Soft Tissue Sarcoma Cell Research Articles

Clinicopathological and molecular features of small round cell sarcoma of bone and soft tissue: a study of 72 cases

Objective: To investigate the clinicopathological, immunohistochemical and molecular features of small round cell sarcoma (SRCS) of the bone and soft tissue, and to compare the diagnostic value of different techniques. Methods: Seventy-two cases of SRCS of the bone and soft tissue diagnosed at People's Hospital, Peking University from January 2016 to March 2020 were recruited and retrospectively analyzed for pathological morphology, immunophenotype and fluorescence in situ hybridization (FISH) data. Next generation sequencing (NGS) was performed on 13 difficult cases. Results: In the study cohort, the patients ranged in age from 4-55 years, with a male predominance. The most Ewing's sarcomas and osteosarcomas occurred in the bone, while CIC-rearranged sarcomas, BCOR-rearranged sarcoma, synovial sarcoma, extraskeletal myxoid chondrosarcoma and FUS-NFATc2 rearranged sarcoma occurred in soft tissue. Histologically, all cases were composed predominantly of small round cells. Most cases were positive for vimentin and CD99, and showed a variable reactivity for neurogenic markers. Muscle marker and epithelial marker were negative for most cases. Combined with clinical features, histopathologic findings, immunophenotype, FISH and NGS, we diagnosed 46 Ewing sarcomas, 14 osteosarcomas, 3 CIC-rearranged sarcomas, 1 BCOR-rearranged sarcoma, 1 synovial sarcoma, 1 clear cell soft tissue sarcoma, 1 extraskeletal myxoid chondrosarcoma, 1 FUS-NFATc2 rearranged sarcoma, and 4 undifferentiated small round cell sarcomas. Conclusions: SRCS of bone and soft tissue is a group of malignant mesenchymal tumors based on morphological features. Most cases can be diagnosed with a combination of clinical characteristics, morphological features and immunohistochemical phenotype, while some cases require such further tests as FISH and NGS technologies, and NGS can be useful in diagnosing and categorizing SRCS.

PO-1921 Radiosensitizing effect of Trabectidin on human soft tissue sarcoma cells

PO-1921 Radiosensitizing effect of Trabectidin on human soft tissue sarcoma cells

Abstract 1946: Synergistic cell death induction in breast cancer cell lines by combining ABT-199 and cisplatin irrespective of estrogen receptor and p53 expression

Abstract 50 Years after the generation of the most studied human breast cancer cell line, MCF-7, many research results fundamentally changed patient outcomes for the better. The mortality of breast cancer in women has decreased significantly, especially in the last 30 years. But the total number of deaths is rising again in the last ten years. Considering that breast cancer is the most common cancer in women and the second most common cancer overall, with incidences of 24,2% and 11,6%, respectively, total numbers of new cases are in the millions every year. First line drugs for the treatment of breast cancer often target microtubules (e.g. Vinblastine and Paclitaxel) or induce DNA damage (e.g. Carboplatin and Cisplatin (CisPt)). In addition to these classic therapeutic drugs new compounds that specifically target cancer-driving molecular alterations have been developed. Among these is the group of BH3-mimetics. The efficacy of several BH3-mimetics is investigated in numerous clinical trials while the BCL-2 specific inhibitor ABT-199 (Venetoclax) is already approved for clinical application in chronic lymphatic leukemia (CLL), acute myeloid leukemia (AML) and small lymphocytic lymphoma (SLL). BH3-mimetics target the apoptosis machinery by interacting with anti-apoptotic Bcl-2 proteins, mimicking the function of pro-apoptotic BH3-only proteins. In turn, the pro-apoptotic potential of pore-forming BCL-2 effectors BAX, BAK and BOK is released inducing mitochondrial outer membrane permeabilization and cell death. Recently we reported synergistic cell death induction by ABT-199 in combination with the proteasome inhibitor Bortezomib (BOZ) in soft tissue sarcoma (STS) cells. ABT-199 and BOZ induce accumulation of BOK and the BH3-only protein NOXA (Muenchow et al. 2020). Here, we combined ABT-199, CisPt and Nutlin-3 in breast cancer cell lines that differ in expression of the estrogen receptor (ER) and p53 activity. Similar to the results in STS cells synergistic induction of apoptotic cell death was observed by FACS analysis of the mitochondrial membrane potential (TMRM) and exposure of phosphatidyl serine (Annexin-V). Western blot analysis revealed that ABT-199 induced accumulation of NOXA and MCL-1, irrespective of the ER status or p53 mutation. Simultaneous treatment with ABT-199 and CisPt or Nutlin-3 further increased expression of NOXA and MCL-1. Interestingly, qRT-PCR revealed enhanced expression of NOXA and MCL-1 mRNA in ABT-199 treated cells indicating transcriptional regulation. However, transcriptional induction of NOXA was also detected independent of p53 activity. Thus, combined therapeutic approaches using CisPt and ABT-199 should be effective irrespective of ER and p53 expression. Nevertheless, in p53 proficient tumors Nutlins might increase therapeutic efficacy. Citation Format: Benjamin Schaefer, Sandra Weller, Tobias Beigl, Kathrin Boepple, Reiner Hoppe, Klaus Schulze-Osthoff, Hans-Georg Kopp, Walter E. Aulitzky, Frank Essmann. Synergistic cell death induction in breast cancer cell lines by combining ABT-199 and cisplatin irrespective of estrogen receptor and p53 expression [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 1946.

