Forschungstag der Universitäts-Kinderkliniken Bern

16. Oktober 2019

Kuppelraum

Hauptgebäude

Universität Bern

Forschungstag der Universitäts-Kinderkliniken Bern

Datum

Mittwoch, 16. Oktober 2019

Ort

Kuppelraum, Hauptgebäude, Universität Bern

Organisation

Birgit Seyfried

Forschungssekretariat

Universitätsspital für Kinderheilkunde

Inselspital, Universitätsspital Bern

Tel: +41 31 632 94 93

E-Mail: birgit.seyfried@insel.ch

9:00Begrüssung

Christa Flück IT-Support, Florian Singer

Orale Präsentationen 1, Neurologie

Chairs: Karen Lidzba, Rahel Kasteler

9:15

Janine Spitzhüttl

Impact of Non-CNS Childhood Cancer on Resting-State Connectivity

and its Association with Cognition

9:27

Leonie Steiner

Resting state functional connectivity and motor functions in patients

with and without hemiparesis after pediatric arterial ischemic stroke

9:39

Celine Hochstrasser (geb. Webland)

A new test for assessing processing speed in children: Swiss validation of

the c-SDMT

9:47Quendresa Thaqi

The impact of age at arterial ischemic stroke on executive functions

9:55

Robin Münger

Focal cerebral arteriopathy in childhood stroke of the posterior circulati-

on – analysis of a population based registry

10:02

Nicola Biesold

Hypoxic-ischemic encephalopathy in adolescents; neuropsychological

impairments and rehabilation outcome - a case report.

10:09

Daniel Brechbühl

Isolated pediatric Neurosarcoidosis presenting as bilateral subacute

complete sensorineural hearing loss

10:16

Cornelia Enzmann

A cross-sectional and longitudinal natural history study of the Swiss

cohort of LAMA2-related congenital muscular dystrophy

10:23Valerie Siegwart

Cognition, behavior, and quality of life in pediatric cancer survivors

10:45 Kaffeepause

Programm Forschungstag der Universitäts-Kinderkliniken Bern

Orale Präsentationen 2, Onkologie

Chairs: Eva Brack, Michele Bernasconi

11:00

Luana Lavieri

Risk prediction of fever in neutropenia in children and adolescents under

chemotherapy for cancer in a prospective, multicenter trial

11:12

Mutlu Kartal-Kaess

High throughput sequencing of CD34+ selected peripheral blood stem

cells in children with various cancer types undergoing autologous stem

cell transplantation

11:24Lukas Meyer-Landolt

Paraneoplastic pruritus in patients with Hodgkin Lymphoma Case report

11:31

Christa König

Continuous monitoring of health data with a wearable device in

pediatric patients undergoing chemotherapy for cancer – a feasibility

pilot study

11:38

Dzhangar Dzhumashev

Screening of peptide ligands for targeted drug delivery to rhabdomyos-

arcoma cells

11:45Andrea Timpanaro

Surfaceome profiling of rhabdomyosarcoma for CAR T cell targets

11:52

Bettina Blank

Challenges of treating osteosarcoma in a 12-years old patient with Sickle

Cell Disease

12:00

Elizaveta Fasler-Kan

Gastrin releasing peptide receptor is overexpressed in Wilms tumour

patients and cell lines

12:15 Lunch

13.30 Gastreferat: Prof. Claudia Kuehni

“Use of hospital routine data for research? Limitations and opportunities”

Short biosketch

Prof. Claudia Kuehni: Medical studies in Bern. Training in paediatrics (FMP Paediatrics) with

subsequent specialisation in paediatric pulmonology in St.Gallen, Delémont and Bern). MSc

in Epidemiology in London (LSHTM, 1996-7), research fellow in Leicester, UK (1997-99).

Current position as research group leader and associate Professor in Paediatric Epidemiolo-

gy at the Institute of Social and Preventive Medicine at the University of Bern.

Claudia Kuehni is an expert in the epidemiology of childhood diseases, especially respirato-

ry diseases, childhood cancer, and rare diseases. Her research focuses on the causes,

treatments and long-term course of common and rare diseases in childhood. She leads

SwissPedRegistry (the registry hub of SwissPednet), and national or multicentre cohort

studies and registries (Leicester Respiratory cohort studies, Swiss Childhood Cancer

Registry, Swiss Rare Disease Registry, Swiss Paediatric Airway Cohort (SPAC), Swiss Primary

Ciliary Dyskinesia Registry, iPCD cohort and many more), and is responsible for the

surveillance of the neonatal screening for Cystic Fibrosis.

Claudia is a member of the research council of the Swiss National Science Foundation

(Division 3), Paediatric section editor for the European Respiratory Journal, and member of

many other boards relevant for research in Child Health.

