HMGA1 regulates trabectedin sensitivity in advanced soft-tissue sarcoma (STS): A Spanish Group for Research on Sarcomas (GEIS) study.

HMGA1 is a structural epigenetic chromatin factor that has been associated with tumor progression and drug resistance. Here, we reported the prognostic/predictive value of HMGA1 for trabectedin in advanced soft-tissue sarcoma (STS) and the effect of inhibiting HMGA1 or the mTOR downstream pathway in trabectedin activity. The prognostic/predictive value of HMGA1 expression was assessed in a cohort of 301 STS patients at mRNA (n = 133) and protein level (n = 272), by HTG EdgeSeq transcriptomics and immunohistochemistry, respectively. The effect of HMGA1 silencing on trabectedin activity and gene expression profiling was measured in leiomyosarcoma cells. The effect of combining mTOR inhibitors with trabectedin was assessed on cell viability in vitro studies, whereas in vivo studies tested the activity of this combination. HMGA1 mRNA and protein expression were significantly associated with worse progression-free survival of trabectedin and worse overall survival in STS. HMGA1 silencing sensitized leiomyosarcoma cells for trabectedin treatment, reducing the spheroid area and increasing cell death. The downregulation of HGMA1 significantly decreased the enrichment of some specific gene sets, including the PI3K/AKT/mTOR pathway. The inhibition of mTOR, sensitized leiomyosarcoma cultures for trabectedin treatment, increasing cell death. In in vivo studies, the combination of rapamycin with trabectedin downregulated HMGA1 expression and stabilized tumor growth of 3-methylcholantrene-induced sarcoma-like models. HMGA1 is an adverse prognostic factor for trabectedin treatment in advanced STS. HMGA1 silencing increases trabectedin efficacy, in part by modulating the mTOR signaling pathway. Trabectedin plus mTOR inhibitors are active in preclinical models of sarcoma, downregulating HMGA1 expression levels and stabilizing tumor growth.

STAT6-targeting antisense oligonucleotides against solitary fibrous tumor.

Solitary fibrous tumor (SFT) is a rare, non-hereditary soft tissue sarcoma thought to originate from fibroblastic mesenchymal stem cells. The etiology of SFT is thought to be due to an environmental intrachromosomal gene fusion between NGFI-A-binding protein 2 (NAB2) and signal transducer and activator protein 6 (STAT6) genes on chromosome 12, wherein the activation domain of STAT6 is fused with the DNA-binding domain of NAB2 resulting in the oncogenesis of SFT. All NAB2-STAT6 fusion variations discovered in SFTs contain the C-terminal of STAT6 transcript, and thus can serve as target site for antisense oligonucleotides (ASOs)-based therapies. Indeed, our in vitro studies show the STAT6 3' untranslated region (UTR)-targeting ASO (ASO 993523) was able to reduce expression of NAB2-STAT6 fusion transcripts in multiple SFT cell models with high efficiency (half-maximal inhibitory concentration: 116-300 nM). Encouragingly, in vivo treatment of SFT patient-derived xenograft mouse models with ASO 993523 resulted in acceptable tolerability profiles, reduced expression of NAB2-STAT6 fusion transcripts in xenograft tissues (21.9%), and, importantly, reduced tumor growth (32.4% decrease in tumor volume compared with the untreated control). Taken together, our study established ASO 993523 as a potential agent for the treatment of SFTs.

Circulating TNF-RII, IP-10 and HGF are associated with severity of COVID-19 in oncologic patients.

The COVID-19 patients showed hyperinflammatory response depending on the severity of the disease but little have been reported about this response in oncologic patients that also were infected with the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Sixty-five circulating cytokines/chemokines were quantified in 15 oncologic patients, just after SARS-CoV-2 infection and fourteen days later, and their levels were compared in patients who required hospitalisation by COVID-19 versus non-hospitalised patients. A higher median age of 72 years (range 61-83) in oncologic patients after SARS-CoV-2 infection was associated with hospitalisation requirement by COVID-19 versus a median age of 49 years (20-75) observed in the non-hospitalised oncologic patients (p = 0.008). Moreover, oncologic patients at metastatic stage or with lung cancer were significantly associated with hospitalisation by COVID-19 (p = 0.044). None of these hospitalised patients required ICU treatment. Higher basal levels of tumour necrosis factor receptor II (TNF-RII), interferon-γ (IFNγ)-induced protein 10 (IP-10) and hepatocyte growth factor (HGF) in plasma were significantly observed in oncologic patients who required hospitalisation by COVID-19. Higher TNF-RII, IP-10 and HGF levels after the SARS-CoV-2 infection in oncologic patients could be used as biomarkers of COVID-19 severity associated with hospitalisation requirements.