Phase II trial of pegylated arginine deiminase in combination with gemcitabine and docetaxel for the treatment of soft tissue sarcoma.

11508 Background: Soft tissue sarcoma (STS) is dependent on extracellular arginine as it often lacks expression of argininosuccinate synthase 1 (ASS1), the urea cycle enzyme needed to produce intracellular arginine. PEGylated arginine deiminase (ADI-PEG 20) is an extracellular arginine-degrading enzyme that causes ASS1 deficient tumors to enter the starvation state. Preclinical data demonstrated that addition of docetaxel (D) with ADI-PEG20 upregulates expression of the transporter for gemcitabine (G), overcoming transporter level resistance, and causing increased cell death. In vivo modeling demonstrated that the combination of ADI-PEG20 with G+D was superior to G+D alone. Therefore, we performed a phase 2 trial testing the addition of ADI-PEG20 to G+D. Methods: We performed an investigator-initiated, phase 2, multicenter, multi-arm clinical trial of ADI-PEG20 with G (90minute infusion)+D in STS, Ewing’s, osteosarcoma and small cell lung cancer. We are reporting Arm A, the STS arm. Eligible patients had STS that would be standardly treated with G+D that had progressed on at least one prior line of therapy with measurable disease by RECIST1.1 and had adequate organ function Based on a historic median PFS of 6.2 months for G+D, we targeted to enroll N = 75 patients in cohort A to detect a 2.8 month improvement with 80% power at a 5% alpha level. Primary endpoint was progression-free survival (PFS). Secondary endpoints included overall survival (OS), clinical benefit rate (CBR), safety, tolerability, cancer related mortality, and correlation with ASS1 expression by IHC. We evaluated PFS by Kaplan-Meier method and estimated overall response rate (ORR). Results: 75 patients were treated and deemed evaluable. The trial underwent two dose reductions by the data safety monitoring board due to prolonged neutropenia and thrombocytopenia preventing the use of day 8 G+D, consistent with preclinical mechanism of action data showing that ADI-PEG 20+D enhanced G uptake. Originally, the G dose was 900mg/m2 reduced first to 750mg/m2 then to 600mg/m2. D was dose reduced at the time of the second dose reduction from 75mg/m2 to 60mg/m2. ADI-PEG20 was given at a fixed intramuscular dose (36 mg/m2) weekly. The need for two dose reductions affected the PFS. The PFS/OS (months) were for the 600mg/m2 group (n = 31) was 6.0/N.D., leiomyosarcoma (LMS) (N = 33) 7.2/22.5, liposarcoma 5.1/17.4, and other (N = 36) 2.8/15.0. Responses were 8% complete (6/75) (3 LMS, 1 synovial and 2 angiosarcoma), 17% partial (13/75), and 43% stable disease (32/75), for an ORR of 25% (19/75) and CBR of 68% (51/75). There was a trend for ASS1 negative tumors to benefit more than ASS1 positive tumors. Conclusions: The combination of ADI-PEG20 with G+D can be safely and effectively given at a dose of 600mg/m2 G and 60mg/m2 D. Future randomized trials of ADI-PEG20 with G+D are planned. Clinical trial information: NCT03449901.