Orale Präsentationen 3, Pneumologie und Physiotherapie

Chairs: Christiane Sokollik, Florian Singer

14:30

Johanna Kurz

Association of lung clearance index and survival in patients with cystic

fibrosis

14:42

Ruth Stauffer-Lacorcia

Improving opportunities for physical activity and participation in

inpatient settings for adolescent cancer patients

14:54

Bettina Frauchiger

Lung clearance index: a promising biomarker to track lung disease

progression during childhood in patients with cystic fibrosis

15:06

Marc-Alexander Oesterreich

Systematic differences in analysis software versions hamper infant lung

function testing

Orale Präsentationen 4, Gastrologie & Endokrinologie/ Metabolik

Chairs: Christa König, Claudia Böttcher, Matthias Gautschi

16:00

Mariia Borsuk

Lack of aquaporin 9 expression in iPSC-derived hepatocytes impairs urea

secretion

16:12

Shaheena Parween

Effect of metformin on ACTH receptor activation and downstream

signaling

16:24

Mara Grassi

Expanding the phenotype of the p.R63W mutation of HNF4A – a case

report

16:31Emanuele Pignatti

New Mechanistic Insights into Androgen Production

16:38

Tanja Zingg

Level of glycemic control in pediatric patients with type 1 diabetes in

Bern: a cross-sectional study

16:45

Luca Cecchini

Trajectory of microbiota maturation in healthy Bern infants - a network

approach

16:52

Efstathios Katharopoulos

Human-induced steroidogenic cells from urine consist a new cell-based

model to study adrenal disorders in a personalised manner

17:00 Schlusswort und Preisverleihung, anschliessend Apéro

15:13

Andras Soti

Join Effects of prenatal smoke exposure and polymorphism in the

MBL2-gene on cord blood levels of mannos-binding lectin

15:30 Kaffeepause

Resting state functional connectivity and motor functions in patients with and

without hemiparesis after pediatric arterial ischemic stroke

Leonie Steiner1*, Stephanie Winkelbeiner2,3*, Andrea Federspiel2, Salome Kornfeld1,4,

Regula Everts1, Sandeep Kamal1, Juan Delgado Rodriguez1, Nedelina Slavova5, Roland

Wiest5, Maja Steinlin1, Sebastian Grunt1

* The authors contributed equally to this work

1 Division of Neuropaediatrics, Development and Rehabilitation, Children’s University

Hospital, Inselspital, Bern, Switzerland.Division of Systems Neuroscience

2 Translational Research Center, University Hospital of Psychiatry and Psychotherapy,

Bern, Switzerland

3 Center for Psychiatric Neuroscience, The Feinstein Institute for Medical Research,

Manhasset, NY, USA

4 Center for Cognition, Learning and Memory, University of Bern, Bern, Switzerland

5 Institute of Diagnostic and Interventional Neuroradiology, University Hospital, Inselspi-

tal, University of Bern, Bern, Switzerland

Introduction: The objective of this cross-sectional observational study was to investigate

the relationship between brain functional connectivity and severity of sensorimotor

impairments in the upper limb in the acute phase post stroke.

Patients and Methods: Patients with arterial ischemic stroke (diagnosed <16 years, >2 years

after diagnosis) and age-matched healthy controls were recruited from the Swiss Neurope-

diatric Stroke Registry. Eight patients with hemiparesis, ten patients without hemiparesis

and thirteen healthy controls underwent and a clinical sensorimotor assessment and

resting-state functional magnetic resonance imaging (rs-fMRI). Functional connectivity of

the motor network -including the primary motor cortex (M1), dorsal premotor cortex

(PMC), supplementary motor area (SMA), prefrontal cortex (PFC) and the superior parietal

lobus (SPL) - was assessed.

Results: The three groups displayed different functional connectivity indices in terms of

interhemispheric and intrahemispheric connectivity (p = 0.001). Moreover, significant

associations were found between sensorimotor outcome and interhemispheric (r = -0.63)

and intrahemispheric (r = -0.51) network indices.

Conclusion: We found different network patterns in patients with hemiparesis, patients

without hemiparesis and healthy controls in the motor network. These findings can help to

better understand the mechanisms of motor recovery. They are essential for a better

understanding of how targeted therapies after childhood stroke may affect network

changes.

A new test for assessing processing speed in children: Swiss validation of the

c-SDMT

Céline Hochstrasser, Ursina Jufer, Li Mei Cao, Marie-Noëlle Klein, Sarah Rieder, Michelle

Steiner, Sandra Bigi, Karen Lidzba

Neuropediatrics

Background: Processing speed (PS) is a marker for cognitive function in patients with

neurological conditions. Associated with neural maturation, it increases during develop-

ment. Traditionally, PS is measured by paper-pencil tasks requiring adequate fine motor

skills, which are often impaired in patients with neurological conditions. Thus, a non-motor

alternative is desirable. The Candian computerized Symbol-Digit Modalities Test (c-SDMT)

requires the patient to verbally associate numbers with symbols. The examiner measures

speed via key-press. It has been standardized in a large Canadian paediatric sample and

validated in patients with paediatric MS.

