Durable tumor regression in genetically altered malignant rhabdoid tumors by inhibition of methyltransferase EZH2.

Inactivation of the switch/sucrose nonfermentable complex component SMARCB1 is extremely prevalent in pediatric malignant rhabdoid tumors (MRTs) or atypical teratoid rhabdoid tumors. This alteration is hypothesized to confer oncogenic dependency on EZH2 in these cancers. We report the discovery of a potent, selective, and orally bioavailable small-molecule inhibitor of EZH2 enzymatic activity, (N-((4,6-dimethyl-2-oxo-1,2-dihydropyridin-3-yl)methyl)-5-(ethyl(tetrahydro-2H-pyran-4-yl)amino)-4-methyl-4'-(morpholinomethyl)-[1,1'-biphenyl]-3-carboxamide). The compound induces apoptosis and differentiation specifically in SMARCB1-deleted MRT cells. Treatment of xenograft-bearing mice with (N-((4,6-dimethyl-2-oxo-1,2-dihydropyridin-3-yl)methyl)-5-(ethyl(tetrahydro-2H-pyran-4-yl)amino)-4-methyl-4'-(morpholinomethyl)-[1,1'-biphenyl]-3-carboxamide) leads to dose-dependent regression of MRTs with correlative diminution of intratumoral trimethylation levels of lysine 27 on histone H3, and prevention of tumor regrowth after dosing cessation. These data demonstrate the dependency of SMARCB1 mutant MRTs on EZH2 enzymatic activity and portend the utility of EZH2-targeted drugs for the treatment of these genetically defined cancers.

A selective inhibitor of EZH2 blocks H3K27 methylation and kills mutant lymphoma cells.

EZH2 catalyzes trimethylation of histone H3 lysine 27 (H3K27). Point mutations of EZH2 at Tyr641 and Ala677 occur in subpopulations of non-Hodgkin's lymphoma, where they drive H3K27 hypertrimethylation. Here we report the discovery of EPZ005687, a potent inhibitor of EZH2 (K(i) of 24 nM). EPZ005687 has greater than 500-fold selectivity against 15 other protein methyltransferases and has 50-fold selectivity against the closely related enzyme EZH1. The compound reduces H3K27 methylation in various lymphoma cells; this translates into apoptotic cell killing in heterozygous Tyr641 or Ala677 mutant cells, with minimal effects on the proliferation of wild-type cells. These data suggest that genetic alteration of EZH2 (for example, mutations at Tyr641 or Ala677) results in a critical dependency on enzymatic activity for proliferation (that is, the equivalent of oncogene addiction), thus portending the clinical use of EZH2 inhibitors for cancers in which EZH2 is genetically altered.

Function-based gene identification using enzymatically generated normalized shRNA library and massive parallel sequencing.

As a general strategy for function-based gene identification, an shRNA library containing approximately 150 shRNAs per gene was enzymatically generated from normalized (reduced-redundance) human cDNA. The library was constructed in an inducible lentiviral vector, enabling propagation of growth-inhibiting shRNAs and controlled activity measurements. RNAi activities were measured for 101 shRNA clones representing 100 human genes and for 201 shRNAs derived from a firefly luciferase gene. Structure-activity analysis of these two datasets yielded a set of structural criteria for shRNA efficacy, increasing the frequencies of active shRNAs up to 5-fold relative to random sampling. The same library was used to select shRNAs that inhibit breast carcinoma cell growth by targeting potential oncogenes. Genes targeted by the selected shRNAs were enriched for 10 pathways, 9 of which have been previously associated with various cancers, cell cycle progression, or apoptosis. One hundred nineteen genes, enriched through this selection and represented by two to six shRNAs each, were identified as potential cancer drug targets. Short interfering RNAs against 19 of 22 tested genes in this group inhibited cell growth, validating the efficiency of this strategy for high-throughput target gene identification.

Interim results from an ongoing, open-label, single-arm trial of odevixibat in progressive familial intrahepatic cholestasis.

