Update on Venous Thromboembolism in Orthopaedic Trauma Surgery.

Venous thromboembolism (VTE) is a notable contributor to the morbidity and mortality of patients with orthopaedic trauma. Several associations have published guidelines on VTE prophylaxis, with a strong predilection toward low-molecular-weight heparin for chemoprophylaxis. However, previous recommendations may be revisited because recent high-level evidence demonstrated aspirin to be noninferior to low-molecular-weight heparin in preventing serious complications of VTE. Direct oral anticoagulants are emerging as safe alternatives to injectable medication and may offer improved patient compliance. There is no consensus on postinjury duration of chemoprophylaxis as an outpatient. Mechanical prophylaxis remains an important adjunctive VTE preventive measure for inpatients with pelvis and extremity fractures. Proteomics and novel laboratory measurements may be able to predict high-risk patients and detect early thrombus formation. Despite high-quality prospective research over the past decade, we still have much to learn about patient- and injury-specific risk factors.

Genomic profiling identifies somatic mutations predicting thromboembolic risk in patients with solid tumors.

Venous thromboembolism (VTE) associated with cancer (CAT) is a well-described complication of cancer and a leading cause of death in patients with cancer. The purpose of this study was to assess potential associations of molecular signatures with CAT, including tumor-specific mutations and the presence of clonal hematopoiesis. We analyzed deep-coverage targeted DNA-sequencing data of >14 000 solid tumor samples using the Memorial Sloan Kettering-Integrated Mutation Profiling of Actionable Cancer Targets platform to identify somatic alterations associated with VTE. End point was defined as the first instance of cancer-associated pulmonary embolism and/or proximal/distal lower extremity deep vein thrombosis. Cause-specific Cox proportional hazards regression was used, adjusting for pertinent clinical covariates. Of 11 695 evaluable individuals, 72% had metastatic disease at time of analysis. Tumor-specific mutations in KRAS (hazard ratio [HR], 1.34; 95% confidence interval (CI), 1.09-1.64; adjusted P = .08), STK11 (HR, 2.12; 95% CI, 1.55-2.89; adjusted P < .001), KEAP1 (HR, 1.84; 95% CI, 1.21-2.79; adjusted P = .07), CTNNB1 (HR, 1.73; 95% CI, 1.15-2.60; adjusted P = .09), CDKN2B (HR, 1.45; 95% CI, 1.13-1.85; adjusted P = .07), and MET (HR, 1.83; 95% CI, 1.15-2.92; adjusted P = .09) were associated with a significantly increased risk of CAT independent of tumor type. Mutations in SETD2 were associated with a decreased risk of CAT (HR, 0.35; 95% CI, 0.16-0.79; adjusted P = .09). The presence of clonal hematopoiesis was not associated with an increased VTE rate. This is the first large-scale analysis to elucidate tumor-specific genomic events associated with CAT. Somatic tumor mutations of STK11, KRAS, CTNNB1, KEAP1, CDKN2B, and MET were associated with an increased risk of VTE in patients with solid tumors. Further analysis is needed to validate these findings and identify additional molecular signatures unique to individual tumor types.

Rivaroxaban treatment of cancer-associated venous thromboembolism: Memorial Sloan Kettering Cancer Center institutional experience.

BACKGROUND: Low-molecular-weight heparin has been the preferred treatment of cancer-associated thrombosis (CAT); however, emerging data support the use of direct oral anticoagulants (DOACs). OBJECTIVES: The Memorial Sloan Kettering Cancer Center Clinical Pathway has served as the institutional guideline for the use of rivaroxaban to treat CAT since 2014. Key elements are to recommend against use of a DOAC in patients with active gastrointestinal (GI) or genitourinary tract lesions, and a prespecified dose reduction in the elderly (75+ years old). We present our institutional experience for treatment of CAT. METHODS: From January 2014 through September 2016, 1072 patients began rivaroxaban treatment for CAT; 91.9% had a solid tumor, 8.1% had hematologic malignancies, and 75% of patients with solid tumors had metastatic disease. All patients with CAT treated with rivaroxaban were included in this analysis, regardless of adherence to the Clinical Pathway. RESULTS: The 6-month cumulative incidence of recurrent venous thromboembolism and major bleeding were 4.2% (95% confidence interval [CI], 2.7%-5.7%) and 2.2% (95% CI, 1.1%-3.2%), respectively. The incidence of clinically relevant non-major bleeding leading to discontinuation of rivaroxaban for at least 7 days was 5.5% (95% CI,  3.7%-7.1%), and 73.3% of major bleeds occurred in the GI tract. The 6-month cumulative mortality rate was 22.2% (95% CI, 19.4%-24.9%). The elderly had similar rates of recurrent thrombosis and bleeding as those aged under 75 years. CONCLUSION: Our institutional experience suggests that in appropriately selected patients, rivaroxaban may be used for treatment of CAT with promising safety and efficacy.

