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This study investigates the effect of Gliding Arc Plasma Activated Water (GAPAW) technique on maize germination and growth variables. The effect of GAPAW was evaluated on corn texture, water absorption in Lab conditions, and the pathway Scanning electron microscopy (SEM) analysis of corn seeds was also carry out. Maize seeds were sown 150 m2 and then watered with tap water and with GAPAW. Seed germination, maize growth and chlorophyll content were evaluated in field conditions in a complete randomized block design with four replicates using the Acid Tolerance Population (ATP) variety. Seed texture treated with 15 min of GAPAW making it darker than those of the control (tap water). Pathway SEM analysis showed no change for the 5 min-GAPAW-treated seeds compared to the control but at 15 min-GAPAW significant changes were observed. Germination was 100% at the 15 min-GAPAW compared to the other treatments (0 and 5 min). Stem length, leaf width, collar diameter, chlorophyll content and water uptake have higher values on plants watered with 15 min-GAPAW compared to others treatments. This application could highlight the germination properties of GAPAW in crop production.

Synovial sarcomas (SS) of the extremities are rare soft tissue sarcomas that are more common in young adults. We deciphered the imaging phenotype of SS with the aim to determine if imaging could provide an incremental value to currently known prognostic factors (PF)-age and histological grade-to predict long-term overall survival (OS).This retrospective multicenter study included consecutive pediatric and adult patients with synovial sarcomas of the extremities from December 2002 to August 2020. Inclusion criteria were (i) a follow-up greater than 5 years and (ii) available pre-therapeutic MRI. A subset analysis included MRI and CT-scan. Clinical, pathological, and imaging variables were collected in all patients. The primary endpoint was to evaluate the association of these variables with OS using univariate and multivariate Cox regressions.Out of 428 patients screened for eligibility, 98 patients (mean age: 37.1 ± 15.2 years) were included (MRI: n = 98/98, CT scan: n = 34/98; 35%). The median OS was 75.25 months (IQR = 55.50-109.12) and thirty-six patients (n = 36/98;37%) died during follow-up. The recurrence rate was 12.2% (n =12/98). SS lesions were mostly grade 2 (57/98; 58%). On MRI, SS had a mean long-axis diameter of 67.5 ± 38.3 mm. On CT scan, 44% (15/34) were calcified. Grade (hazard ratio [HR] = 2.71; 95%CI = 1.30-5.66; p = 0.008), size of the lesions evaluated on MRI (HR = 1.02; 95% CI = 1.01-1.03; p0.001), and calcifications on CT scan (HR = 0.10; 95% CI = 0.02-0.50; p = 0.005) were independent PF of OS.This study demonstrated that imaging biomarkers can be used to predict long-term outcome in patients with SS. Strikingly, the presence of calcifications on CT scan is associated with favorable outcome and provides an incremental value over existing PF such as age, grade, and size.• Beyond its diagnostic value, MRI is a pre-operative prognostic tool in synovial sarcomas of the extremities since the size of the lesion is an important prognostic factor. • Calcifications on CT scans are independently and significantly associated with prolonged overall survival.

Gemcitabine has shown clinical activity against angiosarcoma in small series, alone, or combined with taxanes. We aimed to evaluate its activity as a single-agent in a larger series of patients with advanced angiosarcoma. We retrospectively reviewed the electronic medical records of consecutive adult patients with advanced angiosarcoma treated with single-agent gemcitabine at our institutions from January 2010 to January 2021. Response was evaluated according to RECIST 1.1, and toxicity was graded according to NCI-CTC v5.0. 42 patients were identified. 38 patients (90%) had received prior anthracyclines and weekly paclitaxel, and 9 (21%) had received pazopanib. The best tumor response was partial response (PR) in 16 patients (38%), or stable disease (10 patients, 24%). All 8 patients with cardiac angiosarcoma experienced a PR. Median PFS was 5.4 months (95%CI: 3.1-6.5), and median OS was 9.9 months (95%CI: 6.6-13.4). Single-agent gemcitabine has clinically meaningful activity in advanced, heavily pre-treated angiosarcoma.

EWSR1/FUS-CREB-rearranged mesenchymal neoplasms are an emerging heterogeneous group of soft tissue tumors that encompasses low-grade lesions (angiomatoid fibrous histiocytoma/AFH) and a group of predominantly intra-abdominal aggressive sarcomas with epithelioid morphology and frequent keratin expression. Both entities occasionally harbor EWSR1::ATF1 fusions as alternate to the more frequent EWSR1/FUS::CREB1/CREM fusions. Although EWSR1/FUS-CREB-rearranged epithelioid malignant neoplasms have been described in diverse intra-abdominal sites, none involved the female adnexa. Herein, we describe three cases involving uterine adnexa in young females (41, 39, and 42-year-old); two associated with constitutional inflammatory symptoms. The tumors presented as a serosal surface mass of the ovary without parenchymal involvement (Case 1), as circumscribed nodule within ovarian parenchyma (Case 2), and as a periadnexal mass extending into the lateral uterine wall with lymph node metastasis (Case 3). They were composed of sheets and nests of large epithelioid cells with numerous stromal lymphocytes and plasma cells. The neoplastic cells expressed desmin and EMA, and variably WT1. One tumor expressed in addition AE1/AE3, MUC4, synaptophysin, chromogranin, and ALK. None expressed sex cord-associated markers. RNA sequencing identified EWSR1::ATF1 fusions in two cases and an EWSR1::CREM fusion in one. Exome-based RNA capture sequencing and clustering methods showed high transcriptomic proximity of tumor 1 with soft tissue AFH. This novel subset of female adnexal neoplasms should be included in the differential diagnosis of any epithelioid neoplasm involving female adnexa. Their aberrant immunophenotype can be misleading, underlining a wide spectrum of differential diagnosis.

