Characterization of the MSAP Flap in Head and Neck Surgical Oncology: A 3D Cadaveric Study.

OBJECTIVES: The medial sural artery (MSA) perforator flap is a versatile free flap. However, the cutaneous perforators are not well characterized. The objectives of this pilot anatomical study were to: (1) visualize in three-dimensions, as in-situ, the origin, course, and distribution of the cutaneous perforators, (2) characterize the number and frequency of the perforators, and (3) quantify mean pedicle length. METHODS: Thirteen cadaveric specimens were dissected, digitized, and modeled in 3D. Three-dimensional models and dissection photographs were used to determine the origin, course, number, distribution, and pedicle length of MSA perforators. RESULTS: The most common pattern consisted of three perforators (39% of specimens). The maximum number of perforators identified was four (23%). The majority of specimens (92%) had a cutaneous perforator originating from the lateral branch of the MSA and coursed most frequently in the second (43%) and third (37%) quartiles of the length of the tibia. Mean pedicle length was 19.1 ± 6.9 cm. Perforators originating from the medial branch of the MSA were significantly (p < 0.05) shorter than those from the lateral branch and were found to course only in the first quartile. CONCLUSION: The 3D models constructed in this study provide a comprehensive overview of the location and course of the perforators, enabling measurement of parameters in 3D-space. Anatomical characterization of the MSA perforator flap using 3D analysis can assist reconstructive surgeons in understanding the relevant anatomy and optimizing the surgical technique for flap harvest. LEVEL OF EVIDENCE: N/A Laryngoscope, 2024.

Accuracy, readability, and understandability of large language models for prostate cancer information to the public.

BACKGROUND: Generative Pretrained Model (GPT) chatbots have gained popularity since the public release of ChatGPT. Studies have evaluated the ability of different GPT models to provide information about medical conditions. To date, no study has assessed the quality of ChatGPT outputs to prostate cancer related questions from both the physician and public perspective while optimizing outputs for patient consumption. METHODS: Nine prostate cancer-related questions, identified through Google Trends (Global), were categorized into diagnosis, treatment, and postoperative follow-up. These questions were processed using ChatGPT 3.5, and the responses were recorded. Subsequently, these responses were re-inputted into ChatGPT to create simplified summaries understandable at a sixth-grade level. Readability of both the original ChatGPT responses and the layperson summaries was evaluated using validated readability tools. A survey was conducted among urology providers (urologists and urologists in training) to rate the original ChatGPT responses for accuracy, completeness, and clarity using a 5-point Likert scale. Furthermore, two independent reviewers evaluated the layperson summaries on correctness trifecta: accuracy, completeness, and decision-making sufficiency. Public assessment of the simplified summaries' clarity and understandability was carried out through Amazon Mechanical Turk (MTurk). Participants rated the clarity and demonstrated their understanding through a multiple-choice question. RESULTS: GPT-generated output was deemed correct by 71.7% to 94.3% of raters (36 urologists, 17 urology residents) across 9 scenarios. GPT-generated simplified layperson summaries of this output was rated as accurate in 8 of 9 (88.9%) scenarios and sufficient for a patient to make a decision in 8 of 9 (88.9%) scenarios. Mean readability of layperson summaries was higher than original GPT outputs ([original ChatGPT v. simplified ChatGPT, mean (SD), p-value] Flesch Reading Ease: 36.5(9.1) v. 70.2(11.2), <0.0001; Gunning Fog: 15.8(1.7) v. 9.5(2.0), p < 0.0001; Flesch Grade Level: 12.8(1.2) v. 7.4(1.7), p < 0.0001; Coleman Liau: 13.7(2.1) v. 8.6(2.4), 0.0002; Smog index: 11.8(1.2) v. 6.7(1.8), <0.0001; Automated Readability Index: 13.1(1.4) v. 7.5(2.1), p < 0.0001). MTurk workers (n = 514) rated the layperson summaries as correct (89.5-95.7%) and correctly understood the content (63.0-87.4%). CONCLUSION: GPT shows promise for correct patient education for prostate cancer-related contents, but the technology is not designed for delivering patients information. Prompting the model to respond with accuracy, completeness, clarity and readability may enhance its utility when used for GPT-powered medical chatbots.

