Tuning Transcription Factor Availability through Acetylation-Mediated Genomic Redistribution.

It is widely assumed that decreasing transcription factor DNA-binding affinity reduces transcription initiation by diminishing occupancy of sequence-specific regulatory elements. However, in vivo transcription factors find their binding sites while confronted with a large excess of low-affinity degenerate motifs. Here, using the melanoma lineage survival oncogene MITF as a model, we show that low-affinity binding sites act as a competitive reservoir in vivo from which transcription factors are released by mitogen-activated protein kinase (MAPK)-stimulated acetylation to promote increased occupancy of their regulatory elements. Consequently, a low-DNA-binding-affinity acetylation-mimetic MITF mutation supports melanocyte development and drives tumorigenesis, whereas a high-affinity non-acetylatable mutant does not. The results reveal a paradoxical acetylation-mediated molecular clutch that tunes transcription factor availability via genome-wide redistribution and couples BRAF to tumorigenesis. Our results further suggest that p300/CREB-binding protein-mediated transcription factor acetylation may represent a common mechanism to control transcription factor availability.

Click display: a rapid and efficient in vitro protein display method for directed evolution.

We describe a novel method for in vitro protein display-click display-that does not depend on maintaining RNA integrity during biopanning and yields covalently linked protein-cDNA complexes from double-stranded input DNA within 2 h. The display is achieved in a one-pot format encompassing transcription, translation and reverse transcription reactions in series. Stable linkage between proteins and the encoding cDNA is mediated by a modified DNA linker-ML-generated via a click chemistry reaction between a puromycin-containing oligo and a cDNA synthesis primer. Biopanning of a click-displayed mock library coupled with next-generation sequencing analysis revealed >600-fold enrichment of target binders within a single round of panning. A synthetic library of Designed Ankyrin Repeat Proteins (DARPins) with ∼1012 individual members was generated using click display in a 25-µl reaction and six rounds of library panning against a model protein yielded a panel of nanomolar binders. This study establishes click display as a powerful tool for protein binder discovery/engineering and provides a convenient platform for in vitro biopanning selection even in RNase-rich environments such as on whole cells.

Robot-Assisted Pelvic Dissection for Enlarged Lymph Nodes in Melanoma Improves Recovery with Equivalent Oncological Outcomes to Open Pelvic Dissection.

BACKGROUND: Robot-assisted pelvic lymph node dissection (rPLND) has been reported in heterogenous groups of patients with melanoma, including macroscopic or at-high-risk-for microscopic metastasis. With changing indications for surgery in melanoma, and availability of effective systemic therapies, pelvic dissection is now performed for clinically detected bulky lymph node metastasis followed by adjuvant drug therapy. rPLND has not been compared with open pelvic lymph node dissection (oPLND) for modern practice. METHODS: All patients undergoing pelvic node dissection for macroscopic melanoma at a single institution were reviewed as a cohort, observational study. RESULTS: Twenty-two pelvic lymph node dissections were identified (8 oPLND; 14 rPLND). The number of pelvic lymph nodes removed was similar (median oPLND 6.5 (interquartile range [IQR] 6.0-12.5] versus rPLND 6.0 [3.75-9.0]), with frequent matted nodes (11/22, 50.0%). Operative time (median oPLND 130 min [IQR 95.5-182] versus rPLND 126 min [IQR 97.8-160]) and complications (Clavien-Dindo scale) were similar. Length of hospital stay (median 5.34 days (IQR 3.77-6.94) versus 1.98 days (IQR 1.39-3.50) and time to postoperative adjuvant therapy (median 11.6 weeks [IQR 10.6-18.5] versus 7.71 weeks [IQR 6.29-10.4]) were shorter in the rPLND group. No differences in pelvic lymph node recurrence (p = 0.984), distant metastatic recurrence (p = 0.678), or melanoma-specific survival (p = 0.655) were seen (median follow-up 21.1 months [rPLND] and 25.7 months [oPLND]). CONCLUSIONS: rPLND is an effective way to remove bulky pelvic lymph nodes in melanoma, with a shorter recovery and reduced interval to initiating adjuvant therapy compared with oPLND. This group of patients may especially benefit from neoadjuvant systemic approaches to management.

