Publications by authors named "Simon Hughes"

119 Publications

mRNP granule proteins Fmrp and Dcp1a differentially regulate mRNP complexes to contribute to control of muscle stem cell quiescence and activation.

Skelet Muscle 2021 Jul 8;11(1):18. Epub 2021 Jul 8.

Institute for Stem Cell Science and Regenerative Medicine, Bangalore, India.

Background: During skeletal muscle regeneration, satellite stem cells use distinct pathways to repair damaged myofibers or to self-renew by returning to quiescence. Cellular/mitotic quiescence employs mechanisms that promote a poised or primed state, including altered RNA turnover and translational repression. Here, we investigate the role of mRNP granule proteins Fragile X Mental Retardation Protein (Fmrp) and Decapping protein 1a (Dcp1a) in muscle stem cell quiescence and differentiation.

Methods: Using isolated single muscle fibers from adult mice, we established differential enrichment of mRNP granule proteins including Fmrp and Dcp1a in muscle stem cells vs. myofibers. We investigated muscle tissue homeostasis in adult Fmr1-/- mice, analyzing myofiber cross-sectional area in vivo and satellite cell proliferation ex vivo. We explored the molecular mechanisms of Dcp1a and Fmrp function in quiescence, proliferation and differentiation in a C2C12 culture model. Here, we used polysome profiling, imaging and RNA/protein expression analysis to establish the abundance and assembly status of mRNP granule proteins in different cellular states, and the phenotype of knockdown cells.

Results: Quiescent muscle satellite cells are enriched for puncta containing the translational repressor Fmrp, but not the mRNA decay factor Dcp1a. MuSC isolated from Fmr1 mice exhibit defective proliferation, and mature myofibers show reduced cross-sectional area, suggesting a role for Fmrp in muscle homeostasis. Expression and organization of Fmrp and Dcp1a varies during primary MuSC activation on myofibers, with Fmrp puncta prominent in quiescence, but Dcp1a puncta appearing during activation/proliferation. This reciprocal expression of Fmrp and Dcp1a puncta is recapitulated in a C2C12 culture model of quiescence and activation: consistent with its role as a translational repressor, Fmrp is enriched in non-translating mRNP complexes abundant in quiescent myoblasts; Dcp1a puncta are lost in quiescence, suggesting stabilized and repressed transcripts. The function of each protein differs during proliferation; whereas Fmrp knockdown led to decreased proliferation and lower cyclin expression, Dcp1a knockdown led to increased cell proliferation and higher cyclin expression. However, knockdown of either Fmrp or Dcp1a led to compromised differentiation. We also observed cross-regulation of decay versus storage mRNP granules; knockdown of Fmrp enhances accumulation of Dcp1a puncta, whereas knockdown of Dcp1a leads to increased Fmrp in puncta.

Conclusions: Taken together, our results provide evidence that the balance of mRNA turnover versus utilization is specific for distinct cellular states.
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http://dx.doi.org/10.1186/s13395-021-00270-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8265057PMC
July 2021

Knockout of zebrafish desmin genes does not cause skeletal muscle degeneration but alters calcium flux.

Sci Rep 2021 Apr 5;11(1):7505. Epub 2021 Apr 5.

Department of Medical Biology, Hacettepe University Faculty of Medicine, 06100, Ankara, Turkey.

Desmin is a muscle-specific intermediate filament protein that has fundamental role in muscle structure and force transmission. Whereas human desmin protein is encoded by a single gene, two desmin paralogs (desma and desmb) exist in zebrafish. Desma and desmb show differential spatiotemporal expression during zebrafish embryonic and larval development, being similarly expressed in skeletal muscle until hatching, after which expression of desmb shifts to gut smooth muscle. We generated knockout (KO) mutant lines carrying loss-of-function mutations for each gene by using CRISPR/Cas9. Mutants are viable and fertile, and lack obvious skeletal muscle, heart or intestinal defects. In contrast to morphants, knockout of each gene did not cause any overt muscular phenotype, but did alter calcium flux in myofibres. These results point to a possible compensation mechanism in these mutant lines generated by targeting nonsense mutations to the first coding exon.
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http://dx.doi.org/10.1038/s41598-021-86974-wDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8021586PMC
April 2021

Cellular and molecular pathways controlling muscle size in response to exercise.

FEBS J 2021 Mar 23. Epub 2021 Mar 23.

Randall Centre for Cell and Molecular Biophysics, School of Basic and Medical Biosciences, King's College London, UK.

From the discovery of ATP and motor proteins to synaptic neurotransmitters and growth factor control of cell differentiation, skeletal muscle has provided an extreme model system in which to understand aspects of tissue function. Muscle is one of the few tissues that can undergo both increase and decrease in size during everyday life. Muscle size depends on its contractile activity, but the precise cellular and molecular pathway(s) by which the activity stimulus influences muscle size and strength remain unclear. Four correlates of muscle contraction could, in theory, regulate muscle growth: nerve-derived signals, cytoplasmic calcium dynamics, the rate of ATP consumption and physical force. Here, we summarise the evidence for and against each stimulus and what is known or remains unclear concerning their molecular signal transduction pathways and cellular effects. Skeletal muscle can grow in three ways, by generation of new syncytial fibres, addition of nuclei from muscle stem cells to existing fibres or increase in cytoplasmic volume/nucleus. Evidence suggests the latter two processes contribute to exercise-induced growth. Fibre growth requires increase in sarcolemmal surface area and cytoplasmic volume at different rates. It has long been known that high-force exercise is a particularly effective growth stimulus, but how this stimulus is sensed and drives coordinated growth that is appropriately scaled across organelles remains a mystery.
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http://dx.doi.org/10.1111/febs.15820DOI Listing
March 2021

Case series of intradural disc in recurrence of lumbar disc prolapse.