Background & Aims: PEDFIC 2, an ongoing, open-label, 72-week study, evaluates odevixibat, an ileal bile acid transporter inhibitor, in patients with progressive familial intrahepatic cholestasis. Methods: PEDFIC 2 enrolled and dosed 69 patients across two cohorts; all received odevixibat 120 µg/kg per day. Cohort 1 comprised children from PEDFIC 1, and cohort 2 comprised new patients (any age). We report data through 15 July 2020, with Week 24 of PEDFIC 2 the main time point analysed. This represents up to 48 weeks of cumulative exposure for patients treated with odevixibat from the 24-week PEDFIC 1 study (cohort 1A) and up to 24 weeks of treatment for those who initiated odevixibat in PEDFIC 2 (patients who received placebo in PEDFIC 1 [cohort 1B] or cohort 2 patients). Primary endpoints for this prespecified interim analysis were change from baseline to Weeks 22-24 in serum bile acids (sBAs) and proportion of positive pruritus assessments (≥1-point drop from PEDFIC 2 baseline in pruritus on a 0-4 scale or score ≤1) over the 24-week period. Safety monitoring included evaluating treatment-emergent adverse events (TEAEs). Results: In cohort 1A, mean change from PEDFIC 1 baseline to Weeks 22-24 of PEDFIC 2 in sBAs was -201 µmol/L (p <0.0001). For cohort 1B and cohort 2, mean changes from odevixibat initiation to weeks 22-24 in sBAs were -144 and -104 µmol/L, respectively. The proportion of positive pruritus assessments in the first 24-week period of PEDFIC 2 was 33%, 56%, and 62% in cohorts 1A, 1B, and 2, respectively. Most TEAEs were mild or moderate. No drug-related serious TEAEs occurred. Conclusions: Odevixibat in patients with progressive familial intrahepatic cholestasis was generally well tolerated and associated with sustained reductions in sBAs and pruritus. Clinical Trials Registration: This study is registered at ClinicalTrials.gov (NCT03659916). Impact and Implications: Disrupted bile flow is a hallmark feature of patients with progressive familial intrahepatic cholestasis and can result in build-up of bile constituents in the liver with spill over into the bloodstream; other effects that patients can experience include extremely itchy skin, and because not enough bile reaches the gut, patients can have problems digesting food, which may lead to poor growth. Odevixibat is an orally administered medication that shunts bile acids away from the liver. The current study, called PEDFIC 2, suggested that odevixibat can improve the problematic signs and symptoms of progressive familial intrahepatic cholestasis and was generally safe for patients.

Development of the Patient- and Observer-Reported PRUCISION Instruments to Assess Pruritus and Sleep Disturbance in Pediatric Patients with Cholestatic Liver Diseases.

INTRODUCTION: Understanding how patients experience their disease is a vital step in optimal disease management, and patient- and observer-reported outcome (PRO and ObsRO, respectively) measures can add important details to clinical information that is obtained as novel treatments are developed. Instruments that measure meaningful symptoms and impacts from the perspective of pediatric patients with cholestatic liver disease or their caregivers are needed. This study aimed to identify salient concepts in pediatric cholestatic liver disease, develop novel PRO and ObsRO instruments, and establish the instruments' content validity. METHODS: Relevant signs, symptoms, and impacts of cholestatic liver disease were identified through a literature review, interviews with expert clinicians, and concept elicitation interviews with children and caregivers of children who had progressive familial intrahepatic cholestasis (PFIC), Alagille syndrome, biliary atresia, or primary sclerosing cholangitis. Additional cognitive debriefing interviews with patients and caregivers were performed to ensure that participants could understand the instructions, questions, and response scales of the PRO and ObsRO instruments, with modifications made as necessary to improve comprehension and/or usability. RESULTS: A total of 36 interviews with patients and caregivers were conducted. Pruritus and sleep disturbance (e.g., difficulty falling or staying asleep due to itch) were identified as the most problematic symptom and significant impact, respectively, of the pediatric cholestatic liver diseases assessed. The ObsRO and PRO instruments, called PRUCISION, focus on these key disease features in the morning and evening. Several modifications were made to the draft instruments following cognitive interviews. The final PRUCISION PRO and ObsRO measures are designed as an electronic diary to be completed twice daily. The response scales include pictorial, verbal, and numeric scales. CONCLUSION: Novel PRO and ObsRO PRUCISION instruments were created that evaluate the patient experience of cholestatic pruritus in children with PFIC and other cholestatic liver diseases. The content validity of the PRUCISION instruments is established.

