Prothrombin Complex Concentrate vs Conservative Management in ICH Associated With Direct Oral Anticoagulants.

Importance: Intracerebral hemorrhage (ICH) associated with direct oral anticoagulant (DOAC) use carries extremely high morbidity and mortality. The clinical effectiveness of hemostatic therapy is unclear. Objective: To compare the clinical and radiological outcomes of DOAC-associated ICH treated with prothrombin complex concentrate (PCC) vs conservative management. Design, Setting, and Participants: In this population-based, propensity score-weighted retrospective cohort study, patients who developed DOAC-associated ICH from January 1, 2016, to December 31, 2021, in Hong Kong were identified. The outcomes of patients who received 25 to 50 IU/kg PCC with those who received no hemostatic agents were compared. Data were analyzed from May 1, 2022, to June 30, 2023. Main Outcomes and Measures: The primary outcome was modified Rankin scale of 0 to 3 or returning to baseline functional status at 3 months. Secondary outcomes were mortality at 90 days, in-hospital mortality, and hematoma expansion. Weighted logistic regression was performed to evaluate the association of PCC with study outcomes. In unweighted logistic regression models, factors associated with good neurological outcome and hematoma expansion in DOAC-associated ICH were identified. Results: A total of 232 patients with DOAC-associated ICH, with a mean (SD) age of 77.2 (9.3) years and 101 (44%) female patients, were included. Among these, 116 (50%) received conservative treatment and 102 (44%) received PCC. Overall, 74 patients (31%) patients had good neurological recovery and 92 (39%) died within 90 days. Median (IQR) baseline hematoma volume was 21.7 mL (3.6-66.1 mL). Compared with conservative management, PCC was not associated with improved neurological recovery (adjusted odds ratio [aOR], 0.62; 95% CI, 0.33-1.16; P = .14), mortality at 90 days (aOR, 1.03; 95% CI, 0.70-1.53; P = .88), in-hospital mortality (aOR, 1.11; 95% CI, 0.69-1.79; P = .66), or reduced hematoma expansion (aOR, 0.94; 95% CI, 0.38-2.31; P = .90). Higher baseline hematoma volume, lower Glasgow coma scale, and intraventricular hemorrhage were associated with lower odds of good neurological outcome but not hematoma expansion. Conclusions and Relevance: In this cohort study, Chinese patients with DOAC-associated ICH had large baseline hematoma volumes and high rates of mortality and functional disability. PCC treatment was not associated with improved functional outcome, hematoma expansion, or mortality. Further studies on novel hemostatic agents as well as neurosurgical and adjunctive medical therapies are needed to identify the best management algorithm for DOAC-associated ICH.

A randomised controlled pilot study of a Nintendo Ring Fit Adventure™ balance and strengthening exercise program in community-dwelling older adults with a history of falls.

OBJECTIVES: This pilot study examined the feasibility, acceptability, and effects of a Nintendo Ring Fit Adventure™-based balance and muscle strengthening exercise program in community-dwelling older adults with a history of falls. METHODS: Older adults who have had at least one fall in the past year were randomly assigned to an experimental (n = 21) or control group (n = 21). The experimental group performed 16 exercise sessions in total, lasting 60 min each, twice a week for 8 weeks, whereas the control group received usual care. Feasibility was evaluated based on the scores of participants in the exercises. Acceptance was evaluated using a customised questionnaire examining participants' self-perceived enjoyment, feasibility and improvements. Clinical outcomes including balance (Mini-BESTest), lower limb muscle strength (Five-Time Sit-to-Stand test), mobility (Timed-Up and Go test), dual-task ability (Timed-Up and Go test-Dual Task), fear of falling (Icon-FES) and executive function (Color Trails Test) were evaluated at baseline and 8 weeks. RESULTS: Thirty-one participants (74%) finished the 8-week assessment. The experimental group significantly improved their scores in six out of eight exercises (all p < .031). The mean scores of the self-perceived enjoyment, feasibility and improvement domains of the acceptability questionnaire were 3.46 ± .53, 3.08 ± .59, and 3.47 ± .57 respectively. A significant improvement in the anticipatory subscore of the Mini-BESTest was found in the experimental group compared to the control group (p = .02; Partial eta squared = .14). CONCLUSIONS: The Nintendo Ring Fit Adventure™-based exercise program was feasible, acceptable, and potentially effective in community-dwelling older adults with a history of falls.

