Hexokinase 1 cellular localization regulates the metabolic fate of glucose.

The product of hexokinase (HK) enzymes, glucose-6-phosphate, can be metabolized through glycolysis or directed to alternative metabolic routes, such as the pentose phosphate pathway (PPP) to generate anabolic intermediates. HK1 contains an N-terminal mitochondrial binding domain (MBD), but its physiologic significance remains unclear. To elucidate the effect of HK1 mitochondrial dissociation on cellular metabolism, we generated mice lacking the HK1 MBD (ΔE1HK1). These mice produced a hyper-inflammatory response when challenged with lipopolysaccharide. Additionally, there was decreased glucose flux below the level of GAPDH and increased upstream flux through the PPP. The glycolytic block below GAPDH is mediated by the binding of cytosolic HK1 with S100A8/A9, resulting in GAPDH nitrosylation through iNOS. Additionally, human and mouse macrophages from conditions of low-grade inflammation, such as aging and diabetes, displayed increased cytosolic HK1 and reduced GAPDH activity. Our data indicate that HK1 mitochondrial binding alters glucose metabolism through regulation of GAPDH.

Structural Insights into Pseudokinase Domains of Receptor Tyrosine Kinases.

Despite their apparent lack of catalytic activity, pseudokinases are essential signaling molecules. Here, we describe the structural and dynamic properties of pseudokinase domains from the Wnt-binding receptor tyrosine kinases (PTK7, ROR1, ROR2, and RYK), which play important roles in development. We determined structures of all pseudokinase domains in this family and found that they share a conserved inactive conformation in their activation loop that resembles the autoinhibited insulin receptor kinase (IRK). They also have inaccessible ATP-binding pockets, occluded by aromatic residues that mimic a cofactor-bound state. Structural comparisons revealed significant domain plasticity and alternative interactions that substitute for absent conserved motifs. The pseudokinases also showed dynamic properties that were strikingly similar to those of IRK. Despite the inaccessible ATP site, screening identified ATP-competitive type-II inhibitors for ROR1. Our results set the stage for an emerging therapeutic modality of "conformational disruptors" to inhibit or modulate non-catalytic functions of pseudokinases deregulated in disease.

Systematic review and meta-analysis on the efficacy of Indian polyvalent antivenom against the Indian snakes of clinical significance.

Snakebite in India is a severe problem as it causes a mortality rate of 58,000 and a disability rate of 140,000 every year which is the highest among any other country. Antivenom is the primary therapy for snakebite, and its manufacturing techniques have essentially stayed unaltered for over a century. Indian polyvalent antivenom, a scientifically validated medicine for treating the toxic effects of snakebites, is available against the venom of the so-called Big Four snakes namely Spectacled cobra (Naja naja), Saw-scaled viper (Echis carinatus), Russell's viper (Daboia russelli) and the Common krait (Bungarus caeruleus), responsible for majority of the deaths in India. India hosts many other species of snakes, including cobras, kraits, saw-scaled vipers, sea snakes, and pit vipers, responsible for clinically severe envenomation. Neutralization strategy has been applied to access the efficacy of antivenoms, crucial for reducing snake bite deaths and disabilities. This review aims to conduct a systematic review and meta-analysis on the neutralization efficiency of the Polyvalent Antivenom (PAV) and focus on the factors that may contribute to the poor recognition of the antivenom towards the venom toxins. Reports focusing on the investigation of antivenom efficacy were searched and collected from several databases. Preclinical studies that reported the neutralization efficacy of the commercial antivenom against the medically important snakes of India were included. The articles were screened based on the inclusion criteria and 8 studies were shortlisted for meta-analysis. Pooled proportion was calculated for the antivenom efficacy reported by the studies and was found to be statistically significant with a 95% confidence interval. The heterogenicity in the venom toxicity and neutralization potency of the antivenom was evident in the overall estimate (proportion) and individual data. We provide comprehensive evidence on antivenom efficacy against medically important snakes from various parts of India which may aid in identifying the gaps in snake envenomation therapy and the need for novel potentially improved treatment of snakebites.