Method: 86 neurologically healthy participants (8 - 16 years; 45 male; 48 hospitalized for

non-neurological reasons) were examined from 06/18 – 11/18, 48 of these returned for a

retest two weeks later. All participants performed the written and computerized SDMT and

a nonverbal intelligence screening (TONI-4). Statistical analyses included a comparison of

the Swiss data to the Canadian norms, a multiple regression on the effects of age and

hospitalization on the performance on the (c-)SDMT, ANCOVAs for the effects of age, sex

and IQ, as well as measures of test-retest reliability.

Results: 1. The Swiss data is comparable to the Canadian norms. 2. As expected, age is the

best predictor for performance on (c-)SDMT. 3. Hospitalization influences the performance

on the SDMT but not on the c-SDMT. 4. Sex and IQ do not have an effect. 5. The (c-)SDMT

have an excellent test-retest reliability, however, practice effects are high and have to be

taken into account.

Conclusions: The c-SDMT is a valid and reliable measure of processing speed in children

and adolescents. The Canadian norms can safely be used to evaluate the performance of

Swiss children. The c-SDMT is more feasible for the use in hospitalized populations than the

SDMT. In future studies, we will validate the c-SDMT in selected neurological populations.

The impact of age at arterial ischemic stroke on executive functions

Thaqi, Q.1,2,3, Steinlin, M.1, Roebers, C.2,3, & Everts, R.1,2

1 Division of Neuropediatrics, Development and Rehabilitation, Children’s University

Hospital, Inselspital, University of Bern, 3010 Bern, Switzerland;

2 Swiss Graduate School for Cognition, Learning, and Memory (GSCLM), University of

Bern, 3012 Bern, Switzerland;

3 University of Bern, Department of Psychology, 3012 Bern, Switzerland

The development of executive functions (EF) is closely related to the maturation of the

prefrontal cortex, which is particularly vulnerable to brain injury due to its prolonged

development. EF, such as the ability to inhibit prepotent responses, flexibly shift between

mindsets, and store and manipulate information in short-term memory, have rarely been

investigated following a pediatric arterial ischemic stroke (AIS).

The current study investigated in a cross-sectional study design how the age at AIS

influences the development of EF, namely inhibition, shifting, working memory, and

processing speed after AIS (age at assessment: M = 15.4, range = 8.3 – 22.7; neonatal

stroke < 28 d, early childhood stroke < 7 yr, late childhood stroke > 7 yr).

There was no significant difference between age at stroke groups in fluid intelligence.

Overall, mean values of EF in all groups of patients after AIS were within the normal range.

Patients after neonatal stroke showed significant better shifting and working memory

performance than patients after late childhood stroke. Exploratively, mean values for EF and

processing speed were slightly better in patients after neonatal stroke than in patients after

early and late childhood stroke, except for errors in inhibition and errors in shifting.

The findings in the present study propose that age at AIS has an impact on cognitive

functions. More specific, our results show that a neonatal stroke has a benign impact on

cognitive functions which are not yet initiated at the time of brain injury (i.e. on EF). Thus,

our data point towards better outcome, when AIS occurs before the initiation and establish-

ment of EF development. Shifting and working memory are developing during early and

late childhood. AIS during this critical period of EF development seems to have a more

detrimental impact on outcome.

Focal Cerebral Arteriopathy of Children in Posterior Circulation – Analysis of a

population based registry

Robin Münger, Nedelina Slavova, Iciar Sanchez-Albisua, Maja Steinlin

Department of Pediatric Neurology, University Children’s Hospital, University of Bern,

Switzerland (R.M., I.S., M.S.,) and Department of Neuroradiology, Bern University

Hospital, University of Bern, Switzerland (N.S.).

Background and Purpose: Focal cerebral arteriopathy (FCA) is an important risk factor for

arterial ischemic stroke (AIS) in childhood, known especially in the anterior circulation. AIS

due to FCA in the posterior circulation is less well known and scarcely described. This

population based cohort study aimed to define the incidence of AIS due to FCA-i (infectious

triggered) in the posterior circulation and to compare them to anterior circulation cases.

Methods: We reviewed all cases of childhood AIS (age 1 month to 16 years) in the Swiss

Neuropediatric Stroke Registry from January 2000 until December 2018, with abnormal

magnetic resonance angiography, recent acute Varicella infection, or the presumption of a

vasculitis to identify FCA-i cases. Initial clinical presentation provoking factors, MR imaging

findings and outcome by Pediatric Stroke Outcome Measure (PSOM) were compared

between anterior and posterior cases.

