Indirect comparisons of relative efficacy estimates of zuranolone and selective serotonin reuptake inhibitors for postpartum depression.

AIMS: Estimate relative efficacy of zuranolone, a novel oral, Food and Drug Administration-approved treatment for postpartum depression (PPD) in adults vs. selective serotonin reuptake inhibitors (SSRIs) and combination therapies used for PPD in the United States. MATERIALS AND METHODS: Randomized controlled trials (RCTs) for zuranolone and SSRIs, identified from systematic review, were used to construct evidence networks, linking via common comparator arms. Due to heterogeneity in placebo responses, matching-adjusted indirect comparison (MAIC) was applied, statistically weighting the zuranolone treatment arm of Phase 3 SKYLARK Study (NCT04442503) to the placebo arm of RCTs investigating SSRIs for PPD. MAIC outputs were applied in Bucher indirect treatment comparisons (ITCs) and network meta-analysis (NMA), using Edinburgh Postnatal Depression Scale (EPDS) and 17-item Hamilton Rating Scale for Depression (HAMD-17) change from baseline (CFB) on Days 3, 15, 28 (Month 1), 45, and last observation (Day 45, Week 12/18). RESULTS: Larger EPDS CFB was observed among zuranolone-treated vs. SSRI-treated patients from Day 15 onward. Zuranolone-treated (vs. SSRI-treated) patients exhibited 4.22-point larger reduction in EPDS by Day 15 (95% confidence interval: -6.16, -2.28) and 7.43-point larger reduction at Day 45 (-9.84, -5.02) with Bucher ITC. NMA showed EPDS reduction for zuranolone was 4.52 (-6.40, -2.65) points larger than SSRIs by Day 15 and 7.16 (-9.47, -4.85) larger at Day 45. Lack of overlap between study populations substantially reduced effective sample size post-matching, making HAMD-17 CFB analysis infeasible. LIMITATIONS: Limited population overlap between SKYLARK Study and RCTs reduced feasibility of undertaking HAMD-17 CFB ITCs and may introduce uncertainty to EPDS CFB ITC results. CONCLUSIONS: Analysis showed zuranolone-treated patients with PPD experienced greater symptom improvement than SSRI-treated patients from Day 15 onward, with largest mean difference at Day 45. Adjusting for differences between placebo arms, zuranolone may be associated with greater PPD symptom improvement (measured by EPDS) vs. SSRIs.

Utilization of MG-ADL in myasthenia gravis clinical research and care.

The Myasthenia Gravis Activities of Living (MG-ADL) scale is an 8-item patient-reported scale that measures myasthenia gravis (MG) symptoms and functional status. The objective of the current review is to summarize the psychometric properties of the MG-ADL and published evidence of MG-ADL use. A targeted literature review for published studies of the MG-ADL was conducted using a database and gray literature search. A total of 48 publications and 35 clinical trials were included. Studies indicated that the MG-ADL is a reliable and valid measure that has been used as an outcome in clinical trials and observational studies to measure MG symptoms and response to treatment. While most often used as a secondary endpoint in clinical trials, its use as a primary endpoint has increased in recent years. The most common MG-ADL endpoint is change in MG-ADL score from baseline, although there has been an increase in the analysis of a responder threshold using the MG-ADL. A new concept of minimal symptom expression (MSE) has emerged more recently. Duration of treatment effect is another important construct that is being increasingly evaluated using the MG-ADL. The use of the MG-ADL as a primary endpoint in clinical trials and in responder threshold analyses to indicate treatment improvement has increased in recent years. MSE using the MG-ADL shows promise in helping to determine success of treatment and may be the aspirational goal of MG treatment for the future once validated, particularly given the evolving treatment landscape in MG.

Willingness to Pay for National Health Insurance: A Contingent Valuation Study Among Patients Visiting Public Hospitals in Melaka, Malaysia.