Long-Term Dynamics of Serum α-MSH and α-MSH-Binding Immunoglobulins with a Link to Gut Microbiota Composition in Patients with Anorexia Nervosa.

INTRODUCTION: Immunoglobulins (Ig) reactive with α-melanocyte-stimulating hormone (α-MSH), an anorexigenic neuropeptide, are present in humans and were previously associated with eating disorders. In this longitudinal study involving patients with anorexia nervosa (AN), we determined whether α-MSH in serum is bound to IgG and analyzed long-term dynamics of both α-MSH peptide and α-MSH-reactive Ig in relation to changes in BMI and gut microbiota composition. METHODS: The study included 64 adolescents with a restrictive form of AN, whose serum samples were collected at hospital admission, discharge, and during a 1-year follow-up visit and 41 healthy controls, all females. RESULTS: We found that in both study groups, approximately 40% of serum α-MSH was reversibly bound to IgG and that levels of α-MSH-reactive IgG but not of α-MSH peptide in patients with AN were low at hospital admission but recovered 1 year later. Total IgG levels were also low at admission. Moreover, BMI-standard deviation score correlated positively with α-MSH IgG in both groups studied but negatively with α-MSH peptide only in controls. Significant correlations between the abundance of specific bacterial taxa in the gut microbiota and α-MSH peptide and IgG levels were found in both study groups, but they were more frequent in controls. CONCLUSION: We conclude that IgG in the blood plays a role as an α-MSH-binding protein, whose characteristics are associated with BMI in both patients with AN and controls. Furthermore, the study suggests that low production of α-MSH-reactive IgG during the starvation phase in patients with AN may be related to altered gut microbiota composition.

Primary retroperitoneal lymph node dissection in stage II testicular seminoma: a systematic review.

OBJECTIVE: To conduct a systematic review of the current literature to determine the current role of primary retroperitoneal lymph node dissection (RPLND) in stage II testicular seminoma and its associated oncological, functional and peri-operative outcomes. MATERIALS AND METHODS: A comprehensive literature search was conducted in Medline, Embase, and Scopus for publications from inception until November 2023. The systematic review was registered on PROSPERO (ID CRD42023449781), was conducted according to the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines and utilised the Methodological Index for Non-Randomised Studies (MINORS) tool. RESULTS: Six studies involving 385 patients were analysed, with 48.5% clinical stage IIA and 51.5% stage IIB seminomas. The patients' mean (range) age was 37 (20-64) years. The median operation time was 187 min, median estimated blood loss was 150 mL and median length of hospital stay was 4 days. In all, 6.1% of patients developed complications that were greater or equal to Clavien-Dindo grade 3. Only four studies reported on anejaculation rate (median: 4.9%). Only one study had long-term data, demonstrating a 92% 5-year overall survival for stage IIA/B disease treated with RPLND. The remaining five studies had a median follow-up of between 18.5 and 37 months and reported a mean recurrence rate of 15.6%. Most recurrences (78%) were not within the field of RPLND. Recurrence was associated with higher clinical and pathological lymph node stage, and metachronous or delayed development of retroperitoneal lymphadenopathy (initially stage I disease, as opposed to de novo stage IIA/B disease). DISCUSSION: Primary RPLND, performed by experienced surgeons, has good peri-operative outcomes. Recurrence is more common than with standard treatment, but long-term survival and functional data are limited, although promising.

Pseudoephedrine for ejaculatory dysfunction after retroperitoneal lymph node dissection in testicular cancer.

