Publisher Correction: Modular and tunable biological feedback control using a de novo protein switch.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Modular and tunable biological feedback control using a de novo protein switch.

De novo-designed proteins1-3 hold great promise as building blocks for synthetic circuits, and can complement the use of engineered variants of natural proteins4-7. One such designer protein-degronLOCKR, which is based on 'latching orthogonal cage-key proteins' (LOCKR) technology8-is a switch that degrades a protein of interest in vivo upon induction by a genetically encoded small peptide. Here we leverage the plug-and-play nature of degronLOCKR to implement feedback control of endogenous signalling pathways and synthetic gene circuits. We first generate synthetic negative and positive feedback in the yeast mating pathway by fusing degronLOCKR to endogenous signalling molecules, illustrating the ease with which this strategy can be used to rewire complex endogenous pathways. We next evaluate feedback control mediated by degronLOCKR on a synthetic gene circuit9, to quantify the feedback capabilities and operational range of the feedback control circuit. The designed nature of degronLOCKR proteins enables simple and rational modifications to tune feedback behaviour in both the synthetic circuit and the mating pathway. The ability to engineer feedback control into living cells represents an important milestone in achieving the full potential of synthetic biology10,11,12. More broadly, this work demonstrates the large and untapped potential of de novo design of proteins for generating tools that implement complex synthetic functionalities in cells for biotechnological and therapeutic applications.

De novo design of bioactive protein switches.

Allosteric regulation of protein function is widespread in biology, but is challenging for de novo protein design as it requires the explicit design of multiple states with comparable free energies. Here we explore the possibility of designing switchable protein systems de novo, through the modulation of competing inter- and intramolecular interactions. We design a static, five-helix 'cage' with a single interface that can interact either intramolecularly with a terminal 'latch' helix or intermolecularly with a peptide 'key'. Encoded on the latch are functional motifs for binding, degradation or nuclear export that function only when the key displaces the latch from the cage. We describe orthogonal cage-key systems that function in vitro, in yeast and in mammalian cells with up to 40-fold activation of function by key. The ability to design switchable protein functions that are controlled by induced conformational change is a milestone for de novo protein design, and opens up new avenues for synthetic biology and cell engineering.

IFT80 negatively regulates osteoclast differentiation via association with Cbl-b to disrupt TRAF6 stabilization and activation.

Excess bone loss due to increased osteoclastogenesis is a significant clinical problem. Intraflagellar transport (IFT) proteins have been reported to regulate cell growth and differentiation. The role of IFT80, an IFT complex B protein, in osteoclasts (OCs) is completely unknown. Here, we demonstrate that deletion of IFT80 in the myeloid lineage led to increased OC formation and activity accompanied by severe bone loss in mice. IFT80 regulated OC formation by associating with Casitas B-lineage lymphoma proto-oncogene-b (Cbl-b) to promote protein stabilization and proteasomal degradation of tumor necrosis factor (TNF) receptor-associated factor 6 (TRAF6). IFT80 knockdown resulted in increased ubiquitination of Cbl-b and higher TRAF6 levels, thereby hyperactivating the receptor activator of nuclear factor-κß (NF-κß) ligand (RANKL) signaling axis and increased OC formation. Ectopic overexpression of IFT80 rescued osteolysis in a calvarial model of bone loss. We have thus identified a negative function of IFT80 in OCs.

Risk of ICU Admission and Related Mortality in Patients With Sodium-Glucose Cotransporter 2 Inhibitors and Dipeptidyl Peptidase-4 Inhibitors: A Territory-Wide Retrospective Cohort Study.