Feasibility, safety, and effects of a Nintendo Ring Fit Adventure™ balance and strengthening exercise program in community-dwelling older adults with a history of falls: A feasibility randomized controlled trial.

AIM: This pilot study examined the feasibility, safety, and effects of a Nintendo Ring Fit Adventure™-based exercise program to enhance balance and lower limb muscle strength in community-dwelling older adults with a history of falls. METHODS: In total, 42 older adults who experienced at least one fall in the past year were randomly assigned to an experimental or control group. Participants in the experimental group performed 60-min sessions of the exercise program twice per week for 8 weeks. The control group received usual care. We assessed the feasibility (retention and adherence to the exercise program), safety (number of adverse events), and clinical outcomes: (1) balance (Mini-BESTest); (2) functional lower limb muscle strength (Five-Time Sit-to-Stand test); (3) mobility (Timed-Up and Go test); (4) dual-task ability (Timed-Up and Go test - Dual Task); (5) fear of falling (Icon-FES); and (6) executive function (Color Trails Test). RESULTS: Thirty-one participants (74%) completed the 8-week assessment. No adverse event associated with the exercise program was reported. There was a significant interaction in the anticipatory domain score of the Mini-BESTest between the experimental and control groups over the 8 weeks (P = 0.019). CONCLUSIONS: The Nintendo Ring Fit Adventure™-based exercise program was feasible, safe, and potentially effective in improving anticipatory balance in community-dwelling older fallers. Geriatr Gerontol Int 2024; 24: 334-341.

Association of Alternative Anticoagulation Strategies and Outcomes in Patients With Ischemic Stroke While Taking a Direct Oral Anticoagulant.

BACKGROUND AND OBJECTIVES: Ischemic stroke despite a direct oral anticoagulant (DOAC) is increasingly common and portends a high risk of subsequent ischemic stroke. The efficacy and safety of antithrombotic regimens after the condition are unclear. We aimed to compare the outcomes of patients with ischemic stroke despite DOACs with and without an alternative antithrombotic regimen and determine the risk factors of recurrent ischemic stroke while on anticoagulation. METHODS: In a population-based, propensity score-weighted, retrospective cohort study, we compared the clinical outcomes of DOAC-to-warfarin switch, DOAC-to-DOAC switch (DOACswitch), or addition of antiplatelet agents, with those of unchanged DOAC regimen (DOACsame) among patients with nonvalvular atrial fibrillation (NVAF) who developed the first ischemic stroke despite a DOAC from January 1, 2015, to December 31, 2020, in Hong Kong. The primary outcome was recurrent ischemic stroke. Secondary outcomes were intracranial hemorrhage, acute coronary syndrome, and death. We performed competing risk regression analyses to compare the clinical endpoints and determined the predictors of recurrent ischemic stroke in an unweighted multivariable logistic regression model. RESULTS: During the 6-year study period, among 45,946 patients with AF on a DOAC as stroke prophylaxis, 2,908 patients developed ischemic stroke despite a DOAC. A total of 2,337 patients with NVAF were included in the final analyses. Compared with DOACsame, warfarin (aHR 1.96, 95% CI 1.27-3.02, p = 0.002) and DOACswitch (aHR 1.62, 95% CI 1.25-2.11, p < 0.001) were associated with an increased risk of recurrent ischemic stroke. In the DOACsame group, adjunctive antiplatelet agent was not associated with a reduced risk of recurrent ischemic stroke. Diabetes mellitus, concurrent cytochrome P450/P-glycoprotein (CYP/P-gp) modulators, and large artery atherosclerotic disease (LAD) were predictors of recurrent ischemic stroke. DISCUSSION: In patients with NVAF with ischemic stroke despite a DOAC, the increased risk of recurrent ischemic stroke with switching to warfarin called for caution against such practice, while the increased ischemic stroke with DOAC-to-DOAC switch demands further studies. Adjunctive antiplatelet agent did not seem to reduce ischemic stroke relapse. Because diabetes mellitus, the use of CYP/P-gp modulators, and LAD were predictors of recurrent ischemic stroke, further investigations should evaluate whether strict glycemic control, DOAC level monitoring, and routine screening for carotid and intracranial atherosclerosis may reduce ischemic stroke recurrence in these patients. CLASSIFICATION OF EVIDENCE: This study provides Class II evidence that in patients with NVAF experiencing an ischemic stroke while being treated with a DOAC, continuing treatment with that DOAC is more effective at preventing recurrent ischemic stroke than switching to a different DOAC or to warfarin.