Cellular Radiosensitivity of Soft Tissue Sarcoma.

Currently, all soft tissue sarcomas (STS) are irradiated by the same regimen, disregarding possible subtype-specific radiosensitivities. To gain further insight, cellular radiosensitivity was investigated in a panel of sarcoma cell lines. Fourteen sarcoma cell lines, derived from synovial sarcoma, leiomyosarcoma, fibrosarcoma and liposarcoma origin, were submitted to clonogenic survival assays. Cells were irradiated with single doses from 1-8 Gy and surviving fraction (SF) was calculated from the resulting response data. Alpha/beta (α/β) ratios were inferred from radiation-response curves using the linear-quadratic (LQ)-model. Cellular radiosensitivities varied largely in this panel, indicating a considerable degree of heterogeneity. Surviving fraction after 2 Gy (SF2) ranged from 0.27 to 0.76 with evidence of a particular radiosensitive phenotype in only few cell lines. D37% on the mean data was 3.4 Gy and the median SF2 was 0.52. The median α/β was 4.9 Gy and in six cell lines the α/β was below 4 Gy. A fairly homogeneous radiation response was observed in myxoid liposarcoma cell lines with SF2 between 0.64 and 0.67. Further comparing sarcomas of different origin, synovial sarcomas, as a group, showed the lowest SF2 values (mean 0.35) and was significantly more radiosensitive than myxoid liposarcomas and leiomyosarcomas (P = 0.0084 and 0.024, respectively). This study demonstrates a broad spectrum of radiosensitivities across STS cell lines and reveals subtype-specific radiation responses. The particular cellular radiosensitivity of synovial sarcoma cells supports consideration of the different sarcoma entities in clinical studies that aim to optimize sarcoma radiotherapy.

Oncolytic virotherapy promotes radiosensitivity in soft tissue sarcoma by suppressing anti-apoptotic MCL1 expression.

Soft tissue sarcoma (STS) is a rare cancer that develops from soft tissues in any part of the body. Despite major advances in the treatment of STS, patients are often refractory to conventional radiotherapy, leading to poor prognosis. Enhancement of sensitivity to radiotherapy would therefore improve the clinical outcome of STS patients. We previously revealed that the tumor-specific, replication-competent oncolytic adenovirus OBP-301 kills human sarcoma cells. In this study, we investigated the radiosensitizing effect of OBP-301 in human STS cells. The in vitro antitumor effect of OBP-301 and ionizing radiation in monotherapy or combination therapy was assessed using highly radiosensitive (RD-ES and SK-ES-1) and moderately radiosensitive (HT1080 and NMS-2) STS cell lines. The expression of markers for apoptosis and DNA damage were evaluated in STS cells after treatment. The therapeutic potential of combination therapy was further analyzed using SK-ES-1 and HT1080 cells in subcutaneous xenograft tumor models. The combination of OBP-301 and ionizing radiation showed a synergistic antitumor effect in all human STS cell lines tested, including those that show different radiosensitivity. OBP-301 was found to enhance irradiation-induced apoptosis and DNA damage via suppression of anti-apoptotic myeloid cell leukemia 1 (MCL1), which was expressed at higher levels in moderately radiosensitive cell lines. The combination of OBP-301 and ionizing radiation showed a more profound antitumor effect compared to monotherapy in SK-ES-1 (highly radiosensitive) and HT1080 (moderately radiosensitive) subcutaneous xenograft tumors. OBP-301 is a promising antitumor reagent to improve the therapeutic potential of radiotherapy by increasing radiation-induced apoptosis in STS.

Highlights from Recent Cancer Literature

Highlights from Recent Cancer Literature

Bispecific NKG2D-CD3 and NKG2D-CD16 Fusion Proteins as Novel Treatment Option in Advanced Soft Tissue Sarcomas.