BACKGROUND: The Malaysian public healthcare system is tax funded, with healthcare costs highly subsidized by the Government. The current health financing is not sustainable to cover all the healthcare resources. Introducing a national health insurance (NHI), a prepayment method, would protect the population from financial hardship by reducing out-of-pocket payment, minimizing the barrier to accessing health and achieving equitable healthcare. Studies on the willingness to pay (WTP) for NHI are available in different contexts; however, little was found in Malaysia. In addition, factors affecting the WTP for NHI have been less researched in the Malaysian context. OBJECTIVE: This study investigates the WTP for NHI and factors influencing it in Melaka, Malaysia. METHOD: Using the contingent valuation method, the researchers distributed a pre-tested self-administered questionnaire to 489 respondents in three leading public hospitals from 1 to 9 November 2019. Multi-stage sampling forms the selection of respondents. Respondents were presented with a hypothetical scenario on NHI and asked whether they would be willing to pay for NHI. Chi-square was used to examine the association between categorical independent variables and WTP for NHI, and binary logistic regression was applied to determine the variables with the most predictive effect towards WTP for NHI. RESULTS: Out of 462 usable responses, 344 respondents (74.5%) were willing to pay for NHI. The majority (51.3%) of the respondents were willing to pay 15 Malaysian Ringgit (3.70 US dollars) or more for the NHI scheme. Chi-square analysis revealed that age, marital status, education level, employment status, chronic diseases, and perception towards NHI were significantly associated with WTP for NHI. Binary regression analysis found that marital status and perception towards NHI have the most predictive effect on respondents' WTP for NHI. CONCLUSION: This study revealed that the majority of the respondents are willing to pay for NHI if implemented in Malaysia. The findings lay a foundation for implementing a future NHI framework in Malaysia and reference future health financing research.

Atypical Outcomes of Nasal and Lip Appearance After Unilateral Cleft Lip Repair: Judgment by Professionals, Patients, and Laypeople.

OBJECTIVE: To gain more insight into the assessment of "atypical" nasal and lip appearance outcomes compared to "typical" appearance outcomes after unilateral cleft lip and palate (UCLP) repair, when judged by professionals, patients with repaired UCLP, and laypeople. DESIGN: An online survey containing 3 series of photographs with various degrees of "typical" and "atypical" nasal and lip appearance outcomes after UCLP repair was sent to 30 professionals, 30 patients with repaired UCLP, and 50 laypeople in 2 countries. Participants were instructed to rank the photographs from excellent to poor based on overall appearance. Mean rank positions of photographs were analyzed and differences in mean rank score between "typical" and "atypical" results were assessed using a T-test. Agreement of ranking between the 3 groups was assessed with an analysis of variance analysis. SETTING: Amsterdam UMC, location VUmc, Netherlands and Boston Children's Hospital, Boston, USA. PATIENTS: Photographs of 6- to 18-year-old patients with repaired UCLP. RESULTS: "Atypical" appearance outcomes were ranked significantly less favorably (small nostril: P = 0.00; low vermillion border: P = 0.02; whistling deformity: P = 0.00) compared to "typical" outcomes. Difference between professionals, patients and laypeople in rank positioning the photographs was not statistically significant (P = 0.89). CONCLUSIONS: Noses with a smaller nostril and lips containing a whistling deformity were perceived as poorer outcome compared to the "typical" results. Professionals, patients, and laypeople are in agreement when assessing these outcomes.

Does the Nostril Shape Change After Le Fort I Advancement in Patients With Unilateral Complete Cleft Lip?