INTEGRATE II: randomised phase III controlled trials of regorafenib containing regimens versus standard of care in refractory Advanced Gastro-Oesophageal Cancer (AGOC): a study by the Australasian Gastro-Intestinal Trials Group (AGITG).

BACKGROUND: Advanced gastro-oesophageal cancer (AGOC) carries a poor prognosis. No standard of care treatment options are available after first and second-line therapies. Regorafenib is an oral multi-targeted tyrosine kinase inhibitor targeting angiogenic, stromal, and oncogenic receptor tyrosine kinases. Regorafenib 160 mg daily prolonged progression free survival compared to placebo (INTEGRATE, phase 2). Regorafenib 80 mg daily in combination with nivolumab 3 mg/kg showed promising objective response rates (REGONIVO). METHODS/DESIGN: INTEGRATE II (INTEGRATE IIa and IIb) platform comprises two international phase III randomised controlled trials (RCT) with 2:1 randomisation in favor of experimental intervention. INTEGRATE IIa (double-blind) compares regorafenib 160 mg daily on days 1 to 21 of each 28-day cycle to placebo. INTEGRATE IIb (open label) compares REGONIVO, regorafenib 90 mg days 1 to 21 in combination with intravenous nivolumab 240 mg days 1 and 15 each 28-day cycle with investigator's choice of chemotherapy (control). Treatment continues until disease progression or intolerable adverse events as per protocol. Eligible participants include adults with AGOC who have failed two or more lines of treatment. Stratification is by location of tumour (INTEGRATE IIa only), geographic region, prior VEGF inhibitor and prior immunotherapy use (INTEGRATE IIb only). Primary endpoint is overall survival. Secondary endpoints are progression free survival, objective response rate, quality of life, and safety. Tertiary/correlative objectives include biomarker and pharmacokinetic evaluation. DISCUSSION: INTEGRATE II provides a platform to evaluate the clinical utility of regorafenib alone, as well as regorafenib in combination with nivolumab in treatment of participants with refractory AGOC. TRIAL REGISTRATION: INTEGRATE IIa prospectively registered 1 April 2016 Australia New Zealand Clinical Trial Registry: ACTRN12616000420448 (ClinicalTrials.gov NCT02773524). INTEGRATE IIb prospectively registered 10 May 2021 ClinicalTrials.gov: NCT04879368.

Thromboembolic Risks with Concurrent Direct Oral Anticoagulants and Antiseizure Medications: A Population-Based Analysis.

BACKGROUND AND OBJECTIVE: Drug-drug interactions between direct oral anticoagulants (DOAC) and antiseizure medications via the cytochrome P450 (CYP) or the P-glycoprotein (P-gp) systems may lead to under-anticoagulation. The clinical relevance of these interactions is unclear. We aimed to elucidate the risk of thromboembolism with concurrent DOAC and CYP/P-gp modulating antiseizure medications. METHODS: In this propensity score-weighted population-based retrospective cohort study, we used competing risk regression analyses to determine the risks of ischemic stroke, venous thromboembolism, and death in DOAC recipients taking CYP/P-gp-modulating antiseizure medications (phenytoin, valproate, levetiracetam, carbamazepine, or phenobarbital) versus those taking CYP/P-gp-neutral antiseizure medications (pregabalin, gabapentin, or clobazam). We also performed secondary analyses for the epilepsy and atrial fibrillation subgroups. RESULTS: Among DOAC users, CYP/P-gp-modulating antiseizure medications were collectively associated with an increased risk of ischemic stroke (adjusted hazard ratio 1.28, 95% confidence interval 1.05-1.57, p = 0.017). In addition, phenytoin (adjusted hazard ratio 1.34, 95% confidence interval 1.07-1.68, p = 0.011) and valproate (adjusted hazard ratio 1.38, 95% confidence interval 1.10-1.74, p = 0.006) were associated with increased mortality. In the epilepsy subgroup, the risk of ischemic stroke and venous thromboembolism did not differ between CYP/P-gp-modulating and CYP/P-gp-neutral antiseizure medications. CONCLUSIONS: Although CYP/P-gp-modulating antiseizure medications were associated with an increased risk of ischemic stroke when paired with DOAC in the primary analysis, such a phenomenon was not found among patients with epilepsy who took phenytoin, valproate, or levetiracetam with DOAC. Therefore, these antiseizure medication options among patients with epilepsy with concurrent DOAC should not be restricted solely based on their potential drug-drug interactions. Yet, the increased mortality during concurrent use of DOAC with phenytoin or valproate might call for caution.

