ExoSTING, an extracellular vesicle loaded with STING agonists, promotes tumor immune surveillance.

Cyclic dinucleotide (CDN) agonists of the STimulator of InterferoN Genes (STING) pathway have shown immune activation and tumor clearance in pre-clinical models. However, CDNs administered intratumorally also promote STING activation leading to direct cytotoxicity of many cell types in the tumor microenvironment (TME), systemic inflammation due to rapid tumor extravasation of the CDN, and immune ablation in the TME. These result in a failure to establish immunological memory. ExoSTING, an engineered extracellular vesicle (EV) exogenously loaded with CDN, enhances the potency of CDN and preferentially activates antigen presenting cells in the TME. Following intratumoral injection, exoSTING was retained within the tumor, enhanced local Th1 responses and recruitment of CD8+ T cells, and generated systemic anti-tumor immunity to the tumor. ExoSTING at therapeutically active doses did not induce systemic inflammatory cytokines, resulting in an enhanced therapeutic window. ExoSTING is a novel, differentiated therapeutic candidate that leverages the natural biology of EVs to enhance the activity of CDNs.

A versatile platform for generating engineered extracellular vesicles with defined therapeutic properties.

Extracellular vesicles (EVs) are an important intercellular communication system facilitating the transfer of macromolecules between cells. Delivery of exogenous cargo tethered to the EV surface or packaged inside the lumen are key strategies for generating therapeutic EVs. We identified two "scaffold" proteins, PTGFRN and BASP1, that are preferentially sorted into EVs and enable high-density surface display and luminal loading of a wide range of molecules, including cytokines, antibody fragments, RNA binding proteins, vaccine antigens, Cas9, and members of the TNF superfamily. Molecules were loaded into EVs at high density and exhibited potent in vitro activity when fused to full-length or truncated forms of PTGFRN or BASP1. Furthermore, these engineered EVs retained pharmacodynamic activity in a variety of animal models. This engineering platform provides a simple approach to functionalize EVs with topologically diverse macromolecules and represents a significant advance toward unlocking the therapeutic potential of EVs.

Development and validation of a training and assessment tool for laparoscopic radical nephrectomy. / Desarrollo y validación de una herramienta de entrenamiento y evaluación para la nefrectomía radical laparoscópica.

INTRODUCTION: Laparoscopic radical nephrectomy(LRN) is a cornerstone in managing renal cancer and small renal masses. Twenty-first century surgical training faces challenges, thus must be efficient and safe so surgeons attain relevant skills, protecting patients and operative outcomes. This study aimed to systematically develop a tool for training and assessment in LRN and validate the developed tool for use by trainee urologists. METHODS: This prospective, longitudinal, multi-institutional study was undertaken from September 2014 - June 2015. Healthcare Failure Mode and Effect Analysis was utilised for development and followed by validation where the assessment tool was distributed to five specialists to increase content validity. Four experts were observed as a multi-institutional approach. Hand-assisted, transperitoneal and retroperitoneal approaches were considered. RESULTS: The LRN Assessment Tool comprised four phases, 17 processes, 41 sub-processes. Four surgeons and operating teams were observed across four hospitals for 19.5hours (5.75h hand-assisted, 8.75h trans-peritoneal, 5h retro-peritoneal). After hazard analysis, three checklists were constructed. Those for hand-assisted LRN and transperitoneal LRN contained four phases, 20 processes, 33 sub-processes and that for retroperitoneal LRN contained four phases, 20 processes, 30 sub-processes. These were merged to form one assessment tool. The final result was a four phase LRN Assessment Tool with 17 processes, 41 sub-processes. All participants agreed the final LRN Assessment Tool included pertinent steps. CONCLUSIONS: The LRN Assessment Tool was developed using Healthcare Failure Mode and Effect Analysis risk analysis to ensure hazardous procedural sub-steps were included. Validation ascertained important processes were not overlooked. Full application through a pilot study must be undertaken.