Results: We identified 67 cases of FCA in the SNPSR, with eighteen posterior cases (27%),

resulting in an incidence of 0.08/100 000 (95% CI 0.039 to 0.112) for posterior and

0.28/100 000 (95% CI 0.248-0.317) for all strokes due to FCA-i.

Median age at presentation was significantly different with 7.9 years in the posterior group

and 5.4 years in anterior group. Further significant differences were found in the association

with recent varicella infections (16.7% vs. 49%, p 0.024), clinical manifestation as facial

palsy (22.2% vs. 73.5%, p 0.0002), headache (72.2% vs. 22.5%, p 0.0003), ataxia (61.1%

vs. 14.3%, p 0.0003) and nausea and vomiting (61.1% vs. 20.4%, p 0.003).

Conclusion: Children presenting with posterior stroke due to FCA-i are older, present with

more subtle symptoms but do not differ in outcome. One fourth of FCA-i cases affected the

posterior circulation, this points to the importance to include posterior FCA-i in differential

diagnosis and in future research.

Hypoxic-ischemic encephalopathy in adolescents; neuropsychological impair-

ments and rehabilitation outcome - a case report.

Nicola Biesold (M. Sc. Psychology), Co-Author: Prof. Dr. K. Lidzba

Neuropediatrics, Development and Rehabilitation

This case study investigates the relationship between brain scans and neuropsychological

impairments of patients with hypoxic-ischemic encephalopathy (HIE) and the importance

of multidisciplinary therapies.

HIE, also known as hypoxic brain injury is caused by deprivation of oxygen to the brain.

Thorax traumas with hypoxic encephalopathy in children are very rare, normally caused by

accidents. Yet, professional treatment in the first minutes is crucial for survival and

outcome. Otherwise the grey matter, the most vulnerable structure, can be damaged;

specifically in the thalamus, the basal ganglia, the cerebral cortex, the hippocampus and the

cerebellum. Cognitive deficits, such as memory problems, executive dysfunction and

impairment of attention are likely.

This report describes the case of a 15 years old male adolescent. He got half buried under a

roof avalanche leading to a hypoxic encephalopathy with asphyxia by thorax trauma and

aspiration pneumonia basal on both sides. Out of hospital reanimation was executed after

10 minutes. Resulting consequences were a posttraumatic/post-ischemic delir with

normothermia protocol (1 day) and severe neuropsychological deficits such as a persistent

anterograde amnesia. However, CT- and MRI-scans of the brain gave no evidence of HIE,

specifically in the hippocampus. Prognosis was uncertain. After three days of intensive care

followed by one-month early rehabilitation, a 5-months intensive rehabilitation period,

including multidisciplinary therapies, was carried out. Improvements in all cognitive

domains were registered but memory problems persisted. Nevertheless, reintegration into

work and study environment was successful due to the good interdisciplinary collaboration.

This single case report gives important insights into the treatment of traumatic brain

injuries. It raises the question of how valid brain scans are in terms of neuropsychological

deficits. Furthermore, it helps to discuss and improve current knowledge on how to help

children with traumatic brain injuries to integrate back into their usual environment

successfully.

Key words: hypoxic encephalopathy, thorax trauma, cognitive deficits, children/adolescent

Isolated pediatric Neurosarcoidosis presenting as bilateral subacute complete

sensorineural hearing loss

D. Brechbühl (1), N. Schöbi (2), G. Mantokoudis (3), M. Steinlin (1)

Division of Neuropediatrics (1), Infectiology,(2) Department of Pediatrics, and ENT-de-

partment (3) Inselspital, Bern University Hospital, University of Bern, Bern, Switzerland)

Case Report: A 10-year-old boy was reffered for progressive hearing loss over the last 4

months. At examination he showed bilateral subacute complete sensorineural hearing loss,

but was otherwise besides significant fatigue free of signs and symptoms. A brain MRI

showed generalized cerebral and spinal leptomeningeal enhancement with bilateral

enhancement of the III, VII and VIII cranial nerve. There was also enhancement of the

internal auditory canal with postcontrast enhancement of the cochlea and the vestibulum.

The cerebrospinal fluid (CSF) analysis showed mild increase of monocytic cells (7) and

increase of protein- and was consistent with blood brain barrier dysfunction, his CSF

angiotensin converting enzyme (ACE) was elevated at 10.16 U/l (0 to 2 U/l). A combined

dural, leptomeningeal and cortical biopsy established the diagnosis of neurosarcoidosis,

showing epitheloid granuloma.

After treatment 2 weeks of high dose steroids there was significant decrease of leptomenin-

geal enhancement, but persistent hearing loss. CSF-monitoring showed normalization of

both the CSF ACE and the blood brain barrier. Treatment was enforced by infliximab and

the patient underwent cochlear implantation to prevent hearing deprivation.