VA-ECMO With Pericardiocentesis and Autologous Blood Transfusion - Stabilisation Strategy in Acute Cardiac Free Wall Rupture.

A 68-year old lady present with left ventricular free wall rupture and cardiac arrest post-myocardial infarction. This article illustrates a strategy combining pericardiocentesis with autologous transfusion together with VA-ECMO as a bridge to definitive surgical repair.

Effect of unfractionated heparin and low molecular weight heparin on the clotting of platelet-reduced whole blood: an in-vitro study utilizing thromboelastography.

Treatment of venous thromboembolism with concomitant thrombocytopenia is challenging. The platelet threshold for safe administration of anticoagulants is under debate, with minimum platelet count of 50 × 109/l being recommended as the safe cutoff. However, some evidence suggests administration of anticoagulants may still be safe at platelet levels of 30 × 109/l. Therefore, we developed an in-vitro thromboelastography (TEG) study to examine the effect of therapeutic or prophylactic levels of unfractionated heparin (UFH) and low molecular weight heparin (LMWH) on the clotting profile of platelet-reduced whole blood. Using magnetic bead-based antibody chromatography, platelets were removed to achieve platelet-depleted blood (<10 × 109/l of platelets). Platelet-depleted blood was then mixed with whole blood to produce blood samples with platelet counts of 30 × 109, 50 × 109 and 150 × 109/l. These blood samples were incubated with therapeutic or prophylactic levels of UFH or LMWH in disposable TEG cups. Clotting was initiated with 10 mmol/l calcium and optimized tissue factor levels for each anticoagulant used (2.25 pmol/l for UFH and 2.05 pmol/l for LMWH). Clotting was monitored by TEG at 37 °C for 180 min. The following TEG parameters were evaluated: R (time to clot), maximum amplitude (strength of clot) and area under the curve in 15 min (overall speed and strength of the clot at 15 min of clotting). No statistically significant differences were observed between platelet counts of 30 × 109 and 50 × 109/l for R, maximum amplitude or area under the curve in 15 min for most of the therapeutic and prophylactic doses of UFH and LMWH tested in this study. Use of anticoagulants compromised all of the TEG parameters relative to a normal platelet count of 150 × 109/l, in a dose dependent manner. The current study demonstrates that in-vitro clotting is impaired with use and increasing doses of anticoagulants. Despite this observation, we did not observe a significant difference in clotting between platelet levels of 30 × 109 and 50 × 109/l. Overall, this work provides further insight in the debated use of anticoagulants in patients with venous thromboembolism and concomitant thrombocytopenia, and provides support for possible use of anticoagulants at lower platelet thresholds.

Potential Risk Factors Contributing to Development of Venous Thromboembolism for Total Knee Replacements Patients Prophylaxed With Rivaroxaban: A Retrospective Case-Control Study.

