We used the classification of complications and graft failures proposed by the International Society of Limb Salvage. Factors relating to complications and grafted autograft removal were analyzed. The secondary endpoints were the proportion of bony union and better limb function, evaluated by the Musculoskeletal Tumor Society score. Factors relating to bony union and limb function were also analyzed. Data were investigated in each center by a record review and transferred to Kanazawa University. RESULTS: The cumulative incidence rate of any complication was 42% at 5 years and 51% at 10 years. The most frequent complications were nonunion in 36 patients and infection in 34 patients. Long resection (≥ 15 cm) was associated with an increased risk of any complication based on the multivariate analyses (RR 1.8 [95% CI 1.3 to 2.5]; p < 0.01). There was no difference in the rate of complications among the three devitalizing methods. The cumulative graft survival rates were 87% at 5 years and 81% at 10 years. After controlling for potential confounding variables including sex, resection length, reconstruction type, procedure type, and chemotherapy, we found that long resection (≥ 15 cm) and composite reconstruction were associated with an increased risk of grafted autograft removal (RR 2.5 [95% CI 1.4 to 4.5]; p < 0.01 and RR 2.3 [95% CI 1.3 to 4.1]; p < 0.01). The pedicle freezing procedure showed better graft survival than the extracorporeal devitalizing procedures (94% versus 85% in 5 years; RR 3.1 [95% CI 1.1 to 9.0]; p = 0.03). No difference was observed in graft survival among the three devitalizing methods. Further, 78% (156 of 200 patients) of patients in the intercalary group and 87% (39 of 45 patients) of those in the composite group achieved primary union within 2 years. Male sex and the use of nonvascularized grafts were associated with an increased risk of nonunion (RR 2.8 [95% CI 1.3 to 6.1]; p < 0.01 and 0.28 [95% CI 0.1 to 1.0]; p = 0.04, respectively) in the intercalary group after controlling for confounding variables, including sex, site, chemotherapy, resection length, graft type, operation time, and fixation type. The median Musculoskeletal Tumor Society score was 83% (range 12% to 100%). After controlling for confounding variables including age, site, resection length, event occurrence, and graft removal, age younger than 40 years (RR 2.0 [95% CI 1.1 to 3.7]; p = 0.03), tibia (RR 6.9 [95% CI 2.7 to 17.5]; p < 0.01), femur (RR 4.8 [95% CI 1.9 to 11.7]; p < 0.01), no event (RR 2.2 [95% CI 1.1 to 4.5]; p = 0.03), and no graft removal (RR 2.9 [95% CI 1.2 to 7.3]; p = 0.03) were associated with an increased limb function. The composite graft was associated with decreased limb function (RR 0.4 [95% CI 0.2 to 0.7]; p < 0.01). CONCLUSION: This multicenter study revealed that frozen, irradiated, and pasteurized tumor-bearing autografts had similar rates of complications and graft survival and all resulted in similar limb function. The recurrence rate was 10%; however, no tumor recurred with the devitalized autograft. The pedicle freezing procedure reduces the osteotomy site, which may contribute to better graft survival. Furthermore, tumor-devitalized autografts had reasonable survival and favorable limb function, which are comparable to findings reported for bone allografts. Overall, tumor-devitalized autografts are a useful option for biological reconstruction and are suitable for osteoblastic tumors or osteolytic tumors without severe loss of mechanical bone strength. Tumor-devitalized autografts could be considered when obtaining allografts is difficult and when a patient is unwilling to have a tumor prosthesis and allograft for various reasons such as cost or socioreligious reasons. LEVEL OF EVIDENCE: Level III, therapeutic study.

Imaging characteristics of tenosynovial giant cell tumors on 18F-fluorodeoxyglucose positron emission tomography/computed tomography: a retrospective observational study.