Delays in diagnosis and surgery of sarcoma patients during the COVID-19 outbreak in Spain.

Background and objectives: Social distancing and quarantine implanted during the COVID-19 outbreak could have delayed the accession of oncologic patients to hospitals and treatments. This study analysed the management of sarcoma patients during this period in five Spanish hospitals. Design and methods: Clinical data from adult sarcoma patients, soft tissue and bone sarcomas, managed during the COVID-19 outbreak, from 15 March to 14 September 2020 (Covid cohort), were retrospectively collected and time for diagnosis, surgery and active treatments were compared with sarcoma patients managed during the same pre-pandemic period in 2018 (Control cohort). Results: A total of 126 and 182 new sarcoma patients were enrolled in the Covid and Control cohorts, respectively, who were mainly diagnosed as soft tissue sarcomas (81.0% and 80.8%) and at localized stage (80.2% and 79.1%). A diagnostic delay was observed in the Covid cohort with a median time for the diagnosis of 102.5 days (range 6-355) versus 83 days (range 5-328) in the Control cohort (p = 0.034). Moreover, a delay in surgery was observed in cases with localized disease from the Covid cohort with a median time of 96.0 days (range 11-265) versus 54.5 days (range 2-331) in the Control cohort (p = 0.034). However, a lower delay for neoadjuvant radiotherapy was observed in the Covid cohort with a median time from the diagnosis to the neoadjuvant radiotherapy of 47 days (range 27-105) versus 91 days (range 27-294) in the Control cohort (p = 0.039). No significant differences for adjuvant radiotherapy, neoadjuvant/adjuvant chemotherapy and neoadjuvant/adjuvant palliative chemotherapy were observed between both cohorts. Neither progression-free survival (PFS) nor overall survival (OS) was significantly different. Conclusion: Delays in diagnosis and surgery were retrospectively observed in sarcoma patients during the COVID-19 outbreak in Spain, while the time for neoadjuvant radiotherapy was reduced. However, no impact on the PFS and OS was observed.

Reduction of Tumor Growth with RNA-Targeting Treatment of the NAB2-STAT6 Fusion Transcript in Solitary Fibrous Tumor Models.

Solitary fibrous tumor (SFT) is a rare soft-tissue sarcoma. This nonhereditary cancer is the result of an environmental intrachromosomal gene fusion between NAB2 and STAT6 on chromosome 12, which fuses the activation domain of STAT6 with the repression domain of NAB2. Currently there is not an approved chemotherapy regimen for SFTs. The best response on available pharmaceuticals is a partial response or stable disease for several months. The purpose of this study is to investigate the potential of RNA-based therapies for the treatment of SFTs. Specifically, in vitro SFT cell models were engineered to harbor the characteristic NAB2-STAT6 fusion using the CRISPR/SpCas9 system. Cell migration as well as multiple cancer-related signaling pathways were increased in the engineered cells as compared to the fusion-absent parent cells. The SFT cell models were then used for evaluating the targeting efficacies of NAB2-STAT6 fusion-specific antisense oligonucleotides (ASOs) and CRISPR/CasRx systems. Our results showed that fusion specific ASO treatments caused a 58% reduction in expression of fusion transcripts and a 22% reduction in cell proliferation after 72 h in vitro. Similarly, the AAV2-mediated CRISPR/CasRx system led to a 59% reduction in fusion transcript expressions in vitro, and a 55% reduction in xenograft growth after 29 days ex vivo.

Expression of p53 as a biomarker of pazopanib efficacy in solitary fibrous tumours: translational analysis of a phase II trial.

Background: Solitary fibrous tumours (SFT) are soft tissue sarcomas molecularly defined by the presence of the NAB2::STAT6 intrachromosomal fusion gene. Recently, a prospective phase II trial evaluating the role of the antiangiogenic tyrosine kinase inhibitor pazopanib in SFT has been conducted (NCT02066285). Methods: Here, we analysed the mRNA and protein expression levels of the tumour suppressor and angiogenesis regulator p53 (TP53) in pre-treatment tumour samples from 22 patients with low aggressive (or typical) SFT and 28 patients with high aggressive (26 malignant and 2 dedifferentiated) SFT enrolled in the aforementioned pazopanib phase II trial. These results were correlated with radiological progression-free survival (PFS) and objective response. Univariate and multivariate Cox regression analyses were also performed, including known clinic-pathological prognostic factors. Results: Diffuse immunohistochemistry (IHC) expression of p53 was only found in patients with aggressive SFT and was associated with significantly shorter PFS [hazard ratio (HR): 4.39, 95% confidence interval (CI): 1.19-16.14). TP53 mRNA levels were significantly higher in the low aggressive SFT group. Only in the high aggressive SFT group, relatively higher levels of TP53 were significantly associated with shorter PFS (HR: 4.16, 95% CI: 1.46-11.89) as well as to a lower rate of disease control following treatment with pazopanib. In the multivariate analysis, the only independent prognostic factor in the whole cohort was mitotic count. Conclusion: Diffuse p53 IHC expression and higher TP53 mRNA levels are associated with worse prognosis in the subset of aggressive SFT patients treated with pazopanib.