Rivaroxaban after total knee arthroplasty (TKA) is used to prevent postoperative venous thromboembolism (VTE); however, despite thromboprophylaxis, some patients still develop postoperative VTE. To determine whether tourniquet time, time to initiate rivaroxaban (TTIRIV), or Body Mass Index (BMI) was associated with postoperative VTE. A retrospective case-control study was conducted. Those patients that developed VTE despite prophylaxis (cases) were compared to controls (no VTE). A univariate analysis was conducted (p < 0.05 statistically significant). Seven VTE cases were identified from 234 TKA-patients. Patients with and without VTE had BMI of 40.1 ± 9.1 and 32.8 ± 7.5, respectively (p = 0.064). TTIRIV in VTE and control group was 28.2 ± 4.7 hours and 26.4 ± 4.2 hours, respectively (p = 0.39). Mean tourniquet time in VTE and control group was 65.0 ± 8.7 minutes and 49 ± 8.8 minutes, respectively (p = 0.0007). Statistically significant differences in tourniquet times were noted between VTE and non-VTE group but not for TTIRIV and BMI. Prolonged tourniquet use could pose a potential risk factor for postoperative VTE. Thromboprophylaxis management may need to be adjusted, based on patient-specific factors that could include increasing doses of oral anticoagulants and/or mechanical prophylaxis. However, further large-scale studies are required to establish pathophysiology.

FGFR3 signaling and function in triple negative breast cancer.

BACKGROUND: Triple negative breast cancer (TNBC) accounts for 16% of breast cancers and represents an aggressive subtype that lacks targeted therapeutic options. In this study, mass spectrometry (MS)-based tyrosine phosphorylation profiling identified aberrant FGFR3 activation in a subset of TNBC cell lines. This kinase was therefore evaluated as a potential therapeutic target. METHODS: MS-based tyrosine phosphorylation profiling was undertaken across a panel of 24 TNBC cell lines. Immunoprecipitation and Western blot were used to further characterize FGFR3 phosphorylation. Indirect immunofluorescence and confocal microscopy were used to determine FGFR3 localization. The selective FGFR1-3 inhibitor, PD173074 and siRNA knockdowns were used to characterize the functional role of FGFR3 in vitro. The TCGA and Metabric breast cancer datasets were interrogated to identify FGFR3 alterations and how they relate to breast cancer subtype and overall patient survival. RESULTS: High FGFR3 expression and phosphorylation were detected in SUM185PE cells, which harbor a FGFR3-TACC3 gene fusion. Low FGFR3 phosphorylation was detected in CAL51, MFM-223 and MDA-MB-231 cells. In SUM185PE cells, the FGFR3-TACC3 fusion protein contributed the majority of phosphorylated FGFR3, and largely localized to the cytoplasm and plasma membrane, with staining at the mitotic spindle in a small subset of cells. Knockdown of the FGFR3-TACC3 fusion and wildtype FGFR3 in SUM185PE cells decreased FRS2, AKT and ERK phosphorylation, and induced cell death. Knockdown of wildtype FGFR3 resulted in only a trend for decreased proliferation. PD173074 significantly decreased FRS2, AKT and ERK activation, and reduced SUM185PE cell proliferation. Cyclin A and pRb were also decreased in the presence of PD173074, while cleaved PARP was increased, indicating cell cycle arrest in G1 phase and apoptosis. Knockdown of FGFR3 in CAL51, MFM-223 and MDA-MB-231 cells had no significant effect on cell proliferation. Interrogation of public datasets revealed that increased FGFR3 expression in breast cancer was significantly associated with reduced overall survival, and that potentially oncogenic FGFR3 alterations (eg mutation and amplification) occur in the TNBC/basal, luminal A and luminal B subtypes, but are rare. CONCLUSIONS: These results indicate that targeting FGFR3 may represent a therapeutic option for TNBC, but only for patients with oncogenic FGFR3 alterations, such as the FGFR3-TACC3 fusion. Video abstract.

The AGITG GAP Study: A Phase II Study of Perioperative Gemcitabine and Nab-Paclitaxel for Resectable Pancreas Cancer.