BACKGROUND: 18F-fluorodeoxyglucose positron emission tomography/computed tomography (18F-FDG-PET/CT) is useful for assessing location, metastasis, staging, and recurrence of malignant tumors. Tenosynovial giant cell tumor (TSGCT) is a benign tumor; however, some studies have reported that TSGCTs have a high uptake of FDG. Few studies have reported on the detailed evaluation of TSGCT using 18F-FDG-PET/CT. The purpose of the current study is to evaluate the image characteristics and locations, particularly where possible, with or without, extra-articular invasion from TSGCT of the knee in 18F-FDG-PET/CT could occur. METHODS: We retrospectively reviewed the patients with TSGCT who were diagnosed pathologically either by biopsy or surgical specimen. Furthermore, we evaluated the difference of the maximum standardized uptake value (SUVmax) between diffused TSGCT with extra-articular invasion and TSGCT with intra-articular localization in the knee. RESULTS: The study consisted of 20 patients with TSGCT. The mean SUVmax of TSGCT was 12.0 ± 6.50. There were five patients with TSGCT arising in the knee with extra-articular invasion and six with TSGCT with intra-articular localization. The mean SUVmax of TSGCT with extra-articular invasion and those with intra-articular localization were 14.3 ± 6.00 and 5.94 ± 3.89, respectively. TSGCT with extra-articular invasion had significantly higher SUVmax than TSGCT with intra-articular localization (p < 0.05). CONCLUSIONS: TSGCT revealed high FDG uptake. Furthermore, SUVmax was higher in diffused TSGCT with extra-articular invasion than in intra-articular localized TSGCT; this may reflect its local aggressiveness.

The influence of COVID-19 epidemic on the number of orthopaedic surgeries in Japan.

BACKGROUND: There is limited data on the impact of COVID-19 epidemic on the number of orthopaedic surgeries in Japan. METHODS: We conducted a nationwide hospital survey asking for the monthly number of orthopaedic surgeries performed at each facility from January 2019 to June 2021. Those facilities that had performed at least 100 surgeries in 2019 were included for analyses. The facilities were further grouped by prefecture and by hospital characteristics. A brief health economic evaluation was also performed. Risk ratios were compared using univariate analyses with P < 0.05 considered statistically significant. RESULTS: Questionnaire was sent to 1988 hospitals with 1671 hospitals (84%) responding. The survey data indicated a total number of orthopaedic surgeries decreased in 2020 compared to 2019 (1,061,541 vs 1,119,955 P < 0.01), and also for the first six months of 2021 compared to the same period in 2019 (530,388 vs 550,378 P < 0.01). In 2020, over 50% of all facilities in nearly all of the prefectures saw a decline in surgical procedures. The risk of incurring more than a 25% decease in the number of surgeries was significantly higher in 2020 for class I designated medical institutions compared to those that were not designated for any types of infectious diseases among the institutions with a tertiary emergency medical center in 2020 (crude risk ratio 2.9: 95% CI 1.2-7.4, p = 0.02) and in 2021 (crude risk ratio 4.7: 95% CI 1 0.9-12.1, p < 0.01). The estimated total nationwide decrease of revenue were in the range of approximately ¥29.2 to ¥116.8 billion per year for orthopaedic surgeries alone. CONCLUSION: There was a statistically significant decrease in the number of orthopaedic surgeries in Japan. The magnitude of the decline varied by prefectures and hospital characteristics, with the greater impact imposed on medical institutions with higher classification functions. The estimated immediate health economic impact was sizable.

Veno-accompanying artery fasciocutaneous flaps following wide excision of soft tissue tumors in the lower legs.

BACKGROUND: Soft tissue defects following wide excision of malignant soft tissue tumors (STTs) are sometimes too large for primary closure, especially in the lower legs where available soft tissue is limited. This study aimed to determine the clinical outcomes of reconstruction of a defect after wide excision of an STT with a veno-accompanying artery fasciocutaneous (VAF) flap in the lower leg. METHODS: This study comprised 9 patients with malignant STTs who had undergone reconstructive surgeries using VAF flaps after wide excisions, between October 2010 and September 2017. We retrospectively reviewed and collected data involving age, sex, follow-up period, histological diagnosis, surgical procedures, size and location of defects, size and location of the flaps, venous source of the flaps, direction of the pedicles, closing of donor sites, perioperative chemotherapies, postoperative complications, and the presence of postoperative local recurrence and metastasis. RESULTS: The median follow-up period was 91.5 (range, 15.5-189.0) months. Four patients had defects located around the knee, 3 patients had defects located on the calf, and 2 patients had defects located around the ankle. The mean flap size was 95.6 × 119.4 (range, 50 × 100-130 × 140) mm. Six patients had venous sources from the small saphenous vein and 3 patients had venous sources from the great saphenous vein. The pedicles were proximally based in 4 patients and distally based in 5 patients. All flaps remained viable without any complications. CONCLUSIONS: Our findings showed that the VAF flap was easily elevated and reliable. Furthermore, it was effective in reconstructing soft tissue defects following wide excisions of STTs in the lower leg.