Bile, a greenish liquid that is made in the liver, is released into the gut to help digest food. In cholestatic liver disease (CLD), bile flow is interrupted, and bile can build up in the body. One potential effect of this buildup is pruritus, or itchiness of the skin, which can be so intense that it interferes with daily activities. In this study, interviews were done with doctors, patients, and their caregivers to develop new tools to evaluate the most impactful symptoms of CLD in children. After interviewing five doctors and 36 patients and caregivers, two questionnaires called PRUCISION were developed and refined. During this process, participants were first asked about the frequency, severity, duration, and impact of their or their child's symptoms; pruritus was identified as the most common and disruptive symptom associated with CLD, even interfering with sleep. Then, the wording of the questionnaires was modified to make them easier to understand, particularly for younger children. The researchers also had patients do a card-sorting task to ensure that they understood the picture-based responses used in the questionnaires. Finally, more details were added to the instructions for caregivers to more clearly define scratching behaviors. In summary, the questionnaires developed in this study include the perspective of the patient or their caregiver and may be useful to see if new treatments can impact the most prominent symptoms and impacts associated with CLD.

Validation of the PRUCISION Instruments in Pediatric Patients with Progressive Familial Intrahepatic Cholestasis.

INTRODUCTION: Patients with cholestatic liver disease, including progressive familial intrahepatic cholestasis (PFIC) and Alagille syndrome, may have debilitating pruritus, and reducing pruritus is a key therapeutic goal. However, few instruments are available that adequately measure pruritus in pediatric patients with cholestatic liver disease. The objectives of the current study were to establish the measurement properties of the novel PRUCISION patient-reported outcome (PRO) and observer-reported outcome (ObsRO) instruments and to estimate a threshold for clinically meaningful change in pruritus score. METHODS: The PRO/ObsRO instruments are completed twice daily via electronic diary and include 5-point pictorial responses to assess pruritus. Sleep disturbance and tiredness were quantified using 5-point pictorial responses, yes/no responses, and numerical ratings. Data from PEDFIC 1 (NCT03566238), a phase 3 study evaluating odevixibat efficacy and safety in children with PFIC, were used to assess the psychometric properties of these instruments. Quantitative assessments included evaluation of test-retest reliability, determination of construct validity via convergent and known-group validity analyses, and characterization of sensitivity to change. A threshold for within-patient meaningful change from baseline to week 24 was determined using blinded data from PEDFIC 1 and distribution- and anchor-based analyses. RESULTS: Because the majority of patients in PEDFIC 1 were aged < 8 years (n = 52/62) and thus too young to complete the PRO instrument, which was intended for patients aged ≥ 8 years, the small sample size of patients who completed the PRO precluded a full psychometric analysis of the PRO instrument. The ObsRO was completed by a caregiver of every patient in PEDFIC 1. The ObsRO instrument had acceptable test-retest reliability based on intraclass correlation values (most > 0.75). Convergent validity analyses revealed moderate-to-strong correlations (r ≥ 0.3) between baseline ObsRO pruritus scores and baseline Global Impression of Symptoms (GIS) items. In known-groups validity analyses, there were significant differences between baseline groups defined by the GIS for ObsRO pruritus scores and for some sleep disturbance scores. Week 24 ObsRO scores were in the expected direction in groups defined by the Global Impression of Change scale (i.e., improved or not improved); many mean differences between these groups were significant. Sensitivity to change for the ObsRO PRUCISION instrument was also demonstrated by moderate-to-strong Pearson correlations between change from baseline to weeks 21-24 in ObsRO scores and GIS items (r ≥ 0.3). Based on these analyses, a within-patient change of -1.00 from baseline in ObsRO pruritus score was determined to be clinically meaningful. CONCLUSION: The PRUCISION ObsRO instrument is reliable, valid, and sensitive to change, supporting its use as a tool to measure pruritus and sleep disturbance in patients with PFIC and other pediatric cholestatic liver diseases.

Progressive familial intrahepatic cholestasis (PFIC) is a collection of liver diseases that typically affects very young children. A problematic symptom of PFIC is extremely itchy skin, or pruritus, that can keep patients and their families up at night. The PRUCISION questionnaire was developed to measure the severity of a patient's pruritus and sleep disturbance from the perspective of the patient's caregiver. The current study had two primary goals: (1) to assess whether PRUCISION could reliably measure these symptoms and detect changes over time relative to other established rating scales that assess related concepts, and (2) to identify what score change on PRUCISION could be considered clinically meaningful. To do this, data from a clinical study, called PEDFIC 1, in patients with PFIC were used: patient's scores on PRUCISION from their caregiver's perspective were compared with scores on other established scales, first before any treatment was given in PEDFIC 1, and then again after 24 weeks of treatment with a drug called odevixibat. In general, there was good agreement between PRUCISION scores and scores on other scales. For example, when PRUCISION scores indicated that symptoms improved, this tended to correlate with improvement on other measures. Additionally, these analyses indicated that if the PRUCISION score drops by 1 point or more, that can be considered a clinically important change. Overall, this study found that the caregiver-reported PRUCISION questionnaire is valid for assessing changes in pruritus and sleep symptoms in patients with PFIC, which may benefit patients as new treatments are developed.