Activating mutations drive human MEK1 kinase using a gear-shifting mechanism.

There is an unmet need to classify cancer-promoting kinase mutations in a mechanistically cognizant way. The challenge is to understand how mutations stabilize different kinase configurations to alter function, and how this influences pathogenic potential of the kinase and its responses to therapeutic inhibitors. This goal is made more challenging by the complexity of the mutational landscape of diseases, and is further compounded by the conformational plasticity of each variant where multiple conformations coexist. We focus here on the human MEK1 kinase, a vital component of the RAS/MAPK pathway in which mutations cause cancers and developmental disorders called RASopathies. We sought to explore how these mutations alter the human MEK1 kinase at atomic resolution by utilizing enhanced sampling simulations and free energy calculations. We computationally mapped the different conformational stabilities of individual mutated systems by delineating the free energy landscapes, and showed how this relates directly to experimentally quantified developmental transformation potentials of the mutations. We conclude that mutations leverage variations in the hydrogen bonding network associated with the conformational plasticity to progressively stabilize the active-like conformational state of the kinase while destabilizing the inactive-like state. The mutations alter residue-level internal molecular correlations by differentially prioritizing different conformational states, delineating the various modes of MEK1 activation reminiscent of a gear-shifting mechanism. We define the molecular basis of conversion of this kinase from its inactive to its active state, connecting structure, dynamics, and function by delineating the energy landscape and conformational plasticity, thus augmenting our understanding of MEK1 regulation.

Computational studies of anaplastic lymphoma kinase mutations reveal common mechanisms of oncogenic activation.

Kinases play important roles in diverse cellular processes, including signaling, differentiation, proliferation, and metabolism. They are frequently mutated in cancer and are the targets of a large number of specific inhibitors. Surveys of cancer genome atlases reveal that kinase domains, which consist of 300 amino acids, can harbor numerous (150 to 200) single-point mutations across different patients in the same disease. This preponderance of mutations-some activating, some silent-in a known target protein make clinical decisions for enrolling patients in drug trials challenging since the relevance of the target and its drug sensitivity often depend on the mutational status in a given patient. We show through computational studies using molecular dynamics (MD) as well as enhanced sampling simulations that the experimentally determined activation status of a mutated kinase can be predicted effectively by identifying a hydrogen bonding fingerprint in the activation loop and the αC-helix regions, despite the fact that mutations in cancer patients occur throughout the kinase domain. In our study, we find that the predictive power of MD is superior to a purely data-driven machine learning model involving biochemical features that we implemented, even though MD utilized far fewer features (in fact, just one) in an unsupervised setting. Moreover, the MD results provide key insights into convergent mechanisms of activation, primarily involving differential stabilization of a hydrogen bond network that engages residues of the activation loop and αC-helix in the active-like conformation (in >70% of the mutations studied, regardless of the location of the mutation).

Adolescent Idiopathic Scoliosis: Increased Body Mass Associated with Decreased Bracing Outcomes.

Bracing reduces the need for surgical intervention in patients with adolescent idiopathic scoliosis (AIS). However, bracing outcomes with variable body mass index (BMI) are understudied. The authors sought to determine the association of BMI with bracing outcomes. The authors performed a retrospective cohort study of 104 patients presenting with AIS. Initial Risser score, hours of bracing per day, BMI percentile, and curve magnitude pre- and postbracing were collected. There was no detectable difference between years of brace wear or primary curve magnitude at time of presentation between both groups. Overall, 29% (25/87) of underweight/normal weight patients and 59% (10/17) of overweight/obese patients had curves ≥ 45 degrees at the end of bracing (p = 0.016). Odds of having a curve ≥ 45 degrees after bracing were 3.5 (95% confidence interval: 1.2 to 10.3, p = 0.021) times higher for overweight/obese patients compared with underweight/normal weight patients. Increased overlying adipose tissue may reduce the corrective forces required to straighten the spine. (Journal of Surgical Orthopaedic Advances 33(1):029-032, 2024).