BACKGROUND: While combination therapy with nab-paclitaxel/gemcitabine (nab-gem) is effective in pancreatic ductal adenocarcinoma (PDAC), its efficacy as perioperative chemotherapy is unknown. The primary objective of this multicenter, prospective, single-arm, phase II study was to determine whether neoadjuvant therapy with nab-gem was associated with higher complete resection rates (R0) in resectable PDAC, while the secondary objectives were to determine the utility of radiological assessment of response to preoperative chemotherapy and the safety and efficacy of nab-gem as perioperative therapy. METHODS: Patients were recruited from eight Australian sites, and 42 patients with radiologically defined resectable PDAC and an Eastern Cooperative Oncology Group performance status of 0-2 were enrolled. Participants received two cycles of preoperative nab-paclitaxel 125 mg/m2 and gemcitabine 1000 mg/m2 on days 1, 8, and 15 (28-day cycle) presurgery, and four cycles postoperatively. Early response to chemotherapy was measured with fluorodeoxyglucose-positron emission tomography/computed tomography (FDG-PET/CT) scans on day 15. RESULTS: Preoperative nab-gem was completed by 93% of participants, but only 63% postoperatively. Thirty-six patients had surgery: 6 (17%) were unresectable, 15 (52%) had R0 (≥ 1 mm) resections, 14 (48%) had R1 (< 1 mm) resections, and 1 patient did not have PDAC. Median progression-free survival was 12.3 months and median overall survival (OS) was 23.5 months: R0 patients had an OS of 35 months versus 25.6 months for R1 patients after surgery. Seven patients had not progressed after 43 months. CONCLUSIONS: The GAP trial demonstrated that perioperative nab-gem was tolerable. Although the primary endpoint of an 85% R0 rate was not met, the R0 rate was similar to trials using a > 1 mm R0 resection definition, and survival rates were comparable with recent adjuvant studies.

Proteomic Profiling of Human Prostate Cancer-associated Fibroblasts (CAF) Reveals LOXL2-dependent Regulation of the Tumor Microenvironment.

In prostate cancer, cancer-associated fibroblasts (CAF) exhibit contrasting biological properties to non-malignant prostate fibroblasts (NPF) and promote tumorigenesis. Resolving intercellular signaling pathways between CAF and prostate tumor epithelium may offer novel opportunities for research translation. To this end, the proteome and phosphoproteome of four pairs of patient-matched CAF and NPF were characterized to identify discriminating proteomic signatures. Samples were analyzed by liquid chromatography-tandem mass spectrometry (LC-MS/MS) with a hyper reaction monitoring data-independent acquisition (HRM-DIA) workflow. Proteins that exhibited a significant increase in CAF versus NPF were enriched for the functional categories "cell adhesion" and the "extracellular matrix." The CAF phosphoproteome exhibited enhanced phosphorylation of proteins associated with the "spliceosome" and "actin binding." STRING analysis of the CAF proteome revealed a prominent interaction hub associated with collagen synthesis, modification, and signaling. It contained multiple collagens, including the fibrillar types COL1A1/2 and COL5A1; the receptor tyrosine kinase discoidin domain-containing receptor 2 (DDR2), a receptor for fibrillar collagens; and lysyl oxidase-like 2 (LOXL2), an enzyme that promotes collagen crosslinking. Increased activity and/or expression of LOXL2 and DDR2 in CAF were confirmed by enzymatic assays and Western blotting analyses. Pharmacological inhibition of CAF-derived LOXL2 perturbed extracellular matrix (ECM) organization and decreased CAF migration in a wound healing assay. Further, it significantly impaired the motility of co-cultured RWPE-2 prostate tumor epithelial cells. These results indicate that CAF-derived LOXL2 is an important mediator of intercellular communication within the prostate tumor microenvironment and is a potential therapeutic target.

Characterization of the Src-regulated kinome identifies SGK1 as a key mediator of Src-induced transformation.

Despite significant progress, our understanding of how specific oncogenes transform cells is still limited and likely underestimates the complexity of downstream signalling events. To address this gap, we use mass spectrometry-based chemical proteomics to characterize the global impact of an oncogene on the expressed kinome, and then functionally annotate the regulated kinases. As an example, we identify 63 protein kinases exhibiting altered expression and/or phosphorylation in Src-transformed mammary epithelial cells. An integrated siRNA screen identifies nine kinases, including SGK1, as being essential for Src-induced transformation. Accordingly, we find that Src positively regulates SGK1 expression in triple negative breast cancer cells, which exhibit a prominent signalling network governed by Src family kinases. Furthermore, combined inhibition of Src and SGK1 reduces colony formation and xenograft growth more effectively than either treatment alone. Therefore, this approach not only provides mechanistic insights into oncogenic transformation but also aids the design of improved therapeutic strategies.

Protocol for the P3BEP trial (ANZUP 1302): an international randomised phase 3 trial of accelerated versus standard BEP chemotherapy for adult and paediatric male and female patients with intermediate and poor-risk metastatic germ cell tumours.