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 αv integrin green flourescent protein (GFP) vector. 143B cells expressing αv 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 αv 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 αv integrin-GFP. Six weeks after transplantation, pulmonary metastases expressing αv integrin-GFP could be identified. TAMs expressing RFP accompanied metastasized osteosarcoma cells expressing αv integrin-GFP in the lung. The current study demonstrates the importance of αv integrin interaction with stromal elements in osteosarcoma.

Tumor-targeting Salmonella typhimurium A1-R arrests a doxorubicin-resistant PDGFRA-amplified patient-derived orthotopic xenograft mouse model of pleomorphic liposarcoma.

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.

A combination of irinotecan/cisplatinum and irinotecan/temozolomide or tumor-targeting Salmonella typhimurium A1-R arrest doxorubicin- and temozolomide-resistant myxofibrosarcoma in a PDOX mouse model.

Myxofibrosarcoma (MFS) is the most common sarcomas in elderly patients and is either chemo-resistant or recurs with metastasis after chemotherapy. This recalcitrant cancer in need of improved treatment. We have established a patient-derived orthotopic xenograft (PDOX) of MFS. The MFS PDOX model was established in the biceps femoris of nude mice and randomized into 7 groups of 7 mice each: control; doxorubicin (DOX); pazopanib (PAZ); temozolomide (TEM); Irinotecan (IRN); IRN combined with TEM; IRN combined with cisplatinum (CDDP) and Salmonella typhimurium A1-R (S. typhimurium A1-R). Treatment was evaluated by relative tumor volume and relative body weight. The MFS PDOX models were DOX, PAZ, and TEM resistant. IRN combined with TEM and IRN combined with CDDP were most effective on the MFS PDOX. S. typhimurium A1-R arrested the MFS PDOX tumor. There was no significant body weight loss in any group. The present study suggests that the combination of IRN with either TEM or CDDP, and S. typhimurium have clinical potential for MFS.

Trabectedin arrests a doxorubicin-resistant PDGFRA-activated liposarcoma patient-derived orthotopic xenograft (PDOX) nude mouse model.

BACKGROUND: Pleomorphic liposarcoma (PLPS) is a rare, heterogeneous and an aggressive variant of liposarcoma. Therefore, individualized therapy is urgently needed. Our recent reports suggest that trabectedin (TRAB) is effective against several patient-derived orthotopic xenograft (PDOX) mouse models. Here, we compared the efficacy of first-line therapy, doxorubicin (DOX), and TRAB in a platelet-derived growth factor receptor-α (PDGFRA)-amplified PLPS. METHODS: We used a fresh sample of PLPS tumor derived from a 68-year-old male patient diagnosed with a recurrent PLPS. Subcutaneous implantation of tumor tissue was performed in a nude mouse. After three weeks of implantation, tumor tissues were isolated and cut into small pieces. To match the patient a PDGFRA-amplified PLPS PDOX was created in the biceps femoris of nude mice. Mice were randomized into three groups: Group 1 (G1), control (untreated); Group 2 (G2), DOX-treated; Group 3 (G3), TRAB-treated. Measurement was done twice a week for tumor width, length, and mouse body weight. RESULTS: The PLPS PDOX showed resistance towards DOX. However, TRAB could arrest the PLPS (p < 0.05 compared to control; p < 0.05 compared to DOX) without any significant changes in body-weight. CONCLUSIONS: The data presented here suggest that for the individual patient the PLPS PDOX model could specifically distinguish both effective and ineffective drugs. This is especially crucial for PLPS because effective first-line therapy is harder to establish if it is not individualized.