Supplementary Figure from PBRM1 Deficiency Confers Synthetic Lethality to DNA Repair Inhibitors in Cancer

Supplementary Table from PBRM1 Deficiency Confers Synthetic Lethality to DNA Repair Inhibitors in Cancer

Supplementary tables 1-3 and supplementary figures 1-3

Purpose:Desmoid-type fibromatosis (DF) are locally aggressive neoplasms, with a need for effective systemic treatment in case of progression to avoid the short- and long-term complications of local treatments.Experimental Design:We retrospectively analyzed the outcomes of adult patients with DF treated with oral vinorelbine (90 mg once weekly) at Gustave Roussy Cancer Institute (Villejuif, Paris, France). Only patients with documented progressive disease according to RECIST v1.1 for more than 3 months (±2 weeks) before treatment initiation were included.Results:From 2009 to 2019, 90 out of 438 patients with DF were eligible for this analysis. Vinorelbine was given alone in 56 patients (62%), or concomitantly with endocrine therapy in 34 patients, for a median duration of 6.7 months. A partial response was observed in 29% and stable disease in another 57%. With a median follow-up of 52.4 months, the median time to treatment failure (TTF) was not reached. Progression-free rates at 6 and 12 months were 88.7% and 77.5%, respectively. Concomitant endocrine therapy was associated with longer TTF in women [HR, 2.16; 95% confidence interval (CI), 1.06–4.37; P = 0.03). Among 64 patients with documented CTNNB1 mutational status, p.S45F or p.S45P mutations were associated with longer TTF compared with p.T41A or wild-type tumors (HR, 2.78; 95% CI, 1.23–6.27; P = 0.04). Toxicity profile was favorable, without grade 3–4 toxicity, except for one grade 3 neutropenia.Conclusions:Oral vinorelbine is an effective, affordable, and well-tolerated regimen in patients with advanced, progressive DF. Prolonged activity was observed in patients with tumors harboring CTNNB1 p.S45F or p.S45P mutations.

Supplementary data from PBRM1 Deficiency Confers Synthetic Lethality to DNA Repair Inhibitors in Cancer

Inactivation of Polybromo 1 (PBRM1), a specific subunit of the PBAF chromatin remodeling complex, occurs frequently in cancer, including 40% of clear cell renal cell carcinomas (ccRCC). To identify novel therapeutic approaches to targeting PBRM1-defective cancers, we used a series of orthogonal functional genomic screens that identified PARP and ATR inhibitors as being synthetic lethal with PBRM1 deficiency. The PBRM1/PARP inhibitor synthetic lethality was recapitulated using several clinical PARP inhibitors in a series of in vitro model systems and in vivo in a xenograft model of ccRCC. In the absence of exogenous DNA damage, PBRM1-defective cells exhibited elevated levels of replication stress, micronuclei, and R-loops. PARP inhibitor exposure exacerbated these phenotypes. Quantitative mass spectrometry revealed that multiple R-loop processing factors were downregulated in PBRM1-defective tumor cells. Exogenous expression of the R-loop resolution enzyme RNase H1 reversed the sensitivity of PBRM1-deficient cells to PARP inhibitors, suggesting that excessive levels of R-loops could be a cause of this synthetic lethality. PARP and ATR inhibitors also induced cyclic GMP-AMP synthase/stimulator of interferon genes (cGAS/STING) innate immune signaling in PBRM1-defective tumor cells. Overall, these findings provide the preclinical basis for using PARP inhibitors in PBRM1-defective cancers.Significance:This study demonstrates that PARP and ATR inhibitors are synthetic lethal with the loss of PBRM1, a PBAF-specific subunit, thus providing the rationale for assessing these inhibitors in patients with PBRM1-defective cancer.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) or coronavirus disease 2019 (Covid-19), has uncontrollable effects on many organs. A great number of previously published scientific reports have revealed that patients with diabetes mellitus face a more severe form of Covid-19 with a higher death rate. Here we present the case of a 13-year-old unvaccinated boy who was admitted to an intensive care unit (ICU) with a history of fever, cough, dyspnea, throat pain, nausea, and confusion that progressed to lethargy after 24 h. On clinical examination, he was in a coma with Kussmaul's breathing, and was anuric. His blood biochemical analysis demonstrated hyperglycemia, severe metabolic acidosis, kidney failure, electrolyte disturbances, and inflammation. Chest x-ray showed pneumonia and a pleural effusion. The results of the SARS-CoV-2 real-time polymerase chain reaction were positive. The patient was diagnosed with Covid-19-induced acute respiratory distress syndrome associated with multisystem inflammatory syndrome in children secondary to his acute respiratory failure, acute kidney injury, and new-onset type 1 diabetes mellitus with diabetic ketoacidosis. He was intubated for invasive mechanical ventilation and received a normal saline infusion and continuous insulin infusion (0.1 IU/kg/h) for the treatment of his diabetic ketoacidosis. He was also treated with methylprednisolone, aspirin, and heparin, and underwent continuous renal replacement therapy for acute renal failure for 9 days. The patient was discharged from ICU on day 16 and was followed up regularly as an outpatient with daily treatment, including subcutaneous insulin injection (30 IU/day) and a calcium channel blocker for hypertension (nifedipine 20 mg/day).