PURPOSE: Patients with unilateral cleft lip and palate (UCLP) may require Le Fort I advancement to correct maxillary hypoplasia after reaching skeletal maturity. The underlying cleft anatomy, previous operations, and scarring can affect nostril changes after maxillary advancement. The purpose of the present study was to determine whether Le Fort I advancement affects the nostril configuration (ie, width, axis, shape) in patients with UCLP. The specific aims were to (1) compare cleft and noncleft nostrils in patients with UCLP after maxillary advancement and (2) compare the changes in nostril configuration in patients with UCLP with those in noncleft controls after Le Fort advancement. PATIENTS AND METHODS: A retrospective case-control study of nonsyndromic, skeletally mature patients with UCLP and a case-matched control group without UCLP who had undergone single-piece Le Fort I advancement with alar cinch suture from 2010 to 2014. Patients were included if they had undergone pre- and postoperative 3-dimensional photogrammetry without intervening nasal revision. Three-dimensional anthropometry was used to evaluate changes in nostril axis and width, soft triangle angle, columellar show, and nasal width after orthognathic correction. RESULTS: The present study included 19 patients with UCLP (11 males; mean age, 18.0 years) and 19 noncleft controls (11 males; mean age, 18.7 years; P = .276). The mean sagittal advancement in the patients with UCLP and noncleft controls was 7.5 mm and 6.3 mm, respectively (P = .143). On average, the nostrils widened, the soft triangles flattened, and the columellar show increased. No significant difference was found in the changes to the nostril configuration between the cleft and noncleft sides in the patients with UCLP. No significant differences were found in the nostril changes between patients with UCLP and noncleft controls. CONCLUSIONS: Baseline nostril asymmetry is not altered by Le Fort osteotomy in patients with UCLP because both nostrils respond similarly to the deforming forces of maxillary advancement. Similarly, no differences were found in the nostril changes between the cleft and noncleft controls. These findings can aid proper surgical planning for cleft nasal revisions.

Beware the Alar Base Optical Illusion in Assessment of Unilateral Cleft Lip Nasal Deformity.

BACKGROUND: After repair of unilateral cleft lip, lateral drift of the alar base is expected and may require revision. Although the alar base appears wider than on the noncleft side, by measurement it could be symmetric or narrow. Is this an optical illusion? METHODS: This is a retrospective photographic evaluation of children who underwent repair of unilateral complete cleft lip and palate. On submental standardized photographs, raters judged whether the alar base position was wider, equal to, or narrower than on the noncleft side. The true alar base position was measured by photogrammetry and the frequency of alar base misjudgment was assessed. On control photographs, the cleft side alar base was wider than the noncleft side. On test photographs, the cleft side alar base was narrower than the noncleft side. Possible association between misjudgment and nasal features was also evaluated. Data were analyzed using the Fisher's exact test. RESULTS: Three raters assessed 61 patients aged 6 to 9 years and treated between 1995 and 2015. In 77 percent of control photographs, raters correctly judged the cleft side alar base as wider. In 65 percent of test photographs, raters incorrectly judged the alar base position as wider on the cleft side. Asymmetry of nostril outline and a transverse nostril axis were associated with misjudged photographs (p < 0.05). CONCLUSIONS: Alar base assessment can be altered by an optical illusion. Simple anthropometry counters this misperception. Measurement should be repeated after correction of the deviated anterocaudal septum and elevation of the lower lateral cartilage. Thereafter, a decision is made on altering the alar base position. CLINICAL QUESTION/LEVEL OF EVIDENCE: Diagnostic, II.

Defining the Aesthetic Range of Normal Symmetry for Lip and Nose Features in 5-Year-Old Children Using the Computer-Based Program SymNose.

OBJECTIVE: To provide a normal comparison group against which to judge symmetry results after cleft surgery and to introduce the thin lip correction (TLC) feature in SymNose. A lip-aspect ratio algorithm has been added to the latest version of SymNose to compensate for the higher degree of overlap in thicker lips when compared to thin lips. DESIGN: Retrospective analysis of symmetry in healthy participants, using the computer-based program SymNose on both anteroposterior (AP) and base view images. Photographs of 91 noncleft children were traced twice by 3 independent investigators experienced with SymNose. PARTICIPANTS: Five-year-old healthy participants from a local state school in Tavistock (West Devon, United Kingdom). MAIN OUTCOME MEASURE: Asymmetry expressed as the perimeter mismatch percentage for nose and lip features on AP view images and for nose features on base view images. RESULTS: The perimeter mismatch reference range for the nose (AP view) was 2.65% to 30.91%, for the lip 2.13% to 15.44%, for the nose (base view) 1.69% to 14.84%, for the nostrils 4.68% to 26.6%, and for the width-height ratio 1.15% to 1.80%. The perimeter mismatch percentage for the lip without TLC was significantly higher compared to the perimeter mismatch percentage with TLC (P < .001). CONCLUSION: This article provides a noncleft reference range for all perimeters drawn from SymNose against which to compare results after cleft surgery at 5 years of age. Furthermore, it shows the importance of correcting for variance in lip volume per child.