J Surg Case Rep 2021 Feb 27;2021(2):rjaa611. Epub 2021 Feb 27.

Spinal Surgery, The Royal Orthopaedic Hospital Birmingham, Birmingham B31 2AP, UK.

Intradural disc herniation is a rare entity reported at 0.04-1.1% that occurs most commonly in the lumbar spine particularly at L4-L5 region. There is a paucity of literature due to the rarity of this condition. Intradural disc herniations must be considered in the differential diagnosis of prolapsed intervertebral disc disease especially with recent worsening of symptoms and mismatch of unenhanced magnetic resonance induction (MRI) findings. The confirmation is made with intraoperative findings. An intradural disc herniation is most often diagnosed intraoperatively. Contrast enhanced MRI scan is mandatory for pre-operative diagnosis. We report on two cases presenting to our unit in the form of recurrent intradural disc disease following previous lumbar surgery occurring within 3 months of the index procedure in both cases.
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http://dx.doi.org/10.1093/jscr/rjaa611DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7923311PMC
February 2021

Clinical Outcomes of a Randomized Trial of Adaptive Plan-of-the-Day Treatment in Patients Receiving Ultra-hypofractionated Weekly Radiation Therapy for Bladder Cancer.

Int J Radiat Oncol Biol Phys 2021 06 11;110(2):412-424. Epub 2020 Dec 11.

The Institute of Cancer Research, London, United Kingdom.

Purpose: Hypofractionated radiation therapy can be used to treat patients with muscle-invasive bladder cancer unable to have radical therapy. Toxicity is a key concern, but adaptive plan-of the day (POD) image-guided radiation therapy delivery could improve outcomes by minimizing the volume of normal tissue irradiated. The HYBRID trial assessed the multicenter implementation, safety, and efficacy of this strategy.

Methods: HYBRID is a Phase II randomized trial that was conducted at 14 UK hospitals. Patients with T2-T4aN0M0 muscle-invasive bladder cancer unsuitable for radical therapy received 36 Gy in 6 weekly fractions, randomized (1:1) to standard planning (SP) or adaptive planning (AP) using a minimization algorithm. For AP, a pretreatment cone beam computed tomography (CT) was used to select the POD from 3 plans (small, medium, and large). Follow-up included standard cystoscopic, radiologic, and clinical assessments. The primary endpoint was nongenitourinary Common Terminology Criteria for Adverse Events (CTCAE) grade ≥ 3 (≥G3) toxicity within 3 months of radiation therapy. A noncomparative single stage design aimed to exclude ≥30% toxicity rate in each planning group in patients who received ≥1 fraction of radiation therapy. Local control at 3-months (both groups combined) was a key secondary endpoint.

Results: Between April 15, 2014, and August 10, 2016, 65 patients were enrolled (SP, n = 32; AP, n = 33). The median follow-up time was 38.8 months (interquartile range [IQR], 36.8-51.3). The median age was 85 years (IQR, 81-89); 68% of participants (44 of 65) were male; and 98% of participants had grade 3 urothelial cancer. In 63 evaluable participants, CTCAE ≥G3 nongenitourinary toxicity rates were 6% (2 of 33; 95% confidence interval [CI], 0.7%-20.2%) for the AP group and 13% (4 of 30; 95% CI, 3.8%-30.7%) for the SP group. Disease was present in 9/48 participants assessed at 3 months, giving a local control rate of 81.3% (95% CI, 67.4%-91.1%).

Conclusions: POD adaptive radiation therapy was successfully implemented across multiple centers. Weekly ultrahypofractionated 36 Gy/6 fraction radiation therapy is safe and provides good local control rates in this older patient population.
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http://dx.doi.org/10.1016/j.ijrobp.2020.11.068DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8114997PMC
June 2021

Mef2c factors are required for early but not late addition of cardiomyocytes to the ventricle.

Dev Biol 2021 02 25;470:95-107. Epub 2020 Nov 25.

Randall Centre for Cell and Molecular Biophysics, School of Basic and Medical Biosciences and British Heart Foundation Centre of Research Excellence, Faculty of Life Sciences and Medicine, Guy's Campus, King's College London, London, SE1 1UL, UK. Electronic address:

During heart formation, the heart grows and undergoes dramatic morphogenesis to achieve efficient embryonic function. Both in fish and amniotes, much of the growth occurring after initial heart tube formation arises from second heart field (SHF)-derived progenitor cell addition to the arterial pole, allowing chamber formation. In zebrafish, this process has been extensively studied during embryonic life, but it is unclear how larval cardiac growth occurs beyond 3 days post-fertilisation (dpf). By quantifying zebrafish myocardial growth using live imaging of GFP-labelled myocardium we show that the heart grows extensively between 3 and 5 dpf. Using methods to assess cell division, cellular development timing assay and Kaede photoconversion, we demonstrate that proliferation, CM addition, and hypertrophy contribute to ventricle growth. Mechanistically, we show that reduction in Mef2c activity (mef2ca;mef2cb), downstream or in parallel with Nkx2.5 and upstream of Ltbp3, prevents some CM addition and differentiation, resulting in a significantly smaller ventricle by 3 dpf. After 3 dpf, however, CM addition in mef2ca;mef2cb mutants recovers to a normal pace, and the heart size gap between mutants and their siblings diminishes into adulthood. Thus, as in mice, there is an early time window when SHF contribution to the myocardium is particularly sensitive to loss of Mef2c activity.
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http://dx.doi.org/10.1016/j.ydbio.2020.11.008DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7819464PMC
February 2021

Circadian regulation of muscle growth independent of locomotor activity.