OBJECTIVES: The benefit of sodium-glucose cotransporter 2 (SGLT2) inhibitors in reducing the occurrence rate of adverse cardiac and renal outcomes in patients with type 2 diabetes has been well described in randomized trials. Whether this benefit extends to patients at the most severe end of the disease spectrum requiring admission to the ICU remains to be examined. DESIGN: Retrospective observational study. SETTING: Data were obtained from a territory-wide clinical registry in Hong Kong (Clinical Data Analysis and Reporting System). PATIENTS: All adult patients (age ≥ 18 yr) with type 2 diabetes and newly prescribed SGLT2 inhibitors or dipeptidyl peptidase-4 (DPP-4) inhibitors between January 1, 2015, and December 31, 2019. INTERVENTIONS: None. MEASUREMENTS AND MAIN RESULTS: After 1:2 propensity score matching, a total of 27,972 patients (10,308 SGLT2 inhibitors vs 17,664 DPP-4 inhibitors) were included in the final analysis. The mean age was 59 ± 11 years, and 17,416 (62.3%) were male. The median follow-up period was 2.9 years. The use of SGLT2 inhibitors was associated with decreased ICU admission (286 [2.8%] vs 645 [3.7%]; hazard ratio [HR], 0.79; 95% CI, 0.69-0.91; p = 0.001) and lower risks of all-cause mortality (315 [3.1%] vs 1,327 [7.5%]; HR, 0.44; 95% CI, 0.38-0.49; p < 0.001), compared with DPP-4 inhibitors. The severity of illness upon ICU admission by Acute Physiology and Chronic Health Evaluation IV-predicted risk of death was also lower in SGLT2 inhibitors users. Admissions and mortality due to sepsis were lower in SGLT2 inhibitor users compared with DPP-4 inhibitor users (admissions for sepsis: 45 [0.4%] vs 134 [0.8%]; p = 0.001 and mortality: 59 [0.6%] vs 414 [2.3%]; p < 0.001, respectively). CONCLUSIONS: In patients with type 2 diabetes, SGLT2 inhibitors were independently associated with lower rates of ICU admission and all-cause mortality across various disease categories.

Analysis of localized cAMP perturbations within a tissue reveal the effects of a local, dynamic gap junction state on ERK signaling.

Beyond natural stimuli such as growth factors and stresses, the ability to experimentally modulate at will the levels or activity of specific intracellular signaling molecule(s) in specified cells within a tissue can be a powerful tool for uncovering new regulation and tissue behaviors. Here we perturb the levels of cAMP within specific cells of an epithelial monolayer to probe the time-dynamic behavior of cell-cell communication protocols implemented by the cAMP/PKA pathway and its coupling to the ERK pathway. The time-dependent ERK responses we observe in the perturbed cells for spatially uniform cAMP perturbations (all cells) can be very different from those due to spatially localized perturbations (a few cells). Through a combination of pharmacological and genetic perturbations, signal analysis, and computational modeling, we infer how intracellular regulation and regulated cell-cell coupling each impact the intracellular ERK response in single cells. Our approach reveals how a dynamic gap junction state helps sculpt the intracellular ERK response over time in locally perturbed cells.

Australasian Sleep Association position statement on consensus and evidence based treatment for primary snoring.

Primary snoring impacts a significant portion of the adult population and has the potential to significantly impair quality of life. The purpose of these guidelines is to provide evidence-based recommendations to assist Australasian practitioners in the management of adult patients who present with primary snoring without significant obstructive sleep apnoea. The Timetable, Methodology and Standards by which this Position Statement has been established is outlined in the Appendix S1. The main recommendations are: Weight loss, and reduced alcohol consumption should be recommended, where appropriate If clinical judgement dictates, benzodiazepine and opioid reduction or avoidance may be advised Positional therapy should be considered in supine dominant snorers In dentate patients, Mandibular advancement devices (MAD) should be recommended as a first line treatment following assessment by both an appropriate Dentist and Sleep physician Continuous positive airway pressure (CPAP) devices may be recommended in patients with primary snoring in those already committed to their use or willing to try Surgical treatment of primary snoring by an appropriately credentialled surgeon may be advised and includes nasal (adjunctive), palatal and other interventions This position statement has been designed based on the best available current evidence and our combined expert clinical experience to facilitate the management of patients who present with primary snoring. It provides clinicians with a series of both non-surgical and surgical options with the aim of achieving optimal symptom control and patient outcomes. This is the first such set of recommendations to be established within Australasia and has also been reviewed and endorsed by the Australasian Sleep Association.