Use of αv Integrin Linked to GFP to Image Molecular Dynamics in Trafficking Cancer-Cell Emboli.

Human osteosarcoma cells with an αv integrin green fluorescent protein (GFP) fusion gene were previously established and imaged in vitro and in vivo. In the present study, we imaged the interaction of αv integrin-GFP in osteosarcoma cells and collagen fibers in vascular-trafficking osteosarcoma emboli in nude mice. Human 143B osteosarcoma cells, expressing αv integrin-GFP, were injected by a vascular route in an abdominal skin flap in nude mice. Osteosarcoma cells were fluorescently imaged in the epigastric cranialis vein in the abdominal skin flap by confocal microscopy. Collagen fibers were imaged in reflectance mode. At early stages of tumor embolus-formation, cancer cells adhered firmly to each other, diffusely expressing αv integrin-GFP. Two weeks after injection, collagen fiber scaffolds were visualized at the margins of tumor emboli or within them. Four weeks after injection, cancer cells invading from emboli were strongly expressing αv integrin-GFP, and were aligned along collagen fibers. The results suggest αv integrin and collagen fiber scaffolds are important for tumor embolus formation, which are potential seeds of metastasis. J. Cell. Biochem. 118: 26-30, 2017. © 2016 Wiley Periodicals, Inc.

Analysis of Stroma Labeling During Multiple Passage of a Sarcoma Imageable Patient-Derived Orthotopic Xenograft (iPDOX) in Red Fluorescent Protein Transgenic Nude Mice.

A patient-derived orthotopic xenograft (PDOX) model of undifferentiated pleomorphic sarcoma (UPS) was previously established that acquired red fluorescent protein (RFP)-expressing stroma by growth in an RFP transgenic nude mouse. In the present study, an imageable PDOX model (iPDOX) of UPS was established by orthotopic implantation in the biceps femoris of transgenic RFP nude mice. After the tumors grew to a diameter of 10 mm, they were harvested and the brightest portion of the tumors were subsequently orthotopically transplanted to both RFP and non-colored nude mice. The UPS PDOX tumor was again transplanted to RFP transgenic and non-colored nude mice, and finally a 3rd passage was made in the same manner. Five UPS tumors from each passage in both RFP and non-colored mouse models were harvested. The FV1,000 confocal microscope was used to visualize and quantitate the RFP area of the resected tumors. The average percent fluorescent area in the first passage of RFP mice was 34 ± 22%; in the second passage, 34 ± 20%; and 36 ± 11% in the third passage of RFP transgenic nude mice. The average tumor RFP area in the first passage from RFP mice to non-colored mice was 20 ± 7%; in the second passage, 28 ± 11%; in the third passage was 27 ± 13%. The present results demonstrate the extensive and stable acquisition of stroma by the UPS-tumor growing orthotopically in transgenic RFP nude mice (iPDOX). This model can be used for screening for effective drugs for individual patients and drug discovery. J. Cell. Biochem. 118: 3367-3371, 2017. © 2017 Wiley Periodicals, Inc.

Labeling the Stroma of a Patient-Derived Orthotopic Xenograft (PDOX) Mouse Model of Undifferentiated Pleomorphic Soft-Tissue Sarcoma With Red Fluorescent Protein for Rapid Non-Invasive Imaging for Drug Screening.

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.

3-dimensional tissue is formed from cancer cells in vitro on Gelfoam®, but not on Matrigel™.

Cell and tissue culture can be performed on different substrates such as on plastic, in Matrigel™, and on Gelfoam(®), a sponge matrix. Each of these substrates consists of a very different surface, ranging from hard and inflexible, a gel, and a sponge-matrix, respectively. Folkman and Moscona found that cell shape was tightly coupled to DNA synthesis and cell growth. Therefore, the flexibility of a substrate is important for cells to maintain their optimal shape. Human osteosarcoma cells, stably expressing a fusion protein of α(v) integrin and green fluorescent protein (GFP), grew as a simple monolayer without any structure formation on the surface of a plastic dish. When the osteosarcoma cells were cultured within Matrigel™, the cancer cells formed colonies but no other structures. When the cancer cells were seeded on Gelfoam(®), the cells formed three-dimensional tissue-like structures. The behavior of 143B osteosarcoma cells on Gelfoam(®) in culture is remarkably different from those of these cells in monolayer culture or in Matrigel™. Tissue-like structures were observed only in Gelfoam(®) culture. The data in this report suggest a flexible structural substrate such as Gelfoam(®) provides a more in vivo-like culture condition than monolayer culture or Matrigel(TM) and that Matrigel(TM) does not result in actual three-dimensional culture.