Radiation therapy is a standard treatment for limbs soft tissue sarcomas. Preoperative versus postoperative radiotherapy has been a controversial topic for years. With preoperative irradiation, the treatment volume is more limited, the delivered dose possibly lower and the tumor volume easier to delimit. Only one randomized trial compared these two irradiation sequences. The results in terms of local control and survival were equivalent but the risk of acute postoperative complications was higher if irradiation was administered before surgery. However, in the latest update of this trial, patients who received adjuvant irradiation exhibited more severe late toxicity than those treated preoperatively. In addition, with modern irradiation techniques such as conformal with image-guided intensity modulated radiotherapy and flap coverage techniques, the incidence of complications after preoperative irradiation were lower than historically published rates. Locally advanced proximal sarcomas and the failure of other neoadjuvant treatments are nowadays classical indications for preoperative irradiation. As with other neoadjuvant treatments, induction radiotherapy must be personalized according to the histological subtype, the tumor site and the benefit/risk ratio, which is best appreciated by a multidisciplinary surgical and oncological team in a specialized center in the management of soft-tissue sarcomas.

Extraskeletal myxoid chondrosarcoma (EMC) is a rare soft tissue neoplasm of uncertain lineage characterized by the pathognomonic rearrangement of the NR4A3 gene, which in most cases is fused with EWSR1. Other NR4A3 fusion partners have been described, namely TAF15, FUS, TCF12, and TGF. Some studies suggest that EMCs with non-EWSR1 variant fusion are associated with high-grade morphology and worst clinical behavior compared to EWSR1::NR4A3 tumors, supporting the potential significance of particular fusion variant in EMC. We report a case of a 34-year-old male who presented with calf EMC and subsequently developed a slowly progressive metastatic disease 3 years after diagnosis. Whole-transcriptome analysis with total RNA sequencing enabled identification of a novel fusion transcript LSM14A::NR4A3, expanding the molecular spectrum of EMC.

Sarcoma with EWSR1-PATZ1 gene fusion is an exceedingly rare and newly described Ewing-like sarcoma harboring EWSR1 rearrangements involving fusion partners other than ETS family genes. The clinical, histopathologic and immunophenotypic features of cases reported in literature are fairly diverse and not specific. We report a new case report posing real challenges for histological and molecular diagnosis.

Purpose: Desmoid-type fibromatosis (DF) are locally aggressive neoplasms, with a need for effective systemic treatment in case of progression to avoid the short- and long-term complications of local treatments. Experimental Design: We retrospectively analyzed the outcomes of adult patients with DF treated with oral vinorelbine (90 mg once weekly) at Gustave Roussy Cancer Institute (Villejuif, Paris, France). Only patients with documented progressive disease according to RECIST v1.1 for more than 3 months (±2 weeks) before treatment initiation were included. Results: From 2009 to 2019, 90 out of 438 patients with DF were eligible for this analysis. Vinorelbine was given alone in 56 patients (62%), or concomitantly with endocrine therapy in 34 patients, for a median duration of 6.7 months. A partial response was observed in 29% and stable disease in another 57%. With a median follow-up of 52.4 months, the median time to treatment failure (TTF) was not reached. Progression-free rates at 6 and 12 months were 88.7% and 77.5%, respectively. Concomitant endocrine therapy was associated with longer TTF in women [HR, 2.16; 95% confidence interval (CI), 1.06–4.37; P = 0.03). Among 64 patients with documented CTNNB1 mutational status, p.S45F or p.S45P mutations were associated with longer TTF compared with p.T41A or wild-type tumors (HR, 2.78; 95% CI, 1.23–6.27; P = 0.04). Toxicity profile was favorable, without grade 3–4 toxicity, except for one grade 3 neutropenia. Conclusions: Oral vinorelbine is an effective, affordable, and well-tolerated regimen in patients with advanced, progressive DF. Prolonged activity was observed in patients with tumors harboring CTNNB1 p.S45F or p.S45P mutations.

A 27-year-old woman at 17 weeks gestation was admitted to the intensive care unit (ICU) with a history of fever, dyspnea, and dry cough for 3 days. She was diagnosed with coronavirus disease 2019 (COVID-19) based on her nasopharyngeal swab polymerase chain reaction (PCR) that was positive for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). In the ICU, the patient developed acute respiratory distress syndrome (ARDS) and increased levels of inflammatory markers. She was then intubated for mechanical ventilation and had a treatment for critical COVID-19 illness during pregnancy. She also received three cycles on alternating days of therapeutic plasma exchange (TPE) since she was failing to respond to conventional medical treatment. During hospitalization, the patient's fetus was closely monitored by repetitive ultrasound. After 27 days of hospitalization and 10 days of mechanical ventilation weaning, the patient's respiratory condition improved and her inflammatory biomarkers normalized. She was discharged from the hospital with an apparently healthy 20th week fetus. This case report highlights the role of TPE for treatment of ARDS due to cytokine storm in pregnant women with severe COVID-19 infection. This case emphasizes that careful evaluation of clinical and biological progression of the patient's status is very important and when conventional therapies are failing, alternative therapies such as TPE should be considered.