SPECT/CT Scan: A New Diagnostic Tool in Pain Medicine.

PURPOSE OF REVIEW: The purpose of this review is to evaluate the role of SPECT/CT in identifying facet joint arthropathy and the outcomes of interventions with SPECT/CT as an adjunct. RECENT FINDINGS: A positive finding of facet arthropathy on SPECT/CT is associated with a higher likelihood of a unilateral procedure and a significantly more effective intervention compared with those performed on patients with facet arthropathy diagnosed only by clinical and/or radiologic examination. Surgical treatment of SPECT/CT-positive findings appears to have a good effect; however, due to limitations in the available studies, no strong conclusion can be drawn. SPECT/CT has a good correlation identifying pain generators in chronic neck and back pain. SPECT/CT-targeted facet interventions demonstrate a higher success rate, but SPECT/CT is not recommended as a first-line diagnostic tool prior to diagnostic facet interventions. More robust studies are needed to confirm the higher success of surgical treatment for SPECT/CT-positive facet arthropathy.

Review of Sacroiliac Joint Injection Techniques.

PURPOSE OF REVIEW: The purpose of the review is to evaluate the current evidence on techniques for sacroiliac joint (SIJ) injections using landmark, ultrasound, fluoroscopy, and computed tomography (CT) guidance. METHODS: A literature search was performed to find all relevant retrospective, prospective, and randomized controlled (RCT) studies where SIJ injections were performed under ultrasound, fluoroscopy, and CT guidance. RECENT FINDINGS: A total of eight studies were identified with suitable data for inclusion. There were two RCTs, four prospective, and two retrospective studies included. Case reports or case series were excluded. A total of 420 patients were enrolled across all eight studies. CT guidance provided the most accurate needle placement in the SIJ injections followed by fluoroscopy, which was more accurate than ultrasound. Landmark-guided injections were not accurate. Accurate needle placement in SIJ confirms SIJ-mediated pain and injection of corticosteroids leads to improvement in pain and/or disability outcome measures regardless of guidance technique. Diagnostic CT-guided SIJ injections should be performed prior to consideration of SIJ fusion.

Cardiac implantable electronic device surgery with interruption of warfarin forgoing post-operative bridging therapy in patients with moderate or high thromboembolic risks.

BACKGROUND: For patients taking warfarin and undergoing pacemaker or implantable cardioverter-defibrillator surgery, clinical evidence and guidelines support continuation of warfarin therapy, as opposed to interruption of warfarin therapy with heparin bridging. Interruption of warfarin without post-operative bridging therapy may be a feasible alternative but data is sparse. METHODS: This is a single-arm observational study including adults who had interruption of warfarin therapy without post-operative bridging therapy for cardiac implantable electronic device (CIED) surgery performed between 2010 and 2019 in a tertiary referral hospital. The primary outcome was a composite of all-cause mortality, arterial or venous thromboembolic events. The secondary outcomes were clinically significant device-pocket hematoma and other procedural complications. RESULTS: Of the 411 patients analysed including 257 patients (62.5%) who had mechanical heart valves, the primary outcome developed in 5 (1.2%) patients within 30 days after surgery, including death in 3 (0.7%) patients, transient ischemic attack in 1 (0.2%) patient and non-CNS embolism in 1 (0.2%) patient. Clinically significant hematomas occurred in 24 (5.8%) patients, including 15 (3.7%) requiring additional interruption of anti-coagulation and 6 (1.5%) requiring clot evacuation. Other procedural complications and bleeding events were rare (< 1%). CONCLUSIONS: Warfarin interruption without post-operative bridging therapy for CIED surgery was associated with low thromboembolic risks and acceptable bleeding risk. Randomized controlled trials are required to formulate an optimal approach to anti-coagulation management.

Improving hospital readmission prediction using individualized utility analysis.