Development of the Submental Nasal Appearance Scale for the Assessment of Repaired Unilateral Complete Cleft Lip: A Pilot Study.

OBJECTIVE: To develop the "Submental Nasal Appearance Scale" (SNAS), which is an easy-to-use objectified tool to represent a cleft surgeon's standard for assessment of the nasal appearance from the submental perspective. DESIGN: Eighty-five photographs of patients with unilateral complete cleft lip and palate were selected and cropped, displaying the submental view. Sixty-one photographs were used to develop 5 sets of reference photographs. Three cleft surgeons graded 24 photographs with these sets and subjectively graded the overall nasal appearance as well. Internal agreement for both methods was calculated, as well as correlation between them. The SNAS was created, by only using the combination of sets that showed the highest reliability and correlation. SETTING: Boston Children's Hospital, Boston, Massachusetts. PATIENTS: Six- to 9-year-old patients with unilateral complete cleft lip and palate. RESULTS: The intrarater and interrater reliability was 0.84 and 0.79, respectively, for the SNAS and 0.76 and 0.62, respectively, for the overall appearance assessment. The correlation was 0.74 between the methods. CONCLUSIONS: The SNAS is a reliable tool that reflects a cleft surgeon's standard and could be used independently or in combination with existing rating scales using the frontal and/or lateral view, for assessment after cleft lip repair.

Rating Nasolabial Aesthetics in Unilateral Cleft Lip and Palate Patients: Cropped Versus Full-Face Images.

OBJECTIVE: To determine if cropping facial images affects nasolabial aesthetics assessments in unilateral cleft lip patients and to evaluate the effect of facial attractiveness on nasolabial evaluation. DESIGN: Two cleft surgeons and one cleft orthodontist assessed standardized frontal photographs 4 times; nasolabial aesthetics were rated on cropped and full-face images using the Cleft Aesthetic Rating Scale, and total facial attractiveness was rated on full-face images with and without the nasolabial area blurred using a 5-point Likert scale. SETTING: Cleft Palate Craniofacial Unit of a University Medical Center. PATIENTS: Inclusion criteria: nonsyndromic unilateral cleft lip and an available frontal view photograph around 10 years of age. EXCLUSION CRITERIA: a history of facial trauma and an incomplete cleft. Eighty-one photographs were available for assessment. MAIN OUTCOME MEASURES: Differences in mean CARS scores between cropped versus full-face photographs and attractive versus unattractive rated patients were evaluated by paired t test. RESULTS: Nasolabial aesthetics are scored more negatively on full-face photographs compared to cropped photographs, regardless of facial attractiveness. (Mean CARS score, nose: cropped = 2.8, full-face = 3.0, P < .001; lip: cropped = 2.4, full-face = 2.7, P < .001; nose and lip: cropped = 2.6, full-face = 2.8, P < .001). CONCLUSION: Aesthetic outcomes of the nasolabial area are assessed significantly more positively when using cropped images compared to full-face images. For this reason, cropping images, revealing the nasolabial area only, is recommended for aesthetical assessments.

Exposure to male sexual scents (androstenone) influences women's drinking.