Compression Screw Fixation for Pediatric Olecranon Fractures.

BACKGROUND: Pediatric olecranon fractures can be treated with several methods of fixation. Though postoperative outcomes of various fixation techniques, including cannulated intramedullary screws, have been described in adults, functional and radiographic outcomes of screw fixation in pediatric patients are unclear. In this study, we assessed clinical, radiographic, functional, and patient-reported outcomes of pediatric olecranon fractures treated with compression screw fixation. METHODS: We retrospectively identified 37 patients aged 16 years or younger with a total of 40 olecranon fractures treated with screw fixation at our level-1 trauma center between April 2005 and April 2022. From medical records, we extracted data on demographic characteristics, time to radiographic union, range of elbow motion at final follow-up, and complications during the follow-up period. Patient-reported outcomes were evaluated using the Quick Disabilities of the Arm, Shoulder, and Hand and Patient-Reported Outcomes Measurement Information System Pediatric Upper Extremity Short Form 8a measures. RESULTS: There were no malunions or nonunions at the final mean follow-up of 140 days (range, 26 to 614 d). Four patients had implant failure (11%), of whom 3 experienced fracture union with no loss of fixation or need for revision surgery. One patient underwent a revision for fracture malreduction. Screw prominence was documented in 1 patient. Instrumentation was removed at our institution for 33 of 40 fractures. Mean time to radiographic union was 53 days (range, 20 to 168 d). Postoperative range of motion at the most recent follow-up visit showed a mean extension deficit of 6 degrees (range, 0-30 degrees) and mean flexion of 134 degrees (range, 60-150 degrees). At the final follow-up, the mean (±SD) Quick Disabilities of the Arm, Shoulder, and Hand score was 4.2±8.0, and the mean Patient-Reported Outcomes Measurement Information System score was 37±1.5, indicating good function and patient satisfaction. CONCLUSIONS: All 37 patients in our series had excellent radiographic, functional, and patient-reported outcomes after screw fixation. We observed no cases of nonunion or malunion, growth disturbance, or refracture. These results suggest that screw fixation is a safe and effective option for pediatric olecranon fractures. LEVEL OF EVIDENCE: Level IV, case series.

Consensus-Based Best Practice Guidelines for the Management of Spinal Deformity and Associated Tumors in Pediatric Neurofibromatosis Type 1: Screening and Surveillance, Surgical Intervention, and Medical Therapy.

BACKGROUND: Spinal conditions, such as scoliosis and spinal tumors, are prevalent in neurofibromatosis type 1 (NF1). Despite the recognized importance of their early detection and treatment, there remain knowledge gaps in how to approach these manifestations. The purpose of this study was to utilize the experience of a multidisciplinary committee of experts to establish consensus-based best practice guidelines (BPGs) for spinal screening and surveillance, surgical intervention, and medical therapy in pediatric patients with NF1. METHODS: Using the results of a prior systematic review, 10 key questions that required further assessment were first identified. A committee of 20 experts across medical specialties was then chosen based on their clinical experience with spinal deformity and tumors in NF1. These were 9 orthopaedic surgeons, 4 neuro-oncologists/oncologists, 3 neurosurgeons, 2 neurologists, 1 pulmonologist, and 1 clinical geneticist. An initial online survey on current practices and opinions was conducted, followed by 2 additional surveys via a formal consensus-based modified Delphi method. The final survey involved voting on agreement or disagreement with 35 recommendations. Items reaching consensus (≥70% agreement or disagreement) were included in the final BPGs. RESULTS: Consensus was reached for 30 total recommendations on the management of spinal deformity and tumors in NF1. These were 11 recommendations on screening and surveillance, 16 on surgical intervention, and 3 on medical therapy. Five recommendations did not achieve consensus and were excluded from the BPGs. CONCLUSION: We present a set of consensus-based BPGs comprised of 30 recommendations for spinal screening and surveillance, surgical intervention, and medical therapy in pediatric NF1.

