De novo mapping of α-helix recognition sites on protein surfaces using unbiased libraries.

The α-helix is one of the most common protein surface recognition motifs found in nature, and its unique amide-cloaking properties also enable α-helical polypeptide motifs to exist in membranes. Together, these properties have inspired the development of α-helically constrained (Helicon) therapeutics that can enter cells and bind targets that have been considered "undruggable", such as protein-protein interactions. To date, no general method for discovering α-helical binders to proteins has been reported, limiting Helicon drug discovery to only those proteins with previously characterized α-helix recognition sites, and restricting the starting chemical matter to those known α-helical binders. Here, we report a general and rapid screening method to empirically map the α-helix binding sites on a broad range of target proteins in parallel using large, unbiased Helicon phage display libraries and next-generation sequencing. We apply this method to screen six structurally diverse protein domains, only one of which had been previously reported to bind isolated α-helical peptides, discovering 20 families that collectively comprise several hundred individual Helicons. Analysis of 14 X-ray cocrystal structures reveals at least nine distinct α-helix recognition sites across these six proteins, and biochemical and biophysical studies show that these Helicons can block protein-protein interactions, inhibit enzymatic activity, induce conformational rearrangements, and cause protein dimerization. We anticipate that this method will prove broadly useful for the study of protein recognition and for the development of both biochemical tools and therapeutics for traditionally challenging protein targets.

Golf-Related Craniofacial Injuries in Adults: A NEISS Database Study.

INTRODUCTION: Despite its popularity, there have been no reports outlining adult craniofacial trauma in the setting of golf injuries. Our main objective was to identify and describe trends in head and neck injuries incurred while participating in golf including injury type, anatomic location, and patient disposition. METHODS: A retrospective cohort analysis of the National Electronic Injury Surveillance System (NEISS) was conducted from 2009-2018. Population characteristics of golf-related injuries were recorded, including, age, sex, and race distributions. Distribution of injury anatomic location, injury type, distribution of fracture location, patient disposition was further analyzed. A one way ANOVA was utilized to obtain the mean ages for all injuries and compare them for any statistical difference. To identify statistical significance, a Fisher exact test with a Monte Carlo simulation was performed. RESULTS: A total of 509 golf-related injuries (national estimate 24,425 cases) were recorded over the study period. Lacerations were the most common injury overall (54.2%), while contusions and abrasions were the next most common injuries (27.3%). The most common fracture subtype observed was midface (40.43%), followed by nasal bone (27.66%), mandible (12.77%), skull (12.77%), and cervical spine (6.38%). The highest proportion of patients admitted for further treatment were individuals >70 years of age. Fractures had the highest rate of admission (29.8%). CONCLUSIONS: A better understanding of golf injuries can allow for rapid detection and appropriate treatment when encountered. This knowledge can also help to develop safety precautions by potentially reforming rules and regulations as well as protective equipment.

Bridging gaps in heart valve disease care: Opportunities for quality improvement.

In January 2018, the Society of Cardiovascular and Angiography Interventions (SCAI) and Avalere Health ("Avalere") launched the development of the Heart Valve Initiative to advance the identification and management of patients with heart valve disease. To define the priority gap areas in identification and management of valve disease, a formal environmental scan was conducted to uncover current and planned initiatives that might inform the development of the initiative and bring to light stakeholder activities that seek to address the target patient population. This document summarizes the methodology and key findings from the scan, which will inform the development of interventions, research questions, and quality measure concepts to consider. Understanding that quality improvement initiatives and evidence in cardiovascular care may evolve rapidly, this synopsis represents a snapshot of information gathered from February 2018 to May 2019.

Tryptophan Scanning Reveals Dense Packing of Connexin Transmembrane Domains in Gap Junction Channels Composed of Connexin32.

Tryptophan was substituted for residues in all four transmembrane domains of connexin32. Function was assayed using dual cell two-electrode voltage clamp after expression in Xenopus oocytes. Tryptophan substitution was poorly tolerated in all domains, with the greatest impact in TM1 and TM4. For instance, in TM1, 15 substitutions were made, six abolished coupling and five others significantly reduced function. Only TM2 and TM3 included a distinct helical face that lacked sensitivity to tryptophan substitution. Results were visualized on a comparative model of Cx32 hemichannel. In this model, a region midway through the membrane appears highly sensitive to tryptophan substitution and includes residues Arg-32, Ile-33, Met-34, and Val-35. In the modeled channel, pore-facing regions of TM1 and TM2 were highly sensitive to tryptophan substitution, whereas the lipid-facing regions of TM3 and TM4 were variably tolerant. Residues facing a putative intracellular water pocket (the IC pocket) were also highly sensitive to tryptophan substitution. Although future studies will be required to separate trafficking-defective mutants from those that alter channel function, a subset of interactions important for voltage gating was identified. Interactions important for voltage gating occurred mainly in the mid-region of the channel and focused on TM1. To determine whether results could be extrapolated to other connexins, TM1 of Cx43 was scanned revealing similar but not identical sensitivity to TM1 of Cx32.