BACKGROUND: Bleomycin, etoposide, and cisplatin (BEP) chemotherapy administered every 3 weeks for 4 cycles remains the standard first line treatment for patients with intermediate- and poor-risk metastatic germ cell tumours (GCTs). Administering standard chemotherapy 2-weekly rather than 3-weekly, so-called 'accelerating chemotherapy', has improved cure rates in other cancers. An Australian multicentre phase 2 trial demonstrated this regimen is feasible and tolerable with efficacy data that appears promising. The aim of this trial is to determine if accelerated BEP is superior to standard BEP as first line chemotherapy for adult and paediatric male and female participants with intermediate and poor risk metastatic GCTs. METHODS: This is an open label, randomised, stratified, 2-arm, international multicentre, 2 stage, phase 3 clinical trial. Participants are randomised 1:1 to receive accelerated BEP or standard BEP chemotherapy. Eligible male or female participants, aged between 11 and 45 years with intermediate or poor-risk metastatic GCTs for first line chemotherapy will be enrolled from Australia, the United Kingdom and the United States. Participants will have regular follow up for at least 5 years. The primary endpoint for stage 1 of the trial (n = 150) is complete response rate and for the entire trial (n = 500) is progression free survival. Secondary endpoints include response following treatment completion (by a protocol-specific response criteria), adverse events, health-related quality of life, treatment preference, delivered dose-intensity of chemotherapy (relative to standard BEP), overall survival and associations between biomarkers (to be specified) and their correlations with clinical outcomes. DISCUSSION: This is the first international randomised clinical trial for intermediate and poor-risk metastatic extra-cranial GCTs involving both adult and pediatric age groups open to both males and females. It is also the largest, current randomised trial for germ cell tumours in the world. Positive results for this affordable intervention could change the global standard of care for intermediate and poor risk germ cell tumours, improve cure rates, avoid the need for toxic and costly salvage treatment, and return young adults to long, healthy and productive lives. TRIAL REGISTRATION: ACTRN 12613000496718 on 3rd May 2013 and Clinicaltrials.gov NCT02582697 on 21st October 2015.

Identification of Novel Response and Predictive Biomarkers to Hsp90 Inhibitors Through Proteomic Profiling of Patient-derived Prostate Tumor Explants.

Inhibition of the heat shock protein 90 (Hsp90) chaperone is a promising therapeutic strategy to target expression of the androgen receptor (AR) and other oncogenic drivers in prostate cancer cells. However, identification of clinically-relevant responses and predictive biomarkers is essential to maximize efficacy and treatment personalization. Here, we combined mass spectrometry (MS)-based proteomic analyses with a unique patient-derived explant (PDE) model that retains the complex microenvironment of primary prostate tumors. Independent discovery and validation cohorts of PDEs (n = 16 and 30, respectively) were cultured in the absence or presence of Hsp90 inhibitors AUY922 or 17-AAG. PDEs were analyzed by LC-MS/MS with a hyper-reaction monitoring data independent acquisition (HRM-DIA) workflow, and differentially expressed proteins identified using repeated measure analysis of variance (ANOVA; raw p value <0.01). Using gene set enrichment, we found striking conservation of the most significantly AUY922-altered gene pathways between the discovery and validation cohorts, indicating that our experimental and analysis workflows were robust. Eight proteins were selectively altered across both cohorts by the most potent inhibitor, AUY922, including TIMP1, SERPINA3 and CYP51A (adjusted p < 0.01). The AUY922-mediated decrease in secretory TIMP1 was validated by ELISA of the PDE culture medium. We next exploited the heterogeneous response of PDEs to 17-AAG in order to detect predictive biomarkers of response and identified PCBP3 as a marker with increased expression in PDEs that had no response or increased in proliferation. Also, 17-AAG treatment led to increased expression of DNAJA1 in PDEs that exhibited a cytostatic response, revealing potential drug resistance mechanisms. This selective regulation of DNAJA1 was validated by Western blot analysis. Our study establishes "proof-of-principle" that proteomic profiling of drug-treated PDEs represents an effective and clinically-relevant strategy for identification of biomarkers that associate with certain tumor-specific responses.

Structure of SgK223 pseudokinase reveals novel mechanisms of homotypic and heterotypic association.