Computed tomography alone versus computed tomography and magnetic resonance imaging in the identification of occult injuries to the cervical spine: a meta-analysis.

BACKGROUND: Ruling out injuries of the cervical spine in obtunded blunt trauma patients is controversial. Although computed tomography (CT) readily demonstrates fractures and malalignment, it provides limited direct evaluation of ligamentous integrity, leading some to advocate a magnetic resonance imaging (MRI) in obtunded patients. Thus, the question remains: does adding an MRI provide useful information that alters treatment when a CT scan reveals no evidence of injury? METHODS: Published studies from 2000 to 2008 involving patients undergoing MRI for the purposes of further cervical spine evaluation after a "negative" CT scan were identified via a literature search of online databases. Data from eligible studies were pooled and original scale meta-analyses were performed to calculate overall sensitivity, specificity, positive and negative predictive values, likelihood ratios, and relative risk. The Q-statistic p value was used to evaluate heterogeneity. RESULTS: Eleven studies met the inclusion criteria, yielding data on 1,550 patients with a negative CT scan after blunt trauma subsequently evaluated with a MRI. The MRI detected abnormalities in 182 patients (12%). Ninety traumatic injuries were identified, including ligamentous injuries (86/182), fractures, and dislocations (4/182). In 96 cases (6% of the cohort), the MRI identified an injury that altered management. Eighty-four patients (5%) required continued collar immobilization and 12 (1%) required surgical stabilization. The Q-statistic p value for heterogeneity was 0.99, indicating the absence of heterogeneity among the individual study populations. CONCLUSIONS: Reliance on CT imaging alone to "clear the cervical spine" after blunt trauma can lead to missed injuries. This study supports a role for the addition of MRI in evaluating patients who are obtunded, or unexaminable, despite a negative CT scan.

EZH2 Inhibition by Tazemetostat Results in Altered Dependency on B-cell Activation Signaling in DLBCL.

The EZH2 small-molecule inhibitor tazemetostat (EPZ-6438) is currently being evaluated in phase II clinical trials for the treatment of non-Hodgkin lymphoma (NHL). We have previously shown that EZH2 inhibitors display an antiproliferative effect in multiple preclinical models of NHL, and that models bearing gain-of-function mutations in EZH2 were consistently more sensitive to EZH2 inhibition than lymphomas with wild-type (WT) EZH2 Here, we demonstrate that cell lines bearing EZH2 mutations show a cytotoxic response, while cell lines with WT-EZH2 show a cytostatic response and only tumor growth inhibition without regression in a xenograft model. Previous work has demonstrated that cotreatment with tazemetostat and glucocorticoid receptor agonists lead to a synergistic antiproliferative effect in both mutant and wild-type backgrounds, which may provide clues to the mechanism of action of EZH2 inhibition in WT-EZH2 models. Multiple agents that inhibit the B-cell receptor pathway (e.g., ibrutinib) were found to have synergistic benefit when combined with tazemetostat in both mutant and WT-EZH2 backgrounds of diffuse large B-cell lymphomas (DLBCL). The relationship between B-cell activation and EZH2 inhibition is consistent with the proposed role of EZH2 in B-cell maturation. To further support this, we observe that cell lines treated with tazemetostat show an increase in the B-cell maturation regulator, PRDM1/BLIMP1, and gene signatures corresponding to more advanced stages of maturation. These findings suggest that EZH2 inhibition in both mutant and wild-type backgrounds leads to increased B-cell maturation and a greater dependence on B-cell activation signaling. Mol Cancer Ther; 16(11); 2586-97. ©2017 AACR.

Correction: Preclinical Evidence of Anti-Tumor Activity Induced by EZH2 Inhibition in Human Models of Synovial Sarcoma.

[This corrects the article DOI: 10.1371/journal.pone.0158888.].

Preclinical Evidence of Anti-Tumor Activity Induced by EZH2 Inhibition in Human Models of Synovial Sarcoma.