Rate of Tibiotalocalcaneal (TTC) Fusion Using the Surgical Implant Generation Network (SIGN) Intramedullary Nail in Developing Countries.

Outcomes of the Surgical Implant Generation Network (SIGN) nail have been reported for femur and tibial fractures, but its use in tibiotalocalcaneal arthrodesis (TTCA) is not well studied. Radiographic and clinical outcomes of TTCA using the SIGN database in patients with > 6 months of radiographic follow up were analyzed. Rates of tibiotalar (TT) fusion and subtalar (ST) fusion at final follow up were assessed by two independent reviewers. Of the 62 patients identified, use of the SIGN nail for TCCA resulted in 53% rate of fusion in the TT joint and 20% in the ST joint. Thirty-seven patients (60%) demonstrated painless weight bearing at final follow up. There were no differences in incidence of painless weight bearing between consensus fused and not fused cohorts for TT and ST joints (p > 0.05). There were five implant failures, no cases of infection, and seven cases of reoperation. (Journal of Surgical Orthopaedic Advances 32(3):187-192, 2023).

An interdomain helix in IRE1α mediates the conformational change required for the sensor's activation.

The unfolded protein response plays an evolutionarily conserved role in homeostasis, and its dysregulation often leads to human disease, including diabetes and cancer. IRE1α is a major transducer that conveys endoplasmic reticulum stress via biochemical signals, yet major gaps persist in our understanding of how the detection of stress is converted to one of several molecular outcomes. It is known that, upon sensing unfolded proteins via its endoplasmic reticulum luminal domain, IRE1α dimerizes and then oligomerizes (often visualized as clustering). Once assembled, the kinase domain trans-autophosphorylates a neighboring IRE1α, inducing a conformational change that activates the RNase effector domain. However, the full details of how the signal is transmitted are not known. Here, we describe a previously unrecognized role for helix αK, located between the kinase and RNase domains of IRE1α, in conveying this critical conformational change. Using constructs containing mutations within this interdomain helix, we show that distinct substitutions affect oligomerization, kinase activity, and the RNase activity of IRE1α differentially. Furthermore, using both biochemical and computational methods, we found that different residues at position 827 specify distinct conformations at distal sites of the protein, such as in the RNase domain. Of importance, an RNase-inactive mutant, L827P, can still dimerize with wildtype monomers, but this mutation inactivates the wildtype molecule and renders leukemic cells more susceptible to stress. We surmise that helix αK is a conduit for the activation of IRE1α in response to stress.

Sacral-Alar-Iliac (SAI) Fixation in Children With Spine Deformity: Minimum 10-Year Follow-Up.

INTRODUCTION: Sacral-alar-iliac (SAI) screws are utilized to achieve pelvic fixation in spine deformity patients. The primary purpose of this study is to investigate the long-term outcomes of pediatric patients with scoliosis treated with posterior spinal fusion and SAI fixation at 10-year clinical and radiographic follow-up. METHODS: We reviewed the clinical and radiographic records of patients aged 18 years or below treated for scoliosis with posterior spinal fusion using SAI fixation. Pelvic obliquity and the major coronal curve were determined at the preoperative visit and 6-week, 1-year, 5-year, and 10-year postoperative visits. SAI screw-specific data collected included screw dimensions, rate of screw revision, pain at the SAI screw sites, presence of lucency >2 mm around the screw, screw loosening or breaking, and deep surgical site infections. RESULTS: Ninety-seven of 151 patients (75%) were included. The average age at index surgery was 13.5±3.1 years, and the most common diagnosis was cerebral palsy (67%). The mean duration of follow-up was 11±3 years. The mean pelvic obliquity measured 20±8.0 degrees preoperatively, and 8.7±4.0 degrees at the 10-year follow-up. There were no significant difference in pelvic obliquity when comparing the 10-year follow-up visit with the 6-week postoperative follow-up. Average screw dimensions were 8.4×68.8 mm. By the 10-year follow-up, 4 patients (4%) had at least 1 SAI screw-related complication. Of these patients, 2 (2%) had pain at 1 SAI screw, 4 (4%) had lucency around the screw, and 3 (3%) had broken or loose screws. Two (2%) required SAI screw revision because of late deep wound infection, and underwent exchange with a longer screw. There were no intrapelvic protrusions, vascular, or neurological complications. CONCLUSIONS: SAI screws are a safe and effective method for pelvic fixation in children with spinal deformity. The outcomes at ≥10 years are satisfactory, with low rates of long-term complications and excellent postoperative correction and subsequent maintenance of coronal curvature and pelvic obliquity over time. LEVEL OF EVIDENCE: Level IV.