In a demonstration of a heretofore unknown motivational pathway for alcohol consumption, we recently showed that exposure to scents emitted by human females during the fertile phase of their menstrual cycle could increase men's drinking. The current study examined the reverse: whether exposure to male sexual scents (androstenone) would increase women's drinking. One hundred three female participants were primed with either androstenone or a control prime (plain water) camouflaged as a men's "cologne." They then completed a laboratory assessment of beer consumption and related measures. (Nonalcoholic beer was used for methodological and safety reasons.) Results indicated that females exposed to the androstenone prime drank significantly more than those exposed to the control prime. Social and sexual expectancies taken subsequent to drinking (to avoid unwanted manipulation influences) were correlated with drinking in the primed group but not in the neutral group, supporting the idea that information-processing pathways related to alcohol use had been engaged in the primed group. Few females were ovulating, precluding assessment of the effects of fertility on this process. Because of the centrality of sexual signaling to fundamental evolutionary/biological forces, these results indicate a potentially powerful influence on alcohol consumption that calls for continued investigation. (PsycINFO Database Record

Use and cost comparison of clobazam to other antiepileptic drugs for treatment of Lennox-Gastaut syndrome.

Background: Lennox-Gastaut syndrome (LGS) is a severe form of childhood-onset epilepsy associated with serious injuries due to frequent and severe seizures. Of the antiepileptic drugs (AEDs) approved for LGS, clobazam is a more recent market entrant, having been approved in October 2011. Recent AED budget impact and cost-effectiveness analyses for LGS suggest that adding clobazam to a health plan formulary may result in decreased medical costs; however, research on clinical and economic outcomes and treatment patterns with these AED treatments in LGS is limited. Objectives: To compare the baseline characteristics and treatment patterns of new initiators of clobazam and other AEDs among LGS patients and compare healthcare utilization and costs before and after clobazam initiation among LGS patients. Methods: A retrospective study of probable LGS patients was conducted using the MarketScan® Commercial, Medicare Supplemental, and Medicaid databases (10/1/2010-3/31/2014). Results: In the Commercial/Medicare Supplemental population, clobazam users were younger, had fewer comorbidities, and more prior AED use than non-clobazam users. In the 12 months pre-treatment initiation, clobazam users had significantly more seizure-related inpatient stays and outpatient visits and higher total seizure-related (P < 0.001) and all-cause (P < 0.001) costs than non-clobazam users. Among clobazam users, when compared to the 12 months pre-clobazam initiation, seizure-related medical utilization and costs were lower in the 12 months post-clobazam initiation (P = 0.004). Total all-cause (P < 0.001) and seizure-related (P = 0.029) costs increased post-clobazam initiation mainly due to the increase in outpatient pharmacy costs. Similar results were observed in the Medicaid population. Conclusions: Baseline results suggest a prescribing preference for clobazam in severe LGS patients. Clobazam users had a reduction in seizure-related medical utilization and costs after clobazam initiation. The improvement in medical costs mostly offset the higher prescription costs following clobazam initiation.

Comparison of two- and three-dimensional assessment methods of nasolabial appearance in cleft lip and palate patients: Do the assessment methods measure the same outcome?

OBJECTIVE: For the assessment of the nasolabial appearance in cleft patients, a widely accepted, reliable scoring system is not available. In this study four different methods of assessment are compared, including 2D and 3D asymmetry and aesthetic assessments. METHODS: The data and ratings from an earlier study using the Asher-McDade aesthetic index on 3D photographs and the outcomes of 3D facial distance mapping were compared to a 2D aesthetic assessment, the Cleft Aesthetic Rating Scale, and to SymNose, a computerized 2D asymmetry assessment technique. The reliability and correlation between the four assessment techniques were tested using a sample of 79 patients. RESULTS: The 3D asymmetry assessment had the highest reliability and could be performed by just one observer (Intraclass correlation coefficient (ICC): 0.99). The 2D asymmetry assessment of the nose was highly reliable when performed by just one observer (ICC: 0.89). However, for the 2D asymmetry assessment of the lip more observers were needed. For the 2D aesthetic assessments 3 observers were needed. The 3D aesthetic assessment had the lowest single-observer reliability (ICC: 0.38-0.56) of all four techniques. The agreement between the different assessment methods is poor to very poor. The highest correlation (R: 0.48) was found between 2D and 3D aesthetic assessments. Remarkably, the lowest correlations were found between 2D and 3D asymmetry assessments (0.08-0.17). CONCLUSION: Different assessment methods are not in agreement and seem to measure different nasolabial aspects. More research is needed to establish exactly what each assessment technique measures and which measurements or outcomes are relevant for the patients.