Discusssion: Isolated Neurosarcoidosis is a rare disease mimicking central nervous system

(CNS) infection, inflammatory disorders and neoplasia. Earlier reported manifestations in a

pediatric population include progressive cranial neuropathy (most commonly the VII, but

also involvement of optic, III and VIII nerve are reported). Additional symptoms might be

seizures and hypothalamic dysfunction. Studies showed poor sensitivity and specificity of

CSF-ACE, remaining leptomingeal/brain biopsy the most reliable approach for diagnosis of

Neurosarcoidosis . Early diagnosis would be important to prevent progressive symptoms.

A cross-sectional and longitudinal natural history study of the Swiss cohort of

LAMA2-related congenital muscular dystrophy

Cornelia Enzmann1,2, Andrea Klein1,2,3

1 Pediatric neurology, University Hospital Bern, Inselspital, Switzerland

2 Pediatric neurology, University Children’s Hospital Basel, UKBB, University of Basel,

Switzerland

3 Pediatric neurology, University Hospital Lausanne, CHUV, Lausanne, Switzerland

We report about a planned cross-sectional and longitudinal natural history study. Congeni-

tal muscular dystrophy due to mutations in the LAMA2 gene, also known as Merosin-nega-

tive muscular dystrophy, is one of the most common congenital muscular dystrophies.

LAMA2 encodes for the protein Laminin α2, which belongs to the extracellular matrix.

Missing Laminin α2 leads to an instable connection between the basement membrane and

the sarcolemma of the muscle cell. The disease is diagnosed on the basis of absent or

reduced merosin in immunohistochemistry staining in muscle biopsy, and/or by genetic

testing. Affected children show hypotonia at birth, delayed motor milestones, severe

muscle weakness and usually never achieve independent walking. To date, no pharmacolo-

gical treatment is available, but there are promising results in basic research and a first

clinical trial with omigapil was already conducted in a small cohort. There is little data about

the exact prevalence and the natural disease course.

In this study we aim to increase the knowledge about the natural history of LAMA2-related

MD, provide data about the prevalence in Switzerland and the phenotypic spectrum.

Additionally we want to evaluate different physiotherapeutic assessment tools, to

document clinically relevant changes in MDC1A.

We will evaluate patients with genetically confirmed MDC1A. Data will be collected/ stored

in the existing Swiss neuromuscular registry. A clinically relevant dataset will be collected

over three years with yearly follow-ups. Descriptive statistical analysis will help to characte-

rize the Swiss cohort of MDC1A-patients.

Cognition, behavior, and quality of life in pediatric cancer survivors

Valerie Siegwart1, Valentin Benzing2, Janine S. Spitzhuettl1, 5, Mirko Schmidt2, Michael

Grotzer3, Maja Steinlin1, Kurt Leibundgut4, Claudia Roebers5, Regula Everts1

1 Division of Neuropediatrics, Development and Rehabilitation, University Children`s

Hospital Bern, Inselspital, 3010 Bern, Switzerland

2 Institute of Sport Science, University of Bern, Switzerland

3 University Children`s Hospital Zurich

4 Division of Pediatric Oncology, University Children`s Hospital Bern, Inselspital, 3010

Bern, Switzerland

5 Department of Psychology, University of Bern, Switzerland

Long-term sequelae of cancer and its treatments put pediatric cancer survivors (PCS) at risk

of cognitive and behavioral difficulties, which likely affect quality of life. The aim of the

study was to describe cognition, focusing on executive functions, psychosocial characte-

ristics and health-related quality of life in PCS (with and without central nervous system

(CNS) involvement) and controls. Executive functions are crucial for learning and cognitive

development and contribute strongly to academic achievement and hence merit particular

attention. The influence of processing speed and demographical and clinical risk factors was

considered. 78 PCS, one year or more post-treatment, and 56 controls (aged 7 - 16 years)

were included. Cognitive functioning, behavior and quality of life were assessed using

standardized tests and questionnaires. Results revealed similar fluid intelligence and

significantly poorer executive functions in PCS than controls, with strongest effects in

working memory. Significantly poorer cognitive performance was observed in PCS with

than without CNS involvement reaching its peak in cognitive flexibility. Processing speed

significantly predicted performance in executive functions and CNS involvement was estab-

lished as a key risk factor. Although quality of life was comparable, behavioral difficulties,

such as peer problems, were more pronounced in PCS than controls. Despite evidence for

good intellectual functioning, PCS need to be supervised closely in clinical protocols as

alterations in executive functions may occur years after cancer and likely affect psychosocial

development.