Proc Natl Acad Sci U S A 2020 12 23;117(49):31208-31218. Epub 2020 Nov 23.

Randall Centre for Cell and Molecular Biophysics, King's College London, London SE1 1UL, United Kingdom

Muscle tissue shows diurnal variations in function, physiology, and metabolism. Whether such variations are dependent on the circadian clock per se or are secondary to circadian differences in physical activity and feeding pattern is unclear. By measuring muscle growth over 12-h periods in live prefeeding larval zebrafish, we show that muscle grows more during day than night. Expression of dominant negative CLOCK (ΔCLK), which inhibits molecular clock function, ablates circadian differences and reduces muscle growth. Inhibition of muscle contraction reduces growth in both day and night, but does not ablate the day/night difference. The circadian clock and physical activity are both required to promote higher muscle protein synthesis during the day compared to night, whereas markers of protein degradation, messenger RNAs, are higher at night. Proteasomal inhibitors increase muscle growth at night, irrespective of physical activity, but have no effect during the day. Although physical activity enhances TORC1 activity, and the TORC1 inhibitor rapamycin inhibits clock-driven daytime growth, no effect on muscle growth at night was detected. Importantly, day/night differences in 1) muscle growth, 2) protein synthesis, and 3) expression all persist in entrained larvae under free-running constant conditions, indicating circadian drive. Removal of circadian input by exposure to either permanent darkness or light leads to suboptimal muscle growth. We conclude that diurnal variations in muscle growth and metabolism are a circadian property that is independent of, but augmented by, physical activity, at least during development.
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http://dx.doi.org/10.1073/pnas.2012450117DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7733834PMC
December 2020

Myogenin is an essential regulator of adult myofibre growth and muscle stem cell homeostasis.

Elife 2020 10 1;9. Epub 2020 Oct 1.

Randall Centre for Cell and Molecular Biophysics, King's College London, London, United Kingdom.

Growth and maintenance of skeletal muscle fibres depend on coordinated activation and return to quiescence of resident muscle stem cells (MuSCs). The transcription factor Myogenin (Myog) regulates myocyte fusion during development, but its role in adult myogenesis remains unclear. In contrast to mice, zebrafish are viable, but have hypotrophic muscles. By isolating adult myofibres with associated MuSCs, we found that myofibres have severely reduced nuclear number, but increased myonuclear domain size. Expression of fusogenic genes is decreased, Pax7 upregulated, MuSCs are fivefold more numerous and mis-positioned throughout the length of myofibres instead of localising at myofibre ends as in wild-type. Loss of Myog dysregulates mTORC1 signalling, resulting in an 'alerted' state of MuSCs, which display precocious activation and faster cell cycle entry ex vivo, concomitant with upregulation. Thus, beyond controlling myocyte fusion, Myog influences the MuSC:niche relationship, demonstrating a multi-level contribution to muscle homeostasis throughout life.
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http://dx.doi.org/10.7554/eLife.60445DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7599067PMC
October 2020

Cohort profile: King's Health Partners bladder cancer biobank.

BMC Cancer 2020 Sep 25;20(1):920. Epub 2020 Sep 25.

Translational Oncology & Urology Research (TOUR), School of Cancer and Pharmaceutical Sciences, King's College London, Guy's Hospital, 3rd Floor Bermondsey Wing, London, SE1 9RT, UK.

Background: Bladder cancer (BC) is the 9th most common cancer worldwide, but little progress has been made in improving patient outcomes over the last 25 years. The King's Health Partners (KHP) BC biobank was established to study unanswered, clinically relevant BC research questions. Donors are recruited from the Urology or Oncology departments of Guy's Hospital (UK) and can be approached for consent at any point during their treatment pathway. At present, patients with bladder cancer are approached to provide their consent to provide blood, urine and bladder tissue. They also give access to medical records and linkage of relevant clinical and pathological data across the course of their disease. Between June 2017 and June 2019, 531 out of 997 BC patients (53.3%) gave consent to donate samples and data to the Biobank. During this period, the Biobank collected fresh frozen tumour samples from 90/178 surgical procedures (of which 73 were biopsies) and had access to fixed, paraffin embedded samples from all patients who gave consent. Blood and urine samples have been collected from 38 patients, all of which were processed into component derivatives within 1 to 2 h of collection. This equates to 193 peripheral blood mononuclear cell vials; 238 plasma vials, 224 serum vials, 414 urine supernatant vials and 104 urine cell pellets. This biobank population is demographically and clinically representative of the KHP catchment area.

Conclusion: The King's Health Partners BC Biobank has assembled a rich data and tissue repository which is clinically and demographically representative of the local South East London BC population, making it a valuable resource for future BC research.
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http://dx.doi.org/10.1186/s12885-020-07437-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7519499PMC
September 2020

Fgf-driven Tbx protein activities directly induce and to initiate zebrafish myogenesis.

Development 2020 04 28;147(8). Epub 2020 Apr 28.