OBJECTIVE: To assess the impact of ejaculatory dysfunction (EjD; failure of emission or retrograde ejaculation) on health-related quality of life (HRQoL) after retroperitoneal lymph node dissection (RPLND) for testicular cancer and explore the efficacy of pseudoephedrine hydrochloride as treatment. PATIENTS AND METHODS: In a single arm, phase II trial, patients at ≥6 months after RPLND were invited to complete patient-reported outcome measures (European Organisation for Research and Treatment of Cancer [EORTC] quality of life questionnaire [QLQ]-30-item core, EORTC QLQ-testicular cancer-26, and Brief Male Sexual Function Inventory) evaluating HRQoL and sexual function in follow-up (ACTRN12622000537752/12622000542796). If EjD was reported, post-ejaculatory urine ± semen analysis was undertaken. In eligible patients, pseudoephedrine hydrochloride 60 mg was administered orally every 6 h for six doses. The primary endpoint was sperm count >39 million sperm/ejaculate (>5th centile) following treatment. The trial was powered to detect a clinically relevant 36% achieving sperm count of >39 million sperm/ejaculate. Secondary endpoints included semen volume >1.5 mL, total motile sperm count, safety, and HRQoL impacts. RESULTS: Of the 58 patients enrolled, the median (interquartile range [IQR]) age was 35 (29-41) years, with a median (IQR) of 37 (18-60) months from RPLND. EjD was reported in 33 (57%), including 27/52 (52%) receiving follow-up at our centre. There were no differences in global HRQoL; however, role functioning (P = 0.045), sexual problems (P < 0.005), and sexual enjoyment (P = 0.005) was poorer if EjD was present. In all, 24/33 (73%) patients with EjD consented to pseudoephedrine treatment. Of 22 evaluable patients, four (18%) achieved a sperm count of >39 million/ejaculate (P = 0.20), and four (18%) had a semen volume of >1.5 mL (P = 0.20). There was a mean increase of 105 million sperm/ejaculate (P = 0.051) and 1.47 mL increase in semen volume (P = 0.01). No safety concerns arose. CONCLUSION: Ejaculatory dysfunction is common after RPLND but did not impact global HRQoL in our cohort. Pseudoephedrine improved EjD for some; however, its efficacy was lower than expected. Pseudoephedrine may be considered on an individualised basis.

Severe behavior problems in SYNGAP1-related disorder: A summary of 11 consecutive patients in a tertiary care specialty clinic.

SYNGAP1-related disorder (SYNGAP1-RD) is a neurodevelopmental disorder that is commonly associated with epilepsy, autism spectrum disorder (ASD), and disruptive behaviors. In this study, behavior problems in 11 consecutive patients with SYNGAP1-RD are described and quantified based on a behavioral screening conducted within the context of a multi-disciplinary tertiary care specialty clinic visit. The behavioral phenotype was then compared to published samples of behavior problems in ASD and other genetic cause of epilepsy occurring in the context of neurodevelopmental disorders using results from the Aberrant Behavior Checklist-Community (ABC-C), an empirically derived outcome measure. We report common antecedent and consequent events surrounding problem behavior across individuals. Additionally, we report on the management approach of caregivers and the impact of problem behaviors on the family. Our results suggest a number of commonalities between behavioral profiles in SYNGAP1-RD with ASD and other genetic causes of developmental and epileptic encephalopathies, and also highlight severe behavior problems as a specific behavioral phenotype of SYNGAP1-RD.

Retraction Note: DDX5 and its associated lncRNA Rmrp modulate TH17 cell effector functions.

Change History: This Article has been retracted; see accompanying Retraction. Corrected online 20 January: In this Article, author Frank Rigo was incorrectly listed with a middle initial; this has been corrected in the online versions of the paper.

The combined risk predictive power of frailty and hypoalbuminemia in free tissue flap reconstruction: A cohort study of 34,571 patients from the NSQIP database.