Comparison of UVB and UVC effects on the DNA damage-response protein 53BP1 in human pancreatic cancer.

We have previously demonstrated that ultraviolet (UV) light is effective against a variety of cancer cells expressing fluorescent proteins in vivo as well as in vitro. In the present report, we compared the DNA damage repair (DDR) response of pancreatic cancer cells after UVB or UVC irradiation. The UV-induced DNA damage repair was imaged with green fluorescent protein (GFP) fused to the DDR-related chromatin-binding protein 53BP1 in MiaPaCa-2 human pancreatic cancer cells growing in 3D Gelfoam® histoculture and in superficial tumors grown in nude mice. 53BP1-GFP forms foci during DNA damage repair. A clonogenic assay in 2D monolayer culture initially showed that UVC and UVB inhibited MiaPaCa-2 cell proliferation in a dose-dependent manner, with UVC having more efficacy. Three-dimensional Gelfoam® histocultures and confocal imaging enabled 53BP1-GFP foci to be observed within 1 h after UV irradiation, indicating the onset of DDR response. UVB-induced 53BP1-GFP focus formation was observed up to a depth of 120 µm in MiaPaCa-2 cells on Gelfoam® compared to 80 µm for UVC. UVB-induced 53BP1-GFP focus formation was observed up to a depth of 80 µm in MiaPaCa-2 cells, implanted within skin flaps in mice, at a significantly greater extent than UVC. MiaPaCa-2 cells irradiated by UVB or UVC in the skin-flap mouse model had a significant decrease in tumor growth compared to untreated controls with UVB having more efficacy than UVC. Our results demonstrate that UVB has greater tissue penetration than UVC because of its longer wavelength and has clinical potential for eradicating superficial cancer.

Osteosarcoma cells enhance angiogenesis visualized by color-coded imaging in the in vivo Gelfoam® assay.

We previously described a color-coded imaging model that can quantify the length of nascent blood vessels using Gelfoam® implanted in nestin-driven green fluorescent protein (ND-GFP) nude mice. In ND-GFP mice, nascent blood vessels are labeled with GFP. We report here that osteosarcoma cells promote angiogenesis in the Gelfoam® angiogenesis assay in ND-GFP mice. Gelfoam® was initially transplanted subcutaneously in the flank of transgenic ND-GFP nude mice. Seven days after transplantation of Gelfoam®, skin flaps were made and human 143B osteosarcoma cells expressing green fluorescent protein (GFP) in the nucleus and red fluorescent protein (RFP) in cytoplasm were injected into the transplanted Gelfoam®. The control-group mice had only implanted Gelfoam®. Skin flaps were made at days 14, 21, and 28 after transplantation of the Gelfoam® to allow imaging of vascularization in the Gelfoam® using a variable-magnification small animal imaging system and confocal fluorescence microscopy. ND-GFP expressing nascent blood vessels penetrated and spread into the Gelfoam® in a time-dependent manner in both control and osteosarcoma-implanted mice. ND-GFP expressing blood vessels in the Gelfoam® of the osteosarcoma-implanted mice were associated with the cancer cells and larger and longer than in the Gelfoam®-only implanted mice (P < 0.01). The results presented in this report demonstrate strong angiogenesis induction by osteosarcoma cells and suggest this process is a potential therapeutic target for this disease.

Arthroscopic Resection of Symptomatic Ossicles of the Medial Malleolus Combined With Deltoid Ligament Repairs in Athletes: A Report of Three Cases.