Effective treatments are scarce in non-operable scalp cutaneous angiosarcoma patients. Curative-intent definitive sequential IMRT and plesiobrachytherapy allowed complete response with limited side effect in two elder patients. This could represent a non-invasive therapeutic option for patients with locally advanced presentation.

Basal/squamous (Ba/Sq) subtype represents an intrinsic and robust group in the consensus molecular classification of muscle-invasive bladder cancer (MIBC), with poor outcome and controversial chemosensitivity. We aimed to investigate the spectrum of intratumor heterogeneity (ITH) in the Ba/Sq subtype. First, we validated a 29-gene NanoString CodeSet to predict the Ba/Sq subtype for FFPE samples. We identified heterogeneous Ba/Sq tumors in a series of 331 MIBC FFPE samples using dual GATA3/KRT5/6 immunohistochemistry (IHC). Heterogeneous regions with distinct immunostaining patterns were studied separately for gene expression using the 29-gene CodeSet, for mutations by targeted next-generation sequencing, and for copy number alteration (CNA) by microarray hybridization. Among 83 Ba/Sq tumors identified by GATA3/KRT5/6 dual staining, 19 tumors showed heterogeneity at the IHC level. In one third of the 19 cases, regions from the same tumor were classified in different distinct molecular subtypes. The mutational and CNA profiles confirmed the same clonal origin for IHC heterogeneous regions with possible subclonal evolution. Overall, two patterns of intratumoral heterogeneity (ITH) were observed in Ba/Sq tumors: low ITH (regions with distinct immunostaining, but common molecular subtype and shared CNA) or high ITH (regions with distinct immunostaining, molecular subtype, and CNA). These results showed multilayer heterogeneity in Ba/Sq MIBC. In view of personalized medicine, this heterogeneity adds complexity and should be taken into account for sampling procedures used for diagnosis and treatment choice. © 2021 The Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Probe-based confocal laser endomicroscopy (pCLE) is a noninvasive and real-time imaging technique allowing acquisition of in situ images of the tissue microarchitecture during oral surgery. We aimed to assess the diagnostic performance of pCLE combined with patent blue V (PB) in improving the management of early oral cavity, oro/hypopharyngeal, and laryngeal cancer by imaging squamous cell carcinoma in vivo. The prospective study enrolled 44 patients with early head and neck lesions. All patients underwent white-light inspection or panendoscopy depending on the lesion’s location, followed by pCLE imaging of the tumor core and its margins after topical application of PB. Each zone imaged by pCLE was interpreted at distance of the exam by three pathologists blinded to final histology. Most imaged zones could be presented to pathologists; the final sensitivity and specificity of pCLE imaging in head and neck cancers was 73.2–75% and 30–57.4%, respectively. During imaging, head and neck surgeons encountered some challenges that required resolving, such as accessing lesions with the flexible optical probe, achieving sufficiently precise imaging on the targeted tissues, and heterogeneous tissue staining by fluorescent dye. Final sensitivity scores were reasonable but final specificity scores were low. pCLE zones used to calculate specificity were acquired in areas of tumor margins, and the poor quality of the images acquired in these areas explains the final low specificity scores. Practical adjustments and technical training are needed to analyze head and neck lesions in various anatomical sites in real-time by pCLE.

AIMS Antibody-mediated rejection (AMR) is an emerging and challenging issue in transplantation. Endothelial deposition of C4d and microvascular inflammation (MI) are reliable markers of AMR in renal and cardiac transplantation, but remain controversial in the lung. Our aim was to assess C4d immunohistochemistry and histological patterns for the diagnosis of lung AMR. METHODS AND RESULTS We reviewed 158 transbronchial biopsies (TBBs) (n = 85 clinically indicated, and n = 73 surveillance TBBs) from 48 recipients, blinded to clinical and serological data. C4d was scored as 0, 1+ ( 50%). TBBs were reassessed for MI and acute lung injury (ALI). Donor-specific antibodies (DSAs), acute clinical graft dysfunction and chronic lung allograft graft dysfunction (CLAD) were recorded. C4d3+, C4d2+, C4d1+ and C4d0 occurred respectively in four (2.5%), six (3.8%), 28 (17.7%) and 120 (75.9%) TBBs. MI and ALI were rare but more frequent in C4d1-3+ TBBs than in the absence of C4d. C4d2+ was frequently observed with concomitant infection. Among the surveillance TBBs, only two (2.7%) showed MI. Neither ALI nor C4d3+ was diagnosed on surveillance TBBs. No significant association was found between histopathological findings and DSAs. All four patients with C4d3+ could retrospectively be diagnosed with AMR and developed CLAD. CONCLUSION Although rare, diffuse C4d deposition appears to be a strong indication of acute clinical AMR in lung transplant patients, whereas intermediate C4d2+ requires more investigations. In stable patients, histopathology and C4d may lack the sensitivity to diagnose subclinical AMR. This emphasises the need for a multidisciplinary evaluation of each suspected AMR case, and the need for complementary diagnostic tools.