The catalytic activities of covalent and ATP-dependent chromatin remodeling are central to regulating the conformational state of chromatin and the resultant transcriptional output. The enzymes that catalyze these activities are often contained within multiprotein complexes in nature. Two such multiprotein complexes, the polycomb repressive complex 2 (PRC2) methyltransferase and the SWItch/Sucrose Non-Fermentable (SWI/SNF) chromatin remodeler have been reported to act in opposition to each other during development and homeostasis. An imbalance in their activities induced by mutations/deletions in complex members (e.g. SMARCB1) has been suggested to be a pathogenic mechanism in certain human cancers. Here we show that preclinical models of synovial sarcoma-a cancer characterized by functional SMARCB1 loss via its displacement from the SWI/SNF complex through the pathognomonic SS18-SSX fusion protein-display sensitivity to pharmacologic inhibition of EZH2, the catalytic subunit of PRC2. Treatment with tazemetostat, a clinical-stage, selective and orally bioavailable small-molecule inhibitor of EZH2 enzymatic activity reverses a subset of synovial sarcoma gene expression and results in concentration-dependent cell growth inhibition and cell death specifically in SS18-SSX fusion-positive cells in vitro. Treatment of mice bearing either a cell line or two patient-derived xenograft models of synovial sarcoma leads to dose-dependent tumor growth inhibition with correlative inhibition of trimethylation levels of the EZH2-specific substrate, lysine 27 on histone H3. These data demonstrate a dependency of SS18-SSX-positive, SMARCB1-deficient synovial sarcomas on EZH2 enzymatic activity and suggests the potential utility of EZH2-targeted drugs in these genetically defined cancers.

EPZ011989, A Potent, Orally-Available EZH2 Inhibitor with Robust in Vivo Activity.

Inhibitors of the protein methyltransferase Enhancer of Zeste Homolog 2 (EZH2) may have significant therapeutic potential for the treatment of B cell lymphomas and other cancer indications. The ability of the scientific community to explore fully the spectrum of EZH2-associated pathobiology has been hampered by the lack of in vivo-active tool compounds for this enzyme. Here we report the discovery and characterization of EPZ011989, a potent, selective, orally bioavailable inhibitor of EZH2 with useful pharmacokinetic properties. EPZ011989 demonstrates significant tumor growth inhibition in a mouse xenograft model of human B cell lymphoma. Hence, this compound represents a powerful tool for the expanded exploration of EZH2 activity in biology.

Reaction coupling between wild-type and disease-associated mutant EZH2.

EZH2 and EZH1 are protein methyltransferases (PMTs) responsible for histone H3, lysine 27 (H3K27) methylation. Trimethylation of H3K27 (H3K27me3) is a hallmark of many cancers, including non-Hodgkin lymphoma (NHL). Heterozygous EZH2 point mutations at Tyr641, Ala677, and Ala687 have been observed in NHL. The Tyr641 mutations enhance activity on H3K27me2 but have weak or no activity on unmethylated H3K27, whereas the Ala677 and Ala687 mutations use substrates of all methylation states effectively. It has been proposed that enzymatic coupling of the wild-type and mutant enzymes leads to the oncogenic H3K27me3 mark in mutant-bearing NHL. We show that coupling with the wild-type enzyme is needed to achieve H3K27me3 for several mutants, but that others are capable of achieving H3K27me3 on their own. All forms of PRC2 (wild-type and mutants) display kinetic signatures that are consistent with a distributive mechanism of catalysis.

Synergistic Anti-Tumor Activity of EZH2 Inhibitors and Glucocorticoid Receptor Agonists in Models of Germinal Center Non-Hodgkin Lymphomas.

Patients with non-Hodgkin lymphoma (NHL) are treated today with a cocktail of drugs referred to as CHOP (Cyclophosphamide, Hydroxyldaunorubicin, Oncovin, and Prednisone). Subsets of patients with NHL of germinal center origin bear oncogenic mutations in the EZH2 histone methyltransferase. Clinical testing of the EZH2 inhibitor EPZ-6438 has recently begun in patients. We report here that combining EPZ-6438 with CHOP in preclinical cell culture and mouse models results in dramatic synergy for cell killing in EZH2 mutant germinal center NHL cells. Surprisingly, we observe that much of this synergy is due to Prednisolone - a glucocorticoid receptor agonist (GRag) component of CHOP. Dramatic synergy was observed when EPZ-6438 is combined with Prednisolone alone, and a similar effect was observed with Dexamethasone, another GRag. Remarkably, the anti-proliferative effect of the EPZ-6438+GRag combination extends beyond EZH2 mutant-bearing cells to more generally impact germinal center NHL. These preclinical data reveal an unanticipated biological intersection between GR-mediated gene regulation and EZH2-mediated chromatin remodeling. The data also suggest the possibility of a significant and practical benefit of combining EZH2 inhibitors and GRag that warrants further investigation in a clinical setting.