Parasagittal needle placement approach for lumbar medial branch denervation: a brief technical report.

Radiofrequency denervation of lumbar medial branches is a viable treatment option to manage chronic facetogenic low back pain. Traditionally, lumbar medial branch denervation involves placement of the electrode's active tip at a 20-degree angulation away from the parasagittal plane. However, more recent anatomical studies have provided evidence supporting the feasibility of an alternative parasagittal approach targeting the posterior half of the lateral neck of the superior articular process to capture the lumbar medial branches. Currently, there is a lack of clinical data on the effectiveness of the alternative parasagittal needle placement technique. Therefore, in this brief technical report, the parasagittal needle placement technique and the pain relief outcomes in four consecutive patients following treatment with the parasagittal approach are described.

Quantification of needle angles for lumbar medial branch denervation targeting the posterior half of the superior articular process: an osteological study.

BACKGROUND: Lumbar medial branch radiofrequency ablation (RFA) is a common intervention to manage chronic axial low back pain originating from the facet joints. A more parasagittal approach targeting the posterior half of the lateral neck of superior articular process (SAP) was previously proposed. However, specific needle angles to achieve parallel placement at this target site have not been investigated. OBJECTIVE: To quantify and compare the needle angles, on posterior and lateral views, to achieve parallel placement of electrodes along the medial branch at the posterior half of the lateral neck of SAP at each lumbar vertebral level (L1-L5) and sacrum. DESIGN: Osteological Study. METHODS: Twelve disarticulated lumbosacral spines (n = 72 individual bones) were used in this study. Needles were placed along the periosteum of the posterior half of the lateral neck of SAP, bilaterally and photographed. Mean needle angles for each vertebral level (L1-L5) and sacrum were quantified, and statistical differences were analyzed. RESULTS: The posterior view provided the degrees of lateral displacement from the parasagittal plane (abduction angle), while the lateral view provided the degrees of declination (cranial-to-caudal angle) of the needle. Mean needle angles at each level varied, ranging from 5.63 ± 5.76° to 14.50 ± 14.24° (abduction angle, posterior view) and 40.17 ± 7.32° to 64.10 ± 9.73° (cranial-to-caudal angle, lateral view). In posterior view, a < 10-degree needle angle interval was most frequently identified (57.0% of needle placements). In lateral view, the 40-50-degree (L1-L2), 50-60-degree (L3-L5), and 60-70-degree (sacrum) needle angle intervals occurred most frequently (54.2%, 50.0%, and 41.7% of needle placements, respectively). CONCLUSIONS: Targeting the posterior half of the lateral neck of SAP required <10-degree angulation from parasagittal plane in majority of cases. However, variability of needle angles suggests a standard "one-size-fits-all" approach may not be the optimal technique.

The 3D muscle morphology and intramuscular innervation of the digital bellies of flexor digitorum profundus: Clinical implications for botulinum toxin injection sites.

Spasticity of flexor digitorum profundus is frequently managed with botulinum toxin injections. Knowledge of the 3D morphology and intramuscular innervation of the digital bellies of flexor digitorum profundus is necessary to optimize the injections. The purpose of this study was to digitize and model in 3D the contractile and connective tissue elements of flexor digitorum profundus to determine muscle morphology, model and map the intramuscular innervation and propose sites for botulinum toxin injection. Fiber bundles (FBs)/aponeuroses and intramuscular nerve branches were dissected and digitized in 12 formalin embalmed cadaveric specimens. Cartesian coordinate data were reconstructed into 3D models as in situ to visualize and compare the muscle morphology and intramuscular innervation patterns of the bellies of flexor digitorum profundus. The 3rd, 4th and 5th digital bellies were superficial to the 2nd digital belly and located adjacent to each other in all specimens. Each digital belly had distinct intramuscular innervation patterns. The 2nd digital belly received intramuscular branches from the anterior interosseus nerve (AIN). The superior half of the 3rd digital belly was innervated intramuscularly by the ulnar nerve (n = 4) or by both the anterior interosseus and ulnar nerves (n = 1). The inferior half of the belly received dual innervation from the anterior interosseus and ulnar nerves in 2 specimens, or exclusively from the AIN (n = 2) or the ulnar nerve (n = 1). The 4th digital belly was innervated by intramuscular branches of the ulnar nerve. One main branch, after coursing through the 4th digital belly, entered the lateral aspect of the 5th digital belly and arborized intramuscularly. The morphology of the FBs, aponeuroses and intramuscular innervation of the digital bellies of FDP were mapped and modelled volumetrically in 3D as in situ. Previous studies were not volumetric nor identified the course of the intramuscular nerve branches within each digital belly. Based on the intramuscular innervation of each of the digital bellies, one possible optimized botulinum toxin injection location was proposed. This injection location, at the junction of the superior and middle thirds of the forearm, would be located in dense nerve terminal zones of the anterior interosseus and ulnar nerves. Future anatomical and clinical investigations are necessary to evaluate the efficacy of these anatomical findings in the management of spasticity.