Using the American Thyroid Association Risk-Stratification System to Refine and Individualize the American Joint Committee on Cancer Eighth Edition Disease-Specific Survival Estimates in Differentiated Thyroid Cancer.

BACKGROUND: The eighth edition of the American Joint Committee on Cancer (AJCC) staging system has reclassified up to one third of differentiated thyroid cancer patients into one of the younger prognostic stage groups (<55 years of age at diagnosis, stage I or stage II). This reclassification widens the spectrum of disease in these lower stages without significantly impacting overall disease-specific survival (DSS) for the entire stage group. However, the optimistic DSS estimates in the <55-year-old stage groups may not accurately reflect the prognosis of individual patients with American Thyroid Association (ATA) high-risk features. Therefore, the aim of this study was to integrate the ATA risk classification system into the eighth edition AJCC staging system to refine and individualize DSS estimates for differentiated thyroid cancer patients aged <55 years at diagnosis. METHODS: Using the Memorial Sloan Kettering Cancer Center tumor registry, 4881 adult DTC patients aged <55 years at diagnosis receiving initial therapy between 1980 and 2016 were retrospectively analyzed. Using Memorial Sloan Kettering Cancer Center registry coded data, all patients were assigned an eighth edition AJCC stage (I or II), ATA risk of recurrence (low, intermediate, or high), and age group at diagnosis (younger patients defined as ≤45 years old, older patients defined as 45-55 years old). The primary outcome was 10-year DSS. RESULTS: A total of 122 (2.5%) disease-related deaths were observed in the cohort of 4881 patients during a median follow-up of 6.6 years. Integration of the AJCC stage, ATA risk, and age groups identified six subgroups with differing outcomes: (i) stage I/ATA low risk, younger and older, 100% DSS; (ii) stage I/ATA intermediate risk, younger and older, 98% DSS; (iii) stage I/ATA high risk, younger, 95% DSS; (iv) stage I/ATA high risk, older, 89% DSS; (v) stage II/ATA high risk, younger, 78% DSS; and (vi) stage II/ATA high risk, older, 61% DSS. CONCLUSIONS: Integration of AJCC stage, ATA risk, and age group (i) identifies six subgroups of patients with progressively worse DSS as AJCC stage, ATA risk, and age increases, and (ii) provides a more individualized estimate of DSS, especially in ATA high-risk patients.

Antidepressants appear safe in patients with carcinoid tumor: Results of a retrospective review.

INTRODUCTION: Patients living with neuroendocrine tumors have high rates of depression, often necessitating antidepressants, including selective serotonin reuptake inhibitors (SSRI). Neuroendocrine tumors (NETs) secrete vasoactive substances, including serotonin, which contribute to the cluster of symptoms known as carcinoid syndrome (flushing and diarrhea). Controversy exists over whether or not antidepressants are safe in NET. We aimed to study the safety of antidepressant use in NET patients. METHODS: We conducted a retrospective chart review of patients with well differentiated NET who were also prescribed antidepressants from January 2008 through April 2015. The study took place at Memorial Sloan Kettering Cancer Center and was approved by the hospital's institutional review board. RESULTS: Ninety-two patients were included. There were 16 (17.4%) patients with carcinoid syndrome (10 ileum, 1 duodenum, 1 jejunum and 4 unknown primary); and 76 (82.6%) patients without (41 lung, 9 pancreas, 8 ileal, 5 duodenum, 5 appendix, 2 unknown primary, 1 jejunum and 5 other). Median duration of antidepressant prescription was 11.6 months (range, 0-121) among those with carcinoid syndrome (N = 16) and 14.3 months (range, 0-172) among those without carcinoid syndrome (n = 76). Antidepressants were stopped in 31 cases (33.7%), though the reason was not specified in the majority of cases (n = 18; 58%). None of the patients developed carcinoid syndrome while being prescribed antidepressants. No patients developed carcinoid crisis. CONCLUSION: Our findings do not support previous authors' recommendations that SSRIs must be avoided in NET patients. Several classes of antidepressants appeared safe in NET patients with and without carcinoid syndrome.