ISG15 as a prognostic biomarker in solitary fibrous tumour.

BACKGROUND: Solitary fibrous tumour (SFT) is a rare mesenchymal malignancy that lacks robust prognostic and predictive biomarkers. Interferon-stimulated gene 15 (ISG15) is a ubiquitin-like modifier, associated with tumour progression, and with poor survival of SFT patients, as previous published by our group. Here, we describe the role of ISG15 in the biology of this rare tumour. METHODS: ISG15 expression was assessed by immunohistochemistry in tissue microarrays from SFT patients and tested for correlation with progression-free survival and overall survival (OS). The effects of ISG15 knockdown or induction were investigated for cancer stem cell (CSC) characteristics and for drug sensitivity in unique in vitro models of SFT. RESULTS: The prognostic value of ISG15 for OS was validated at protein level in malignant SFT patients, prospectively treated with pazopanib and enrolled in GEIS-32 trial. In SFT in vitro models, ISG15 knockdown lead to a decrease in the expression of CSC-related genes, including SOX2, NANOG, ALDH1A1, ABCB1 and ABCC1. Likewise, ISG15 downregulation decreased the clonogenic/ tumoursphere-forming ability of SFT cells, while enhancing apoptotic cell death after doxorubicin, pazopanib or trabectedin treatment in 3D cell cultures. The regulation of CSC-related genes by ISG15 was confirmed after inducing its expression with interferon-ß1; ISG15 induction upregulated 1.28- to 451-fold the expression of CSC-associated genes. CONCLUSIONS: ISG15 is a prognostic factor in malignant SFT, regulating the expression of CSC-related genes and CSCs maintenance. Our results suggest that ISG15 could be a novel therapeutic target in SFT, which could improve the efficacy of the currently available treatments.

Efficacy of Eribulin Plus Gemcitabine Combination in L-Sarcomas.

Although the overall survival of advanced soft-tissue sarcoma (STS) patients has increased in recent years, the median progression-free survival is lower than 5 months, meaning that there is an unmet need in this population. Among second-line treatments for advanced STS, eribulin is an anti-microtubule agent that has been approved for liposarcoma. Here, we tested the combination of eribulin with gemcitabine in preclinical models of L-sarcoma. The effect in cell viability was measured by MTS and clonogenic assay. Cell cycle profiling was studied by flow cytometry, while apoptosis was measured by flow cytometry and Western blotting. The activity of eribulin plus gemcitabine was evaluated in in vivo patient-derived xenograft (PDX) models. In L-sarcoma cell lines, eribulin plus gemcitabine showed to be synergistic, increasing the number of hypodiploid events (increased subG1 population) and the accumulation of DNA damage. In in vivo PDX models of L-sarcomas, eribulin combined with gemcitabine was a viable scheme, delaying tumour growth after one cycle of treatment, being more effective in leiomyosarcoma. The combination of eribulin and gemcitabine was synergistic in L-sarcoma cultures and it showed to be active in in vivo studies. This combination deserves further exploration in the clinical context.

A Novel NFIX-STAT6 Gene Fusion in Solitary Fibrous Tumor: A Case Report.