Pelvic Osteotomy in Patients With Previous Sacral-Alar-Iliac (SAI) Fixation.

INTRODUCTION: Patients with neuromuscular disease are at high risk for developing hip dysplasia and scoliosis. The purpose of this study was to investigate the technical challenges and outcomes of pelvic osteotomy in patients with prior sacral-alar-iliac (SAI) fixation. METHODS: We reviewed clinical and radiographic records of patients aged 18 years and below who underwent pelvic osteotomy after SAI fixation. We recorded technical challenges during the osteotomy, time from SAI fixation to osteotomy, type of osteotomy, migration index, and distance from the SAI screw to the acetabulum. A 2-sample Wilcoxon rank-sum test was used to assess the data. RESULTS: Nineteen patients were included. Technical challenges were defined as having greater intraoperative fluoroscopy times and noted difficult osteotomy in the operative report. The mean time from SAI fixation to pelvic osteotomy was 2.2±1.5 years. For all 12 Chiari osteotomies, the ilium could not be laterally displaced; however, medial displacement of the distal segment of the osteotomy allowed adequate coverage. All 7 Dega osteotomies were performed by cutting the cortex at the tip of the SAI screw. The screw improved proximal leverage and provided a strong buttress for bone graft. The mean migration index before pelvic osteotomy was 59±19%, and at most recent follow-up was 13±4%. Twelve patients, who had a noted complicated osteotomy, had SAI screws that were ≤1.87 cm ( P <0.01) from the acetabulum and significantly increased intraoperative fluoroscopy time (1.76 vs. 1.18 min, P <0.01). CONCLUSIONS: The presence of SAI screws may cause iliac osteotomies to be technically challenging if the tip of the SAI screw is ≤1.87 cm to the acetabulum. When initially implanting SAI screws in neuromuscular patients, surgeons should attempt to place screw tips ∼2 cm from the acetabulum in the event these patients require subsequent pelvic osteotomy. LEVEL OF EVIDENCE: Level IV.

Connective Tissue Disease Patients Maintain Greater Distraction During Growth Friendly Surgery.

STUDY DESIGN: Multicenter retrospective study. BACKGROUND: Recent studies have demonstrated diminishing returns in patients with early onset scoliosis (EOS) undergoing repeated lengthening of growing rods. Little is known about whether this same phenomenon occurs in patients with lax connective tissue disease (CTD). The primary purpose of this study is to investigate whether EOS patients with connective tissue laxity disorders have diminishing returns during growth friendly surgery. METHODS: CTD EOS patients below 10 years old, underwent growth friendly spine surgery with distal anchors and at least 1 proximal spine anchor, and had minimum follow-up of 5 years were included in this study. Coronal T1-S1 height at preindex surgery, postindex, and every available lengthening was assessed. Mean coronal height change during early set distractions and late set distractions were calculated for the cohort. To account for varying distraction intervals, we normalized the distractions by the time interval. The outcome parameter was T1-S1 height gain, mm/year. RESULTS: Twenty-one CTD patients were included in this study. Total coronal height (T1-S1) was 26.7MHCcm before index, 32.2 cm at D1-D3, 34.7 cm at D4-D6, and 36.7 cm at D7-L10. There were no significant differences in coronal height gains between early and late distractions (P=0.70). Moreover, when normalized for time, there was no significant difference in net gain per year at different lengthening time points for the CTD group, P=0.59. CONCLUSION: There is no evidence of diminishing returns in coronal T1-S1 height gain in patients with EOS in the setting of CTD. LEVEL OF EVIDENCE: Level III.