Toward Improving Electrocardiogram (ECG) Biometric Verification using Mobile Sensors: A Two-Stage Classifier Approach.

Electrocardiogram (ECG) signals sensed from mobile devices pertain the potential for biometric identity recognition applicable in remote access control systems where enhanced data security is demanding. In this study, we propose a new algorithm that consists of a two-stage classifier combining random forest and wavelet distance measure through a probabilistic threshold schema, to improve the effectiveness and robustness of a biometric recognition system using ECG data acquired from a biosensor integrated into mobile devices. The proposed algorithm is evaluated using a mixed dataset from 184 subjects under different health conditions. The proposed two-stage classifier achieves a total of 99.52% subject verification accuracy, better than the 98.33% accuracy from random forest alone and 96.31% accuracy from wavelet distance measure algorithm alone. These results demonstrate the superiority of the proposed algorithm for biometric identification, hence supporting its practicality in areas such as cloud data security, cyber-security or remote healthcare systems.

A Cross-Sectional Study on the Use of, Preference for, and Perceived Reliability of Mass Media for Drug-Related Information Among the General Public in Sarawak.

BACKGROUND: Because of the large size of Sarawak and the high proportion of people residing in rural areas in this Malaysian state, disseminating drug-related information there can be challenging. It is, therefore, important to recognize the type of mass media for drug-related information that are salient to the people of Sarawak. This study was aimed at identifying the use of and the preferences for mass media to obtain drug-related information among the public in Sarawak. We also aimed to recognize the media perceived as the most reliable for drug-related information. METHODS: This was a cross-sectional study using a self-administered questionnaire carried out from September to October 2013. Survey respondents were recruited from 4 divisions in Sarawak: Kuching, Sibu, Miri, and Bintulu. RESULTS: A total of 433 completed questionnaires were obtained at the end of the study period. All respondents had access to common mass media such as television (89.8%, 389/433), radio (68.6%, 297/433), and the Internet (66.1%, 286/433). Among all respondents, television (71.4%, 309/433) was noted as the most preferred media for drug-related information. Compared with rural respondents, urban respondents were significantly more likely to have access to and prefer the Internet to obtain drug-related information. On the other hand, rural respondents were more likely to have access to and prefer radio for such information compared to their urban counterparts. CONCLUSIONS: Television can be an important and attractive choice of mass media in a quality use of medicines (QUM) campaign. The Internet can be used to disseminate drug-related information in urban areas, whereas radio can be used in a QUM campaign targeting the rural public.

The use and reliability of SymNose for quantitative measurement of the nose and lip in unilateral cleft lip and palate patients.

INTRODUCTION: It is essential to have a reliable assessment method in order to compare the results of cleft lip and palate surgery. In this study the computer-based program SymNose, a method for quantitative assessment of the nose and lip, will be assessed on usability and reliability. METHODS: The symmetry of the nose and lip was measured twice in 50 six-year-old complete and incomplete unilateral cleft lip and palate patients by four observers. For the frontal view the asymmetry level of the nose and upper lip were evaluated and for the basal view the asymmetry level of the nose and nostrils were evaluated. RESULTS: A mean inter-observer reliability when tracing each image once or twice was 0.70 and 0.75, respectively. Tracing the photographs with 2 observers and 4 observers gave a mean inter-observer score of 0.86 and 0.92, respectively. The mean intra-observer reliability varied between 0.80 and 0.84. CONCLUSIONS: SymNose is a practical and reliable tool for the retrospective assessment of large caseloads of 2D photographs of cleft patients for research purposes. Moderate to high single inter-observer reliability was found. For future research with SymNose reliable outcomes can be achieved by using the average outcomes of single tracings of two observers.