Soft tissue sarcoma (STS) constitutes a rare group of heterogeneous malignancies. Effective treatment options for most subtypes of STS are still limited. As a result, especially in metastatic disease, prognosis is still dismal. The ligands for the activating immunoreceptor NKG2D (NKG2DL) are commonly expressed in STS, but generally absent in healthy tissues. This provides the rationale for utilization of NKG2DL as targets for immunotherapeutic approaches. We here report on the preclinical characterization of bispecific fusion proteins (BFP) consisting of the extracellular domain of the NKG2D receptor fused to Fab-fragments directed against CD3 (NKG2D-CD3) or CD16 (NKG2D-CD16) for treatment of STS. After characterization of NKG2DL expression patterns on various STS cell lines, we demonstrated that both NKG2D-CD16 and NKG2D-CD3 induce profound T and NK cell reactivity as revealed by analysis of activation, degranulation and secretion of IFNγ as well as granule associated proteins, resulting in potent target cell lysis. In addition, the stimulatory capacity of the constructs to induce T and NK cell activation was analyzed in heavily pretreated STS patients and found to be comparable to healthy donors. Our results emphasize the potential of NKG2D-CD3 and NKG2D-CD16 BFP to target STS even in an advanced disease.

Tertiary lymphoid structures marker CXCL13 is associated with better survival for patients with advanced-stage bladder cancer treated with immunotherapy

IntroductionImmune checkpoint inhibitors (ICIs) have proved to be an effective treatment for up to 40% of muscle-invasive bladder cancer (MIBC), but there is still a need for better performing biomarkers allowing to improve prediction of response to ICI. Response to immunotherapy in soft-tissue sarcoma, melanoma and renal cell carcinoma have been recently linked to the presence of tertiary lymphoid structures (TLS) in the tumour. TLS are organised aggregates of T, B and dendritic cells, participating in adaptive antitumor immune response. The chemokine CXCL13 is involved in the formation of TLS, and is reported as a reliable transcriptomic marker of TLS. ObjectivesIn this study, we sought to assess whether CXCL13 transcript expression can be a prognostic biomarker for ICI-treated MIBC patients and also investigated whether it can serve a biomarker of TLS in MIBC. MethodsWe analysed transcriptomic data from three publicly available MIBC cohorts and evaluated pathological slides from the TCGA-BLCA cohort for TLS presence and stage of maturation. ResultsWe showed that CXCL13 was independently associated with both prolonged survival (HR = 0.8, 95% CI [0.68–0.94]) and objective response (p < 0.0001) in patients treated with ICI, at the difference of others immunological signatures. However, it was not a predictor for non–ICI-treated MIBC, suggesting a predictive effect of ICI efficacy. Finally, we validated that CXCL13 expression was correlated with tumour TLS in TCGA data set (p < 0.001), and can serve as a marker of TLS in bladder cancer. ConclusionThese results support that CXCL13 expression, as a surrogate for tumour TLS, is a relevant candidate predictive biomarker of response to ICI for patients with advanced-stage bladder cancer.

MNK1 and MNK2 enforce expression of E2F1, FOXM1, and WEE1 to drive soft tissue sarcoma

Soft tissue sarcoma (STS) is a heterogeneous disease that arises from connective tissues. Clinical outcome of patients with advanced tumors especially de-differentiated liposarcoma and uterine leiomyosarcoma remains unsatisfactory, despite intensive treatment regimens including maximal surgical resection, radiation, and chemotherapy. MAP kinase-interacting serine/threonine-protein kinase 1 and 2 (MNK1/2) have been shown to contribute to oncogenic translation via phosphorylation of eukaryotic translation initiation factor 4E (eIF4E). However, little is known about the role of MNK1/2 and their downstream targets in STS. In this study, we show that depletion of either MNK1 or MNK2 suppresses cell viability, anchorage-independent growth, and tumorigenicity of STS cells. We also identify a compelling antiproliferative efficacy of a novel, selective MNK inhibitor ETC-168. Cellular responsiveness of STS cells to ETC-168 correlates positively with that of phosphorylated ribosomal protein S6 (RPS6). Mirroring MNK1/2 silencing, ETC-168 treatment strongly blocks eIF4E phosphorylation and represses expression of sarcoma-driving onco-proteins including E2F1, FOXM1, and WEE1. Moreover, combination of ETC-168 and MCL1 inhibitor S63845 exerts a synergistic antiproliferative activity against STS cells. In summary, our study reveals crucial roles of MNK1/2 and their downstream targets in STS tumorigenesis. Our data encourage further clinical translation of MNK inhibitors for STS treatment.

Senolytic Targeting of Bcl-2 Anti-Apoptotic Family Increases Cell Death in Irradiated Sarcoma Cells.