Translatability of In Vitro Stress for Predicting Deamidation and Oxidation Biotransformation on Biotherapeutics.

Biotransformation leading to single residue modifications (e.g., deamidation, oxidation) can contribute to decreased efficacy/potency, poor pharmacokinetics, and/or toxicity/immunogenicity for protein therapeutics. Identifying and characterizing such liabilities in vivo are emerging needs for biologics drug discovery. In vitro stress assays involving PBS for deamidation or AAPH for oxidation are commonly used for predicting liabilities in manufacturing and storage and are sometimes considered a predictive tool for in vivo liabilities. However, reports discussing their in vivo translatability are limited. Herein, we introduce a mass spectrometry workflow that characterizes in vivo oxidation and deamidation in pharmacokinetically relevant compartments for diverse protein therapeutic modalities. The workflow has low bias of <10% in quantitating degradation in the relevant pharmacokinetic concentration range for monkey and rabbit serum/plasma (1-100 µg/mL) and allows for high sequence coverage (∼85%) for discovery/monitoring of amino acid modifications. For oxidation and deamidation, the assay was precise, with percent coefficient of variation of <8% at 1-100 µg/mL and ≤6% method-induced artifacts. A high degree of in vitro and in vivo correlation was observed for deamidation on the six diverse protein therapeutics (seven liability sites) tested. In vivo translatability for oxidation liabilities were not observed for the 11 molecules tested using in vitro AAPH stress. One of the molecules dosed in eyes resulted in a false positive and a false negative prediction for in vivo oxidation following AAPH stress. Finally, peroxide stress was also tested but resulted in limited success (1 out of 4 molecules) in predicting oxidation liabilities.

High-Throughput Analyses of Therapeutic Antibodies Using High-Field Asymmetric Waveform Ion Mobility Spectrometry Combined with SampleStream and Intact Protein Mass Spectrometry.

Intact protein mass spectrometry (MS) coupled with liquid chromatography was applied to characterize the pharmacokinetics and stability profiles of therapeutic proteins. However, limitations from chromatography, including throughput and carryover, result in challenges with handling large sample numbers. Here, we combined intact protein MS with multiple front-end separations, including affinity capture, SampleStream, and high-field asymmetric waveform ion mobility spectrometry (FAIMS), to perform high-throughput and specific mass measurements of a multivalent antibody with one antigen-binding fragment (Fab) fused to an immunoglobulin G1 (IgG1) antibody. Generic affinity capture ensures the retention of both intact species 1Fab-IgG1 and the tentative degradation product IgG1. Subsequently, the analytes were directly loaded into SampleStream, where each injection occurs within ∼30 s. By separating ions prior to MS detection, FAIMS further offered improvement in signal-overnoise by ∼30% for denatured protein MS via employing compensation voltages that were optimized for different antibody species. When enhanced FAIMS transmission of 1Fab-IgG1 was employed, a qualified assay was established for spiked-in serum samples between 0.1 and 25 µg/mL, resulting in ∼10% accuracy bias and precision coefficient of variation. Selective FAIMS transmission of IgG1 as the degradation surrogate product enabled more sensitive detection of clipped species for intact 1Fab-IgG1 at 5 µg/mL in serum, generating an assay to measure 1Fab-IgG1 truncation between 2.5 and 50% with accuracy and precision below 20% bias and coefficient of variation. Our results revealed that the SampleStream-FAIMS-MS platform affords high throughput, selectivity, and sensitivity for characterizing therapeutic antibodies from complex biomatrices qualitatively and quantitatively.