Risk prediction of fever in neutropenia in children and adolescents under

chemotherapy for cancer in a prospective, multicenter trial

Luana Lavieri, Christa Koenig, Roland A Ammann

Division of Pediatric Hematology/Oncology, Department of Pediatrics, Inselspital, Bern

University Hospital, University of Bern, Bern, Switzerland

Background: Fever in severe chemotherapy-induced neutropenia (FN) is the most frequent

potentially lethal complication for children and adolescents with cancer. Risk prediction of

FN would allow for targeted preventive therapies during chemotherapy. This study aimed to

develop risk prediction rules for FN, and FN with safety relevant events (SRE), defined as

serious medical complications (including death, admission to an intensive care unit, severe

sepsis) or bacteremia.

Methods: This multicenter trial prospectively collected data of pediatric patients between 1

to 17.99 years under chemotherapy for cancer. It was designed to compare different

temperature levels for the definition of fever. Patients characteristics and potential risk

factors for FN were collected. Characteristics related to patients (gender, age group), to

diagnosis (type of malignancy, relapse status, bone marrow involvement) and to therapy

and course of disease (chemotherapy intensity, present central venous access device, time

since diagnosis, prior FN episodes) will be analysed with univariate and multivariate mixed

Poisson regression.

Results: Six centers included 269 pediatric patients and 360 episodes of FN, including 72

with SRE, 56 with bacteremia and 30 with serious medical complications. In univariate

analysis, diagnosis of AML, higher chemotherapy intensity, bone marrow involvement and

shorter time since diagnosis were identified as risk factors for FN and FN with SRE. Further

analysis will be added to these preliminary results.

Conclusion: Established and validated pediatric rules predicting the risk to develop FN

during chemotherapy are important to allow specific prophylactic therapies, such as

prophylactic antimicrobial therapy, granulocyte colony-stimulating factor (G-CSF), or other

procedures to protect patients at high risk to develop FN. This study will develop models to

predict the risk for FN, what will allow more specific treatment of individual patients.

High throughput sequencing of CD34+ selected peripheral blood stem cells in

children with various cancer types undergoing autologous stem cell transplantation

Mutlu Kartal-Kaess1,2, Naomi Azur Porret3, Raphael Joncourt3, Vera Ulrike Bacher3,4,

Thomas Pabst5, Gabriela M Baerlocher2,3, Jochen Roessler1,2, Axel Karow1,2

1 Division of Pediatric Hematology & Oncology, Department of Pediatrics, Inselspital,

University Hospital, University of Bern, Bern, Switzerland

2 Department for BioMedical Research, University of Bern, Bern, Switzerland.

3 Department of Hematology and Central Hematology Laboratory, Inselspital, Bern

University Hospital, University of Bern, Bern, Switzerland.

4 Center of Laboratory Medicine (ZLM)/University Institute of Clinical Chemistry,

Inselspital, Bern University Hospital, Bern, Switzerland

5 Department of Medical Oncology, Inselspital, Bern University Hospital, Bern, Switzer-

land

Clonal haematopoiesis of indeterminate potential (CHIP) is present in patients (pts) with

therapy-related myeloid neoplasms at the time of primary cancer diagnosis and before

treatment. Such clones expand under selective pressure from cytotoxic treatment for

primary cancer and can subsequently give rise to overt myeloid neoplasms. CHIP being an

age-related condition, it remains unclear how in children/adolescents cellular stressors such

as cytotoxic therapy influence the expansion of clones carrying mutations in genes

commonly affected in CHIP. Thus, the incidence of CHIP after chemotherapy is less clear in

the paediatric setting.

We retrospectively analysed our cohort of paediatric pts undergoing autologous stem cell

transplantation following intensive chemotherapy for mostly solid tumours. We aimed to

investigate whether the stress from cytotoxic therapy promotes the expansion of clones

with mutations in “CHIP-genes”.

We applied a sensitive error-corrected ion torrent NGS based approach on CD34+ selected

stem cell samples of thirty-three (n=33) children and adolescents stored after stem cell

harvest. The panel comprised 18 genes so far described to be implicated in CHIP including

ASXL1, ATM, CBL, CHEK2, DNMT3A, GNAS, GNB1, IDH1, IDH2, JAK2, MYD88 (L265P),

PPM1D, SF3B1, SRSF2, STAT3, TET2, TP53, U2AF1.

We found genomic aberrations in 6/33 (18%) pts with a high incidence of variants mainly in

ATM but also in CHEK2 with variant allele frequencies (VAFs) ~50% indicating potentially

germline variants. We found CHIP typical findings in 1/33 pts (3%) with a mutation in TP53

(VAF 18%) potentially reflecting CHIP. This patient’s stem cells were harboring two

additional variants of unknown significance (VUS) in ATM and CHEK2. Thus, germline

variants and their accumulation might predispose to the development of solid tumors and/

or pave the way to clonal haematopoiesis. In conclusion, we did not find CHIP at the

reported high frequencies in adults. Further studies of more paediatric cancer pts are

needed.