Solitary fibrous tumor is a rare subtype of soft-tissue sarcoma with a wide spectrum of histopathological features and clinical behaviors, ranging from mildly to highly aggressive tumors. The defining genetic driver alteration is the gene fusion NAB2-STAT6, resulting from a paracentric inversion within chromosome 12q, and involving several different exons in each gene. STAT6 (signal transducer and activator of transcription 6) nuclear immunostaining and/or the identification of NAB2-STAT6 gene fusion is required for the diagnostic confirmation of solitary fibrous tumor. In the present study, a new gene fusion consisting of Nuclear Factor I X (NFIX), mapping to 19p13.2 and STAT6, mapping to 12q13.3 was identified by targeted RNA-Seq in a 74-year-old female patient diagnosed with a deep-seated solitary fibrous tumor in the pelvis. Histopathologically, the neoplasm did not display nuclear pleomorphism or tumor necrosis and had a low proliferative index. A total of 378 unique reads spanning the NFIXexon8-STAT6exon2 breakpoint with 55 different start sites were detected in the bioinformatic analysis, which represented 59.5% of the reads intersecting the genomic location on either side of the breakpoint. Targeted RNA-Seq results were validated by RT-PCR/ Sanger sequencing. The identification of a new gene fusion partner for STAT6 in solitary fibrous tumor opens intriguing new hypotheses to refine the role of STAT6 in the sarcomatogenesis of this entity.

A Comprehensive Review on Solitary Fibrous Tumor: New Insights for New Horizons.

Solitary fibrous tumor (SFT) is a rare mesenchymal, ubiquitous tumor, with an incidence of 1 new case/million people/year. In the 2020 WHO classification, risk stratification models were recommended as a better tool to determine prognosis in SFT, to the detriment of "typical" or "malignant" classic terms. The risk for metastasis is up to 35-45%, or even greater, in series with a longer follow-up. Over the last few decades, advances in immunohistochemistry and molecular diagnostics identified STAT6 nuclear protein expression and the NAB2-STAT6 fusion gene as more precise tools for SFT diagnosis. Recent evidence taken from retrospective series and from two prospective phase II clinical trials showed that antiangiogenics are active and their sequential use from first line should be considered, except for dedifferentiated SFT for which chemotherapy is the best option. Since the fusion transcript driver's first description in 2013, new insights have been brought on key molecular events in SFT. This comprehensive review mainly focuses on the superior efficacy of antiangiogenics over chemotherapeutic agents in SFT, provides the current knowledge of key molecules that could co-drive the SFT behavior, and suggests new target candidates that deserve to be explored in preclinical and clinical research in SFT.

Cancer Stem Cells in Soft-Tissue Sarcomas.

Soft tissue sarcomas (STS) are a rare group of mesenchymal solid tumors with heterogeneous genetic profiles and clinical features. Systemic chemotherapy is the backbone treatment for advanced STS; however, STS frequently acquire resistance to standard therapies, which highlights the need to improve treatments and identify novel therapeutic targets. Increases in the knowledge of the molecular pathways that drive sarcomas have brought to light different molecular alterations that cause tumor initiation and progression. These findings have triggered a breakthrough of targeted therapies that are being assessed in clinical trials. Cancer stem cells (CSCs) exhibit mesenchymal stem cell (MSC) features and represent a subpopulation of tumor cells that play an important role in tumor progression, chemotherapy resistance, recurrence and metastasis. In fact, CSCs phenotypes have been identified in sarcomas, allied to drug resistance and tumorigenesis. Herein, we will review the published evidence of CSCs in STS, discussing the molecular characteristic of CSCs, the commonly used isolation techniques and the new possibilities of targeting CSCs as a way to improve STS treatment and consequently patient outcome.

CUL4A, ERCC5, and ERCC1 as Predictive Factors for Trabectedin Efficacy in Advanced Soft Tissue Sarcomas (STS): A Spanish Group for Sarcoma Research (GEIS) Study.

A translational study was designed to analyze the expression of nucleotide excision repair (NER) and homologous recombination (HR) genes as potential predictive biomarkers for trabectedin in soft-tissue sarcoma (STS). This study is part of a randomized phase II trial comparing trabectedin plus doxorubicin versus doxorubicin in advanced STS. Gene expression levels were evaluated by qRT-PCR, while CUL4A protein levels were quantified by immunohistochemistry. Expression levels were correlated with patients' progression-free survival (PFS) and overall survival (OS). Gene expression was also evaluated in cell lines and correlated with trabectedin sensitivity. In doxorubicin arm and in the whole series, which includes samples from both arms, no significant differences in terms of PFS were observed amongst the analyzed genes. In the group treated with trabectedin plus doxorubicin, the median of PFS was significantly longer in cases with CUL4A, ERCC1, or ERCC5 overexpression, while BRCA1 expression did not correlated with PFS. Gene expression had no prognostic influence in OS. CUL4A protein levels correlated with worse PFS in doxorubicin arm and in the whole series. In cell lines, only overexpression of ERCC1 was significantly correlated with trabectedin sensitivity. In conclusion, CUL4A, ERCC5, and mainly ERCC1 acted as predictive factors for trabectedin efficacy in advanced STS.