KRAS G12C Drug Development: Discrimination between Switch II Pocket Configurations Using Hydrogen/Deuterium-Exchange Mass Spectrometry.

KRAS G12C, the most common RAS mutation found in non-small-cell lung cancer, has been the subject of multiple recent covalent small-molecule inhibitor campaigns including efforts directed at the guanine nucleotide pocket and separate work focused on an inducible pocket adjacent to the switch motifs. Multiple conformations of switch II have been observed, suggesting that switch II pocket (SIIP) binders may be capable of engaging a range of KRAS conformations. Here we report the use of hydrogen/deuterium-exchange mass spectrometry (HDX MS) to discriminate between conformations of switch II induced by two chemical classes of SIIP binders. We investigated the structural basis for differences in HDX MS using X-ray crystallography and discovered a new SIIP configuration in response to binding of a quinazoline chemotype. These results have implications for structure-guided drug design targeting the RAS SIIP.

Genetic variation, expression and ontogeny of sulfotransferase SULT2A1 in humans.

Sulfotransferases (SULTs) are enzymes involved in the metabolism of several endogenous molecules. The activity and expression exhibit inter- and intra-individual variations due to age and genetic variation. The aims of this study were to compare the gene expression of SULT2A1 in fetal and adult livers, to study the intra-individual tissue distribution, and investigate if expression is associated with a SULT2A1 copy number variation polymorphism. In contrast to other drug metabolizing enzyme systems the expression of SULT2A1 did not differ between fetal and adult liver samples and it was not affected by maternal smoking or gestational age. Gene expression in relation to sex could not be assessed as the sex of the fetuses was unknown. SULT2A1 was consistently expressed in livers and adrenals, being seven times more abundant in adrenals, but was absent in the lungs. The SULT2A1 copy number variation was proportional to gene expression in liver and adrenals. Our results show that SULT2A1 is important in the first trimester; particularly in the adrenals.

Therapeutic targeting of oncogenic K-Ras by a covalent catalytic site inhibitor.

We report the synthesis of a GDP analogue, SML-8-73-1, and a prodrug derivative, SML-10-70-1, which are selective, direct-acting covalent inhibitors of the K-Ras G12C mutant relative to wild-type Ras. Biochemical and biophysical measurements suggest that modification of K-Ras with SML-8-73-1 renders the protein in an inactive state. These first-in-class covalent K-Ras inhibitors demonstrate that irreversible targeting of the K-Ras guanine-nucleotide binding site is potentially a viable therapeutic strategy for inhibition of Ras signaling.

Anterior compartment mesh: a descriptive study of mesh anchoring failure.

OBJECTIVES: To define types of support failure after anterior compartment mesh placement and to investigate any association with predictors of recurrence. METHODS: This was a retrospective study on patients ≥ 3 months after anterior mesh placement. All patients underwent a standardized interview, clinical examination using the International Continence Society Pelvic Organ Prolapse Quantification system (ICS POP-Q) and three-dimensional/four-dimensional (3D/4D) translabial ultrasound. Mesh failure was defined as recurrence of anterior/central compartment prolapse seen on ultrasound. Failures were classified as anterior, global and apical. Their association with hiatal area on Valsalva maneuver and levator avulsion was tested. RESULTS: Three hundred and one patients were seen initially, of whom five were excluded because of missing data, leaving 296. Mean follow-up was 1.8 (range, 0.3-5.6) years. Mean age was 65 (range, 32-88) years. One hundred and thirty-nine (47%) women were fitted with a Perigee mesh, 66 (22%) with an Anterior Prolift mesh and 91 (31%) with an Anterior Elevate mesh. Recurrent symptoms (lump/drag) were reported in 65 (22%), a recurrent cystocele was noted in 128 clinically (43%) and in 105 on ultrasound (35%). Avulsion was diagnosed in 117 patients (40%). Mean hiatal area on Valsalva was 33.3 (range, 14.1-60.0) cm2. Mesh failure was diagnosed in 112 patients (38%), comprising global failure in 81 (27%), apical failure in 23 (8%) and anterior failure in eight (3%). Apical and global failures were significantly associated with hiatal area, associations that remained after controlling for potential confounders. CONCLUSIONS: Mesh failure, i.e. anterior or central compartment recurrent prolapse, was noted in 38% of patients on average 1.8 years after placement of anterior compartment mesh. Global and apical failures together constituted 93% of all mesh failures, both types of failure being significantly associated with hiatal area on Valsalva maneuver.