OBJECTIVE: Machine learning (ML) models for allocating readmission-mitigating interventions are typically selected according to their discriminative ability, which may not necessarily translate into utility in allocation of resources. Our objective was to determine whether ML models for allocating readmission-mitigating interventions have different usefulness based on their overall utility and discriminative ability. MATERIALS AND METHODS: We conducted a retrospective utility analysis of ML models using claims data acquired from the Optum Clinformatics Data Mart, including 513,495 commercially-insured inpatients (mean [SD] age 69 [19] years; 294,895 [57%] Female) over the period January 2016 through January 2017 from all 50 states with mean 90 day cost of $11,552. Utility analysis estimates the cost, in dollars, of allocating interventions for lowering readmission risk based on the reduction in the 90-day cost. RESULTS: Allocating readmission-mitigating interventions based on a GBDT model trained to predict readmissions achieved an estimated utility gain of $104 per patient, and an AUC of 0.76 (95% CI 0.76, 0.77); allocating interventions based on a model trained to predict cost as a proxy achieved a higher utility of $175.94 per patient, and an AUC of 0.62 (95% CI 0.61, 0.62). A hybrid model combining both intervention strategies is comparable with the best models on either metric. Estimated utility varies by intervention cost and efficacy, with each model performing the best under different intervention settings. CONCLUSION: We demonstrate that machine learning models may be ranked differently based on overall utility and discriminative ability. Machine learning models for allocation of limited health resources should consider directly optimizing for utility.

Respiratory failure, clinical course and community management of COVID-19 patients in a large Australian cohort.

BACKGROUND: Coronavirus disease 2019 (COVID-19) has wreaked health and economic damage globally. A thorough understanding of the characteristics of COVID-19 patients in Australia plus the strategies that successfully 'flatten the curve' are vitally important to contain this pandemic. AIM: To describe the clinical characteristics and outcomes of COVID-19 patients in the Sutherland Shire, and the management model adopted to manage these patients. METHODS: A retrospective case series of COVID-19 patients monitored in the Sutherland Shire between 19 March and 15 May 2020 was performed. Demographic, clinical and outcome data of COVID-19 inpatients at the Sutherland Hospital and demographic data of COVID-19 patients in the Sutherland Shire community were obtained. The Sutherland Hospital COVID-19 Management Model involved close collaboration among the Sutherland Fever Clinic, Sutherland COVID-19 community telemonitoring team (CTAC) and Sutherland COVID-19 inpatient team. RESULTS: Ninety-nine COVID-19 cases (median age, 49 years, 50 (51%) male) were monitored in Sutherland Shire, with 19 cases (median age, 54 years, 10 (53%) male) requiring inpatient management. Common comorbidities included obesity, asthma, hypertension and Type 2 diabetes mellitus. Six (32%) patients required supplemental oxygen and three (16%) patients required intensive care admission. There was one mortality. The CTAC team identified five (5%) patients requiring admission, and three (3%) patients requiring re-admission. The majority of COVID-19 source was from overseas travel (67%), with nine (9%) cases having unknown source. CONCLUSION: A comprehensive COVID-19 management model is needed to successfully manage COVID-19 patients in both outpatient and inpatient settings in order to 'squash the curve'.

Failure to thrive - an overlooked manifestation of KMT2B-related dystonia: a case presentation.