The Tri-Institutional Pain Registry-Analysis of Outpatient Pain Management at a Specialized Cancer Center.

Objectives: The Outpatient Pain Clinics at Memorial Sloan Kettering Cancer Center participated in developing a pain registry to gain insight on the referral and management of cancer pain as related to demographic information, cancer history, prescription records, and interventional pain procedures stored in the institutional database. Methods: Five cohorts (subsets of one another) were defined and compared to describe demographics and differences in management and outcomes by age, race, sex, and cancer type. Clinic patients were compared with the entire institution to determine factors associated with better pain relief and reduced side effects. Results: A small percentage were referred to a pain specialist. A total of 1,043 patients completed 3,544 surveys. Compared with the institution, there were higher proportions of patients age 51 to 60 years, nonwhites, and patients with thoracic, abdominal, and head and neck cancers. Medical management controlled pain with three drug categories in 40% of visits. Short-acting opioids were the only category that statistically provided good pain relief with fewer side effects. Pain scores were improved with increasing opioid dose. Management differed by sex, age, and race; women consistently had lower doses of opioids, poorer pain control, more side effects, and were prescribed a greater variety of medications. Conclusions: A limited set of medications was required to manage most patients in the clinic, supporting the continued place of opioids and the World Health Organization analgesic ladder in managing cancer pain. Women may need a more nuanced approach for obtaining the best balance of pain relief and side effects.

Rivaroxaban for Stroke Prevention in Patients With Nonvalvular Atrial Fibrillation and Active Cancer.

Rivaroxaban is broadly used for the primary prevention of stroke and systemic embolism in the general population with nonvalvular atrial fibrillation (AF). However, there is little published evidence on the safety and efficacy of rivaroxaban for AF in patients with active cancer. The aim of this study was to assess the safety and efficacy of rivaroxaban in patients with active cancer and AF. The use of rivaroxaban in patients with cancer at the Memorial Sloan Kettering Cancer Center is monitored in the setting of a Quality Assessment Initiative. Patients with active cancer and AF, treated with rivaroxaban from January 1, 2014, to March 31, 2016, are included in this analysis. Clinical end points were defined a priori and assessed through text searches of medical records. A total of 163 evaluable patients were identified. After adjusting for competing risks, the estimated 1-year cumulative incidence of ischemic stroke was 1.4% (95% CI 0% to 3.4%) and major bleeding was 1.2% (95% CI 0% to 2.9%). The risk of clinically relevant nonmajor bleeding leading to discontinuation of anticoagulation at 1 year was 14.0% (95% CI 4.2% to 22.7%). The cumulative incidence of mortality was 22.6% (95% CI 12.2% to 31.7%) at 1 year, reflecting an active cancer population. One patient died after developing an acute ischemic cerebrovascular insult. In conclusion, the safety and efficacy of rivaroxaban treatment for nonvalvular AF in patients with active cancer is comparable to the results of the Rivaroxaban Once Daily Oral Direct Factor Xa Inhibition Compared with Vitamin K Antagonism for Prevention of Stroke and Embolism Trial in Atrial Fibrillation (ROCKET-AF) study in the general population.

18F-FDG-PET/CT for systemic staging of patients with newly diagnosed ER-positive and HER2-positive breast cancer.