Simple SummaryLimited volumetric change after pre-operative radiotherapy (RT) suggests that sarcomas generally do not undergo cell death. Senolytic drugs represent a highly promising field as a new therapy approach to drive senescent cancer cells towards cell death to enhance treatment response. Here, we demonstrate that the Bcl-2 family of anti-apoptotic proteins in irradiated senescent sarcoma cells represents a senotherapeutic target to improve the cell death response in RT. This study paves the way for new treatment options in soft tissue sarcoma management.Radiotherapy (RT) is a key component of cancer treatment. Most of the time, radiation is given after surgery but for soft-tissue sarcomas (STS), pre-surgical radiation is commonly utilized. However, despite improvements in RT accuracy, the rate of local recurrence remains high and is the major cause of death for patients with STS. A better understanding of cell fates in response to RT could provide new therapeutic options to enhance tumour cell killing by RT and facilitate surgical resection. Here, we showed that irradiated STS cell cultures do not die but instead undergo therapy-induced senescence (TIS), which is characterized by proliferation arrest, senescence-associated β-galactosidase activity, secretion of inflammatory cytokines and persistent DNA damage. STS-TIS was also associated with increased levels of the anti-apoptotic Bcl-2 family of proteins which rendered cells targetable using senolytic Bcl-2 inhibitors. As oppose to radiation alone, the addition of senolytic agents Venetoclax (ABT-199) or Navitoclax (ABT-263) after irradiation induced a rapid apoptotic cell death in STS monolayer cultures and in a more complex three-dimensional culture model. Together, these data suggest a new promising therapeutic approach for sarcoma patients who receive neoadjuvant RT. The addition of senolytic agents to radiation treatments may significantly reduce tumour volume prior to surgery and thereby improve the clinical outcome of patients.

Preliminary results of the phase II ALTER-H003 trial: Anlotinib plus toripalimab as a first-line treatment for patients with unresectable hepatocellular carcinoma.

314 Background: Hepatocellular carcinoma (HCC) is particularly prevalent in China because of the high prevalence of chronic hepatitis B infection. In the last decade, sorafenib was the first molecular target drug approved for the treatment of advanced liver cancer. Until recently, the combination of angiogenic inhibitor with anti-PD-1 /L1 monoclonal antibody has represented significant efficacy over sorafenib monotherapy in the first-line regimen of unresectable HCC. Nevertheless, there are still unmet needs in clinical practice due to limited choice of drugs. Anlotinib is a novel oral multi-targeted tyrosine kinase inhibitor that targets VEGF receptors 1/2/3, FGF receptors 1-4, PDGF receptors α and β, and c-kit, and has been approved for standard treatment of non-small cell lung cancer, small cell lung cancer and soft tissue sarcoma in China. Toripalimab, a humanized IgG4 mAb against PD-1, is the first Chinese-produced PD-1 inhibitor marketed, which has been approved as a second-line treatment for metastasis melanoma. This study aims to evaluate the efficacy and safety of anlotinib plus toripalimab as the first-line treatment for unresectable HCC. Methods: This was a single-arm, multicenter, phase II trial. 30 patients with unresectable HCC, Child-Pugh ≤7 and ECOG PS &lt;2 will be enrolled if they had not been treated with prior treatment. Pts received anlotinib (12 mg, p.o., qd, d1-14, q3w) and toripalimab (240 mg, iv, d1, q3w) until disease progression or unacceptable toxicity. The primary endpoint was objective response rate (ORR) assessed by investigator according to mRECIST and irRECIST. Secondary endpoints included progression-free survival (PFS), overall survival (OS), disease control rate (DCR), duration of response (DoR) and safety. Results: By the cutoff date of September 21, 2020, 11 pts were enrolled and of those 8 pts were evaluable. 9 pts (81.8%) experienced treatment related adverse events (TRAEs) and grade 3 TRAEs occurred in 5 pts (45.5%). No grade 4 or above TRAEs occurred. The most common TRAEs were decreased appetite (n =6, 54.5%) and fatigue (n =5, 45.5%).Among 8 evaluable pts, unconfirmed ORR was 25% (95% CI 0.032-0.651) with 1 unconfirmed CR and 1 unconfirmed PR, and DCR was 87.5% (95% CI 0.473-0.997) according to mRECIST. Conclusions: Anlotinib in combination with toripalimab showed tolerant toxicity and preliminary anti-tumor efficacy in first-line regimen for uHCC patients. Furthermore, it is needed to be proved in update results and large scale studies. Clinical trial information: ChiCTR1900028295.