The mammalian pseudokinase SgK223, and its structurally related homologue SgK269, are oncogenic scaffolds that nucleate the assembly of specific signalling complexes and regulate tyrosine kinase signalling. Both SgK223 and SgK269 form homo- and hetero-oligomers, a mechanism that underpins a diversity of signalling outputs. However, mechanistic insights into SgK223 and SgK269 homo- and heterotypic association are lacking. Here we present the crystal structure of SgK223 pseudokinase domain and its adjacent N- and C-terminal helices. The structure reveals how the N- and C-regulatory helices engage in a novel fold to mediate the assembly of a high-affinity dimer. In addition, we identified regulatory interfaces on the pseudokinase domain required for the self-assembly of large open-ended oligomers. This study highlights the diversity in how the kinase fold mediates non-catalytic functions and provides mechanistic insights into how the assembly of these two oncogenic scaffolds is achieved in order to regulate signalling output.

Phosphoproteomic Profiling Reveals ALK and MET as Novel Actionable Targets across Synovial Sarcoma Subtypes.

Despite intensive multimodal treatment of sarcomas, a heterogeneous group of malignant tumors arising from connective tissue, survival remains poor. Candidate-based targeted treatments have demonstrated limited clinical success, urging an unbiased and comprehensive analysis of oncogenic signaling networks to reveal therapeutic targets and personalized treatment strategies. Here we applied mass spectrometry-based phosphoproteomic profiling to the largest and most heterogeneous set of sarcoma cell lines characterized to date and identified novel tyrosine phosphorylation patterns, enhanced tyrosine kinases in specific subtypes, and potential driver kinases. ALK was identified as a novel driver in the Aska-SS synovial sarcoma (SS) cell line via expression of an ALK variant with a large extracellular domain deletion (ALKΔ2-17). Functional ALK dependency was confirmed in vitro and in vivo with selective inhibitors. Importantly, ALK immunopositivity was detected in 6 of 43 (14%) of SS patient specimens, one of which exhibited an ALK rearrangement. High PDGFRα phosphorylation also characterized SS cell lines, which was accompanied by enhanced MET activation in Yamato-SS cells. Although Yamato-SS cells were sensitive to crizotinib (ALK/MET-inhibitor) but not pazopanib (VEGFR/PDGFR-inhibitor) monotherapy in vitro, synergistic effects were observed upon drug combination. In vivo, both drugs were individually effective, with pazopanib efficacy likely attributable to reduced angiogenesis. MET or PDGFRα expression was detected in 58% and 84% of SS patients, respectively, with coexpression in 56%. Consequently, our integrated approach has led to the identification of ALK and MET as promising therapeutic targets in SS. Cancer Res; 77(16); 4279-92. ©2017 AACR.

In-vitro assessment of the effect of dabigatran on thrombosis of adult and neonatal plasma: comparisons using thromboelastography and microscopic visualization of fibrin clot structure.

: Thromboelastography (TEG) is a global assay used for evaluating features of clot formation in vitro. Dabigatran is a reversible direct inhibitor of thrombin that has not been studied in neonates using a sophisticated global assay, such as TEG. Neonatal hemostasis differs from adult hemostasis in both quantitative and qualitative characteristics. Our aim was to compare the TEG clotting profile of neonatal and adult platelet-poor plasma when exposed to different concentrations of dabigatran. We used commercially collected adult pooled plasma and neonatal cord blood collected from placentas of healthy full term newborns. Platelet-poor plasma was isolated, pooled, and frozen. Prior to experiment, plasma was thawed and filtered. A reaction mixture of CaCl2, corn trypsin inhibitor, tissue factor, and dabigatran in imidazole buffer was mixed with plasma in a TEG cup. Time to clot initiation (R-time), speed of clot strengthening (α-angle), and maximum clot strength (maximal amplitude) were measured. Scanning electron microscopy was performed to evaluate fibrin clot structure. Without dabigatran, there was no significant difference in TEG measurements between neonatal and adult samples. However, neonatal plasma clotting with dabigatran had slower onset, slower speed, and weaker clots that were more porous with thicker fibers, compared with adult plasma clotting. Thus, neonatal plasma may be more sensitive to dabigatran as assessed by our in-vitro TEG study.