OBJECTIVES: This study assesses 18F-FDG-PET/CT for patients with newly diagnosed estrogen receptor-positive/human epidermal growth factor receptor-negative (ER+/HER2-) and human epidermal growth factor receptor-positive (HER2+) breast cancer. METHODS: In this Institutional Review Board-approved retrospective study, our Healthcare Information System was screened for patients with ER+/HER2- and HER2+ breast cancer who underwent 18F-FDG-PET/CT prior to systemic or radiation therapy. The initial stage was determined from mammography, ultrasound, magnetic resonance imaging, and/or surgery.18F-FDG-PET/CT was evaluated to identify unsuspected extra-axillary regional nodal and distant metastases. The proportion of patients upstaged overall and stratified by stage and receptor phenotypes was calculated along with confidence intervals (CI). RESULTS: A total of 238 patients with ER+/HER2- and 245 patients with HER2+ who met inclusion criteria were evaluated. For patients with ER+/HER2-breast cancer, 18F-FDG-PET/CT revealed unsuspected distant metastases in 3/71 (4%) initial stage IIA, 13/95 (14%) stage IIB, and 15/57 (26%) stage III. For patients with HER2+ breast cancer, 18F-FDG-PET/CT revealed unsuspected distant metastases in 3/72 (4%) initial stage IIA, 13/93 (14%) stage IIB, and 13/59 (22%) stage III. The overall upstaging rate for IIB was 14% (95% confidence interval (CI): 9-20%). CONCLUSIONS: 18F-FDG-PET/CT revealed distant metastases in 14% (95% CI: 9-20%) of patients with stage IIB ER+/HER2- and HER2+ breast cancer, which is similar to upstaging rates previously seen in patients with stage IIB triple-negative breast cancer (15%, 95% CI: 9-24%). The detection of unsuspected distant metastases in these patients alters treatment and prognosis. NCCN guidelines should consider adding patients with stage IIB breast cancer for consideration of systemic staging with 18F-FDG-PET/CT at the time of initial diagnosis.

Enoxaparin dose reduction for thrombocytopenia in patients with cancer: a quality assessment study.

The development of thrombocytopenia in the setting of therapeutic anticoagulation for venous thromboembolic disease (VTE) is common in cancer patients, but guidelines for management are based on limited past data and have not been validated. In 2011, Memorial Sloan Kettering Cancer Center (MSKCC) implemented the following guidelines in this setting: administer full dose enoxaparin for a platelet count > 50,000/mcL, half-dose enoxaparin for a platelet count of 25,000-50,000/mcL, and hold anticoagulation for a platelet count < 25,000/mcL. We now report validation of safety and efficacy of these guidelines. As a Quality Assessment Initiative, we evaluated our guidelines for adult cancer patients at MSKCC who were on therapeutic-dose enoxaparin for VTE during the years 2011 through 2013 and experienced at least one 7-day period of thrombocytopenia (platelet count ≤ 50,000/mcL). We assessed adherence to the enoxaparin dose modification guidelines, major bleeding, clinically relevant non-major bleeding, recurrent VTE, and mortality during the thrombocytopenic episodes. We identified 99 patients with 140 episodes of thrombocytopenia of 7 or more days. The median duration of these thrombocytopenic episodes was 12 days. The enoxaparin dose was modified in 133 of the 140 episodes (95%), reflecting satisfactory adherence to our institutional guidelines. There were no recurrent VTE events or major bleeding episodes when the anticoagulant dose was reduced or held. In this cohort, there was only one major bleeding episode, a trauma-associated retroperitoneal hemorrhage that occurred on the third day of a thrombocytopenic episode, prior to enoxaparin dose modification. There were 13 clinically relevant non-major bleeding episodes. Lastly, 10 patients died of cancer-related causes during an episode of thrombocytopenia. This Quality Assessment Initiative supports the safety and efficacy of our guidelines for therapeutic enoxaparin dose modification.

High rate of thiamine deficiency among inpatients with cancer referred for psychiatric consultation: results of a single site prevalence study.

OBJECTIVE: Thiamine deficiency (TD) is increasingly recognized in medically ill patients. The prevalence of TD among cancer patients is unknown. This study aims to characterize the prevalence of TD among inpatients with cancer. METHODS: Retrospective chart review of patients admitted to a large cancer center who were referred for psychiatric consultation and whose serum thiamine concentration was measured. Patients with alcohol use were excluded. RESULTS: Among 217 patients with various cancer types, TD was found in 55.3%. Risk factors included fluorouracil-based chemotherapy, significant weight loss, and undergoing active cancer treatment. Almost all patients were normal weight, overweight, or obese, and few had concomitant vitamin B12 or folate deficiency. A total of 17.5% were receiving multivitamin supplementation. Nearly half (49.8%) did not receive empiric treatment with thiamine and among those who did, treatment delay occurred in the majority of cases (59.6%). Measurement of serum thiamine concentration preceded psychiatric consultation in only 10.6% of cases. CONCLUSIONS: Our findings suggest that TD is highly prevalent among inpatients with cancer, even among normal and overweight individuals, in the absence of other vitamin deficiencies, and while receiving multivitamin supplements. Several potential risk factors were identified, including active cancer treatment. Evaluation of TD was most commonly not initiated by oncologists. Failure to treat and treatment delay were common. Given these findings, oncologists must be vigilant about detecting TD among inpatients with cancer. Copyright © 2016 John Wiley & Sons, Ltd.