INTRODUCTION: Significant morbidity and mortality are hallmarks of the functional decline seen in physically frail patients. The modified frailty index 5 (mFI-5) represents a risk predictor score that has been validated as a comorbidity-based scale in surgery. Serum albumin levels of <3.5 g/dL (hypoalbuminemia) have also been implicated with poor postoperative outcomes. However, the association between these two parameters remains to be investigated. We aimed to elucidate the interdependence of preoperative albumin levels and frailty, as evaluated by the mFI-5 score, and its reliability to prognosticate postoperative results in free flap reconstruction (FFR). METHODS: We conducted a multicenter, retrospective cohort study and accessed the ACS National Surgical Quality Improvement Program (ACS-NSQIP) from 2008 to 2021. We identified all adult patients (≥18 years of age) who underwent a FFR. We extracted perioperative data and lab values including albumin. Multivariable linear and logistic regression analyses were performed to identify independent risk predictors. Main outcomes involved mortality, length of hospital stay, reoperation, medical and surgical complications, and discharge destination within the 30-day postoperative period. RESULTS: A total of 34,571 patients were included in the study, with an average age of 53.9 years (standard deviation [SD] 12.2) and an average body mass index (BMI) of 28.8 (SD 6.1). Of these patients, 7484 were male (21.6%), whereas 22,363 (64.7%) had no frailty (mFI = 0). Additionally, 9466 patients had a frailty score of 1 (27.4%), 2505 had a score of 2 (7.2%), 226 had a score of 3 (0.7%), and 11 had a score of 4 or higher (0.0%). Albumin levels were available for 16,250 patients (47.0%), and among them, 1334 (8.2%) had hypoalbuminemia. Regression analyses showed that higher mFI scores were independent predictors of any, surgical, and medical complications, as well as increased rates of reoperations, unplanned readmissions, and prolonged hospital stays. Hypoalbuminemia independently predicted any, surgical, and medical complications, and higher mortality, reoperation, and longer hospital stay. When both frailty and albumin levels (mFI-5 and albumin) were considered together, this combined assessment was found to be a more accurate predictor of all major outcomes (any, medical and surgical complications, mortality, and reoperation). Further, our analysis identified a weak negative correlation between serum albumin levels and mFI scores (Spearman R: -.1; p < .0001). CONCLUSION: In conclusion, this cohort study highlights the association of hypoalbuminemia with adverse postoperative outcomes, including those not directly related to frailty. Simultaneously, higher mFI scores independently predicted outcomes not associated with hypoalbuminemia. Stemming from these findings, we recommend considering both serum albumin levels and frailty in patients receiving FFR. This perioperative algorithm may help provide more individualized planning including multidisciplinary care and pre and posthabilitation.

The free vastus lateralis-And conjoined vastus lateralis anterolateral thigh/tensor fascia lata flap for oncological chest wall reconstruction.

INTRODUCTION: A reconstructive option for extensive chest wall reconstruction is the free myocutaneous vastus lateralis muscle (VL) flap which can be performed in isolation or in conjunction with a fasciocutaneus anterolateral thigh (cVLALT) and/or myofasciocutaneous tensor fascia lata flap (cVLTFL). We aimed to directly compare the outcomes of these reconstructive options. METHODS: Patients who underwent oncological chest wall reconstruction with a free VL, cVLALT, or cVLTFL flap between February 2010 and 2022 were included in this retrospective study. Patient demographics, surgical characteristics, as well as medical and reconstructive outcomes, were evaluated. The operative outcomes between myocutaneous VL, cVLALT, and cVLTFL flap reconstructions were compared. RESULTS: A total of 41 patients underwent chest wall reconstruction with a free myocutaneous VL (n = 25; 61%), cVLALT (n = 14; 34%), or cVLTFL Three acute flap thromboses occurred in the entire cohort (3/41, 7%), with one myocutaneous VL flap failing because of recurrent venous thrombosis during the salvage procedure. Total flap necrosis was seen in two cases (5%; VL flap: n = 1; cVLALT flap: n = 1), and partial flap necrosis in one VL flap (1/25, 4%) and in the distal ALT portion of three cVLALT flaps (3/14, 21%). No significant difference was seen between isolated VL and conjoined VL flaps regarding the partial (p = .28) or total flap necrosis rate (p = .9). CONCLUSION: The free (conjoined) VL flap provides reliable outcomes for obliterating dead space achieving durable reconstruction of complex chest wall defects.

Formation of a conceptual framework during the development of a patient-reported outcome measure for early gastrointestinal recovery: phase I of the PRO-diGi study.

