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Adoptive immunotherapy can induce sustained therapeutic effects in some cancers. Antitumor T cell grafts are often individually prepared in vitro from autologous T cells, which requires an intensive workload and increased costs. The quality of the generated T cells can also be variable, which affects the therapy's antitumor efficacy and toxicity. Standardized production of antitumor T cell grafts from third-party donors will enable widespread use of this modality if allogeneic T cell responses are effectively controlled. Here, we generated HLA class I, class II, and T cell receptor triple knockout (tKO) T cells by simultaneous knockout of the B2M, CIITA and TRAC genes through Cas9/sgRNA ribonucleoprotein electroporation. Although HLA deficient T cells were targeted by natural killer cells, they persisted better than HLA sufficient T cells in the presence of allogeneic peripheral blood mononuclear cells (PBMC) in immunodeficient mice.When transduced with a CD19 chimeric antigen receptor (CAR) and stimulated by tumor cells, tKO CAR-T cells persisted better when cultured with allogeneic PBMC compared with TRAC and B2M double-knockout T cells. The CD19 tKO CAR-T cells did not induce graft-versus-host disease, but retained antitumor responses. These results demonstrated the benefit of HLA class I, class II, and TCR deletion in enabling allogeneic-sourced T cells to be used for off-the-shelf adoptive immunotherapy. SynopsisAntitumor T cell grafts concurrently ablated of HLA class I, class II, and TCR molecules evade allogeneic T cell responses and can be used as a universal T cell source for adoptive cancer immunotherapy.

Epithelioid sarcoma (ES) is a relatively rare, highly malignant soft tissue sarcoma. The mainstay of treatment is resection or amputation. Currently other therapeutic options available for this disease are limited. Therefore, a novel therapeutic option needs to be developed. In the present study, we established a new human ES cell line (ESX) and analyzed the characteristics of its cancer stem-like cells/cancer-initiating cells (CSCs/CICs) based on ALDH1 activity. We demonstrated that a subpopulation of ESX cells with high ALDH1 activity (ALDHhigh cells) correlated with enhanced clonogenic ability, sphere-formation ability, and invasiveness in vitro and showed higher tumorigenicity in vivo. Next, using gene expression profiling, we identified CD109, a GPI-anchored protein upregulated in the ALDHhigh cells. CD109 mRNA was highly expressed in various sarcoma cell lines, but weakly expressed in normal adult tissues. CD109-positive cells in ESX predominantly formed spheres in culture, whereas siCD109 reduced ALDH1 expression and inhibited the cell proliferation in vitro. Subsequently, we evaluated the expression of CD109 protein in 80 clinical specimens of soft tissue sarcoma. We found a strong correlation between CD109 protein expression and the prognosis (P = 0.009). In conclusion, CD109 might be a CSC/CIC marker in epithelioid sarcoma. Moreover, CD109 is a promising prognostic biomarker and a molecular target of cancer therapy for sarcomas including ES.

Object External supports serve as a traditional treatment option for osteoporotic vertebral fractures (OVFs). However, the role of external supports in the treatment of OVF remains inconclusive. The purpose of this study was to determine the role of a rigid external support in the healing of OVFs by prospectively evaluating union (fracture settling) rates and prognostic variables for patients suffering from an incident OVF. Methods Fifty-five patients with acute back pain were enrolled in this study after being diagnosed with an OVF based on MRI findings. Patients were treated using a plastic thoracolumbosacral orthosis (TLSO) and underwent follow-up at 2, 3, and 6 months. Vertebrae were referred to as “settled” when there was no dynamic mobility on sitting lateral and supine lateral radiographs. At the time of the 3- and 6-month follow-up visits, the patients were divided into 2 groups, the “settled group” and the “unsettled group.” Patients in these groups were compared with regard to clinical and radiographic features. Results Of the 55 patients enrolled, 53 patients were followed up for 6 months. There were 14 men and 39 women with an average age of 75.3 years. Fracture settling of the affected vertebra was defined in 54.7% of the patients at 2 months, in 79.2% at 3 months, and in 88.7% at 6 months. All 5 components of the Japanese Orthopaedic Association Back Pain Evaluation Questionnaire improved significantly both at 3 months and 6 months. Patients in the unsettled group exhibited a statistically greater likelihood of having fractures at the thoracolumbar junction, Type A3 fractures, and fractures with a diffuse low-intensity area on T2-weighted MRI studies at 3 months. In contrast, at 6 months, the only statistically significant difference between the groups was patient age. Conclusions The biomechanical disadvantages of OVFs (location, type, and size) adversely influencing the fracture healing were overcome by the treatment using a TLSO within 6 months. The authors' findings show that a TLSO plays a biomechanical role in the healing of OVFs.

Immunohistochemical CD109 expression in myxofibrosarcoma was associated with poor prognosis.

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Osteosarcoma (OS) is a highly malignant bone tumor and the prognosis for non‐responders to chemotherapy remains poor. Previous studies have shown that human sarcomas contain sarcoma‐initiating cells (SIC), which have the characteristics of high tumorigenesis and resistance to chemotherapy. In the present study, we characterized SIC of a novel OS cell line, screened for SIC‐related genes, and tried to regulate the proliferation of OS by metabolic interference. Initially, we established a new human OS cell line (OS13) and isolated clones showing higher tumorigenesis as SIC (OSHIGH) and counterpart clones. OSHIGH cells showed chemoresistance and their metabolism highly depended on aerobic glycolysis and suppressed oxidative phosphorylation. Using RNA‐sequencing, we identified LIN28B as a SIC‐related gene highly expressed in OSHIGH cells. mRNA of LIN28B was expressed in sarcoma cell lines including OS13, but its expression was not detectable in normal organs other than the testis and placenta. LIN28B protein was also detected in various sarcoma tissues. Knockdown of LIN28B in OS13 cells reduced tumorigenesis, decreased chemoresistance, and reversed oxidative phosphorylation function. Combination therapy consisting of a glycolysis inhibitor and low‐dose chemotherapy had antitumor effects. In conclusion, manipulation of glycolysis combined with chemotherapy might be a good adjuvant treatment for OS. Development of immunotherapy targeting LIN28B, a so‐called cancer/testis antigen, might be a good approach.

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