Selective inhibition of EZH2 by EPZ-6438 leads to potent antitumor activity in EZH2-mutant non-Hodgkin lymphoma.

Mutations within the catalytic domain of the histone methyltransferase EZH2 have been identified in subsets of patients with non-Hodgkin lymphoma (NHL). These genetic alterations are hypothesized to confer an oncogenic dependency on EZH2 enzymatic activity in these cancers. We have previously reported the discovery of EPZ005678 and EPZ-6438, potent and selective S-adenosyl-methionine-competitive small molecule inhibitors of EZH2. Although both compounds are similar with respect to their mechanism of action and selectivity, EPZ-6438 possesses superior potency and drug-like properties, including good oral bioavailability in animals. Here, we characterize the activity of EPZ-6438 in preclinical models of NHL. EPZ-6438 selectively inhibits intracellular lysine 27 of histone H3 (H3K27) methylation in a concentration- and time-dependent manner in both EZH2 wild-type and mutant lymphoma cells. Inhibition of H3K27 trimethylation (H3K27Me3) leads to selective cell killing of human lymphoma cell lines bearing EZH2 catalytic domain point mutations. Treatment of EZH2-mutant NHL xenograft-bearing mice with EPZ-6438 causes dose-dependent tumor growth inhibition, including complete and sustained tumor regressions with correlative diminution of H3K27Me3 levels in tumors and selected normal tissues. Mice dosed orally with EPZ-6438 for 28 days remained tumor free for up to 63 days after stopping compound treatment in two EZH2-mutant xenograft models. These data confirm the dependency of EZH2-mutant NHL on EZH2 activity and portend the utility of EPZ-6438 as a potential treatment for these genetically defined cancers.

Pain intensity and patients' acceptance of surgical complication risks with lumbar fusion.

STUDY DESIGN: Cross-sectional study with prospective recruitment. OBJECTIVE: To determine the relationship of pain intensity (back and leg) on patients' acceptance of surgical complication risks when deciding whether or not to undergo lumbar spinal fusion. SUMMARY OF BACKGROUND DATA: To formulate informed decisions regarding lumbar fusion surgery, preoperative discussions should include a review of the risk of complications balanced with the likelihood of symptom relief. Pain intensity has the potential to influence a patient's decision to consent to lumbar fusion. We hypothesized that pain intensity is associated with a patient's acceptance of surgical complication risks. METHODS: Patients being seen for the first time by a spine surgeon for treatment of a nontraumatic or non-neoplastic spinal disorder completed a structured questionnaire. It posed 24 scenarios, each presenting a combination of risks of 3 complications (nerve damage, wound infection, and nonunion) and probabilities of symptom relief. For each scenario, the patient indicated whether he or she would or would not consent to a fusion for low back pain (LBP). The sum of the scenarios in which the patient responded that he or she would elect surgery was calculated to represent acceptance of surgical complication risks. A variety of other data were also recorded, including age, sex, education level, race, history of nonspinal surgery, duration of pain, and history of spinal injections. Data were analyzed using bivariate and multivariate regression analyses. RESULTS: The mean number of scenarios accepted by 118 enrolled subjects was 10.2 (median, 8; SD, 8.5; range, 0-24, or 42.5% of scenarios). In general, subjects were more likely to accept scenarios with lower risks and higher efficacy. Spearman rank correlation estimates demonstrated a moderate association between the LBP intensity and acceptance of surgical complication risks (r = 0.37, P = 0.0001) whereas leg pain intensity had a weak but positive correlation (r = 0.19, P = 0.04). In bivariate analyses, a history of spinal injections was strongly associated with patients' acceptance of surgical complication risks and willingness to proceed with surgery (54.5% of scenarios accepted for those who had a history of spinal injections vs. 27.6% for those with no history of spinal injections, P = 0.0001). White patients were more willing to accept surgery (45.9% of scenarios) than non-white patients (28.4%, P = 0.03). With the available numbers, age, sex, history of nonspinal surgery, education, and the duration of pain demonstrated no clear association with acceptance of surgical complication risks. Although education overall was not an influential factor, more educated males had greater risk tolerance than less educated males whereas more educated females had less risk tolerance than less educated females (P = 0.023). In multivariate analysis, LBP intensity remained a highly statistically significant correlate (P = 0.001) of the proportion of scenarios accepted, as did a history of spinal injections (P = 0.001) and being white (0.03). CONCLUSION: The current investigation indicates that the intensity of LBP is the most influential factor affecting a patient's decision to accept risk of complication and symptom persistence when considering lumbar fusion. This relationship has not been previously shown for any surgical procedure. These data could potentially change the manner in which patients are counseled to make informed choices about spinal surgery. With growing interest in adverse events and complications, these data could be important in establishing guidelines for patient-directed surgical decision making.