Randall Centre for Cell and Molecular Biophysics, New Hunt's House, Guy's Campus, King's College London, SE1 1UL, UK

Skeletal muscle derives from dorsal mesoderm formed during vertebrate gastrulation. Fibroblast growth factor (Fgf) signalling cooperates with Tbx transcription factors to promote dorsal mesoderm formation, but their role in myogenesis has been unclear. Using zebrafish, we show that dorsally derived Fgf signals act through Tbx16 and Tbxta to induce slow and fast trunk muscle precursors at distinct dorsoventral positions. Tbx16 binds to and directly activates the and genes, which are required for commitment to myogenesis. Tbx16 activity depends on Fgf signalling from the organiser. In contrast, Tbxta is not required for expression, but binds a specific site upstream of that is not bound by Tbx16 and drives (dependent on Fgf signals) expression in adaxial slow precursors, thereby initiating trunk myogenesis. After gastrulation, when similar muscle cell populations in the post-anal tail are generated from tailbud, declining Fgf signalling is less effective at initiating adaxial myogenesis, which is instead initiated by Hedgehog signalling from the notochord. Our findings suggest a hypothesis for ancestral vertebrate trunk myogenic patterning and how it was co-opted during tail evolution to generate similar muscle by new mechanisms.This article has an associated 'The people behind the papers' interview.
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http://dx.doi.org/10.1242/dev.184689DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7197714PMC
April 2020

Overall survival, disease-specific survival and local recurrence outcomes in patients with muscle-invasive bladder cancer treated with external beam radiotherapy and brachytherapy: a systematic review.

BJU Int 2020 06 1;125(6):780-791. Epub 2020 Apr 1.

Guy's Cancer, Guy's and St. Thomas' NHS Foundation Trust, London, UK.

Introduction: Neoadjuvant chemotherapy followed by radical cystectomy (RC) and pelvic lymph node dissection is the standard radical management for muscle-invasive bladder cancer (MIBC). However, major pelvic surgery is not suitable for all patients and combined modality therapy (CMT) offers an alternative for patients who want to retain their bladder. Brachytherapy (BT), as part of CMT, has been offered in selective cases of bladder cancer.

Objectives: To evaluate the clinical effectiveness of BT for solitary urinary bladder tumours in terms of survival, local recurrence (LR) rates, and adverse events.

Methods: A systematic review was conducted using defined search terms using online databases. Articles that discussed the use of BT as part of multi-modality treatments for MIBC were included.

Results: Searches returned 112 articles of which 20 were deemed suitable for analysis. In all, 15 of the 20 articles reported overall survival (OS) at 5 years, 2747 patients were at risk and 1670 were alive after 5 years (60%): seven studies reported OS at 10 years, with 817 patients at risk and 350 alive at 10 years (42%). Disease-specific survival at 5 years was reported in four studies, with 371 patients at risk and 279 alive (75%) at 5 years. LR rates were reported across all 20 studies and ranged from 0% to 32%.

Conclusion: Brachytherapy as part of CMT for MIBC is not a standard technique. It is an effective treatment in experienced centres for a selected patient population who wish to preserve their bladder. In such patients, CMT-BT is well tolerated with an acceptable safety profile.
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http://dx.doi.org/10.1111/bju.15047DOI Listing
June 2020

Maternal Larp6 controls oocyte development, chorion formation and elevation.

Development 2020 02 26;147(4). Epub 2020 Feb 26.

Randall Centre for Cell and Molecular Biophysics, New Hunt's House, Guy's Campus, King's College London, London SE1 1UL, UK

La-related protein 6 (Larp6) is a conserved RNA-binding protein found across eukaryotes that has been suggested to regulate collagen biogenesis, muscle development, ciliogenesis, and various aspects of cell proliferation and migration. Zebrafish have two Larp6 family genes: and Viable and fertile single and double homozygous and zygotic mutants revealed no defects in muscle structure, and were indistinguishable from heterozygous or wild-type siblings. However, mutant females produced eggs with chorions that failed to elevate fully and were fragile. Eggs from single mutant females showed minor chorion defects, but chorions from eggs laid by double mutant females were more defective than those from single mutants. Electron microscopy revealed defective chorionogenesis during oocyte development. Despite this, maternal zygotic single and double mutants were viable and fertile. Mass spectrometry analysis provided a description of chorion protein composition and revealed significant reductions in a subset of zona pellucida and lectin-type proteins between wild-type and mutant chorions that paralleled the severity of the phenotype. We conclude that Larp6 proteins are required for normal oocyte development, chorion formation and egg activation.
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http://dx.doi.org/10.1242/dev.187385DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7055395PMC
February 2020

The management impact of gallium-tris(hydroxypyridinone) prostate-specific membrane antigen (Ga-THP-PSMA) PET-CT imaging for high-risk and biochemically recurrent prostate cancer.

Eur J Nucl Med Mol Imaging 2020 03 23;47(3):674-686. Epub 2019 Dec 23.

Cancer Imaging Department, School of Biomedical Engineering and Imaging Sciences, King's College London, London, UK.

Purpose: To determine the impact on clinical management of patients with high-risk (HR) prostate cancer at diagnosis and patients with biochemical recurrence (BCR) using a new kit form of Ga-prostate-specific membrane antigen (PSMA), namely tris(hydroxypyridinone) (THP)-PSMA, with positron emission tomography-computed tomography (PET-CT).

Methods: One hundred eighteen consecutive patients (50 HR, 68 BCR) had management plans documented at a multidisciplinary meeting before Ga-THP-PSMA PET-CT. Patients underwent PET-CT scans 60-min post-injection of Ga-THP-PSMA (mean 159 ± 21.2 MBq). Post-scan management plans, Gleason score, prostate-specific antigen (PSA) and PSA doubling time (PSAdt) were recorded.