Minimizing the Risk of Vascular Injury: Anatomy of the Popliteal Artery in the Pediatric Knee.

BACKGROUND: Displaced pediatric tibial tubercle fractures are commonly stabilized with screws directed posteriorly toward neurovascular structures. Here, we (1) characterize the variation of the popliteal artery among pediatric patients; and (2) recommend a safe screw trajectory for fixation of tibial tubercle fractures. METHODS: We retrospectively identified 42 patients (42 knees; 29 female) aged 12-17 years with lower-extremity magnetic resonance imaging (MRI) at a tertiary academic center. The mean patient age was 14.5 (range: 12-17) years, and the mean body mass index value was 19.1 (range: 14.9-25.1). We included patients with open physes or visible physeal scars and excluded those with prior instrumentation or lower-extremity injury. Using sagittal MRI, we measured the distances from 5 levels each on the anterior and posterior tibial cortex to the popliteal artery (level 1, midpoint of proximal tibial epiphysis; level 2, the proximal extent of the tubercle; level 3, tubercle prominence; level 4, 2 cm distal to the proximal extent of the tubercle; level 5, 4 cm distal to the proximal extent of the tubercle). Using coronal MRI, we measured the width of the tibia at each level and the distance from the lateral-most and medial-most cortex to the artery. RESULTS: The popliteal artery was laterally positioned in all knees. The mean distance between the artery and lateral-most aspect of the tibia at each level ranged from 1.9 to 2.4 cm, and from 2.3 to 3.9 cm from the medial-most aspect of the tibia. The mean distance that a screw can advance before vascular injury was 5.1 cm at level 1. The shortest mean distance to the popliteal artery was 1.7 cm, at level 5. There is minimal distance between the posterior tibial cortex and the artery at all levels. CONCLUSIONS: Understanding the position of the popliteal artery in pediatric patients can help when stabilizing tibial tubercle fractures. Because the artery is close to the posterior cortex, a drill exiting in line with the popliteal artery risks vascular injury. Therefore, we recommend that screws exit within the medial 60% of the tibia. LEVEL OF EVIDENCE: IV.

Discovery of 2-amino-3-amido-5-aryl-pyridines as highly potent, orally bioavailable, and efficacious PERK kinase inhibitors.

The protein kinase R (PKR)-like endoplasmic reticulum kinase (PERK) is one of the three endoplasmic reticulum (ER) transmembrane sensors of the unfolded protein response (UPR) that regulates protein synthesis, alleviates cellular ER stress and has been implicated in tumorigenesis and prolonged cancer cell survival. In this study, we report a series of 2-amino-3-amido-5-aryl-pyridines that we have identified as potent, selective, and orally bioavailable PERK inhibitors. Amongst the series studied herein, compound (28) a (R)-2-Amino-5-(4-(2-(3,5-difluorophenyl)-2-hydroxyacetamido)-2-ethylphenyl)-N-isopropylnicotinamide has demonstrated potent biochemical and cellular activity, robust pharmacokinetics and 70% oral bioavailability in mice. Given these data, this compound (28) was studied in the 786-O renal cell carcinoma xenograft model. We observed dose-dependent, statistically significant tumor growth inhibition, supporting the use of this tool compound in additional mechanistic studies.

Interlocking Fixation in Fassier-Duval Rods: Performance and Success Factors.