Innervation of thumb carpometacarpal joint: implications for diagnostic block and denervation procedures.

INTRODUCTION: Osteoarthritis (OA) of the thumb carpometacarpal (CMC) joint is a common disorder that negatively impacts hand function. Denervation of the thumb CMC joint has emerged as a viable treatment option. However, the innervation pattern of the thumb CMC joint is controversial. Therefore, the objective of this study was to identify the articular branches supplying the thumb CMC joint and to document their relationship to anatomical landmarks to provide the foundation for image-guided diagnostic block and denervation procedures. METHODS: In 10 formalin-embalmed upper limb specimens articular branches supplying the thumb CMC joint were dissected from their origin to termination. A frequency map documenting the number of articular branches was generated. The frequency map enabled visualization and comparison of the relative area of innervation of the thumb CMC joint by each articular branch. RESULTS: The thumb CMC joint received innervation from six nerves. These were the deep branch of ulnar nerve (DBUN), dorsal articular nerve (DAN) of the first interosseus space, thenar branch of median nerve (TBMN), palmar cutaneous branch of median nerve (PCBMN), lateral antebrachial cutaneous nerve (LACN) and superficial branch of the radial nerve (SBRN) and/or their branches. Each nerve was found to innervate different aspects of the joint. The DBUN and DAN were found to innervate the posteromedial aspect of the thumb CMC joint, the TBMN and PCBMN anterior/anteromedial aspects, LACN posterolateral/lateral/anterior aspects and SBRN posterolateral/anterolateral aspects. CONCLUSIONS: The thumb CMC joint was innervated by articular branches originating from the SBRN, DAN, LACN, PCBMN, TBMN and DBUN. The documented anatomical relationships provide the foundation to inform selective diagnostic block and denervation of the thumb CMC joint. Further investigations are needed to assess the clinical implications of the current study.

Identification of GDC-1971 (RLY-1971), a SHP2 Inhibitor Designed for the Treatment of Solid Tumors.

Protein tyrosine phosphatase SHP2 mediates RAS-driven MAPK signaling and has emerged in recent years as a target of interest in oncology, both for treating with a single agent and in combination with a KRAS inhibitor. We were drawn to the pharmacological potential of SHP2 inhibition, especially following the initial observation that drug-like compounds could bind an allosteric site and enforce a closed, inactive state of the enzyme. Here, we describe the identification and characterization of GDC-1971 (formerly RLY-1971), a SHP2 inhibitor currently in clinical trials in combination with KRAS G12C inhibitor divarasib (GDC-6036) for the treatment of solid tumors driven by a KRAS G12C mutation.

Dissection, digitization, and three-dimensional modelling: a high-fidelity anatomical visualization and imaging technology.

Technological advances have enabled the development of a novel technique of dissection, digitization and three-dimensional modelling of skeletal muscle and other tissues including neurovascular structures as in situ over the last 25 years. Meticulous serial dissection followed by digitization is used to collect Cartesian coordinate data of the contractile and connective tissue elements throughout the entire muscle volume. The Cartesian coordinate can then be used to construct high-fidelity three-dimensional models that capture the spatial arrangement of the contractile and connective tissue elements as in situ enabling detailed studies of the arrangement of the fiber bundles and their attachment sites to aponeuroses, tendon, and bone. In the laboratory, we have concurrently developed a computational methodology to quantify architectural parameters, including fiber bundle length, pennation angle, volume, physiological cross-sectional area in three-dimensional space. In this paper, a flexor digitorum superficialis specimen will be used to demonstrate the high-fidelity outcomes of dissection, digitization, and three-dimensional modelling. This three-step methodology provides a unique opportunity to study muscle architecture in three dimensions, as in situ. Knowledge translation from the anatomy laboratory to the clinical setting has been highly successful.

Quantifying KRAS G12C Covalent Drug Inhibitor Activity in Mouse Tumors Using Mass Spectrometry.