Results: HR group: 12/50 (24%) patients had management changed (9 inter-modality, 3 intra-modality). Patients with PSA < 20 μg/L had more frequent management changes (9/26, 34.6%) compared with PSA > 20 μg/L (3/24, 12.5%). Gleason scores > 8 were associated with detection of more nodal (4/16, 25% vs 5/31, 16.1%) and bone (2/16, 12.5% vs 2/31, 6.5%) metastases. BCR group: Clinical management changed in 23/68 (34%) patients (17 inter-modality, 6 intra-modality). Forty out of 68 (59%) scans were positive. Positivity rate increased with PSA level (PSA < 0.5 μg/L, 0%; PSA 0.5-1.0 μg/L, 35%; PSA 1.0-5.0 μg/L, 69%; PSA 5.0-10.0 μg/L, 91%), PSAdt of < 6 months (56% vs 45.7%) and Gleason score > 8 (78.9% vs 51.2%).

Conclusions: Ga-THP-PSMA PET-CT influences clinical management in significant numbers of patient with HR prostate cancer pre-radical treatment and is associated with PSA. Management change also occurs in patients with BCR and is associated with PSA and Gleason score, despite lower scan positivity rates at low PSA levels < 0.5 μg/L.
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http://dx.doi.org/10.1007/s00259-019-04643-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7005085PMC
March 2020

Non-uniform Expansion of Epstein-Barr Virus-Specific T-Cells Following Donor Lymphocyte Infusion for Post-transplant Lymphoproliferative Disease.

Front Immunol 2019 29;10:2489. Epub 2019 Oct 29.

Institute for Immunology and Immunotherapy, University of Birmingham, Birmingham, United Kingdom.

Epstein-Barr virus (EBV)-associated post-transplant lymphoproliferative disease (PTLD) is a life-threatening complication of T-lymphocyte deplete allogeneic hematopoietic stem cell transplantation (allo-HSCT). For patients with PTLD refractory to Rituximab, donor lymphocyte infusion (DLI) is established as a successful option for salvage therapy. However, although lymphocyte expansion has been correlated with good clinical outcome following DLI, the specificity and functional characteristics of EBV-specific T-cell responses remain poorly characterized. Here we describe two patients with Rituximab-refractory PTLD complicating T-cell deplete allo-HSCT, both of whom were successfully rescued with 1 × 10/Kg unselected stem cell donor-derived DLI. Prospective analyses revealed that complete clinical and radiological responses were associated with expansion of T and NK cells. Furthermore, EBV MHC tetramer, and interferon gamma analyses revealed a marked increase in EBV-specific T-cell frequency from 4 weeks after DLI. Reactivity was demonstrated against a range of EBV latent and lytic antigens, including those detected in tumor biopsy material. The immunodominant EBV-specific T cell response expanding following infusion matched the dominant response present in the DLI preparations prior to administration. Furthermore, differences in the repertoire of subdominant antigen-specific T-cells were also detected, suggesting that antigen-encounter can shape the immune response. These results demonstrate the value of prospectively studying T-cell responses, by facilitating the identification of important specificities required for clinical efficacy. Applying this approach on a larger scale promises to yield data which may be essential for the optimization of future adoptive immunotherapeutic strategies for PTLD.
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http://dx.doi.org/10.3389/fimmu.2019.02489DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6828838PMC
October 2020

Graham Roberts Study protocol: first 'trials within cohort study' for bladder cancer.

BMJ Open 2019 09 26;9(9):e029468. Epub 2019 Sep 26.

Translational Oncology and Urology Research, King's College London, London, UK.

Introduction: Given the need for more bladder cancer research and the recently observed advantages of introducing the trials within cohort (TwiCs) design, the set-up of the Graham Roberts Study (Roberts Study) will provide valuable infrastructure to answer a wide variety of research questions of a clinical, mechanistic, as well as supportive care nature in the area of bladder cancer.

Methods: Using the TwiCs design, we will recruit patients aged 18 or older who are willing and able to provide signed informed consent and have a diagnosis of new or recurrent bladder cancer into this prospective cohort study. All patients must have a basic understanding of the English language. The following questionnaires will be collected at baseline and every 12 months subsequently: Functional Assessment of Chronic Illness Therapy for Bladder Cancer, the Functional Assessment of Chronic Illness Therapy-Fatigue, the Patient Heath Questionnaire-9, the standardised instrument for a generic health status (EQ-5D-5L), a Short Questionnaire to Assess Health-Enhancing Physical Activity and the Hertfordshire Short Questionnaire to Assess Diet Quality.

Ethics And Dissemination: Due to the nature of this study, we obtained full ethical clearance from the London-Fulham Research Ethics Committee (17/LO1975). All participants must provide full informed consent before recruitment onto the study. The results of this study will be published in peer-reviewed journals and data collected as part of the study will be made available to potential collaborators on an application basis.
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http://dx.doi.org/10.1136/bmjopen-2019-029468DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6773355PMC
September 2019

EGF receptor (EGFR) inhibition promotes a slow-twitch oxidative, over a fast-twitch, muscle phenotype.

Sci Rep 2019 06 25;9(1):9218. Epub 2019 Jun 25.

National Heart and Lung Institute, Imperial College London, London, UK.

A low quadriceps slow-twitch (ST), oxidative (relative to fast-twitch) fiber proportion is prevalent in chronic diseases such Chronic Obstructive Pulmonary Disease (COPD) and is associated with exercise limitation and poor outcomes. Benefits of an increased ST fiber proportion are demonstrated in genetically modified animals. Pathway analysis of published data of differentially expressed genes in mouse ST and FT fibers, mining of our microarray data and a qPCR analysis of quadriceps specimens from COPD patients and controls were performed. ST markers were quantified in C2C12 myotubes with EGF-neutralizing antibody, EGFR inhibitor or an EGFR-silencing RNA added. A zebrafish egfra mutant was generated by genome editing and ST fibers counted. EGF signaling was (negatively) associated with the ST muscle phenotype in mice and humans, and muscle EGF transcript levels were raised in COPD. In C2C12 myotubes, EGFR inhibition/silencing increased ST, including mitochondrial, markers. In zebrafish, egfra depletion increased ST fibers and mitochondrial content. EGF is negatively associated with ST muscle phenotype in mice, healthy humans and COPD patients. EGFR blockade promotes the ST phenotype in myotubes and zebrafish embryos. EGF signaling suppresses the ST phenotype, therefore EGFR inhibitors may be potential treatments for COPD-related muscle ST fiber loss.
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http://dx.doi.org/10.1038/s41598-019-45567-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6592914PMC
June 2019

NICE guidelines on prostate cancer 2019.