Introduction: Desmoplastic small round cell tumor (DSRCT) is a rare and very aggressive subtype of sarcoma, affecting predominantly young males. DSRCT is molecularly characterized by the t(11;22)(q24;q12) chromosomal translocation, which fuses EWSR1 to WT1 and encodes an aberrant chimeric transcription factor. A few additional secondary mutations have been reported, notably in genes encoding proteins involved in chromatin remodeling and DNA repair. No therapeutic advance have been done in the past 20 years, and the five-year overall survival is still below 15% in metastatic cases, despite aggressive poly-chemotherapy and extensive surgical debulking. Thus, we aimed to identify actionable targeted dependencies to propose novel therapeutic approaches, using drug screening and molecular biology. Material and Methods: Drug sensitivity screen evaluating 80 cancer-relevant small molecule inhibitors in dose-response was performed on the JN-DSRCT-1 (JN1) cell line. Revalidation was performed on JN1 and in a Champions Oncology PDX-derived in-house-generated DSRCT cell line (R) using multiple clinical compounds. Mechanistic dissection was performed using flow cytometry, immunofluorescence, western blotting, DNA fiber assay, RT-qPCR, and dot blot to evaluate markers of DNA damage response, replication stress and cell-autonomous innate immune signaling. Results: The drug screen identified that JN1 cells were particularly sensitive to one ATR inhibitor (ATRi) and multiple PARPi inhibitors (PARPi), which scored #6, 7, 9 and 20 of the hits. Sensitivity to ATRi was revalidated in the JN1 and R cell lines, whereas sensitivity to PARPi was only revalidated in the JN1 cell line - consistent with the very low PARP1 expression in that cell line. Sensitivity to ATRi and PARPi was partially reversed upon EWS-WT1 silencing. Combination experiments revealed that both drugs were synergistic in the JN1 cell line. Exposure to PARP inhibitor increased the number of γH2AX and RAD51 foci, suggesting that JN1 cells were not deficient in homologous recombination. Further, PARPi or ATRi exposure decreased replication fork speed and increased R-loops accumulation, whereas EWS-WT1 silencing increased replication fork speed. Resistance to PARPi and to ATRi was restored upon exogenous expression of the R-loop resolution enzyme RNAseH1. The combination of PARPi and ATRi also induced micronuclei formation and activated the cGAS-STING cytosolic innate immune sensing pathway, together with increased expression of CCL5, CXCL10 and PD-L1. Conclusion: DSRCT cells are sensitive to ATR and PARP inhibitors, through a mechanism that, at least in part, may involve enhanced replication stress and increased R-loop formation. Our data provide the preclinical rationale for assessing ATRi combined with PARPi in patients with DSRCT, as a cytotoxic therapy or as an immunomodulatory approach to enhance cell-autonomous immunogenicity. Citation Format: Asuka KAWAI-KAWACHI, Madison LENORMAND, Clémence HENON, Thomas EYCHENNE, Leo COLMET-DAAGE, Nicolas DORVAULT, Marlene GARRIDO, Clémence ASTIER, Carine NGO, Helen PEMBERTON, Aditi GULATI, Stephen PETTITT, Roman CHABANON, Christopher J LORD, Sophie POSTEL-VINAY. Identification of ATR inhibitors as therapeutic opportunities in Desmoplastic small round cell tumors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 6277.

Introduction: Cholangiocarcinoma (CCA) is a rare tumor accounting for less than 2% of all human malignancies. Mutations in the mammalian Switch/Sucrose Non-Fermentable (SWI/SNF) chromatin remodeling complex have been identified in approximately 20% of all human cancers and more than 40% of CCA. If the role of SWI/SNF on the tumor biology and microenvironment is being uncovered in other tumor types, it remains vastly unknown in CCA. Here, we wanted to investigate, using clinical samples, the association between SWI/SNF defects, the molecular landscape and tumor immune microenvironment in CCA. Material and Methods: Mutation profiling of 103 patients with CCA from Gustave Roussy (GR) was assessed with FoundationOne® CDx 324-gene NGS panel. Formalin-fixed paraffin-embedded tumor tissues from 11 patients with CCA were analyzed for SWI/SNF (PBRM1, ARID1A, SMARCB1 and SMARCA4) and Polycomb-DUB (BAP1) subunits, as well as lymphocytic markers (CD4 and CD8). Stainings were performed using a VENTANA BenchMark ULTRA. Absolute count of positive cells per mm2 was determined by digital image analysis with the Definiens Developer XD™ by selecting at least 5 independent tumoral zones. Pairwise comparisons were performed using the Mann-Whitney test. Results: Among the patients for whom NGS data was available, the most frequently altered genes were TP53 (27%), CDKN2A (17%), KRAS (16%), ARID1A (12%), FGFR2 (12%) and PBRM1 (10%). Subunits of the SWI/SNF complex and BAP1 were mutated in 25% and 10% of cases, respectively. Tumor samples with ARID1A mutations (n=5) showed strongly decreased ARID1A expression compared to WT tumor samples (n=2) (mean = 308.03 positive nuclei (p.n)/mm2 versus 6096.82 p.n/mm2 respectively; P Conclusion: Mutations in the ARID1A and PBRM1 subunits of the SWI/SNF complex are associated with a loss of protein expression. In contrast to what reported in other tumor types, ARID1A and PBRM1 mutations correlated with poorly lymphocytic TIME in CCA. Decreased PBRM1 expression is further associated with decreased ARID1A expression. Independent revalidation and further study in larger tumor series are warranted. Citation Format: Clémence Astier, Jean-Yves Scoazec, Virginie Marty, Olivia Bawa, Nicolas Signolle, Carine Ngo, Francesco Facchinetti, Antoine Hollebecque, Sophie Postel-Vinay. Characterization of SWI/SNF complex gene mutations, protein expression and tumor immune microenvironment in cholangiocarcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 5714.