INTRODUCTION: Pediatric patients with osteogenesis imperfecta (OI) can be treated with intramedullary Fassier-Duval rod (FDR) systems for limb deformity or recurrent fractures. Single-interlocking pins can improve epiphyseal fixation, but there is a paucity of literature examining incidence of rod migration or pin backout long-term. The purpose of this study is to quantify rates of rod migration and pin backout in OI patients treated with single-interlocking FDRs. METHODS: A retrospective chart review was performed on pediatric patients treated at a tertiary care center across a 15-year period. Inclusion criteria to select patients was: (1) Pediatric patients (below 18 y of age); (2) Patients with confirmed OI; and (3) Patients with lower extremity fractures or deformity treated with FDRs with distal interlocking pins. Age at time of surgery, rates of obturator migration and pin backout and prominence were collected. We recorded if pin tips were bent by the surgeon during the procedure. Bivariate statistics were used to analyze risk factors for pin backout and prominence. RESULTS: Twenty-four single-interlocking pin FDRs (21 tibia, 3 femur) were identified. The mean age at index surgery was 5.7±3.4 years, with the mean follow-up time of 7.2±4.7 years. Fourteen (58%) rods underwent revision surgery. Obturator proximal migration was observed in 3/24 rods (13%). No cases of obturator distal migration were observed (0/24, 0%). Mean proximal obturator migration was 2.16±1.8 cm. Revision for pin backout was observed in 10 (42%) rods and pin prominence in 11 (46%) extremities. Bending interlocking pins on at least 1 end was associated with decreased pin backout (P=0.01) and prominence (P=0.04). CONCLUSIONS: Even with distal interlocking pins, the obturator of FDRs can still migrate over time. Pin backout is a common indication for revision surgery. Bending interlocking pins can decrease rates of pin backout and prominence. LEVEL OF EVIDENCE: Level III.

Computational algorithms for in silico profiling of activating mutations in cancer.

Methods to catalog and computationally assess the mutational landscape of proteins in human cancers are desirable. One approach is to adapt evolutionary or data-driven methods developed for predicting whether a single-nucleotide polymorphism (SNP) is deleterious to protein structure and function. In cases where understanding the mechanism of protein activation and regulation is desired, an alternative approach is to employ structure-based computational approaches to predict the effects of point mutations. Through a case study of mutations in kinase domains of three proteins, namely, the anaplastic lymphoma kinase (ALK) in pediatric neuroblastoma patients, serine/threonine-protein kinase B-Raf (BRAF) in melanoma patients, and erythroblastic oncogene B 2 (ErbB2 or HER2) in breast cancer patients, we compare the two approaches above. We find that the structure-based method is most appropriate for developing a binary classification of several different mutations, especially infrequently occurring ones, concerning the activation status of the given target protein. This approach is especially useful if the effects of mutations on the interactions of inhibitors with the target proteins are being sought. However, many patients will present with mutations spread across different target proteins, making structure-based models computationally demanding to implement and execute. In this situation, data-driven methods-including those based on machine learning techniques and evolutionary methods-are most appropriate for recognizing and illuminate mutational patterns. We show, however, that, in the present status of the field, the two methods have very different accuracies and confidence values, and hence, the optimal choice of their deployment is context-dependent.

Renal phenotype in mice lacking the Kir5.1 (Kcnj16) K+ channel subunit contrasts with that observed in SeSAME/EAST syndrome.