Mortality in elderly patients with hyperostotic disease of the cervical spine after fracture: an age- and sex-matched study.

BACKGROUND CONTEXT: Several reports indicate that patients with ankylosing spondylitis (AS) and diffuse idiopathic skeletal hyperostosis (DISH) have increased mortality after cervical spine fractures. However, outcomes of the fractured hyperostotic cervical spine are incompletely described, and there are limited data regarding the covariable effects of patient age and medical comorbidities on mortality. PURPOSE: To determine mortality associated with cervical fractures in patients with hyperostotic disease. STUDY DESIGN: Retrospective case-control study. PATIENT SAMPLE: Forty-three patients identified through a registry as having fractures of the cervical spine in the setting of hyperostotic disease. These patients were matched to 43 controls who did not carry the diagnosis of hyperostotic disease. OUTCOME MEASURES: Mortality at 3 months and 1, 2, and 3 years after fracture. METHODS: An institutional database was used to identify all cervical fractures sustained by patients aged 65 years and older from 1991 to 2006. Demographic information, date of injury, associated injuries, treatment type, presence of AS or DISH, and comorbidities were abstracted from medical records and radiographs. Mortality was ascertained using the National Death Index. Patients with AS or DISH were matched to controls who did not carry the diagnosis of hyperostotic disease. Risks of mortality were calculated at 3 months, 1 year, 2 years, and 3 years. Kaplan-Meier methods, logistic regression analysis, the two independent sample t test, and the Fisher exact test were used to compare mortalities between the two groups. Statistical significance was determined as p values <.05. RESULTS: Forty-three patients were identified as having fractures in the setting of hyperostotic disease of the cervical spine. Twenty-seven individuals had DISH, and 16 had AS. The average age of both the study group and controls was 80 years, with an age range of 68 to 94. There was no significantly increased risk of mortality between the overall study group and control group at 3 months (p=.20), 1 (p=.22), 2 (p=.15), or 3 years (p=.50) after injury. Compared with controls, subgroup analysis of patients with AS showed a statistically increased risk of mortality at 3 months (p<.01) and at 1 and 2 years (p<.01 at both time points). When compared with individuals with DISH, those with AS had an increased risk of mortality at time points up to 2 years after fracture. Patients with DISH did not have an increased mortality risk at any time point when compared with controls. CONCLUSIONS: The effect of fracture on mortality appears to be greatest in those with AS. Patients with DISH did not demonstrate an increased risk of mortality compared with age- and sex-matched controls. Future studies of patients with hyperostotic disease should analyze patients with DISH and AS separately instead of as a single homogenous group. LEVEL OF EVIDENCE: Level IV.

Type II odontoid fractures of the cervical spine: do treatment type and medical comorbidities affect mortality in elderly patients?