Background The aim of the study is to evaluate functional and oncological outcomes of patients undergoing abdominal wall soft tissue tumors (AWSTT) surgery. Methods All consecutive patients that underwent surgery for malignant and intermediate AWSTT from 1999 to 2019 were retrospectively analyzed. Results Ninety-two patients were identified, 20 (22%) operated on for a desmoid tumor and 72 (78%) for a soft tissue sarcoma (STS). Fifty-two patients (57%) had in toto resection of the abdominal wall (from the skin to the peritoneum) and 9 (10%) required simultaneous visceral resection. The closure was direct in 28 patients (30%) and requiring a mesh, a flap or a combination of the two in respectively 42, 16, and 6 patients (47%, 17%, 6%). The postoperative complications rate was 26%. Thirteen patients (14%) developed an incisional hernia after a median delay of 27 months. After a median follow-up of 40 months, out of the 72 patients operated on for STS, 7 (10%) developed local recurrence and 11 (15%) distant recurrence. The median recurrence-free and overall survivals were 61 and 116, months respectively. Conclusions Management of AWSTT requires extensive surgery but allows good local control with an acceptable rate of incisional hernia.

Inactivation of Polybromo 1 (PBRM1), a specific subunit of the PBAF chromatin remodeling complex, occurs frequently in cancer, including 40% of clear cell renal cell carcinomas (ccRCC). To identify novel therapeutic approaches to targeting PBRM1-defective cancers, we used a series of orthogonal functional genomic screens that identified PARP and ATR inhibitors as being synthetic lethal with PBRM1 deficiency. The PBRM1/PARP inhibitor synthetic lethality was recapitulated using several clinical PARP inhibitors in a series of in vitro model systems and in vivo in a xenograft model of ccRCC. In the absence of exogenous DNA damage, PBRM1-defective cells exhibited elevated levels of replication stress, micronuclei, and R-loops. PARP inhibitor exposure exacerbated these phenotypes. Quantitative mass spectrometry revealed that multiple R-loop processing factors were downregulated in PBRM1-defective tumor cells. Exogenous expression of the R-loop resolution enzyme RNase H1 reversed the sensitivity of PBRM1-deficient cells to PARP inhibitors, suggesting that excessive levels of R-loops could be a cause of this synthetic lethality. PARP and ATR inhibitors also induced cyclic GMP-AMP synthase/stimulator of interferon genes (cGAS/STING) innate immune signaling in PBRM1-defective tumor cells. Overall, these findings provide the preclinical basis for using PARP inhibitors in PBRM1-defective cancers. Significance: This study demonstrates that PARP and ATR inhibitors are synthetic lethal with the loss of PBRM1, a PBAF-specific subunit, thus providing the rationale for assessing these inhibitors in patients with PBRM1-defective cancer.

Radiation therapy is a standard treatment for limbs soft tissue sarcomas. Preoperative versus postoperative radiotherapy has been a controversial topic for years. With preoperative irradiation, the treatment volume is more limited, the delivered dose possibly lower and the tumor volume easier to delimit. Only one randomized trial compared these two irradiation sequences. The results in terms of local control and survival were equivalent but the risk of acute postoperative complications was higher if irradiation was administered before surgery. However, in the latest update of this trial, patients who received adjuvant irradiation exhibited more severe late toxicity than those treated preoperatively. In addition, with modern irradiation techniques such as conformal with image-guided intensity modulated radiotherapy and flap coverage techniques, the incidence of complications after preoperative irradiation were lower than historically published rates. Locally advanced proximal sarcomas and the failure of other neoadjuvant treatments are nowadays classical indications for preoperative irradiation. As with other neoadjuvant treatments, induction radiotherapy must be personalized according to the histological subtype, the tumor site and the benefit/risk ratio, which is best appreciated by a multidisciplinary surgical and oncological team in a specialized center in the management of soft-tissue sarcomas.

Therapeutic options in patients with metastatic osteosarcoma are limited and effective systemic treatments are needed in this setting. The aim of this case series was to assess the efficacy and toxicity of oral metronomic etoposide in adult patients with progressive metastatic osteosarcoma. We retrospectively reviewed the electronic records of patients treated with oral metronomic etoposide (25 mg thrice daily, 3 weeks out of 4) from December 2002 to December 2018 at Gustave Roussy (Villejuif, France). The primary endpoint was progression‐free rate (PFR) at 4 months; secondary endpoints were: best response (according to RECIST v1.1), progression‐free survival (PFS), overall survival (OS) and safety. With a median follow‐up of 9.8 months, 37 patients were eligible for this analysis: 68% males, median age 42 (range: 21–75), 19% with synchronous metastases, 92% with lung metastases, median PS: 1 (range: 0–3). Median number of previous treatment lines in the metastatic setting was 1 (range: 0–4). Progression‐free rate at 4 months was 40.3% (95% CI: 24.5–56.2). Best response was partial response in 11% and stable disease in 35% of patients (disease control rate: 46%). Median PFS was 3.1 months (95% CI: 2.5–4.7) and median OS was 9.8 months (95% CI: 5.1–12.3). Toxicity profile was acceptable, with 13% grade 3 haematological toxicities (anaemia and neutropenia), without any grade 3–4 non‐haematological toxicity. In our experience, oral metronomic etoposide demonstrated effective palliation along with acceptable toxicity in patients with progressive metastatic osteosarcoma., Oral metronomic etoposide demonstrated effective palliation (progression‐free rate at 4 months was 40.3% (95%CI: 24.5‐56.2), along with acceptable toxicity in adult patients with metastatic osteosarcoma.