AIM: Patients admitted to hospital for abdominal surgery often experience gastrointestinal dysfunction. Many studies have reported outcomes following gastrointestinal dysfunction, yet there is no unified definition of recovery or a validated patient-reported outcome measure (PROM). The first stage of PROM development requires formation of a conceptual framework to identify key themes to patients. The aim of this study was to utilize semistructured interviews to identify core themes and concepts relevant to patients to facilitate development of a conceptual framework. METHOD: Adult patients admitted to hospital for major gastrointestinal, urological or gynaecological surgery, in an emergency or elective setting, were eligible to participate. Patients treated nonoperatively for small bowel obstruction were also eligible. Interviews were conducted by telephone, audio-recorded, transcribed, coded and analysed using NVivo software by two researchers and reviewed by lay members of the steering group. Interviews continued until data saturation was reached. Ethical approval was gained prior to interviews (21/WA/0231). RESULTS: Twenty nine interviews were completed (17 men, median age 64 years) across three specialties (20 gastrointestinal, six gynaecological, three urological). Two overarching themes of 'general recovery' and 'gastrointestinal symptoms' were identified. General recovery included three themes: 'life impact', 'mental impact', including anxiety, and 'physical impact', including fatigue. Gastrointestinal symptoms included three themes: 'abdominal symptoms' such as pain, 'diet and appetite' and 'expulsory function', such as stool frequency. A total of 18 gastrointestinal symptoms were identified during patient recovery-many of which lasted several weeks following discharge. CONCLUSION: This study reports a range of gastrointestinal and nongastrointestinal symptoms experienced by patients during early gastrointestinal recovery. Identified symptoms have been synthesized into a conceptual framework to enable development of a definitive PROM for early gastrointestinal recovery.

Insect biomass density: measurement of seasonal and daily variations using an entomological optical sensor.

Insects are major actors in Earth's ecosystems and their recent decline in abundance and diversity is alarming. The monitoring of insects is paramount to understand the cause of this decline and guide conservation policies. In this contribution, an infrared laser-based system is used to remotely monitor the biomass density of flying insects in the wild. By measuring the optical extinction caused by insects crossing the 36-m long laser beam, the Entomological Bistatic Optical Sensor System used in this study can evaluate the mass of each specimen. At the field location, between July and December 2021, the instrument made a total of 262,870 observations of insects for which the average dry mass was 17.1 mg and the median 3.4 mg. The daily average mass of flying insects per meter cube of air at the field location has been retrieved throughout the season and ranged between near 0 to 1.2 mg/m3. Thanks to its temporal resolution in the minute range, daily variations of biomass density have been observed as well. These measurements show daily activity patterns changing with the season, as large increases in biomass density were evident around sunset and sunrise during Summer but not during Fall.

27-GAUGE PARS PLANA/PLICATA VITRECTOMY FOR PEDIATRIC VITREORETINAL SURGERY.

PURPOSE: To report on the feasibility of 27-gauge (G) vitrectomy for pediatric patients. METHODS: This study is an international, multicenter, retrospective, interventional case series. Participants were patients 17 years or younger who underwent 27-G vitrectomy for various indications. RESULTS: The records of 56 eyes from 47 patients were reviewed. Mean age was 5.7 ± 5.2 years. Diagnoses included retinopathy of prematurity (Stages 3 with vitreous hemorrhage, 4A, 4B, and 5), Terson's syndrome, traumatic macular hole, posterior capsular opacification, endophthalmitis, and others. Instruments used were the 27-G infusion, 27-G vitreous cutter, 27-G light pipe, and 27-G internal limiting membrane forceps. Instrument bending was noted in one (1.8%) case. There were no cases with intraoperative complications, infusion issues, or postoperative endophthalmitis. There were 67/145 (46%) sclerotomies that required suturing, of which most (51/145) were sutured out of precaution. There were four cases (7.1%) that required conversion to a larger gauge and three cases (5.3%) that developed postoperative hypotony. Mean visual acuity improved from logarithm of the minimum angle of resolution 1.32 (20/420) to 0.72 (20/105), after a mean follow-up of 125.1 days (P = 0.01). Anatomic success was achieved in 96.4% of eyes after a single surgery. CONCLUSION: Twenty-seven-gauge vitrectomy was safe and feasible in selected pediatric vitreoretinopathies. Further studies are warranted to examine indications and outcomes.