The heteromeric inwardly rectifying Kir4.1/Kir5.1 K(+) channel underlies the basolateral K(+) conductance in the distal nephron and is extremely sensitive to inhibition by intracellular pH. The functional importance of Kir4.1/Kir5.1 in renal ion transport has recently been highlighted by mutations in the human Kir4.1 gene (KCNJ10) that result in seizures, sensorineural deafness, ataxia, mental retardation, and electrolyte imbalance (SeSAME)/epilepsy, ataxia, sensorineural deafness, and renal tubulopathy (EAST) syndrome, a complex disorder that includes salt wasting and hypokalemic alkalosis. Here, we investigated the role of the Kir5.1 subunit in mice with a targeted disruption of the Kir5.1 gene (Kcnj16). The Kir5.1(-/-) mice displayed hypokalemic, hyperchloremic metabolic acidosis with hypercalciuria. The short-term responses to hydrochlorothiazide, an inhibitor of ion transport in the distal convoluted tubule (DCT), were also exaggerated, indicating excessive renal Na(+) absorption in this segment. Furthermore, chronic treatment with hydrochlorothiazide normalized urinary excretion of Na(+) and Ca(2+), and abolished acidosis in Kir5.1(-/-) mice. Finally, in contrast to WT mice, electrophysiological recording of K(+) channels in the DCT basolateral membrane of Kir5.1(-/-) mice revealed that, even though Kir5.1 is absent, there is an increased K(+) conductance caused by the decreased pH sensitivity of the remaining homomeric Kir4.1 channels. In conclusion, disruption of Kcnj16 induces a severe renal phenotype that, apart from hypokalemia, is the opposite of the phenotype seen in SeSAME/EAST syndrome. These results highlight the important role that Kir5.1 plays as a pH-sensitive regulator of salt transport in the DCT, and the implication of these results for the correct genetic diagnosis of renal tubulopathies is discussed.

Preference Signaling Survey of Program Directors-After the Match.

INTRODUCTION: The 2022 to 2023 orthopaedic residency cycle implemented a preference signaling program (PSP), allowing applicants to send "signals" to up to 30 programs to demonstrate their genuine interest. With the conclusion of the 2022 to 2023 cycle, the primary purpose of this study was to analyze program director (PD) perceptions of the PSP after the match cycle and provide a retrospective evaluation of the effects of the PSP on the orthopaedic resident selection process. METHODS: A 21-question survey was distributed to 98 PDs (32.7% response rate). Contact information was obtained from a national database. RESULTS: Most respondents (96.9%) participated in the American Orthopaedic Association's PSP. The majority (93.7%) view preference signaling as a positive change. Most PDs (56.2%) reported a decreased number in applications received compared with previous years. Receiving a preference signal was ranked among the most important factors in resident selection, and most PDs agreed that preference signaling should be used to screen applicants (84.4%) and differentiate similar applicants (96.8%). Moreover, 65.6% of PDs indicated that they would not rank or invite applicants to interview without a signal or completion of a formal away rotation. PDs report that in the 2022 to 2023 cycle, 98.5% of applicants who matched at their program had sent a preference signal. DISCUSSION: Preference signaling was one of the most important factors assessed during its inaugural application cycle and is anticipated to remain a key tool for screening and differentiating candidates. Applicants should strategically select signal recipients to enhance their success in the match.

Single Stage Combined Approach Sagittal En-Block Spondylectomy for L3 Vertebral Chondrosarcoma: A Technical Note.

BACKGROUND: Primary malignant tumors of the spine are rare which most commonly occur in lumbar and thoracic vertebra. Here, we report a rare case of retroperitoneal chondrosarcoma of the L3 vertebra which was managed with sagittal en-block spondylectomy following chemoradiation. CASE PRESENTATION: A 26-year-old lady was evaluated for abdominal pain with contrast enhanced computer tomogram of abdomen and pelvis which revealed a soft tissue retroperitoneal mass arising from L3 vertebra. She underwent laparotomy and biopsy which revealed chondrosarcoma and she received chemoradiation over a period of 28 weeks, 6 days. After re-imaging she underwent single stage combined approach sagittal en-block spondylectomy of retroperitoneal chondrosarcoma of L3 vertebra with right nephrectomy and spine reconstruction. She was followed for a period of 3 years, there was no evidence of recurrence in follow-up CECT abdomen and pelvis. She has no gait abnormality or spinal deformity. CONCLUSION: Sagittal en-block spondylectomy is a preferred surgical approach for eccentrically placed spinal tumors which offers better oncological and functional outcomes.