We report three cases of young athletes with symptomatic ossicles of the medial malleolus who underwent arthroscopic resections of the ossicle combined with deltoid ligament repair. A differential diagnosis of the ossicles beneath the medial malleolus, accessory ossification center, avulsion fracture resulting in pseudoarthrosis, and accessory bone such as os subtibiale has been proposed. However, it is difficult to differentiate them clearly. Most of these ossicles are asymptomatic, although they can cause chronic medial ankle pain, especially in young athletes who require surgical treatment. All three patients had pain in the distal part of the medial malleolus, which restricted their sports activities. Plain radiographs of all three cases revealed a well-defined, round-shaped bony lesion beneath the medial malleolus. Ultrasonographic imaging, magnetic resonance imaging, and arthroscopic findings revealed that ligament attachment to the ossicle varies in volume and type according to the cases. In other words, the mechanisms through which the existence of the ossicles affects the stability of the ankle joint and foot alignment are different in each case, indicating that deltoid ligament repair is necessary according to the cases. In all three cases, we performed arthroscopic resections of ossicles combined with deltoid ligament repairs, achieving favorable short-term clinical outcomes.

Rare coexistence of multiple osteochondromas and solitary osteoid osteoma: A case report.

Multiple osteochondromas (MOs) are inherited in an autosomal-dominant manner, with a penetrance of ~96 and 100% in female and male patients, respectively. Osteochondromas primarily involve the metaphyses and diaphyses of long bones, including the ribs. Osteoid osteomas account for ~3 and 11% of all bone tumors and benign bone tumors, respectively. Furthermore,1 the male-to-female ratio is 2-3:1, and they generally occur in the long bones of the lower extremities, with the femoral neck being the most frequent site. The present study describes the case of a 16-year-old male patient with a bony mass around the left knee joint and pain in the left calf. Radiography revealed MOs in the upper and lower extremities, while computed tomography showed a nidus in the cortex of the tibial shaft. The patient's family history included the presence of MOs, and the patient was diagnosed with MOs and a solitary osteoid osteoma. Surgical excision of the osteochondroma and curettage of the osteoid osteoma in the proximal tibia and tibial shaft, respectively, were performed simultaneously. Postoperative pathological examination revealed osteochondroma and osteoid osteoma. Furthermore, the pain resolved, and no recurrence was observed 7 months post-operation. To the best of our knowledge, no reports exist on coexisting MOs and osteoid osteoma; therefore, the present study describes the first case of such a condition. Marginal excision for osteochondroma and curettage for osteoid osteoma effectively improved the symptoms.

Differential Diagnosis of Lipomatous Tumors Using 18F-Fluorodeoxygulcose Positron Emission Tomography/Computed Tomography: A Retrospective Observational Study.

Background/Aim: Lipomatous tumors, including lipomas, atypical lipomatous tumors (ALTs), myxoid liposarcomas (MLs), and dedifferentiated liposarcomas (DLs), are often diagnosed using magnetic resonance imaging (MRI). Differential diagnosis of lipomas and ALTs by MRI is often challenging. 18F-fluorodeoxyglucose positron emission tomography/computed tomography (18F-FDG PET/CT) has recently been used for the diagnosis and evaluation of tumor staging and recurrence of soft tissue tumors. The maximum standardized uptake value (SUVmax) is positively associated with malignant grade in several cancers. This study aimed to evaluate SUVmax of 18F-FDG PET/CT in the differential diagnosis of lipomatous tumors. Patients and Methods: Patients who underwent 18F-FDG PET/CT for the diagnosis of lipomatous tumors between January 2013 and September 2021 were included in the study. Patients with lipomatous tumors, confirmed by pathological diagnosis or surgical specimens, were evaluated for lipomatous tumor SUVmax. Results: This study included 44 patients with lipomas (n=19), ALTs (n=12), MLs (n=9), and DLs (n=4). The mean SUVmax of lipomas, ALTs, MLs, and DLs was 0.99±1.41, 1.92±0.95, 5.21±4.94, and 9.29±1.43, respectively. Lipomas showed a significantly lower SUVmax than did ALTs, MLs, and DLs (p<0.05). ALTs demonstrated a significantly lower SUVmax than did MLs and DLs (p<0.05). No significant differences were observed between MLs and DLs. Conclusion: Lipomas or ALTs had a significantly lower SUVmax than lipomatous sarcomas. Lipomas had a significantly lower SUVmax than ALTs, aiding in their preoperative differentiation. 18F-FDG-PET/CT could serve as a potent tool for the differential diagnosis of lipomatous tumors.