Enzymatic Carboxylation of Resorcinol in Aqueous Triethanolamine at Elevated CO2 Pressure.

The fixation of CO2 by enzymatic carboxylation for production of valuable carboxylic acids is one way to recycle carbon. Unfortunately, this type of reaction is limited by an unfavourable thermodynamic equilibrium. An excess of the C1 substrate is required to increase conversions. Solvents with a high CO2 solubility, such as amines, can provide the C1 substrate in excess. Here, we report on the effect of CO2 pressures up to 1100 kPa on the enzymatic carboxylation of resorcinol in aqueous triethanolamine. Equilibrium yields correlate to the bicarbonate concentration. However, inhibition is observed at elevated pressure, severely reducing the enzyme activity. The reaction yields were reduced at higher pressures, whereas at ambient pressure, higher yields were achieved. Overall, CO2 pressures above 100 kPa have been demonstrated to be counterproductive for improving the biotransformation, as productivity decreases rapidly for only a modest improvement in conversion. It is expected that CO2 carbamylation intensifies at elevated CO2 pressures, causing the inhibition of the enzyme. To further increase the reaction yield, the in situ product precipitation is tested by the addition of the quaternary ammonium salt tetrabutylammonium bromide.

Perfusion of the proximal scaphoid pole: correlation between preoperative ge-MRI and intraoperative findings.

BACKGROUND: Gadolinium enhanced MRI (ge-MRI) is considered as gold standard for perfusion evaluation in case of scaphoid nonunion (SNU). However, its clinical value and specificity is still not clearly evaluated. This study compares preoperative ge-MRI-based perfusion assessment and intraoperative proximal pole (PP) perfusion after scaphoid reconstruction by vascularized bone grafts. In addition, the postoperative osseous consolidation (OC) was correlated to intraoperative perfusion findings. METHODS: Between 08/2010 and 01/2020, 60 of 271 patients with scaphoid nonunion received a vascularized radius bone graft for reconstruction. Medical reports were checked for intra-op perfusion findings. Consolidation rate was assessed at mean follow-up of 3 months by CT evaluation. In 50 cases (83.2%), complete medical and radiological history could be obtained. Preoperative ge-MRI was reevaluated by a blinded radiologist for advanced analysis of sensitivity and specificity. RESULTS: Preoperative ge-MRI (initial finding, IF) showed 23 avascular, 20 malperfused, and seven vital PP. Blinded radiological follow-up (second finding, SF) revealed 14 avascular, 28 malperfused, and 8 vital PP, with a concordance of 65.3% (n = 35). After correlation with the intra-op findings, a specificity of preoperative ge-MRI of 76.5% (IF) and 88.2 (SF), respectively, was revealed for exclusion of avitality. For detection of malperfusion, there was a sensitivity of 92.7% (IF) and 85.4% (SF), respectively. Complete OC was seen 12 weeks postoperatively in 37 (73.5%), partial OC in 9 (18.3%), and nonunion in 4 cases (8.2%) on CT-scans. Of the 41 malperfused/avascular PP, 31 (75.6%) progressed to complete and 6 (14.6%) to partial (at least 2 adjacent CT-layers of 2 mm) OC, with 4 nonunions. CONCLUSION: The sensitivity and specificity of ge-MRI for detection/ exclusion of malperfusion/avitality of the PP was lower than expected. Therewith, the intraoperative assessment of PP perfusion regains a high value in decision-making for the appropriate graft. We recommend preservation of the dorsal radial vascular plexus initially until the vascularity of the proximal pole has been estimated. Patient education for all contingencies and retraction options should be obtained.

A Retrospective Matched-Pair Analysis of Long-term Outcomes after Successful Lower Extremity Free Flap Salvage.