Is levator avulsion a predictor of cystocele recurrence following anterior vaginal mesh placement?

OBJECTIVE: Levator avulsion has been shown to be a predictor of cystocele recurrence following anterior colporrhaphy. The aim of this study was to determine if levator avulsion is a risk factor for prolapse recurrence following anterior colporrhaphy with mesh. METHODS: This was a retrospective analysis of data obtained from three surgical audits for subjective and objective outcomes following anterior colporrhaphy with mesh. Recurrence was defined as cystocele ≥ Stage 2 on the prolapse quantification system of the International Continence Society; symptoms of vaginal lump/bulge; or cystocele on ultrasound, defined as maximum bladder descent to ≥ 10 mm below the symphysis pubis. Levator avulsion was diagnosed using tomographic ultrasound imaging. RESULTS: Two hundred and nine patients were followed up at a mean of 2.2 years (range, 3 months to 5.6 years) after anterior vaginal mesh placement. 24% (51/209) had recurrent prolapse symptoms, 33% (68/209) clinical cystocele recurrence ≥ Stage 2, and 26% (54/209) a recurrent cystocele on ultrasound. Twenty-eight out of 80 (35%) women with levator avulsion had significant sonographic cystocele recurrence (odds ratio (OR), 2.24 (95% confidence interval (CI), 1.13-4.43)). This finding was confirmed after adjusting for potential predictors of prolapse recurrence on multivariate logistic regression (OR, 2.13 (95% CI, 1.04-4.39); P = 0.04). CONCLUSION: Levator avulsion doubles the risk of cystocele recurrence after anterior colporrhaphy with transobturator mesh.

The sacrospinous ligament: conveniently effective or effectively convenient?

The sacrospinous ligament has been used for over 50 years as a convenient structure for treating vaginal vault and more recently, uterine prolapse. The procedure has evolved over the years and its efficacy has been hotly debated with invariable comparisons made to abdominal sacral colpopexy. Mesh surgery has introduced a newer dimension to the debate. This review is an attempt to clarify the anatomy, reflect on various techniques and offer a critique on the current 'status' of the sacrospinous ligament.

Microscopic alterations of vaginal tissue in women with pelvic organ prolapse.

The aim of this study was to identify any histological changes between the prolapsed and non-prolapsed vaginal skin to enable us to answer the broader question of whether to excise or re-suture the prolapsed tissue during vaginal reconstructive surgery. Two tissue samples were taken from 20 women who underwent prolapse surgery, one prolapsed and another non-prolapsed tissue. The samples were analysed blind histologically by a pathologist. Our study showed statistically significant differences in three tissue components namely myofibroblast differentiation (0.047), elastin (0.048) and collagen (0.095), depending on whether the tissue was prolapsed or non-prolapsed. However, the study had certain limitations and practical difficulties faced in histological approach of tissue analysis. Histology alone is an insufficient indicator to answer our question and definitive understanding of the prolapsed tissue may result from a randomised study on a larger study population.

Inducible dopaminergic glutathione depletion in an α-synuclein transgenic mouse model results in age-related olfactory dysfunction.