The Oncolytic Activity of Myxoma Virus against Soft Tissue Sarcoma Is Mediated by the Overexpression of Ribonucleotide Reductase.

Background:Myxoma virus (MYXV) is an oncolytic poxvirus that lacks the gene for 1 of the subunits of ribonucleotide reductase (RR), a crucial DNA synthesis and repair enzyme. The overexpression of RR has been implicated in the invasiveness of several cancers, including soft tissue sarcomas (STS). The purpose of the study was to investigate the oncolytic efficacy of MYXV in STS with different levels of RR expression.Methods:The oncolytic effect of recombinant MYXV was evaluated in 4 human STS cell lines, LS141 (a dedifferentiated liposarcoma), DDLS8817 (a dedifferentiated liposarcoma), RDD2213 (recurrent dedifferentiated liposarcoma), and HSSYII (a synovial sarcoma) using infectivity and cytotoxicity assays. Following the overexpression of RRM2 by cDNA transfection and silencing of RRM2 by siRRM2 in these STS cell lines, the RRM2 expression levels were analyzed by Western blot.Results:We observed a direct correlation between viral oncolysis and RRM2 mRNA levels (R = 0.96) in STS. Higher RRM2 expression was associated with a more robust cell kill. Silencing the RRM2 gene led to significantly greater cell survival (80%) compared with the control group (P = .003), whereas overexpression of the RRM2 increased viral oncolysis by 33% (P < .001).Conclusions:Our results show that the oncolytic effects of MYXV correlate directly with RR expression levels and are enhanced in STS cell lines with naturally occurring or artificially induced high expression levels of RR. Myxoma virus holds promise in the treatment of advanced soft tissue cancer, especially in tumors overexpressing RR.

Feline spindle cell soft tissue sarcoma: A case report

Feline spindle cell soft tissue sarcoma: A case report

Variability in tumor margin reporting for soft tissue sarcoma and cutaneous mast cell tumors in dogs: A systematic review.

To identify which classification systems have been used for tumor margin reporting and to determine whether factors (publication year, tumor type, and specialty of the contributing authors) influenced trends in margin reporting within literature describing canine soft tissue sarcoma (STS) and cutaneous mast cell tumors (MCT). Systematic literature review. Eligible articles were identified through electronic database searches performed for STS and MCT. Data abstracted from relevant studies included publication year, author list, specialty of contributing authors, criteria used to report the planned surgical margins, and the status of histologic margins. Categorization of papers was based on the classification systems used to report surgical and histologic tumor margins. Fifty-three articles were included, 11 on STS, 37 on MCT, and five that included both tumor types. Criteria for classifying the planned surgical margins were described in only 50.9% of studies. Articles that listed a veterinary surgeon as a contributing author (P = .01) and STS articles compared to MCT papers (P = .01) were more likely to report surgical margins. Most (56.6%) studies reported the status of histologic margins dichotomously as "complete" or "incomplete." Although a previously published consensus statement recommended that quantitative criteria be used to report histologic margins, only 7.5% of articles used quantitative methods. Classification systems used for reporting tumor margins were highly variable among studies. The findings of this review provide evidence that a standardized classification system for reporting surgical and histologic tumor margins is required in veterinary medicine. A universal system may support more consistent reporting of neoplastic biopsy specimens and allow for more meaningful comparisons across research studies.

CSPG4-Specific CAR.CIK Lymphocytes as a Novel Therapy for the Treatment of Multiple Soft-Tissue Sarcoma Histotypes.

