Metastasis is the leading cause of death in patients with osteosarcoma. Some of these patients fail to respond to chemotherapy and die of metastasis within a short period. Therefore, it is important to identify novel biomarkers to improve the diagnosis and treatment of osteosarcoma. TRIM7 is a member of the tripartite motif (TRIM) family protein that is involved in various pathological conditions including cancer; however, its role in osteosarcoma remains elusive. Cell proliferation, invasion and migration were measured by CCK-8 and Transwell. Immunoprecipitation and mass spectrometry analysis were used to identify candidate proteins associated with TRIM7. Immunoprecipitation, immunofluorescence, pull down and ubiquitination assay were performed to examine the regulation between TRIM7 and its candidate protein. m6A modification of TRIM7 was measured by RNA immunoprecipitation. TRIM7 expression was upregulated in osteosarcoma tissues and was an independent risk factor in predicting poor prognosis. TRIM7 regulates osteosarcoma cell migration and invasion through ubiquitination of breast cancer metastasis suppressor 1 (BRMS1). Moreover, chemoresistance was readily observed in osteosarcoma cells and in patient-derived xenograft (PDX) mice with higher TRIM7 levels. Loss of TRIM7 m6A modification was observed in osteosarcoma tissues. METTL3 and YTHDF2 were the main factors involved in the aberrant m6A modification of TRIM7. Overall, our findings show that TRIM7 plays a key role in regulating metastasis and chemoresistance in osteosarcoma through ubiquitination of BRMS1. This work was financially supported by grants of NSFC (81001192, 81672658 and 81972521) and National Key Research Project of Science and Technology Ministry (2016YFC0106204).

Background: Tripartite motif-containing proteins (TRIM) play a crucial role in carcinogenesis. Little attention has been focused on the possible functions of TRIM6 on carcinogenesis. Methods: The expression levels of TRIM6 were assessed in colorectal cancer (CRC) samples. TRIM6 expression was knocked down in CRC cell lines, and subjected to Cell counting kit-8 (CCK-8), bromodeoxyuridine (BrdU) incorporation and cell cycle assays. Immunoprecipitation and proteomics analysis was performed to identify potential associated proteins of TRIM6. Results: TRIM6 expression was up-regulated in CRC samples and TRIM6 expression may be an independent prognostic marker for CRC. Knocking down TRIM6 expression suppressed CRC cell proliferation, induced cell cycle arrested at G2/M phase and increased sensitivity to 5-fluorouracil and oxaliplatin. TIS21, an anti-proliferative protein involved in the regulation of G2/M arrest, was identified as an interaction partner of TRIM6. Moreover, CRC cells with TRIM6 overexpression showed decreased TIS21 protein stability. TIS21 ubiquitination was increased in CRC cells overexpressing TRIM6, but not in those overexpressing TRIM6 E3 catalytic mutant (C15A). Further, Lys5 was essential for TRIM6 mediated TIS21 ubiquitination. TIS21 overexpression reversed the induced effects of TRIM6 overexpression on CRC cell proliferation, and the levels of forkhead box M1 (FoxM1), phosphorylated FoxM1, Cyclin B1 and c-Myc. Thiostrepton, a specific inhibitor for FoxM1, was less effective in anti-proliferative activity against CRC cells with lower level of TRIM6 in vitro and in vivo. Conclusions: Our study suggests that TRIM6 promotes the progression of CRC via TIS21/FoxM1.

Purpose: We investigated the safety and preliminary efficacy of anti–PD-L1 antibody (ZKAB001) as maintenance therapy for localized patients with high-grade osteosarcoma to reduce the risk of recurrence and metastasis. Patients and Methods: This open-label Phase I/II study was divided into dose-escalation Phase I and expansion Phase II. Phase I used a 3+3 design with ZKAB001 at three escalating doses ranging: 5, 10, 15 mg/kg every 2 weeks in 9 patients with localized high-grade osteosarcoma and Phase II tested 10 mg/kg in 12 patients for up to 24 cycles. Primary endpoints were safety and tolerability assessed using CTCAE4.0.3. Results: Between October 2018 and 2019, 21 eligible patients were enrolled and accepted ZKAB001 treatment: 9 in the dose-escalation phase, and 12 in expansion phase. Six patients with disease progression withdrew from this study and follow-up is ongoing. The MTD was not defined in Phase I. All doses had a manageable safety profile. The recommended dose in Phase II was set at 10 mg/kg. Most frequent immune-related adverse events were thyroiditis (76.2%) and dermatitis (42.9%). Only 1 (4.8%) of 21 patients had a Grade 3 skin rash. The median 3-year event-free survival (EFS) and overall survival (OS) were not established; however, 24-month EFS was 71.4% (95% confidence interval, 47.2–86.0) and 2-year OS was 100%. Preliminary efficacy data showed EFS benefits in patients with PD-L1 positive or an MSI-H sub-population. Conclusions: Switching to maintenance using ZKAB001 showed an acceptable safety profile and provided preliminary evidence of clinical activity in localized patients with osteosarcoma.

Background: Immunotherapy has provided an effective method for the treatment of many cancers. However, its efficacy in osteosarcoma is not satisfactory so far.Case Presentation: Here, we presented a case of osteosarcoma treated with sequential chemotherapy and immunotherapy and showed promising therapeutic potential. The 29-year-old female patient presented 9^th rib osteosarcoma with suspected right lung lower lobe metastasis. Surgery was performed to remove the primary lesion, and a series of chemotherapies were given afterward in consideration of the response and tolerance. The right lung lower lobe metastasis was under control first but progressed (PD) 9 months after the initiation of therapy. The lesion was surgically removed and subsequent chemotherapy was implemented. The patient had good tolerance with chemotherapy and maintained well for approximately 11 months before the discovery of 11^th rib and right lung upper lobe metastases. Surgery was then performed on both lesions and achieved complete response. Post-surgical brief chemotherapy and subsequent long-term immunotherapy (pembrolizumab) maintained continuous remission for 33 months. The patient survived for 60 months with well-controlled disease from the time of confirmed diagnosis. Genetic alterations of all primary and metastatic lesions were investigated by whole-exome sequencing (WES). Substantial similarity in mutational landscape between the primary lesion and 11^th rib metastasis and between the two lung metastases were revealed, while substantial heterogeneity was found between the rib lesions and lung metastases. The tumor mutational burden (TMB) for the 9^th rib primary lesion, the metastatic 11^th rib lesion, and the metastatic right upper and lower lobe nodule tissues was 8.02, 2.38, 4.61, and 0.14 mutations/Mb, respectively. The primary lesion exhibited the most diverse copy number variation (CNV) changes among all lesions. Furthermore, pathway enrichment analysis also suggested significant heterogeneity among the lesions.Conclusions: Surgery with sequential chemotherapy and maintenance immunotherapy was shown to have good response for the first time on osteosarcoma patient who had high TMB tumor lesions and good tolerance for chemotherapy and immunotherapy.