Paraneoplastic pruritus in patients with Hodgkin Lymphoma. A report of two

cases.

Lukas R. Meyer-Landolt, Tobias Dantonello, Jochen K. Rössler

University Hospital of Bern, Department of Pediatric Hematology and Oncology UZKJO

Introduction: Pruritus is a common clinical complaint encountered in pediatric practice,

most causes being dermatological, allergic or infectious in nature. A rare but important

cause for pruritus is malignancy. We report two cases of patients with Hodgkin lymphoma,

in whom pruritus was the first hallmark of disease leading to diagnosis.

Case presentation: The first patient was a 17y old male who developed severe pruritus over

several weeks. No other systemic symptoms were noted. Antihistamine and topical therapy

showed no effect. Further evaluation showed a large mediastinal mass. Aprepitant, a

neurokinin 1 receptor agonist showed some alleviation of itching in case 1.

The second patient was a 14y old female who presented with cervical lymph node

enlargement for further evaluation. In retrospect, she had suffered from plantar pruritus for

4 weeks. Imaging showed a large mediastinal mass leading to respiratory compromise.

Biopsy and/or lymph node extirpation led to the diagnosis of Hodgkin lymphoma in all

patients. After commencing chemotherapy according to standard protocol resolution of the

lymphoma led to resolution of the pruritus.

Discussion: Paraneoplastic phenomena are clinical symptoms linked to malignancy but not

directly mediated through the neoplastic cells. Chronic pruritus can well be of paraneopla-

stic origin in the pediatric population, described in haematological malignancies and

especially Hodgkin lymphoma. The mechanisms are poorly understood. Inflammatory

cytokines, especially IL-31 and IL-6 have been associated with pruritus and are closely

related with the pathophysiology of lymphoma.

Malignancy should be included in the differential diagnosis of chronic pruritus encountered

in everyday clinical practice, especially when the symptom does not respond to standard

treatment. Pruritus can be the first hallmark of disease before life-threatening complications

develop.

Aprepitant is a promising treatment to alleviate paraneoplastic itching, the most important

therapy being the treatment of the underlying disease.

Continuous monitoring of health data with a wearable device in pediatric patients

undergoing chemotherapy for cancer – a feasibility pilot study

Christa Koenig, Roland A Ammann, Jochen K Roessler, Eva Brack

Division of Pediatric Hematology/Oncology, Department of Pediatrics, Inselspital, Bern

University Hospital, University of Bern, Bern, Switzerland

Background: Pediatric patients with chemotherapy-induced neutropenia are at great risk to

develop severe infections. Delay of diagnosis and treatment can result in increased

mortality. It has been shown that infections can trigger changes of vital signs, as heart rate

variability, very early in their course and before further clinical symptoms. Continuous

health data monitoring may detect such changes earlier than discrete measurements.

Non-invasive on-skin wearable devices (WD) could therefore serve as additional diagnostic

tools. We aim to assess the feasibility of continuous monitoring of heart rate in pediatric

patients undergoing chemotherapy for cancer using a WD.

Methods: This feasibility pilot study will include twenty patients undergoing chemotherapy

for cancer, aged 1 month to 17.99 years. The WD investigated is the Everion® by Biovotion,

a light device, worn with an elastic band on the upper arm or thigh. The primary outcome is

≥acceptable quality of monitored heart rate, during a cumulative duration of ≥18/24h per

day during ≥7 consecutive days. Secondary outcomes include other vital signs, their

comparison with discrete measurements, acceptability by patients and parents, reasons not

to wear the WD, side effects, effort for the investigators and accuracy of measurements by

skin types and during activity. Start of this study is planed for October 2019.

Conclusion: No study has proven feasibility of continuous monitoring of health data in

pediatric patients undergoing chemotherapy for cancer. It is not known if children tolerate

the WD and if data quality sustains. Neither is it known, if parents and patients can handle a

WD in outpatient settings. Knowledge gained from this study will enable potential future

studies, which may identify patterns predicting imminent fever or infection in the near

future. This may lead to earlier diagnosis in patients at high risk for severe infection and the

distinction from patients at lower risk.