No effective therapy is available for unresectable soft-tissue sarcomas (STS). This unmet clinical need prompted us to test whether chondroitin sulfate proteoglycan 4 (CSPG4)-specific chimeric antigen receptor (CAR)-redirected cytokine-induced killer lymphocytes (CAR.CIK) are effective in eliminating tumor cells derived from multiple STS histotypes in vitro and in immunodeficient mice. The experimental platform included patient-derived CAR.CIK and cell lines established from multiple STS histotypes. CAR.CIK were transduced with a retroviral vector encoding second-generation CSPG4-specific CAR (CSPG4-CAR) with 4-1BB costimulation. The functional activity of CSPG4-CAR.CIK was explored in vitro, in two- and three-dimensional STS cultures, and in three in vivo STS xenograft models. CSPG4-CAR.CIK were efficiently generated from patients with STS. CSPG4 was highly expressed in multiple STS histotypes by in silico analysis and on all 16 STS cell lines tested by flow cytometry. CSPG4-CAR.CIK displayed superior in vitro cytolytic activity against multiple STS histotypes as compared with paired unmodified control CIK. CSPG4-CAR.CIK also showed strong antitumor activity against STS spheroids; this effect was associated with tumor recruitment, infiltration, and matrix penetration. CSPG4-CAR.CIK significantly delayed or reversed tumor growth in vivo in three STS xenograft models (leiomyosarcoma, undifferentiated pleomorphic sarcoma, and fibrosarcoma). Tumor growth inhibition persisted for up to 2 weeks following the last administration of CSPG4-CAR.CIK. This study has shown that CSPG4-CAR.CIK effectively targets multiple STS histotypes in vitro and in immunodeficient mice. These results provide a strong rationale to translate the novel strategy we have developed into a clinical setting.

The application of aluminium phthalocyanine AlPs-4-mediated photodynamic therapy against human soft tissue sarcoma (RMS) cell line

The main challenge in the cancer treatment is the on-target drug delivery to the affected cells. Various therapies have been designed to target the affected cells efficiently but still the success is awaited. An iron and cobalt nanocomposite for the effective drug delivery to target cells was designed. The photodynamic effect of anticancer drugs loaded with iron oxide and cobalt ferrite nanomaterials coated with polyvinyl alcohol (PVA) was studied. The iron oxide nanoparticles (IONPs) and cobalt ferrite (CF) NPs without the loaded drugs were characterized by UV, XRD, FTIR, SEM and EDX techniques. The photodynamic effect of the photosensitizer, doxorubicin, and dacarbazine loaded nanomaterials were screened against human rhabdomyosarcoma (RMS) cells after incubation for 3 h, 24 h, and 48 h using MTT assay. The combination of photodynamic therapy (PDT) with chemo drugs is studied over different doses. When RMS cells were exposed to nanomaterials loaded with chemo drugs and PDT alone, it resulted in less cell killing compared to chemo drugs followed by PDT. These results revealed that in the case of combined treatment (combination therapy) the cell viability decreases as compared to individual treatment (monotherapy). The in vitro studies showed positive results which give a new pathway for the in vivo studies.

Multi-omics analysis of copy number variations of RNA regulatory genes in soft tissue sarcoma

AimsRNA regulatory genes were closely associated with tumorigenesis and prognosis in multiple tumors. Copy number variation (CNV) is a frequent characteristic in soft tissue sarcomas (STS). However, little is known regarding their possible roles in STS. Main methodsRNA sequence profiles and CNV data of 255 STS patients were downloaded from the Cancer Genome Atlas (TCGA). The correlation analysis involved CNVs of RNA regulatory genes, patient survival, immune infiltration, and DNA methylation. Drug sensitivity (IC50) was analyzed and validated by MTT assays in STS cell lines. Key findingsCNV events were frequently observed in all kinds (m6A, m5C, ac4C, m1A, m3C, m6Am, m7G, and Ψ) of RNA regulatory genes. Diploid copy number (CN) of METTL4 was associated with better overall survival (OS) in STS and the subtypes (leiomyosarcoma, LMS; dedifferentiated liposarcoma, DDLPS). In STS and LMS, diploid CN of METTL4 was significantly associated with higher infiltration fraction of resting mast cells. In STS and DDLPS, diploid CN of METTL4 possessed lower methylation level in CpG site of cg12105018, which represented better OS. Besides, sensitive drugs for STS cell lines were analyzed according to lower IC50 for the loss CN of METTL4. Temozolomide and Olaparib were identified. Further validation by MTT assays demonstrated that GCT was the most sensitive cell line to both Temozolomide and Olaparib. SignificanceCNV of METTL4 could be a prognostic biomarker for STS by potentially influencing mast cell infiltration and DNA methylation. Besides, STS with loss CN of METTL4 would be sensitive to Temozolomide and Olaparib.