BJU Int 2019 07;124(1)

Guy's and St. Thomas NHS Foundation Trust, London, UK.

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http://dx.doi.org/10.1111/bju.14815DOI Listing
July 2019

Scleraxis genes are required for normal musculoskeletal development and for rib growth and mineralization in zebrafish.

FASEB J 2019 08 17;33(8):9116-9130. Epub 2019 May 17.

Randall Centre for Cell and Molecular Biophysics, School of Basic and Medical Biosciences, Faculty of Life Sciences and Medicine, King's College London, London, United Kingdom.

Tendons are an essential part of the musculoskeletal system, connecting muscle and skeletal elements to enable force generation. The transcription factor scleraxis marks vertebrate tendons from early specification. -null mice are viable and have a range of tendon and bone defects in the trunk and limbs but no described cranial phenotype. We report the expression of zebrafish scleraxis orthologs: scleraxis homolog ( and in cranial and intramuscular tendons and in other skeletal elements. Single mutants for either or generated by clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 9 (Cas9), are viable and fertile as adult fish. Although mutants show no obvious phenotype, mutant embryos have defects in cranial tendon maturation and muscle misalignment. Mutation of both scleraxis genes results in more severe defects in cranial tendon differentiation, muscle and cartilage dysmorphogenesis and paralysis, and lethality by 2-5 wk, which indicates an essential function of scleraxis for craniofacial development. At juvenile and adult stages, ribs in mutants fail to mineralize and/or are small and heavily fractured. mutants also have smaller muscle volume, abnormal swim movement, and defects in bone growth and composition. Scleraxis function is therefore essential for normal craniofacial form and function and vital for fish development.-Kague, E., Hughes, S. M., Lawrence, E. A., Cross, S., Martin-Silverstone, E., Hammond, C. L., Hinits, Y. Scleraxis genes are required for normal musculoskeletal development and for rib growth and mineralization in zebrafish.
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http://dx.doi.org/10.1096/fj.201802654RRDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6662971PMC
August 2019

Total spondylectomy.

J Spine Surg 2018 Sep;4(3):663-665

Royal Orthopaedic Hospital, The woodlands, Bristol Road South, Birmingham, UK.

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http://dx.doi.org/10.21037/jss.2018.06.12DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6261760PMC
September 2018

Myogenin promotes myocyte fusion to balance fibre number and size.

Nat Commun 2018 10 12;9(1):4232. Epub 2018 Oct 12.

Randall Centre for Cell and Molecular Biophysics, King's College London, London, SE1 1UL, UK.

Each skeletal muscle acquires its unique size before birth, when terminally differentiating myocytes fuse to form a defined number of multinucleated myofibres. Although mice in which the transcription factor Myogenin is mutated lack most myogenesis and die perinatally, a specific cell biological role for Myogenin has remained elusive. Here we report that loss of function of zebrafish myog prevents formation of almost all multinucleated muscle fibres. A second, Myogenin-independent, fusion pathway in the deep myotome requires Hedgehog signalling. Lack of Myogenin does not prevent terminal differentiation; the smaller myotome has a normal number of myocytes forming more mononuclear, thin, albeit functional, fast muscle fibres. Mechanistically, Myogenin binds to the myomaker promoter and is required for expression of myomaker and other genes essential for myocyte fusion. Adult myog mutants display reduced muscle mass, decreased fibre size and nucleation. Adult-derived myog mutant myocytes show persistent defective fusion ex vivo. Myogenin is therefore essential for muscle homeostasis, regulating myocyte fusion to determine both muscle fibre number and size.
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http://dx.doi.org/10.1038/s41467-018-06583-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6185967PMC
October 2018

2018: A Spacer Odyssey.

Authors:
Simon R Hughes

BJU Int 2018 09;122(3):353-354

Oncology Department, Guy's and St. Thomas' NHS Trust and Translational Urology Oncology Research Group, School of Cancer and Pharmaceutical Sciences, King's College London, London, UK.

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http://dx.doi.org/10.1111/bju.14356DOI Listing
September 2018

Correction to: Association of invasion-promoting tenascin-C additional domains with breast cancers in young women.

Breast Cancer Res 2018 08 1;20(1):80. Epub 2018 Aug 1.

Department of Cancer Studies and Molecular Medicine, University of Leicester, Infirmary Close, Robert Kilpatrick Clinical Sciences Building, Leicester Royal Infirmary, Leicester, LE2 7LX, UK.

After the publication of this work [1] an error was noticed in Fig. 6 (b). In the MCF-7/Vector columns, the same image was used accidentally for the 0 h and 24 h time points. Both images were taken from the 0 h time point.
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http://dx.doi.org/10.1186/s13058-018-0997-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6071433PMC
August 2018

Urologic Complications Following Pelvic Radiotherapy.

Urology 2018 Dec 20;122:1-9. Epub 2018 Jul 20.