Sarcoma with EWSR1-PATZ1 gene fusion is an exceedingly rare and newly described Ewing-like sarcoma harboring EWSR1 rearrangements involving fusion partners other than ETS family genes. The clinical, histopathologic and immunophenotypic features of cases reported in literature are fairly diverse and not specific. We report a new case report posing real challenges for histological and molecular diagnosis.

Objective Urothelial carcinoma (UC) is heterogeneous, and variant histologies (VH) are more frequent than initially reported. Reporting VH is recommended by several guidelines because of prognostic and therapeutic implications. We evaluated the concordance of VH between the first transurethral resections of the bladder (TURBs) and the following radical cystectomy (RC). This paper is the first to compare VH with a central pathology review between TURB and RC. Material and methods In this retrospective study, we only included those patients who underwent TURB with VH and then RC between 01/2010 and 12/2013 at our institution. The presence of VH in both TURB and RC was assessed and compared according to the 2016 World Health Organization (WHO) classification by a central pathology review. Results Among 110 patients who had the initial TURB/RC, 54 (49.1%) were diagnosed with VH, 48 (43%) had a single pattern, and six had (5%) multiple histological patterns. Squamous differentiation was the most common single VH (31%). Twenty patients with UC (18%) showed discordance between TURB and RC, especially in micropapillary versus nested (n=3) cases. Concordant histology between TURB/RC was seen in 82% of the cases. Conclusion Discrepancies can be seen between TURB and RC when reporting VH, which can be problematic for selection of therapy and management. TURB alone might be insufficient to evaluate the presence of VH, especially in VH with heavy therapeutic implications, such as micropapillary carcinomas. Nevertheless, concordance with a central review by an experienced uropathologist when applying the WHO 2016 classification is 82%.

Resume Nous rapportons le cas d’une patiente de 65 ans ayant consulte pour dysphonie. Le scanner ORL et la laryngoscopie montraient une lesion tumorale endolaryngee supraglottique droite. La biopsie de la lesion montrait un infiltrat lymphoide dense du chorion sans necrose ni ulceration de la muqueuse. L’infiltrat etait riche en lymphocytes T reactionnels CD3+, CD5+, CD4+, CD8+ et plasmocytes. Des cellules lymphoides de plus grande taille plus atypiques exprimaient CD56 et les marqueurs de cytotoxicite TIA1 et granzyme B. L’hybridation in situ avec la sonde EBER pour la recherche d’EBV montrait de nombreuses cellules positives. Il s’agit d’un lymphome T/NK extraganglionnaire de type nasal, dont la localisation primitive laryngee est exceptionnelle. L’abondance de l’infiltrat inflammatoire reactionnel peut faire evoquer a tort une pathologie inflammatoire et faire retarder le diagnostic.

e23547 Background: Gemcitabine has shown clinical activity against angiosarcoma in single cases or small series, as a single agent or in combination with taxanes. We aimed to evaluate its activity in a larger, homogeneous series of pts with advanced angiosarcoma. Methods: We retrospectively reviewed the electronic medical records of consecutive adult pts with advanced angiosarcoma treated with gemcitabine(1000 mg/m2 IV days 1, 8 and 15 of a 28-days cycle) at our institutions from January 2010 to January 2021. All cases were confirmed by an expert pathologist from the French network of Sarcoma pathologists (RRePS). Response was evaluated according to RECIST 1.1, and survival was estimated using the Kaplan-Meier method. Results: Fourty-two pts [median age : 52 years (range : 25-90), median PS : 1 (range : 0-2), 36 women)] were identified. Ten (24%) had locally advanced disease; the median number of metastatic sites was 2 (range : 1-4) in the remaining 32 pts. Primary tumor site was breast/chest wall (12), face/scalp (9), heart (8), other viscera (7) or limbs (7). Eleven (26%) occurred in previously irradiated field. All pts but 4 had received prior anthracyclines (peri-operatively or in the metastatic setting) and weekly paclitaxel, and 9 (21%) had received pazopanib. Gemcitabine was given as second, third and fourth line in 7 (17%), 20 (47%) and 15 (36%) pts, respectively. Best tumor response by RECIST 1.1 was PR (16 pts, 38%), SD (10 pts, 24%) or PD (16 pts, 38%). All pts with cardiac angiosarcoma experienced a PR, as well as 4 of 10 pts with angiosarcoma arising in irradiated field and 3 of the 4 pts with CNS metastases. Median PFS was 5.4 months (95%CI : 3.1-6.5), and median OS was 9.9 months (95%CI : 6.6-13.4). Two febrile neutropenia occurred. No new safety signal was observed. Ongoing studies are exploring whether candidate biomarkers such as hENT1 expression could help predicting the benefit of gemcitabine in pts with angiosarcoma of the heart and large vessels. Conclusions: Single agent gemcitabine has clinically meaningful activity in advanced, heavily pre-treated angiosarcoma (including cardiac angiosarcoma). This series may serve as a benchmark for future clinical trials in angiosarcoma.