STUDY DESIGN: Retrospective cohort study. OBJECTIVE: To determine the influence of age, comorbidities, and treatment type on mortality in elderly patients with acute Type II odontoid fractures. SUMMARY OF BACKGROUND DATA: Prior studies have documented increased morbidity and mortality among geriatric patients sustaining odontoid fractures. However, there is limited data regarding the effect of patient age, medical comorbidities, and treatment selection on mortality after Type II odontoid (C2) fractures in the elderly. METHODS: An institutional registry was used to identify all Type II odontoid fractures sustained by patients aged 65 and older from 1991 to 2006. Demographic information, date of injury, associated injuries, treatment type, and comorbidities were abstracted from medical records. Mortality was ascertained using the National Death Index. Risks of mortality and their associated 95% confidence intervals (CIs) were calculated at 3 months, 1 year, 2 years, and 3 years. Multivariable Cox proportional hazard regression was used to evaluate independent factors affecting mortality stratified by age (65-74 years, 75-84 years, ≥ 85 years) and treatment type (operative or nonoperative treatment, and halo or collar immobilization). RESULTS: Of 156 patients identified with Type II odontoid fracture, the average age was 82 years (SD = 7.8; Range: 65-101). One hundred and twelve patients (72%) were treated nonoperatively. At 3 years postinjury, there was a 39% (95% CI: 32-47) mortality rate for the entire cohort. Mortality for the operative group was 11% (95% CI: 2-21) at 3 months and 21% (95% CI: 9-32) at 1 year compared with 25% (95% CI: 17-33) at 3 months and 36% (95% CI: 27-45) at 1 year in the nonoperative group. The Cox regression model showed that the protective effect of surgery was seen in patients aged 65 to 74 years, in whom the hazard ratio associated with surgery for mortality after odontoid fracture was 0.4 (95% CI: 0.1-1.5). Those aged 75 to 84 years had a hazard ratio of 0.8 (95% CI: 0.3-2.3), and patients 85 years or older had a hazard ratio of 1.9 (95% CI: 0.6-6.1; P value for interaction between age and treatment = 0.09) with operative treatment having a protective effect in patients aged 65 to 74 years. CONCLUSION: In a cohort of elderly patients, Type II odontoid fractures were associated with a high rate of mortality, regardless of intervention.

Mortality in elderly patients after cervical spine fractures.

BACKGROUND: Despite an increased risk of cervical spine fractures in older patients, little is known about the mortality associated with these fractures and there is no consensus on the optimal treatment. The purposes of this study were to determine the three-month and one-year mortality associated with cervical spine fractures in patients sixty-five years of age or older and to evaluate potential factors that may influence mortality. METHODS: We performed a retrospective review of all cervical spine fractures in patients sixty-five years of age or older from 1991 to 2006 at two institutions. Information regarding age, sex, race, treatment type, neurological involvement, injury mechanism, comorbidity, and mortality were collected. Overall risk of mortality and mortality stratified by the above factors were calculated at three months and one year. Cox proportional-hazard regression was performed to identify independent correlates of mortality. RESULTS: Six hundred and forty patients were included in our analysis. The mean age was eighty years (range, sixty-five to 101 years). Two hundred and ninety-four patients (46%) were male, and 116 (18%) were nonwhite. The risk of mortality was 19% at three months and 28% at one year. The effect of treatment on mortality varied with age at three months (p for interaction = 0.03) but not at one year (p for interaction = 0.08), with operative treatment being associated with less mortality for those between the ages of sixty-five and seventy-four years. A higher Charlson comorbidity score, male sex, and neurological involvement were all associated with increased risk of mortality. CONCLUSIONS: Operative treatment of cervical spine fractures is associated with a lower mortality rate at three months but not at one year postoperatively for patients between sixty-five and seventy-four years old at the time of fracture.

E. coli virulence factor hemolysin induces neutrophil apoptosis and necrosis/lysis in vitro and necrosis/lysis and lung injury in a rat pneumonia model.

Enteric gram-negative bacilli, such as Escherichia coli are the most common cause of nosocomial pneumonia. In this study a wild-type extraintestinal pathogenic strain of E. coli (ExPEC)(CP9) and isogenic derivatives deficient in hemolysin (Hly) and cytotoxic necrotizing factor (CNF) were assessed in vitro and in a rat model of gram-negative pneumonia to test the hypothesis that these virulence factors induce neutrophil apoptosis and/or necrosis/lysis. As ascertained by in vitro caspase-3/7 and LDH activities and neutrophil morphology, Hly mediated neutrophil apoptosis at lower E. coli titers (1 x 10(5-6) cfu) and necrosis/lysis at higher titers (> or =1 x 10(7) cfu). Data suggest that CNF promotes apoptosis but not necrosis or lysis. We also demonstrate that annexin V/7-amino-actinomycin D staining was an unreliable assessment of apoptosis using live E. coli. The use of caspase-3/7 and LDH activities and neutrophil morphology supported the notion that necrosis, not apoptosis, was the primary mechanism by which neutrophils were affected in our in vivo gram-negative pneumonia model using live E. coli. In addition, in vivo studies demonstrated that Hly mediates lung injury. Neutrophil necrosis was not observed when animals were challenged with purified lipopolysaccharide, demonstrating the importance of using live bacteria. These findings establish that Hly contributes to ExPEC virulence by mediating neutrophil toxicity, with necrosis/lysis being the dominant effect of Hly on neutrophils in vivo and by lung injury. Whether Hly-mediated lung injury is due to neutrophil necrosis, a direct effect of Hly, or both is unclear.