Department of Urology, Guy's Hospital, London, United Kingdom. Electronic address:

Radiotherapy is widely used as a curative treatment for pelvic malignancies. The location of these organs leads to unavoidable exposure of the bladder, urethra, and distal ureters to radiation and may cause subsequent development of radiation cystitis, fistulae, strictures, and secondary malignancy. As cancer survival improves, an ever-increasing number of patients is living with the long-term complications of radiotherapy. Symptoms are not only debilitating for the patient but also pose a treatment challenge to the urologist. This review provides a comprehensive overview of the pathophysiology, clinical presentation, and current management strategies for the urologic complications following pelvic radiotherapy.
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http://dx.doi.org/10.1016/j.urology.2018.07.017DOI Listing
December 2018

Imaging αβ integrin expression in skeletal metastases with Tc-maraciclatide single-photon emission computed tomography: detection and therapy response assessment.

Eur J Nucl Med Mol Imaging 2018 06 2;45(6):898-903. Epub 2018 Feb 2.

Cancer Imaging Department, School of Biomedical Engineering and Imaging Sciences, King's College London, London, UK.

Purpose: Osteoclast activity is an important factor in the pathogenesis of skeletal metastases and is a potential therapeutic target. This study aimed to determine if selective uptake of Tc-maraciclatide, a radiopharmaceutical targeting αβ integrin, occurs in prostate cancer (PCa) bone metastases and to observe the changes following systemic therapy.

Methods: The study group comprised 17 men with bone-predominant metastatic PCa who underwent whole-body planar and single-photon emission computed tomography/computed tomography (SPECT/CT) imaging with Tc-maraciclatide before (n = 17) and 12 weeks after (n = 11) starting treatment with abiraterone. Tumour to normal bone (T:N) ratios, tumour to muscle (T:M) ratios and CT Hounsfield units (HU) were measured in up to five target metastases in each subject. An oncologist blinded to study scans assessed clinical responses up to 24 weeks using conventional criteria.

Results: Before treatment, metastases showed specific Tc-maraciclatide accumulation (mean planar T:N and T:M ratios 1.43 and 3.06; SPECT T:N and T:M ratios 3.1 and 5.19, respectively). Baseline sclerotic lesions (389-740 HU) showed lower T:M ratios (4.22 vs. 7.04, p = 0.02) than less sclerotic/lytic lesions (46-381 HU). Patients with progressive disease (PD; n = 5) showed increased planar T:N and T:M ratios (0.29 and 12.1%, respectively) and SPECT T:N and T:M ratios (11.9 and 20.2%, respectively). Patients without progression showed decreased planar T:N and T:M ratios (0.27 and -8.0%, p = 1.0 and 0.044, respectively) and SPECT T:N and T:M ratios (-21.9, and -27.2%, p = 0.3 and 0.036, respectively). The percentage change in CT HU was inversely correlated with the percentage change in SPECT T:M ratios (r = -0.59, p = 0.006).

Conclusions: Tc-maraciclatide accumulates in PCa bone metastases in keeping with increased αβ integrin expression. Greater activity in metastases with lower CT density suggests that uptake is related to osteoclast activity. Changes in planar and SPECT T:M ratios after 12 weeks of treatment differed between patients with and without PD and Tc-maraciclatide imaging may be a potential method for assessing early response.
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http://dx.doi.org/10.1007/s00259-017-3926-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5915496PMC
June 2018

Mef2 and the skeletal muscle differentiation program.

Semin Cell Dev Biol 2017 12 22;72:33-44. Epub 2017 Nov 22.

Randall Division of Cell and Molecular Biophysics, New Hunt's House, Guy's Campus, King's College London, London SE1 1UL UK.

Mef2 is a conserved and significant transcription factor in the control of muscle gene expression. In cell culture Mef2 synergises with MyoD-family members in the activation of gene expression and in the conversion of fibroblasts into myoblasts. Amongst its in vivo roles, Mef2 is required for both Drosophila muscle development and mammalian muscle regeneration. Mef2 has functions in other cell-types too, but this review focuses on skeletal muscle and surveys key findings on Mef2 from its discovery, shortly after that of MyoD, up to the present day. In particular, in vivo functions, underpinning mechanisms and areas of uncertainty are highlighted. We describe how Mef2 sits at a nexus in the gene expression network that controls the muscle differentiation program, and how Mef2 activity must be regulated in time and space to orchestrate specific outputs within the different aspects of muscle development. A theme that emerges is that there is much to be learnt about the different Mef2 proteins (from different paralogous genes, spliced transcripts and species) and how the activity of these proteins is controlled.
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http://dx.doi.org/10.1016/j.semcdb.2017.11.020DOI Listing
December 2017

Response Rate to Chemotherapy After Immune Checkpoint Inhibition in Metastatic Urothelial Cancer.

Eur Urol 2018 02 13;73(2):149-152. Epub 2017 Sep 13.

Barts Cancer Institute, Queen Mary University of London, London, UK. Electronic address:

Immune checkpoint inhibitors (ICIs) are active in metastatic urothelial carcinoma (MUC). They have joined chemotherapy (CT) as a standard of care. Here, we investigate the activity of CT after progression on ICIs. Two cohorts of sequential patients with MUC were described (n=28). Cohort A received first-line ICIs followed by CT after progression. Cohort B received CT after failure of first-line platinum-based CT followed by ICIs. Response rate (RR) to CT was assessed using Response Evaluation Criteria in Solid Tumors (RECIST v1.1) by a designated radiologist. Best RR for cohort A was 64%. Two patients experienced clinical progression and died before the first radiographic assessment. RR for cohort B was 21%, which was significantly lower than that for cohort A. Progression of disease occurred in 43% of cohort B patients by the end of CT. These data suggest a lack of cross resistance between CT and ICIs in MUC. Therefore, the sequencing of these drugs is likely to be important to maximise outcomes. This is particularly true after first-line ICIs as subsequent CT has significant activity.