11528 Background: Chordoma is a rare tumor with no approved therapy. Preclinical studies have shown expression of EGFR and activated EGFR family kinases (EGFR, HER2 and HER4). Erlotinib and other anti-EGFR agents (gefitinib and cetuximab) have shown clinical activity in advanced chordoma in single case reports or small series. We aimed to evaluate the activity of erlotinib in a larger, homogeneous series of pts with advanced chordoma. Methods: We retrospectively reviewed the electronic medical records of consecutive adult pts with advanced chordoma progressive over 6 months (+/- 2 weeks, according to RECIST 1.1), treated with erlotinib (150 mg daily) at Gustave Roussy (Villejuif, France) following multidisciplinary tumor board discussion, from January 2010 to January 2021. All cases were confirmed by an expert pathologist. Response was evaluated according to RECIST 1.1, and survival was estimated using the Kaplan-Meier method. Results: Thirty-one pts [median age : 60 years (range : 32-88), median PS : 2 (range : 1-3), 30 males)] were identified. Twenty-seven (87%) had locally advanced disease; the median number of metastatic sites was 1 (range : 1-2) in the remaining 4 pts. Primary tumor site was sacral (25), lumbar (3) or cervical (3). All pts but 6 had undergone prior surgery, and 29 (94%) had undergone radiotherapy of the primary tumor. Eight pts had received previous systemic treatments (imatinib in 4, sorafenib and regorafenib in 2 each). Best tumor response by RECIST 1.1 was PR (4 pts, 13%), SD (14 pts, 45%) or PD (13 pts, 42%). Median PFS was 6.2 months (95%CI : 4.5-9.8), and median OS was 15.9 months (95%CI : 10.6-20.2). Fourteen pts (45%) remained progression-free after 1 year, and three (10%) after two years under erlotinib. Grade 3 diarrhea occurred in 4 pts (13%) and grade 3 skin rash in 13 pts (42%). Twelve pts (39%) required dose reduction to 100 mg daily due to multiple grade 2 toxicities. Ongoing studies are exploring whether candidate biomarkers such as EGFR and HER2 expression or amplification, and their mutational status could help predicting the benefit of erlotinib in pts with advanced chordoma. Conclusions: Erlotinib has clinically meaningful but unpredictable activity in advanced chordoma. Molecular profiling would probably be of high interest in this setting. This series may serve as a benchmark for future clinical trials in chordoma.

In lung transplantation, there is a increased recognition on the deleterious role of DSA for chronic dysfunction of pulmonary graft. In order to optimize the therapeutic stategies and to guide clinical trials, the standardized definition of humoral reject (HR) criteria in lung transplantation proves to be necessary. The deposit of C4d complement, confirming the activation of secondary complement in interaction between DSA and target cells, is a reliable biomarker of HR in renal and cardiac transplantations. However, with the same histopathological criteria from HR of kidney and heart, which is less robust when applying in lung transplantation. The sometime contradictory results of C4d are in part linked to variability between medical centers and in the interpretation of this marker. Although no definition of the criteria of HR had been established, ISHLT (International Society for Heart and Lung Transplantation) recommends a multidisciplinary approach to HR for lung transplantation, including the tripod diagnosis: 1) presence of clinical dysfunction, 2) presence of DSA and 3 ) presence of histopathological lesions of C4d with endothelial deposit in the alveolar capillaries.

We report the case of a 65-year-old woman who presented with a dysphonia. ENT tomography and laryngoscopy showed an endolaryngeal tumoral lesion extended to the right supraglottis. Biopsy of the lesion revealed dense lymphoid infiltrate in the lamina propria, without necrosis or ulceration of the mucosa. The infiltrate showed many CD3+, CD5+, CD4+, CD8+ lymphocytes and plasmocytes. Larger lymphoid cells with cytologic atypia expressed CD56 and cytotoxicity markers such as TIA1 and granzyme B. In situ hybridization for EBV revealed numerous positive cells. The diagnosis of extranodal NK/T cell lymphoma was proposed. The primary laryngeal localization of this disease is exceptionally rare. Heavy admixture of inflammatory cells may mimic inflammatory process and delay the diagnosis.

Background. Endothelial C4d deposition is used as a reliable marker to diagnose antibody-mediated rejection (AMR) in renal and cardiac transplantation, but remains controversial in lung transplantation. Objectives. This study was planned to study the utility of C4d by immunohistochemistry (IHC) for the diagnosis of AMR in lung allograft transbronchial biopsies (TBBx). Methods. We evaluated C4d staining by IHC in a blinded fashion, on 158 consecutive formalin-fixed paraffin embedded (FFPE) TBBx, from a cohort of 48 lung transplant recipients. Capillary C4d staining was scored as 0, 1 ( 50% capillaries). C4d scoring was compared with microvascular inflammation (MI), acute lung injury (ALI), acute cellular rejection (ACR), concomitant donor specific antibodies (DSA) and graft dysfunction. Results. C4d3, C4d2, C4d1 and C4d0 occurred in 4 (2.5%), 6 (3.8%), 28 (17.7%) and 120 (75.9%) TBBx, respectively. Capillary staining ≥C4d2 was significantly associated with MI (50% vs 7.4%; p=0.001). C4d3 occurred more frequently with DSA (5.5% in DSA group vs 1.5% in non-DSA group) and was significantly associated with graft dysfunction (p=0.03). Histopathologic findings alone did not correlate with DSA. Three out of 4 patients with C4d3 developed bronchiolitis obliterans syndrome (BOS). Conclusions. Diffuse C4d deposition in endothelial capillaries was correlated with MI and graft dysfunction. It was closely related, but not significantly, with DSA and development of BOS in lung transplant recipients. Diffuse C4d deposition in the lung might be a specific marker for AMR, although lacking sensitivity.