The growing opportunities recognized for covalent drug inhibitors, like KRAS G12C inhibitors, are driving the need for mass spectrometry methods that can quickly and robustly measure therapeutic drug activity in vivo for drug discovery research and development. Effective front-end sample preparation is critical for proteins extracted from tumors but is generally labor intensive and impractical for large sample numbers typical in pharmacodynamic (PD) studies. Herein, we describe an automated and integrated sample preparation method for the measurement of activity levels of KRAS G12C drug inhibitor alkylation from complex tumor samples involving high throughput detergent removal and preconcentration followed by quantitation using mass spectrometry. We introduce a robust assay with an average intra-assay coefficient of variation (CV) of 4% and an interassay CV of 6% obtained from seven studies, enabling us to understand the relationship between KRAS G12C target occupancy and the therapeutic PD effect from mouse tumor samples. Further, the data demonstrated that the drug candidate GDC-6036, a KRAS G12C covalent inhibitor, shows dose-dependent target inhibition (KRAS G12C alkylation) and MAPK pathway inhibition, which correlate with high antitumor potency in the MIA PaCa-2 pancreatic xenograft model.

Intramuscular aponeuroses and fiber bundle morphology of the five bellies of flexor digitorum superficialis: A three-dimensional modeling study.

Restoring balanced function of the five bellies of flexor digitorum superficialis (FDS) following injury requires knowledge of the muscle architecture and the arrangement of the contractile and connective tissue elements. No three-dimensional (3D) studies of FDS architecture were found in the literature. The purpose was to (1) digitize/model in 3D the contractile/connective tissue elements of FDS, (2) quantify/compare architectural parameters of the bellies and (3) assess functional implications. The fiber bundles (FBs)/aponeuroses of the bellies of FDS were dissected and digitized (MicroScribe® Digitizer) in 10 embalmed specimens. Data were used to construct 3D models of FDS to determine/compare the morphology of each digital belly and quantify architectural parameters to assess functional implications. FDS consists of five morphologically and architecturally distinct bellies, a proximal belly, and four digital bellies. FBs of each belly have unique attachment sites to one or more of the three aponeuroses (proximal/distal/median). The proximal belly is connected through the median aponeurosis to the bellies of the second and fifth digits. The third belly exhibited the longest mean FB length (72.84 ± 16.26 mm) and the proximal belly the shortest (30.49 ± 6.45 mm). The third belly also had the greatest mean physiological cross-sectional area, followed by proximal/second/fourth/fifth. Each belly was found to have distinct excursion and force-generating capabilities based on their 3D morphology and architectural parameters. Results of this study provide the basis for the development of in vivo ultrasound protocols to study activation patterns of FDS during functional activities in normal and pathologic states.

Quantification of Needle Angles for Traditional Lumbar Medial Branch Radiofrequency Ablation: An Osteological Study.

BACKGROUND: Clinical outcomes following lumbar medial branch radiofrequency ablation (RFA) have been inconsistent. One possible reason is less-than-optimal placement of the electrode along the medial branch at the lateral neck of superior articular process (SAP). Needle angles that define optimal placement (i.e., parallel to the medial branch) may be helpful for consistent technical performance of RFA. Despite its importance, there is a lack of anatomical studies that quantify RFA needle placement angles. OBJECTIVE: To quantify and compare needle angles to achieve parallel placement along the medial branch as it courses on the middle two-quarters of the lateral neck of the SAP at the L1-L5 vertebrae. DESIGN: Osteological Study. METHODS: Ten lumbar vertebral columns were used in this study. Needles were placed along the periosteum of the middle two-quarters of the lateral neck of SAP. Mean needle angles for L1-L5 were quantified and compared using posterior (n = 100) and lateral (n = 100) photographs. RESULTS: Mean needle angles varied ranging from 29.29 ± 17.82° to 47.22 ± 16.27° lateral to the parasagittal plane (posterior view) and 33.53 ± 10.23° to 49.19 ± 10.69° caudal to the superior vertebral endplate (lateral view). Significant differences in mean angles were found between: L1/L3 (P = .008), L1/L4 (P = .003), and L1/L5 (P = .040) in the posterior view and L1/L3 (P = .042), L1/L4 (P < .001), L1/L5 (P < .001), L2/L4 (P = .004), and L2/L5 (P = .004) in lateral view. CONCLUSIONS: Variability of needle angles suggest a standard "one-size-fits-all" approach may not be the optimal technique. Future research is necessary to determine optimal patient-specific needle angles from a more detailed and granular analysis of fluoroscopic landmarks.