Exposure to female fertility pheromones influences men's drinking.

Research has shown that humans consciously use alcohol to encourage sexual activity. In the current study, we investigated whether decision making about alcohol use and sex can be cued outside of awareness by recently revealed sexual signaling mechanisms. Specifically, we examined if males exposed without their knowledge to pheromones emitted by fertile females would increase their alcohol consumption, presumably via neurobehavioral information pathways that link alcohol to sex and mating. We found that men who smelled a T-shirt worn by a fertile female drank significantly more (nonalcoholic) beer, and exhibited significantly greater approach behavior toward female cues, than those who smelled a T-shirt worn by a nonfertile female. These findings reveal previously unknown influences on human alcohol consumption, augment the research base for pheromone cuing of sexual behavior in humans, and raise the possibility that other, as yet unknown, pathways of behavioral influence may be operating hidden from view.

Decisional capacity among youth with HIV: results from the MacArthur Competence Tool for Treatment.

We assessed the decisional capacity (DC) of 72 youth with HIV, ages 13-24, using the MacArthur Competence Tool for Treatment, a structured interview that assesses DC along the following dimensions: understanding, appreciation, reasoning, and the ability to express a choice. Using previously established cutoff scores, results suggested 100% of youth were competent in the area of appreciation, but only 62% and 60% were competent in the areas of understanding and reasoning, respectively. Additional descriptive analyses reveal more detailed information regarding specific strengths and weaknesses within each of the dimensions of decisional capacity. These findings have important implications for health literacy initiatives, medical education, and treatment for youth with HIV, and support the need for adherence and secondary prevention interventions that include a decisional capacity component.

Enhanced expression and purification of membrane proteins by SUMO fusion in Escherichia coli.

Severe acute respiratory syndrome coronavirus (SARS-CoV) membrane protein and 5-lipoxygenase-activating protein (FLAP) are among a large number of membrane proteins that are poorly expressed when traditional expression systems and methods are employed. Therefore to efficiently express difficult membrane proteins, molecular biologists will have to develop novel or innovative expression systems. To this end, we have expressed the SARS-CoV M and FLAP proteins in Escherichia coli by utilizing a novel gene fusion expression system that takes advantage of the natural chaperoning properties of the SUMO (small ubiquitin-related modifier) tag. These chaperoning properties facilitate proper protein folding, which enhances the solubility and biological activity of the purified protein. In addition to these advantages, we found that SUMO Protease 1, can cleave the SUMO fusion high specificity to generate native protein. Herein, we demonstrate that the expression of FLAP and SARS-CoV membrane proteins are greatly enhanced by SUMO fusions in E. coli.

Expression and purification of SARS coronavirus proteins using SUMO-fusions.

Severe acute respiratory syndrome coronavirus (SARS-CoV) proteins belong to a large group of proteins that is difficult to express in traditional expression systems. The ability to express and purify SARS-CoV proteins in large quantities is critical for basic research and for development of pharmaceutical agents. The work reported here demonstrates: (1) fusion of SUMO (small ubiquitin-related modifier), a 100 amino acid polypeptide, to the N-termini of SARS-CoV proteins dramatically enhances expression in Escherichia coli cells and (2) 6x His-tagged SUMO-fusions facilitate rapid purification of the viral proteins on a large scale. We have exploited the natural chaperoning properties of SUMO to develop an expression system suitable for proteins that cannot be expressed by traditional methodologies. A unique feature of the system is the SUMO tag, which enhances expression, facilitates purification, and can be efficiently cleaved by a SUMO-specific protease to generate native protein with a desired N-terminus. We have purified various SARS-CoV proteins under either native or denaturing conditions. These purified proteins have been used to generate highly specific polyclonal antibodies. Our study suggests that the SUMO-fusion technology will be useful for enhancing expression and purification of the viral proteins for structural and functional studies as well as for therapeutic uses.