Safe and effective use of rivaroxaban for treatment of cancer-associated venous thromboembolic disease: a prospective cohort study.

Low-molecular weight heparin (LMWH) has been the standard of care for treatment of venous thromboembolism (VTE) in patients with cancer. Rivaroxaban was approved in 2012 for the treatment of pulmonary embolism (PE) and deep vein thrombosis (DVT), but no prior studies have been reported specifically evaluating the efficacy and safety of rivaroxaban for cancer-associated thrombosis (CAT). Under a Quality Assessment Initiative (QAI), we established a Clinical Pathway to guide rivaroxaban use for CAT and now report a validation analysis of our first 200 patients. A 200 patient cohort with CAT (PE or symptomatic, proximal DVT), whose full course of anticoagulation was with rivaroxaban, were accrued. In competing risk analysis, primary endpoints at 6 months included new or recurrent PE or symptomatic proximal lower extremity DVT, major bleeding, clinically-relevant non-major bleeding leading to discontinuation of rivaroxaban, or death. In competing risk analysis, the 6 months cumulative incidence of new or recurrent VTE was 4.4 % (95 % CI = 1.4-7.4 %), major bleeding was 2.2 % (95 % CI = 0-4.2 %) and all-cause mortality 17.6 % (95 % CI = 11.7-23.0 %). In this cohort of 200 patients with active cancer and CAT the rates of new or recurrent VTE and major bleeding were comparable to the cancer subgroup analysis from the EINSTEIN studies. The results of our Clinical Pathway provide guidance on Rivaroxaban use for treatment of CAT, and suggest that safety and efficacy is preserved, compared with past-published experience with LMWH.

Acute liver effects, disposition and metabolic fate of [14C]-fenclozic acid following oral administration to normal and bile-cannulated male C57BL/6J mice.

The distribution, metabolism, excretion and hepatic effects of the human hepatotoxin fenclozic acid were investigated following single oral doses of 10 mg/kg to normal and bile duct-cannulated male C57BL/6J mice. Whole body autoradiography showed distribution into all tissues except the brain, with radioactivity still detectable in blood, kidney and liver at 72 h post-dose. Mice dosed with [14C]-fenclozic acid showed acute centrilobular hepatocellular necrosis, but no other regions of the liver were affected. The majority of the [14C]-fenclozic acid-related material recovered was found in the urine/aqueous cage wash, (49%) whilst a smaller portion (13%) was eliminated via the faeces. Metabolic profiles for urine, bile and faecal extracts, obtained using liquid chromatography and a combination of mass spectrometric and radioactivity detection, revealed extensive metabolism of fenclozic acid in mice that involved biotransformations via both oxidation and conjugation. These profiling studies also revealed the presence of glutathione-derived metabolites providing evidence for the production of reactive species by mice administered fenclozic acid. Covalent binding to proteins from liver, kidney and plasma was also demonstrated, although this binding was relatively low (less than 50 pmol eq./mg protein).

(18)F-FDG-PET/CT for systemic staging of newly diagnosed triple-negative breast cancer.