Brief report: citrullination within the atherosclerotic plaque: a potential target for the anti-citrullinated protein antibody response in rheumatoid arthritis.

OBJECTIVE: To investigate whether citrullinated proteins within the atherosclerotic plaque can be targeted by anti-citrullinated protein antibodies (ACPAs), forming stimulatory immune complexes that propagate the progression of atherosclerosis. METHODS: Protein lysates prepared from atherosclerotic segments of human aorta were assessed for the presence of citrulline-modified proteins, and specifically citrullinated fibrinogen (Cit-fibrinogen), by immunoprecipitation and/or immunoblotting followed by mass spectrometry. Immunohistochemical analysis of coronary artery plaque was performed to determine the presence of citrullinated proteins and peptidylarginine deiminase type 4 (PAD-4). Serum levels of anti-cyclic citrullinated peptide (anti-CCP), anti-citrullinated vimentin (anti-Cit-vimentin), and anti-Cit-fibrinogen antibodies were measured in 134 women with seropositive rheumatoid arthritis; these subjects had previously been characterized for the presence of subclinical atherosclerosis, by electron beam computed tomography scanning. RESULTS: Western blot analysis of atherosclerotic plaque lysates demonstrated several citrullinated proteins, and the presence of Cit-fibrinogen was confirmed by immunoprecipitation and mass spectrometry. Immunohistochemical analysis showed colocalization of citrullinated proteins and PAD-4 within the coronary artery plaque. In age-adjusted regression models, antibodies targeting Cit-fibrinogen and Cit-vimentin, but not CCP-2, were associated with an increased aortic plaque burden. CONCLUSION: Citrullinated proteins are prevalent within atherosclerotic plaques, and certain ACPAs are associated with the atherosclerotic burden. These observations suggest that targeting of citrullinated epitopes, specifically Cit-fibrinogen, within atherosclerotic plaques could provide a mechanism for the accelerated atherosclerosis observed in patients with RA.

Examination of care processes and treatment optimization for head and neck cancer patients in a community setting "hub and hub" model.

Objective: To examine referral pattern, the timing of diagnostic/staging processes, and treatment initiation for new head and neck cancer patients in a community setting. Methods: Patients with a newly diagnosed previously untreated diagnosis of head neck cancer managed at Asplundh Cancer Pavilion/Abington Memorial Hospital from October 2018 to March 2020. Source of referral and preceding workup were examined as well as intervals between initial head and neck consult and various timepoints of treatment initiation. Results: One hundred and five patients were included in the study. The primary referral sources were external general otolaryngology (56.3%). Oral surgery and dermatology obtained tissue biopsy approximately 80% of the time before referral. The average time from the ordering of initial staging positron emission tomography/computed tomography to finalized results was 14 days (range: 10-25 days). Patients referred from dermatology and oral surgery were more likely to require single modality care, namely definitive surgical management. Time to treatment initiation average was 37 days (range: 29-41 days). Patients with longer treatment times noted significantly higher times to both radiation and medical oncology consults (48.42 vs. 18.13 days; P < 0.001). Conclusions: No notable differences in treatment initiation times were identified based on referral source or extent of workup performed before head/neck surgery consult. It appears the largest opportunities for improvement in terms of reducing overall treatment length exist in the optimization of radiation initiation time.

Vocal Fold Paresis and Voice Outcomes following Vocal Fold Mass Excision.