Clear Cell Sarcoma of the Premaxillary Soft Tissue: A Case Report

Abstract Casestudy: Clear cell sarcoma of soft tissue (CCSST) is a rare translocation-associated sarcoma which uncommonly presents in the head and neck region. Because the tumor expresses markers of melanocytic differentiation, it can be difficult to distinguish from malignant melanoma and epithelioid malignant peripheral nerve sheath tumor (MPNST), necessitating clinical correlation and ancillary testing specifically fluorescence in situ hybridization (FISH) for EWSR1 rearrangement. Here we report a case of CCSST involving the premaxillary soft tissue. A 61-year-old male without significant past medical history presented with a right premaxillary subcutaneous mass which was slowly enlarging over the course of three years. During surgical resection of the mass, the infraorbital nerve was noted to course into the tumor. Histologic sections revealed a 2.5 cm mass composed of a nodular proliferation of epithelioid tumor cells with pale eosinophilic to vacuolated cytoplasm, large vesicular nuclei with prominent nucleoli, areas of punctate necrosis, and a mitotic rate of 10 mitoses per 10 high power fields. Scattered multinucleated tumor giant cells were also present. No pigment was identified. By immunohistochemistry, the tumor cells were diffusely positive for SOX-10 and S-100 and negative for HMB45, MART1, MITF, inhibin, calretinin, p63 and smooth muscle actin. FISH was positive for EWSR1 rearrangement. Altogether, the morphologic, immunohistochemical and cytogenetic findings supported the diagnosis of CCSST. Because the surgical margins were negative for malignancy, the patient did not require additional surgery but will be receiving adjuvant radiation therapy. Although rare in the head and neck, CCSST should be considered in the differential diagnosis when dealing with an epithelioid soft tissue tumor that expresses any melanocytic markers, and a definitive distinction from malignant melanoma and epithelioid MPNST can be achieved by utilizing FISH for EWSR1 rearrangement.

Impact of Wnt/β-Catenin Inhibition on Cell Proliferation through CDC25A Downregulation in Soft Tissue Sarcomas.

Simple SummaryGrowing evidence suggests that Wnt signaling may be crucial for tumorigenesis and progression of soft tissue sarcomas (STS). Inhibitors of this pathway are currently in clinical trials or pre-clinical studies in order to validate its utility in different neoplasia. One of this inhibitors, PRI-724, is showing promising results for advanced pancreatic adenocarcinoma or ovarian cancer. We found that PRI-724 is able to suppress cell viability/proliferation and to increase cell death rates of soft tissue sarcomas cells in vitro. CDC25A, a target gene of Wnt signaling pathway, is essential for STS proliferation because its downregulation via siRNA was able to mimic the effect of PRT-724 on cell cycle arrest and evaluation of NCBI/GenBank data confirmed its overexpression in STS patients’ samples. Moreover, in vitro administration of PRI-724 along with standard STS chemotherapeutic drugs improved the efficacy of chemotherapy, suggesting that Wnt inhibition could be a promising new therapeutic strategy in STS.The Wnt signaling pathway is an important cellular mechanism for regulating differentiation processes as well as cell cycle events, and different inhibitors of this pathway, for example, PRI-724, are showing promising results in clinical trials for treatment of advanced pancreatic adenocarcinoma or ovarian cancer. Growing evidence suggests that Wnt signaling may also be crucial for tumorigenesis and progression of soft tissue sarcomas (STS), a malignant neoplasm with few therapeutic options at an advanced state. Our study with several STS cell lines and primary cultures shows that inhibition of Wnt/β-catenin signaling with PRI-724 is able to suppress cell viability/proliferation and to increase cell death rates. TCF/β-catenin-mediated transcriptional activity is decreased in treated cells, leading to downregulation of its target genes CCND1 and CDC25A. The latter was critical because its downregulation via siRNA was able to mimic the effect of PRI-724 on cell cycle arrest and cell death induction. An evaluation of NCBI/GenBank data confirmed that CDC25A mRNA is elevated in STS patients. Importantly, PRI-724 in combination with standard STS chemotherapeutics doxorubicin or trabectedin enhanced their antitumoral effect in a synergistic manner according to isobolographic analysis, suggesting that Wnt inhibition through PRI-724 could be a beneficial combination regime in patients with advanced STS.