Patient Summary: In this report, we studied the effect of chemotherapy in metastatic bladder cancer, which relapsed after immune checkpoint inhibitors. We found that the activity of chemotherapy was maintained despite previous exposure to immune therapy. This underlines the importance of sequencing these agents to maximise outcomes.
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http://dx.doi.org/10.1016/j.eururo.2017.08.022DOI Listing
February 2018

Myotome adaptability confers developmental robustness to somitic myogenesis in response to fibre number alteration.

Dev Biol 2017 11 5;431(2):321-335. Epub 2017 Sep 5.

Randall Division of Cell and Molecular Biophysics, New Hunt's House, Guy's Campus, King's College London, London SE1 1UL, UK. Electronic address:

Balancing the number of stem cells and their progeny is crucial for tissue development and repair. Here we examine how cell numbers and overall muscle size are tightly regulated during zebrafish somitic muscle development. Muscle stem/precursor cell (MPCs) expressing Pax7 are initially located in the dermomyotome (DM) external cell layer, adopt a highly stereotypical distribution and thereafter a proportion of MPCs migrate into the myotome. Regional variations in the proliferation and terminal differentiation of MPCs contribute to growth of the myotome. To probe the robustness of muscle size control and spatiotemporal regulation of MPCs, we compared the behaviour of wild type (wt) MPCs with those in mutant zebrafish that lack the muscle regulatory factor Myod. Myod mutants form one third fewer multinucleate fast muscle fibres than wt and show a significant expansion of the Pax7 MPC population in the DM. Subsequently, myod mutant fibres generate more cytoplasm per nucleus, leading to recovery of muscle bulk. In addition, relative to wt siblings, there is an increased number of MPCs in myod mutants and these migrate prematurely into the myotome, differentiate and contribute to the hypertrophy of existing fibres. Thus, homeostatic reduction of the excess MPCs returns their number to normal levels, but fibre numbers remain low. The GSK3 antagonist BIO prevents MPC migration into the deep myotome, suggesting that canonical Wnt pathway activation maintains the DM in zebrafish, as in amniotes. BIO does not, however, block recovery of the myod mutant myotome, indicating that homeostasis acts on fibre intrinsic growth to maintain muscle bulk. The findings suggest the existence of a critical window for early fast fibre formation followed by a period in which homeostatic mechanisms regulate myotome growth by controlling fibre size. The feedback controls we reveal in muscle help explain the extremely precise grading of myotome size along the body axis irrespective of fish size, nutrition and genetic variation and may form a paradigm for wider matching of organ size.
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http://dx.doi.org/10.1016/j.ydbio.2017.08.029DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5667637PMC
November 2017

CRISPR/Cas9 editing reveals novel mechanisms of clustered microRNA regulation and function.

Sci Rep 2017 08 17;7(1):8585. Epub 2017 Aug 17.

King's British Heart Foundation Centre, King's College London, London, UK.

MicroRNAs (miRNAs) are important regulators of diverse physiological and pathophysiological processes. MiRNA families and clusters are two key features in miRNA biology. Here we explore the use of CRISPR/Cas9 as a powerful tool to delineate the function and regulation of miRNA families and clusters. We focused on four miRNA clusters composed of miRNA members of the same family, homo-clusters or different families, hetero-clusters. Our results highlight different regulatory mechanisms in miRNA cluster expression. In the case of the miR-497~195 cluster, editing of miR-195 led to a significant decrease in the expression of the other miRNA in the cluster, miR-497a. Although no gene editing was detected in the miR-497a genomic locus, computational simulation revealed alteration in the three dimensional structure of the pri-miR-497~195 that may affect its processing. In cluster miR-143~145 our results imply a feed-forward regulation, although structural changes cannot be ruled out. Furthermore, in the miR-17~92 and miR-106~25 clusters no interdependency in miRNA expression was observed. Our findings suggest that CRISPR/Cas9 is a powerful gene editing tool that can uncover novel mechanisms of clustered miRNA regulation and function.
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http://dx.doi.org/10.1038/s41598-017-09268-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5561095PMC
August 2017

The Cannabinoid Receptor Interacting Proteins 1 of zebrafish are not required for morphological development, viability or fertility.

Sci Rep 2017 07 7;7(1):4858. Epub 2017 Jul 7.

Randall Division of Cell and Molecular Biophysics, New Hunt's House, Guy's Campus, King's College London, London, SE1 1UL, UK.

The Cannabinoid Receptor Interacting Protein 1 (Cnrip1) was discovered as an interactor with the intracellular region of Cannabinoid Receptor 1 (CB1R, also known as Cnr1 or CB1). Functional assays in mouse show cannabinoid sensitivity changes and Cnrip1 has recently been suggested to control eye development in Xenopus laevis. Two Cnrip1 genes are described in zebrafish, cnrip1a and cnrip1b. In situ mRNA hybridisation revealed accumulation of mRNA encoding each gene primarily in brain and spinal cord, but also elsewhere. For example, cnrip1b is expressed in forming skeletal muscle. CRISPR/Cas9 genome editing generated predicted null mutations in cnrip1a and cnrip1b. Each mutation triggered nonsense-mediated decay of the respective mRNA transcript. No morphological or behavioural phenotype was observed in either mutant. Moreover, fish lacking both Cnrip1a and Cnrip1b both maternally and zygotically are viable and fertile and no phenotype has so far been detected despite strong evolutionary conservation over at least 400 Myr.
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http://dx.doi.org/10.1038/s41598-017-05017-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5501828PMC
July 2017
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