Parkinson's disease (PD) involves both motor and non-motor disturbances. Non-motor features include alterations in sensory olfactory function which may constitute a viable biomarker for the disorder. It is not clear what causes olfactory dysfunction but it appears to coincide with the development of synucleopathy within the olfactory bulb (OB). Elevation in alpha-synuclein (a-syn) is indeed a risk factor for development of the sporadic disorder. The multifactorial nature of the idiopathic disease combined with variability in its presentation suggests that it is likely to be influenced by several factors and that in vivo models that explore the synergistic effect of alpha-synuclein elevation with other potential contributing factors are likely to be of importance in understanding the disease etiology. Using a dual transgenic (DTg) mouse model of dopaminergic alpha-synuclein overexpression coupled with doxycycline (Dox)-inducible glutathione (GSH) depletion in these same cells, we demonstrate an age-related loss in behavioral olfactory function coupled with a significant neurodegeneration of glomerular dopaminergic neurons. This is accompanied by increase in alpha-synuclein levels in non-dopaminergic cells in the granule cell layer (GCL). In addition, isolated olfactory bulb synaptosomes from dual transgenic lines with Dox consistently showed a slight but significant reduction in maximum mitochondrial respiration compared to controls. These results suggest that in the presence of increased oxidative stress, increased alpha-synuclein expression within dopaminergic OB neurons results in neurodegeneration in the glomerular layer (GL) and increased alpha-synuclein levels in the granular cell layer which coincide with olfactory dysfunction.

Thyroid function and pregnancy: before, during and beyond.

Thyroid disturbances are common in women during the reproductive years of their lives. Autoimmunity and altered iodine status together account for a high proportion of the abnormalities. Autoimmune thyroid disease is present in around 4% of young females, and up to 15% are at risk because they are thyroid antibody-positive. There is a strong relationship between thyroid immunity on the one hand and infertility, miscarriage, and thyroid disturbances in pregnancy and postpartum on the other hand. Suboptimal iodine status affects a large proportion of the world's population, and pregnancy further depletes iodine stores. There is controversy surrounding the degree to which iodine should be supplemented and the duration of supplementation. Recent studies have helped to clarify the relationship between maternal thyroid status and neuropsychological development of the child. The role of other environmental factors including smoking and selenium status is also now recognised. Universal screening for thyroid hormone abnormalities is not routinely recommended at present. However, measurement of thyroid function and autoantibodies should certainly be considered in those who are at high risk of thyroid disease and in those whose pregnancy is otherwise high risk. The practicing clinician needs to be aware of the thyroid changes which accompany pregnancy.

Stress urinary incontinence after transobturator mesh for cystocele repair.

Cystocele repair can lead to de novo stress urinary incontinence (SUI) or exacerbate pre-existing SUI. This study was designed to determine the effect of cystocele repair by transobturator mesh on SUI. In a retrospective observational study, we saw 93 patients after a transobturator mesh procedure. Of those, 57 women had not undergone a concomitant anti-incontinence procedure. We analyzed their clinical data and ultrasound datasets. At a median follow-up of 9 months, 21 of 24 preoperatively stress incontinent women reported cure/improvement, one patient reported worsened SUI. Seven of 33 preoperatively continent women complained of de novo SUI. There is a net positive effect on SUI (McNemar chi(2) exact test p = 0.013) after transobturator mesh. A narrower gap between symphysis pubis and mesh was associated with a positive outcome (p = 0.015 on ANOVA). Transobturator mesh for cystocele repair appears to have a net positive effect on SUI.

Prospective evaluation of the safety and efficacy of the Apogee system for treatment of vault prolapse.

This is a prospective study evaluating the efficacy and safety of the Apogee vault repair system in the management of vault prolapse. All patients who underwent this procedure between October 2004 and December 2005 in Townsville, Australia are included in the study. Preoperative and postoperative clinical assessments were done using the pelvic organ prolapse quantification (POPQ) system and patients were followed up at 6 weeks, 12 weeks, 6 months, 12 months and then biannually. Of the 35 patients operated, there were no intraoperative complications and only one patient (2.8%) had a recurrence of vault prolapse at 12 months. Nine patients (25%) had mesh exposure treated successfully with outpatient trimming and oestrogen. This medium term study shows that the Apogee procedure is safe with minimal complications for the management of vault prolapse.