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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.

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 tables 1-3 and supplementary figures 1-3

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

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).

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.

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.

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.

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.

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%.

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.