BACKGROUND: Pedicle occlusion with total free flap loss after microvascular lower extremity reconstruction is a considerably rare yet devastating complication. Fortunately, in the majority of cases, emergency salvage takebacks of compromised free flaps are initiated in a timely manner. In this report, we present our analysis of long-term outcomes following transient vascular compromise mitigated through successful free flap salvage in the lower extremity. METHODS: We performed a single-center retrospective matched-pair analysis of 46 patients with lower extremity free flap reconstructions. Cases underwent successful revisions of microvascular compromise (n = 23), whereas controls had uneventful postoperative courses (n = 23). Patient-reported outcome questionnaires and physical evaluations were used to assess general quality of life, functional outcomes, and cosmesis (Lower Extremity Functional Scale [LEFS], Lower Limb Outcomes Questionnaire [LLOQ], Short Form 36 (SF-36), Vancouver Scar Scale [VSS]). The mean follow-up time was 4.4 years. RESULTS: The health-related quality of life assessed by the SF-36 did not differ significantly between both groups in any of the subscales (p ≥ 0.15 for all subscales). Functional outcomes did not show significant differences between both groups according to the LEFS (p = 0.78) and LLOQ (p = 0.45). The overall scar appearance assessed by the VSS showed significantly poorer cosmesis in the re-exploration group (p = 0.014). CONCLUSION: Salvage of compromised free flaps in the lower extremity yields similar long-term outcomes compared to noncompromised free flaps with regard to function and quality of life. However, free flap revisions may lead to impaired scar formation. This study provides further evidence that the opportunity for urgent re-exploration is indispensable.

A longitudinal study evaluating interim assessment of neoadjuvant chemotherapy for bladder cancer.

OBJECTIVES: To evaluate the usefulness of radiological re-staging after two and four cycles of neoadjuvant chemotherapy (NAC), the impact of re-staging on further patient management, and the correlation between clinical and final pathological tumour stage at radical cystectomy (RC). PATIENTS AND METHODS: We conducted a longitudinal, single-centre, cohort study of prospectively collected consecutive patients who underwent NAC and RC for urothelial muscle-invasive bladder cancer between July 2001 and December 2017. Patients underwent repeated computed tomography scans for re-staging after two cycles of NAC and after completion of NAC before RC. RESULTS: Of 180 patients, 110 had ≥four cycles of NAC and had complete imaging available. In the entire cohort, further patient management was only changed in 2/180 patients (1.1%) after two cycles of NAC based on radiological findings. Patients who were stable after two cycles but then downstaged after at least four cycles of NAC had a similarly lowered risk of death (hazard ratio [HR] 0.53). Only one patient downstaged after two cycles was subsequently upstaged after four cycles. Clinical downstaging was observed in 51 patients (46%), 55 patients (50%) had no change in clinical stage and four patients (3.6%) were clinically upstaged. Patients clinically downstaged after four cycles of NAC had a lower risk of death (HR 0.49, 95% confidence interval 0.25-0.94; P = 0.033) compared to those with no change or upstaged after completion of NAC. CONCLUSIONS: Re-staging of muscle-invasive bladder cancer after two cycles of NAC offers little additional information, rarely changes patient management, and may therefore be omitted, whereas re-staging after completion of NAC by CT is a strong predictor of overall survival.

A comparative study of peri-operative outcomes for 100 consecutive post-chemotherapy and primary robot-assisted and open retroperitoneal lymph node dissections.

PURPOSE: To describe and compare differences in peri-operative outcomes of robot-assisted (RA-RPLND) and open (O-RPLND) retroperitoneal lymph node dissection performed by a single surgeon where chemotherapy is the standard initial treatment for Stage 2 or greater non-seminomatous germ cell tumour. METHODS: Review of a prospective database of all RA-RPLNDs (28 patients) and O-RPLNDs (72 patients) performed by a single surgeon from 2014 to 2020. Peri-operative outcomes were compared for patients having RA-RPLND to all O-RPLNDs and a matched cohort of patients having O-RPLND (20 patients). Further comparison was performed between all patients in the RA-RPLND group (21 patients) and matched O-RPLND group (18 patients) who had previous chemotherapy. RA-RPLND was performed for patients suitable for a unilateral template dissection. O-RPLND was performed prior to the introduction of RA-RPLND and for patients not suitable for RA-RPLND after its introduction. RESULTS: RA-RPLND showed improved peri-operative outcomes compared to the matched cohort of O-RPLND-median blood loss (50 versus 400 ml, p < 0.00001), operative duration (150 versus 195 min, p = 0.023) length-of-stay (1 versus 5 days, p < 0.00001) and anejaculation (0 versus 4, p = 0.0249). There was no statistical difference in complication rates. RA-RPLND had lower median lymph node yields although not significant (9 versus 13, p = 0.070). These improved peri-operative outcomes were also seen in the post-chemotherapy RA-RPLND versus O-RPLND analysis. There were no tumour recurrences seen in either group with median follow-up of 36 months and 60 months, respectively. CONCLUSIONS: Post-chemotherapy RA-RPLND may have decreased blood loss, operative duration, hospital length-of-stay and anejaculation rates in selected cases and should, therefore, be considered in selected patients. Differences in oncological outcomes require longer term follow-up.