PURPOSE: National Comprehensive Cancer Network guidelines recommend (18)F-FDG-PET/CT, in addition to standard staging procedures, for systemic staging of newly diagnosed stage III breast cancer patients. However, factors in addition to stage may influence PET/CT utility. As breast cancers that are negative for estrogen receptor, progesterone receptor, and human epidermal growth factor receptor (triple-negative breast cancer, or TNBC) are more aggressive and metastasize earlier than other breast cancers, we hypothesized that receptor expression may be one such factor. This study assesses (18)F-FDG-PET/CT for systemic staging of newly diagnosed TNBC. METHODS: In this Institutional Review Board-approved retrospective study, our Healthcare Information System was screened for patients with TNBC who underwent (18)F-FDG-PET/CT in 2007-2013 prior to systemic or radiation therapy. Initial stage was determined from mammography, ultrasound, magnetic resonance imaging, and/or surgery, if performed prior to (18)F-FDG-PET/CT. (18)F-FDG-PET/CT was evaluated to identify unsuspected extra-axillary regional nodal and distant metastases, as well as unsuspected synchronous malignancies. Kaplan Meier survival estimates were calculated for initial stage IIB patients stratified by whether or not stage 4 disease was detected by (18)F-FDG-PET/CT. RESULTS: A total of 232 patients with TNBC met inclusion criteria. (18)F-FDG-PET/CT revealed unsuspected distant metastases in 30 (13 %): 0/23 initial stage I, 4/82 (5 %) stage IIA, 13/87 (15 %) stage IIB, 4/23 (17 %) stage IIIA, 8/14 (57 %) stage IIIB, and 1/3 (33 %) stage IIIC. Twenty-six of 30 patients upstaged to IV by (18)F-FDG-PET/CT were confirmed by pathology, with the remaining four patients confirmed by follow-up imaging. In addition, seven unsuspected synchronous malignancies were identified in six patients. Initial stage 2B patients who were upstaged to 4 by (18)F-FDG-PET/CT had significantly shorter survival compared to initial stage 2B patients who were not (3-year Kaplan Meier estimate 0.33, 95 % CI 0.13-0.55 versus 0.97, CI 0.76-0.93, p < .0001). CONCLUSION: F-FDG-PET/CT revealed distant metastases in 15 % of patients with stage IIB TNBC. Stage IIB patients upstaged to 4 by (18)F-FDG-PET/CT had significantly shorter survival than those who were not, consistent with (18)F-FDG-PET/CT detecting an increased burden of disease. This study provides further evidence that populations of patients with stage IIB breast cancer, such as TNBC, should be considered for systemic staging with (18)F-FDG-PET/CT at the time of initial diagnosis.

Automated eligibility screening and monitoring for genotype-driven precision oncology trials.

The Information Systems Department at Memorial Sloan Kettering Cancer Center developed the DARWIN Cohort Management System (DCMS). The DCMS identifies and tracks cohorts of patients based on genotypic and clinical data. It assists researchers and treating physicians in enrolling patients to genotype-matched IRB-approved clinical trials. The DCMS sends automated, actionable, and secure email notifications to users with information about eligible or enrolled patients before their upcoming appointments. The system also captures investigators input via annotations on patient eligibility and preferences on future status updates. As of August 2015, the DCMS is tracking 159,893 patients on both clinical operations and research cohorts. 134 research cohorts have been established and track 64,473 patients. 51,192 of these have had one or more genomic tests including MSK-IMPACT, comprising the pool eligible for genotype-matched studies. This paper describes the design and evolution of this Informatics solution.

α-Synuclein-mediated inhibition of ATF6 processing into COPII vesicles disrupts UPR signaling in Parkinson's disease.

The unfolded protein response (UPR) monitors the folding environment within the endoplasmic reticulum (ER). Accumulation of misfolded proteins within the ER activates the UPR resulting in the execution of adaptive or non-adaptive signaling pathways. α-Synuclein (α-syn) whose accumulation and aggregation define the pathobiology of Parkinson's disease (PD) has been shown to inhibit ER-Golgi transit of COPII vesicles. ATF6, a protective branch of the UPR, is processed via COPII mediated ER-Golgi transit following its activation via ER stress. Using cellular PD models together with biochemical reconstitution assays, we showed that α-syn inhibited processing of ATF6 directly through physical interactions and indirectly through restricted incorporation into COPII vesicles. Impaired ATF6 signaling was accompanied by decreased ER-associated degradation (ERAD) function and increased pro-apoptotic signaling. The mechanism by which α-syn inhibits ATF6 signaling expands our understanding of the role ER stress and the UPR play in neurodegenerative diseases such as PD.

The metabolic fate of [14C]-fenclozic acid in the hepatic reductase null (HRN) mouse.

1. The distribution, metabolism, excretion and hepatic effects of fenclozic acid were investigated following a single oral dose of 10 mg/kg to hepatic reductase null (HRN) mice. 2. The majority of the [(14)C]-fenclozic acid was eliminated via the urine/aqueous cage wash, (55%) with a smaller portion excreted in the faeces, (5%). The total recovery of radioactivity in the excreta over the 72 h period studied was ca. 60%. 3. Metabolism of fenclozic acid in the HRN mice was entirely to the carboxylic acid function and was dominated by amino acid conjugation to glycine and taurine, with lesser amounts of an acyl glucuronide. 4. Whole body autoradiography of mice showed general distribution into all tissues except the brain. Radioactivity was still detectable in the kidney and liver of the HRN mice at 72 h post-dose. Covalent binding studies showed evidence of binding to kidney, liver and plasma proteins however, the degree of binding was less than 50 pmol equiv/mg protein for all tissues. 5. The HRN mouse appears to be a useful in vivo model for the study of the Phase II conjugation metabolism of fenclozic acid in the absence of hepatic cytochrome P450-related oxidative metabolism.