BACKGROUND: KMT2B-related dystonia is a recently described form of childhood onset dystonia that may improve with deep brain stimulation. Prior reports have focused on neurologic features including prominent bulbar involvement without detailing general health consequences that may result from orolingual dysfunction. We describe a family with novel KMT2B mutation with several members with failure to thrive to highlight this non-neurologic, but consequential impact of mutation in this gene. CASE PRESENTATION: We present a case of a 15-year old female who was admitted and evaluated for failure to thrive. On exam, she had severe speech dysfluency, limited ability to protrude the tongue, and generalized dystonia involving the oromandibular region, right upper and left lower extremity with left foot inversion contracture. The proband and her parents underwent whole genome sequencing. A previously undescribed variant, c.4960 T > C (p.Cys1654Arg), was identified in the KMT2B gene in the proband and mother, and this variant was subsequently confirmed in two maternal cousins, one with failure to thrive. Literature review identified frequent reports of prominent bulbar involvement but failure to thrive is rarely mentioned. CONCLUSION: Failure to thrive is a common pediatric clinical condition that has consequences for growth and development. In the presence of an abnormal neurologic exam, a search for a specific underlying genetic etiology should be pursued. With this case series, we highlight an unusual potentially treatable cause of failure to thrive, reinforce the importance of precise molecular diagnosis for patients with failure to thrive and an abnormal neurologic exam, and underscore the importance of cascade screening of family members.

Complications Associated with Lumbar Transforaminal Epidural Steroid Injections.

PURPOSE OF REVIEW: Low back pain with radicular symptoms is a common cause of disability in the adult population in the USA. Lumbar transforaminal epidural steroid injection (TFESI) is one of the most frequently used intervention for lumbar radiculitis. The purpose of this review is to evaluate complications associated with lumbar TFESI. RECENT FINDINGS: Based on the literature review, the reported rate of minor complications was between 2.4 and 9.6%. The major complications including spinal abscess, spinal cord infarct, and epidural hematoma were documented as case reports. Some patients with spinal cord infarct had permanent neurologic deficits, while the other patients had recovery of neurological function after surgical or medical intervention. This review identifies both the minor and major complications related to lumbar transforaminal epidural steroid injections. According to this review, most complications are minor. Lumbar TFESI can be considered a safe treatment in the management of lumbar radicular pain. However, pain specialists should be aware of the potentially devastating major complications. Early recognition and treatment of complications are crucial for improving the outcome.

Review of Ilioinguinal Nerve Blocks for Ilioinguinal Neuralgia Post Hernia Surgery.

PURPOSE OF REVIEW: The purpose of this review is to evaluate the current evidence on ultrasound-guided ilioinguinal nerve blocks for ilioinguinal neuralgia post hernia surgery. METHODS: A literature search was performed to find all relevant case reports, case series, prospective or retrospective cohort studies, and randomized controlled trials (RCTs) where ultrasound-guided or landmark-based ilioinguinal nerve blocks were used for ilioinguinal neuralgia post-inguinal hernia surgery. RECENT FINDINGS: A total of six studies were identified with suitable data for inclusion. Three studies were retrospective, two studies were prospective, and one study was a randomized controlled trial. A total of 133 subjects were enrolled across these studies. Approximately 55-70% had a beneficial analgesic response to treatment. No major complications were reported in these studies. Ultrasound- and landmark-based ilioinguinal nerve blocks are safe and effective for pain relief post inguinal hernia surgery. Although there were two studies that did not show a statically significant difference in both techniques, the ultrasound-guided injection has the advantage of direct visualization of pathology, more accurate needle placement, and decreased risks of intravascular injections.

Incorporating machine learning and social determinants of health indicators into prospective risk adjustment for health plan payments.

BACKGROUND: Risk adjustment models are employed to prevent adverse selection, anticipate budgetary reserve needs, and offer care management services to high-risk individuals. We aimed to address two unknowns about risk adjustment: whether machine learning (ML) and inclusion of social determinants of health (SDH) indicators improve prospective risk adjustment for health plan payments. METHODS: We employed a 2-by-2 factorial design comparing: (i) linear regression versus ML (gradient boosting) and (ii) demographics and diagnostic codes alone, versus additional ZIP code-level SDH indicators. Healthcare claims from privately-insured US adults (2016-2017), and Census data were used for analysis. Data from 1.02 million adults were used for derivation, and data from 0.26 million to assess performance. Model performance was measured using coefficient of determination (R2), discrimination (C-statistic), and mean absolute error (MAE) for the overall population, and predictive ratio and net compensation for vulnerable subgroups. We provide 95% confidence intervals (CI) around each performance measure. RESULTS: Linear regression without SDH indicators achieved moderate determination (R2 0.327, 95% CI: 0.300, 0.353), error ($6992; 95% CI: $6889, $7094), and discrimination (C-statistic 0.703; 95% CI: 0.701, 0.705). ML without SDH indicators improved all metrics (R2 0.388; 95% CI: 0.357, 0.420; error $6637; 95% CI: $6539, $6735; C-statistic 0.717; 95% CI: 0.715, 0.718), reducing misestimation of cost by $3.5 M per 10,000 members. Among people living in areas with high poverty, high wealth inequality, or high prevalence of uninsured, SDH indicators reduced underestimation of cost, improving the predictive ratio by 3% (~$200/person/year). CONCLUSIONS: ML improved risk adjustment models and the incorporation of SDH indicators reduced underpayment in several vulnerable populations.