Genetics of testicular cancer: a review.

PURPOSE OF REVIEW: Testicular germ cell tumours (TGCTs) are the most common solid malignant cancer diagnosed in young males and the incidence is increasing. Understanding the genetic basis of this disease will help us to navigate the challenges of early detection, diagnosis, treatment, surveillance, and long-term outcomes for patients. RECENT FINDINGS: TGCTs are highly heritable. Current understanding of germline risk includes the identification of one moderate-penetrance predisposition gene, checkpoint kinase 2 ( CHEK2 ), and 78 low-to-moderate-risk single nucleotide polymorphisms identified in genome-wide-associated studies, which account for 44% of familial risk. Biomarker research in TGCTs has been challenging for multiple reasons: oncogenesis is complex, actionable mutations are uncommon, clonal evolution unpredictable and tumours can be histologically and molecularly heterogeneous. Three somatic mutations have thus far been identified by DNA exome sequencing, exclusively in seminomas: KIT, KRAS and NRAS . Several genetic markers appear to be associated with risk of TGCT and treatment resistance. TP53 mutations appear to be associated with platinum resistance. MicroRNA expression may be a useful biomarker of residual disease and relapse in future. SUMMARY: The biology of testicular germ cells tumours is complex, and further research is needed to fully explain the high heritability of these cancers, as well as the molecular signatures which may drive their biological behaviour.

The prognostic role of extended preoperative hypercoagulability work-up in high-risk microsurgical free flaps: a single-center retrospective case series of patients with heterozygotic factor V Leiden thrombophilia.

INTRODUCTION: Hypercoagulability is associated with an increased risk of microvascular complications and free flap failures. The authors present their experience and approach to diagnosing and treating patients with heterozygotic factor V Leiden (hFVL) thrombophilia undergoing free flap reconstruction. METHODS: Between November 2009 and June 2018, 23 free flap surgeries were performed in 15 hypercoagulable patients with hFVL. According to the timing of perioperative hypercoagulability work-up, they were grouped into flaps with established diagnoses prior to surgery (Group A) versus flaps with unknown diagnoses prior to surgery (Group B). Baseline characteristics and perioperative complications were compared between both groups, including revision surgeries due to microvascular thromboses, acute bleedings, hematomas, flap necroses, and reconstructive failures. RESULTS: HFVL mutations had been confirmed preoperatively in 14 free flap surgeries (61%, Group A), whereas in 9 free flap surgeries (39%, Group B), mutations were only diagnosed postoperatively after the occurrence of microvascular thromboses had warranted extended hypercoagulability work-up. The overall rate of intraoperative flap thromboses was 9% (n = 2), whereas the overall rate of postoperative flap thromboses was 43% (n = 10). The corresponding salvage rates were 100% (n = 2/2) for intraoperative and 40% (n = 4/10) for postoperative pedicle thromboses. A total of five free flaps were lost (22%). Upon comparison, flaps with an unconfirmed diagnosis prior to surgery were at ten times higher risk for developing total necroses (flaps lost in Group B = 4/9 versus Group A = 1/14; OR: 10.4; 95% CI 1.0, 134.7; p = 0.03). CONCLUSION: Meticulous preoperative work-up of patients with any history of hypercoagulability can help reduce free flap loss rates, thus improving surgical outcomes and increasing patient safety.