Arthroscopic debridement and microfracture for bilateral osteochondral lesions on the lateral process of the talus involving the subtalar joint: A case report.

RATIONALE: Osteochondral lesions on the lateral process of the talus involving the subtalar joint are rare; the optimal surgical treatment remains to be clarified as there are few reports. Additionally, bilateral cases are extremely rare. Therefore, the clinical outcomes of the surgical treatment for bilateral osteochondral lesions on the lateral process of the talus involving the subtalar joint have not been fully elucidated. PATIENT CONCERNS: A 16-year-old boy who played soccer presented to our hospital with bilateral hindfoot pain. The symptoms persisted even after 3 months of conservative treatment. The patient and family requested surgical treatment to relieve the symptoms. DIAGNOSES: The patient was diagnosed with bilateral osteochondral lesions on the lateral process of the talus, involving the subtalar joint based on computed tomography and magnetic resonance imaging findings. INTERVENTIONS: Arthroscopic debridement and microfracture were performed bilaterally. OUTCOMES: Postoperative computed tomography and magnetic resonance imaging of both feet revealed remodeling of the subchondral bone. The patient returned to play at the pre-injury level with no pain. LESSONS: This report describes a case of bilateral osteochondral lesions on the lateral process of the talus, involving the subtalar joint. Arthroscopic debridement and microfracture were effective in relieving symptoms and the subchondral bone remodeling. To the best of our knowledge, this is the first report of arthroscopic treatment of osteochondral lesions of the lateral process of the talus involving the subtalar joint.

Reduced Malignancy of Super Methotrexate-resistant Osteosarcoma Cells With Dihydrofolate Reductase Amplification Despite Paradoxical Gain of Oncogenic PI3K/AKT/mTOR and c-MYC expression.

BACKGROUND/AIM: Methotrexate (MTX) resistance in osteosarcoma leads to a very poor prognosis. In the present study, in order to further understand the basis and ramifications of MTX resistance in osteosarcoma, we selected an osteosarcoma cell line that has a 5,500-fold-increased MTX IC50 Materials and Methods: The super MTX-resistant 143B osteosarcoma cells (143B-MTXSR) were selected from MTX-sensitive parental human 143B osteosarcoma cells (143B-P) by continuous culture with step-wise increased amounts of MTX. To compare the malignancy of 143B-MTXSR and 143B-P, colony-formation capacity was compared with clonogenic assays on plastic and in soft agar. In addition, tumor growth was compared with orthotopic xenograft mouse models of osteosarcoma. Expression of dihydrofolate reductase (DHFR), phosphoinositide 3-kinase (PI3K)/protein kinase B (AKT)/mammalian target of rapamycin (mTOR), and myelocytomatosis oncogene (MYC) was examined with western immunoblotting and compared in 143B-MTXSR and 143B-P cells. RESULTS: 143B-MTXSR had a 5,500-fold increase in the MTX IC50 compared to the parental 143B-P cells. Expression of DHFR was increased 10-fold in 143B-MTXSR compared to 143B-P (p<0.01). 143B-MTXSR cells had reduced colony-formation capacity on plastic (p=0.032) and in soft agar (p<0.01) compared to 143B-P and reduced tumor growth in orthotopic xenograft mouse models (p<0.001). These results demonstrate that 143B-MTXSR had reduced malignancy. 143B-MTXSR also showed an increased expression of PI3K (p<0.01), phosphorylated (activated) AKT (p=0.031), phosphorylated mTOR (p=0.043), and c-MYC (p=0.024) compared to 143B-P. CONCLUSION: The present study demonstrates that the increased expression of DHFR, PI3K/AKT/mTOR and c-MYC appears to be linked to super MTX resistance and, paradoxically, to reduced malignancy. The present results suggest that DHFR may be a powerful tumor suppressor when highly amplified.