OBJECTIVE: To correlate the surgical results of vocal fold mass excision with pre-operative existence of vocal fold paresis. METHODS: Data were collected on 66 patients who underwent excision of benign vocal fold masses from 2015 to 2020. The pre- and post-operative strobovideolaryngoscopy (SVL) examinations for all patients included were evaluated blindly by three otolaryngologists using THE Voice-Vibratory Assessment with Laryngeal Imaging (VALI) Form for scar severity, mucosal wave, free edge contour, glottal closure, and phase closure. The success of mass excision surgery was determined based on the presence of the following criteria post-operatively: 1) improved mucosal wave motion 2) improved phase closure or glottic closure 3) improved free edge contour and 4) lack of worsening of vocal fold scar severity. Surgery was considered successful if 3 or 4 criteria were met, partially successful if 1 or 2 criteria were met, and unsuccessful if no criteria were met. The percent recruitment of the thyroarytenoid, posterior cricoarytenoid (PCA), and cricothyroid muscles were used evaluated the severity of paresis as mild (70-99% recruitment), moderate (40-60% recruitment), or severe (0-39% recruitment). VHI-10 scores were used as subjective measures of pre- and post-operative voice. RESULTS: Sixty-six patients (26 male, 40 female) were included in this study, with a mean age of 37.25 ± 16.6 (range 18-78). Twelve patients had no evidence of VF paresis noted during the initial clinical evaluation; and 52 patients had paresis and had undergone laryngeal EMG. 81% of these patients had mild paresis, 12.8% had moderate paresis, and 5.8% had severe paresis. Based on pre- and post-operative strobovideolaryngoscopy, there was improvement in mucosal wave in 44.9% of cases, improvement of phase or glottic closure in 85.4% of cases, improved free edge contour in 95.5% of cases, and worsening of scar in 38.5% of cases. 39.6% of surgeries were fully successful, 33.3% of surgeries were partially successful, and 27.1% were not successful. There was a significant correlation between female gender and vocal fold paresis (P = 0.048). Paresis severity did not correlate with complete or partial surgical success (P = 0.956), pre-operative VHI-10 scores (P = 0.519), post-operative VHI-10 scores (P = 0.563), or strobovideolaryngoscopy parameters. Unilateral and bilateral paresis did not correlate with any other parameter of surgical success (P >0.05). CONCLUSION: This study suggests that there is no correlation between pre-operative vocal fold paresis and voice outcomes after mass excision surgery, that the majority of mass excision surgeries (72.9%) are successful based on improvement in stroboscopic parameters, and that the proportion of patients with moderate and severe paresis is consistent across all laryngeal nerves.

Caregiver perception of speech quality in patients with ankyloglossia: Comparison between surgery and non-treatment.

OBJECTIVES: Our objective was to determine if there are differences in caregiver perception of speech for patients that undergo frenulectomy compared to patients that were not surgically treated. INTRODUCTION: Ankyloglossia is a controversial topic without standardized treatment guidelines. Although there have been several studies suggesting that breastfeeding does improve after frenulectomy, there is a paucity of literature that investigates the relationship between ankyloglossia and speech. METHODS: A retrospective chart review was conducted to identify patients with ankyloglossia. Patients were categorized into surgical treatment (frenulectomy) and non-treatment groups. Caregivers of both groups were contacted by phone and surveyed on speech quality and tongue mobility using Likert scores. Mann Whitney-U testing was used to determine if there were significant differences in perceived speech quality between the surgical and non-surgical groups. RESULTS: The caregivers of seventy-seven patients participated in the phone survey: 46 (60%) children in the surgical group and 31 (40%) children in the non-surgical group who participated in the phone survey. There were no differences in difficulty with speech (p = 0.484) and tongue mobility (p = 0.064) between the two groups. However, patients that underwent surgical intervention for ankyloglossia reported less difficulty with tongue tasks (p < 0.001) compared to those who were not surgically treated. Additionally, 50% of patients that underwent surgery had a documented family history of ankyloglossia which was significantly higher than 16.1% in the non-surgical group (p = 0.002). CONCLUSIONS: It appears that children with ankyloglossia might have similar speech quality following frenulectomy in comparison to speech quality without treatment. Children who undergo frenulectomy may experience improvements in tongue tasks. This data should encourage further research on the management of speech concerns in children with ankyloglossia.

Speech and Language Outcomes in Patients with Ankyloglossia Undergoing Frenulectomy: A Retrospective Pilot Study.

Ankyloglossia is a controversial topic with no standardized treatment guidelines. A retrospective chart review was conducted to identify children who underwent lingual frenulectomy for speech and language impairment. Impairment severity was recorded pre- and postoperatively as mild, mild to moderate, moderate, moderate to severe, or severe. Variables were tested with chi-square analysis for their statistical relationship to improvements in speech and language. Children with preoperative moderate and moderate-to-severe speech and language impairment attained better speech and language outcomes after frenulectomy as compared with children with mild and mild-to-moderate impairment (100% vs 82%, P = .015). Sutured closure after frenulectomy was associated with better speech and language improvements (100% vs 83%, P = .033). One could consider observation of patients with mild and mild-to-moderate speech and language impairments. Sutured closure might result in better improvements in speech and language impairments. This pilot study sheds light on the potential impact of a larger study currently underway.