(Re)Drawing the Line: Australian Regulation of Human-Animal Interspecies Embryos.

The mixing of human and animal cellular and genetic material is a promising area of science, but inherent societal and safety concerns make such mixing in embryos particularly controversial. The sensitive nature of this research, coupled with science's rapid development, creates problems for policymakers responsible for deciding what practices are and are not permitted in Australia. Australia's regulation in this area, last significantly amended in 2006, is in urgent need of reform. This article investigates what is happening in this fast moving area and the regulatory reforms necessary for Australian scientists to participate.

Leptin is a physiological regulator of skeletal muscle angiogenesis and is locally produced by PDGFRα and PDGFRß expressing perivascular cells.

Skeletal muscle capillarity is characteristically reduced in mature leptin receptor-deficient (Leprdb) mice, which has been attributed to the capillary loss that occurs secondary to metabolic dysfunction. Despite wide recognition of leptin as a pro-angiogenic molecule, the contribution of this adipokine has largely been overlooked in peripheral tissues. Moreover, prior documentation of leptin production within skeletal muscle indicates a potential paracrine role in maintaining local tissue homeostasis. Thus, we hypothesized that leptin is a physiological local paracrine regulator of skeletal muscle angiogenesis and that its production may be modulated by nutrient availability. Leprdb mice exhibited impaired angiogenesis during normal developmental maturation of skeletal myocytes, corresponding with an inability to increase vascular endothelial growth factor-A (VEGFA) mRNA and protein levels between 4 and 13 weeks. In cultured murine and human skeletal myocytes, recombinant leptin increased VEGFA mRNA levels. Leptin mRNA was detectable in skeletal muscle, increasing with prolonged high-fat feeding in mice, and with adiposity in human subjects. Platelet-derived growth factor receptor (PDGFR)α- and PDGFRß- expressing perivascular cell populations were identified as leptin producing within skeletal muscle of mice and humans. Furthermore, in response to 2 weeks of high-fat feeding, PDGFRß+ but not PDGFRα+ cells increased leptin production. We conclude that leptin is a physiological regulator of the capillary network in skeletal muscle and stimulates VEGFA production by skeletal myocytes. PDGFRß expressing perivascular cells exhibit the capacity to act as local "nutrient-sensors" that couple nutrient status to leptin production in skeletal muscle.

SAR optimization studies on modified salicylamides as a potential treatment for acute myeloid leukemia through inhibition of the CREB pathway.

Disruption of cyclic adenosine monophosphate response element binding protein (CREB) provides a potential new strategy to address acute leukemia, a disease associated with poor prognosis, and for which conventional treatment options often carry a significant risk of morbidity and mortality. We describe the structure-activity relationships (SAR) for a series of XX-650-23 derived from naphthol AS-E phosphate that disrupts binding and activation of CREB by the CREB-binding protein (CBP). Through the development of this series, we identified several salicylamides that are potent inhibitors of acute leukemia cell viability through inhibition of CREB-CBP interaction. Among them, a biphenyl salicylamide, compound 71, was identified as a potent inhibitor of CREB-CBP interaction with improved physicochemical properties relative to previously described derivatives of naphthol AS-E phosphate.