Ischaemic stroke as the first presentation of antineutrophilic cytoplasmic autoantibody-associated vasculitis.

The diagnosis of antineutrophil cytoplasmic autoantibody-associated vasculitis in first-episode strokes is particularly challenging, especially in patients lacking features of systemic vasculitis. We present the case of a 71-year-old woman with positive myeloperoxidase antineutrophil cytoplasmic antibodies and negative proteinase 3 autoantibodies. The patient presented with 1 week history of pyramidal weakness in both upper and lower limbs, hyperreflexia, and clonus. Magnetic resonance imaging of the brain demonstrated widespread bihemispheric cortical and deep white matter acute infarcts, which are consistent with features of stroke secondary to vasculitis. Myeloperoxidase antineutrophilic cytoplasmic autoantibody-positive vasculitis diseases are more commonly associated with renal, pulmonary, and cutaneous manifestations; however, in our patient, the central nervous system features predominated. This case highlights the challenges of diagnosing primary central nervous system vasculitis, in this case, an atypical myeloperoxidase antineutrophilic cytoplasmic autoantibody-positive disease without the classical disease course and clinical signs.

Direct Tie2 Agonists Stabilize Vasculature for the Treatment of Diabetic Macular Edema.

Purpose: Diabetic macular edema (DME) is the leading cause of vision loss and blindness among working-age adults. Although current intravitreal anti-vascular endothelial growth factor (VEGF) therapies improve vision for many patients with DME, approximately half do not achieve the visual acuity required to drive. We therefore sought additional approaches to resolve edema and improve vision for these patients. Methods: We explored direct agonists of Tie2, a receptor known to stabilize vasculature and prevent leakage. We identified a multivalent PEG-Fab conjugate, Tie2.1-hexamer, that oligomerizes Tie2 and drives receptor activation and characterized its activities in vitro and in vivo. Results: Tie2.1-hexamer normalized and stabilized intercellular junctions of stressed endothelial cell monolayers in vitro, suppressed vascular leak in mice under conditions where anti-VEGF alone was ineffective, and demonstrated extended ocular exposure and robust pharmacodynamic responses in non-human primates. Conclusions: Tie2.1-hexamer directly activates the Tie2 pathway, reduces vascular leak, and is persistent within the vitreal humor. Translational Relevance: Our study presents a promising potential therapeutic for the treatment of DME.

Assessment of KRAS G12C Target Engagement by a Covalent Inhibitor in Tumor Biopsies Using an Ultra-Sensitive Immunoaffinity 2D-LC-MS/MS Approach.

KRAS is one of the most frequently mutated oncogenes, with KRAS G12C recently becoming an actionable target for small molecule intervention. GDC-6036 is an investigational KRAS G12C inhibitor that acts by irreversibly binding to the switch II pocket of KRAS G12C when in the inactive GDP-bound state, thereby blocking GTP binding and activation. Assessing target engagement is an essential component of clinical drug development, helping to demonstrate mechanistic activity, guide dose selection, understand pharmacodynamics as it relates to clinical response, and explore resistance. Here, we report the development of an ultra-sensitive approach for assessing KRAS G12C engagement. Immunoaffinity enrichment with a commercially available anti-RAS antibody was combined with a targeted 2D-LC-MS/MS technique to quantify both free and GDC-6036-bound KRAS G12C proteins. A KRAS G12C-positive non-small cell lung cancer xenograft model was dosed with GDC-6036 to assess the feasibility of this assay for analyzing small core needle biopsies. As predicted, dose-dependent KRAS G12C engagement was observed. To date, a sensitivity of 0.08 fmol/µg of total protein has been achieved for both free and GDC-6036-bound KRAS G12C with as little as 4 µg of total protein extracted from human tumor samples. This sub-fmol/µg level of sensitivity provides a powerful potential approach to assess covalent inhibitor target engagement at the site of action using core needle tumor biopsies from clinical studies.