Identification of the sites of tau hyperphosphorylation and activation of tau kinases in synucleinopathies and Alzheimer's diseases.

OBJECTIVE: Most neurodegenerative diseases contain hyperphosphorylated Tau [p-Tau]. We examined for the first time epitopes at which Tau is hyperphosphorylated in Parkinson's disease, dementia with Lewy bodies and Alzheimer's disease, and also select Tau kinases. METHODS: Postmortem frontal cortex from Parkinson's disease, dementia with Lewy bodies, Alzheimer's disease and striata from Parkinson's disease, were analyzed by immunoblots using commercially available antibodies against 20 different phospho-epitopes of Tau. Major Tau kinases were also screened. Results in diseased tissues were compared to nondiseased controls. RESULTS: In Alzheimer's disease, Tau was hyperphosphorylated at all the 20 epitopes of p-Tau. In dementia with Lewy bodies, p-Tau formation occurred at 6 sites sharing 30% overlap with Alzheimer's disease, while in Parkinson's frontal cortex, an area which does not degenerate, Tau hyperphosphorylation was seen at just 3 epitopes, indicating 15% overlap with Alzheimer's disease. In Parkinson's disease striatum, an area which undergoes considerable neurodegeneration, Tau was hyperphosphorylated at 10 epitopes, sharing 50% overlap with Alzheimer's disease. Between frontal cortex of Parkinson's disease and dementia with Lewy bodies, there were only two p-Tau epitopes in common. In striata of Parkinson's disease, there were 3 clusters of Tau hyperphosphorylated at 3 contiguous sites, while two such clusters were detected in dementia with Lewy bodies; such clusters disrupt axonal transport of mitochondria, cause microtubule remodeling and result in cell death. p-GSK-3ß, a major Tau kinase, was activated in all brain regions examined, except in dementia with Lewy bodies. Activation of other Tau kinases was seen in all brain regions, with no clear pattern of activation. INTERPRETATION: Our studies suggest that the three neurodegenerative diseases each have a signature-specific profile of p-Tau formation which may be useful in understanding the genesis of the diseases and for the development of a panel of specific biomarkers.

Paraquat, but not maneb, induces synucleinopathy and tauopathy in striata of mice through inhibition of proteasomal and autophagic pathways.

SNCA and MAPT genes and environmental factors are important risk factors of Parkinson's disease [PD], the second-most common neurodegenerative disease. The agrichemicals maneb and paraquat selectively target dopaminergic neurons, leading to parkinsonism, through ill-defined mechanisms. In the current studies we have analyzed the ability of maneb and paraquat, separately and together, to induce synucleinopathy and tauopathy in wild type mice. Maneb was ineffective in increasing α-synuclein [α-Syn] or p-Tau levels. By contrast, paraquat treatment of mice resulted in robust accumulation of α-Syn and hyperphosphorylation of Tau in striata, through activation of p-GSK-3ß, a major Tau kinase. Co-treatment with maneb did not enhance the effects of paraquat. Increased hyperacetylation of α-tubulin was observed in paraquat-treated mice, suggesting cytoskeleton remodeling. Paraquat, but not maneb, inhibited soluble proteasomal activity on a peptide substrate but this was not associated with a decreased expression of 26S proteasome subunits. Both paraquat and maneb treatments increased levels of the autophagy inhibitor, mammalian target of rapamycin, mTOR, suggesting impaired axonal autophagy, despite increases in certain autophagic proteins, such as beclin 1 and Agt12. Autophagic flux was also impaired, as ratios of LC3 II to LC3 I were reduced in treated animals. Increased mTOR was also observed in postmortem human PD striata, where there was a reduction in the LC3 II to LC3 I ratio. Heat shock proteins were either increased or unchanged upon paraquat-treatment suggesting that chaperone-mediated autophagy is not hampered by the agrichemicals. These studies provide novel insight into the mechanisms of action of these agrichemicals, which indicate that paraquat is much more toxic than maneb, via its inhibitory effects on proteasomes and autophagy, which lead to accumulation of α-Syn and p-Tau.