Screening of peptide ligands for targeted drug delivery to rhabdomyosarcoma

cells

Dzhangar Dzhumashev, Andrea Timpanaro, Michele Bernasconi, Jochen Rössler

Department of pediatric Hematology and Oncology, Inselspital, Bern University Hospital,

Department for BioMedical Research (DBMR), University of Bern

Rhabdomyosarcoma (RMS) is the most frequent pediatric soft tissue sarcoma. Surgery and

conventional multimodal therapy are not efficient for patients with recurrence or metasta-

ses. Potential approach to improve current therapies is encapsulation of therapeutic agents

into actively targeted nanoparticles. In this study, we aimed to evaluate peptides as

targeting ligands for nanoparticles. We selected from the literature peptides described to be

ligands for proteins that are overexpressed in RMS. These proteins are Integrins, Neuropilin

1, EGFR, Nucleolin, Transferrin. In addition, RMS-targeting peptides, TmR, RMS-P3, RMS-I

and RMS-II, previously discovered by phage display screening, were included.

Biotinylated peptides were conjugated to streptavidin-coated fluorescent Quantum Dots

(Thermo Fisher). Experiments were performed on RMS cell lines and as control on

myoblasts and fibroblasts. In total, 17 different peptides were evaluated. For microscopic

analysis, 10nM QD-peptide conjugates were incubated for 1h or overnight (12h) with cells

seeded on slides (Ibidi). QD were washed and cells were fixed with 2% paraformaldehyde

(PFA). For flow cytometry analysis, cells were detached with Accutase and incubated with

20nM QD-peptide conjugates for 1h at 37°C in suspension, washed and fixed with 0.5%

PFA. The cell penetrating TAT peptide was used as a positive control.

Preliminary results revealed remarkable intracellular uptake of QD-TmR conjugates by flow

cytometry and fluorescent imaging after 12h incubation in cell culture medium. The uptake

of QD-TmR conjugates was higher than non-conjugated QDs or QDs conjugated with

negative control (CmR peptide). Binding of F3 (Nucleolin-binding) peptide was detected by

flow cytometry after 1h incubation. Internalization was further studied qualitatively by

fluorescent imaging. In conclusion, our data suggests that QDs are a convenient tool to

evaluate nanoparticle targeting potential of peptides binding to cancer-specific receptors.

At least two peptides were shown to be promising for further development of liposomal

drug delivery system for RMS.

Surfaceome profiling of rhabdomyosarcoma for CAR T cell targets

Andrea Timpanaro, Dzhangar Dzhumachev, Michele Bernasconi, Jochen Rössler

Department of pediatric Hematology and Oncology, Inselspital, Bern University Hospital,

Department for BioMedical Research (DBMR), University of Bern

Pediatric rhabdomyosarcoma (RMS) is the most common soft tissue sarcoma in children

and adolescents. Although overall 5-year survival rates have improved with the combined

use of surgery, radiation therapy and chemotherapy, in patients with metastatic and

aggressive disease little progress has been made and the prognosis remains poor.

Cell surface proteins are ideal targets to develop more effective and less invasive cancer

treatments. A novel attractive therapy targeting surface proteins is represented by Chimeric

Antigen Receptor (CAR) T cells. CARs are fusion proteins composed of an antigen

recognition domain, a spacer, a transmembrane domain and one or more intracellular signa-

ling domains. CARs targeting CD19 have achieved significant clinical success against

leukemias, but surface targets for RMS are scarce and no effective CAR T cells for RMS are

available so far.

The aim of this study is to investigate comprehensively RMS surfaceome to identify novel

targets. The final goal is to create effective CAR T cells directed against selected surface

proteins.

In order to achieve our objectives, RMS surfaceome was investigated in a panel of ten RMS

cell lines. Primary myoblasts and a fibroblast cell line were used as controls. Surface proteins

were isolated by two different protocols and the surfaceome of each cell line explored by

LC-MS. We used a set of predicted cell surface proteins, derived from public databases, to

filter the results and to select the most expressed putative antigens. Among these, we found

CD276, known to be overexpressed on RMS. Anti-CD276 CARs are being established.

To conclude, our validated protocols and bioinformatics enrichment of LC-MS data revealed

candidates exclusively expressed on RMS. CD276 CARs, used as positive control, will help

to improve the design of novel CARs against the selected targets to eradicate RMS.

Challenges of treating osteosarcoma in a 12-years old patient with Sickle Cell

Disease

Blank, Bettina et al

Introduction: Cancer in patients with SCD is very rare and treatment including chemothera-

py extremely challenging. We report on a boy with homozygote SCD (HbSS) with knee pain

diagnosed for high-grade osteosarcoma of the distal femur successfully treated at our

center.

Case report: The 12-years old SCD patient presented progressive left knee pain that lasted

since several months. He had a history of multiple vaso-occlusive crisis treated by hydroxyu-

rea since the age of two years. A work-up including imaging and a biopsy revealed

osteosarcoma of the distal femur with pulmonary metastases. He received chemotherapy

including cisplatin/doxorubicin and HD-MTX. Hydroxyurea was stopped to avoid drug-in-

teractions and additional bone marrow toxicity. Monthly erythrocyte apheresis was

conducted to reduce HbS < 30%. Furthermore, because of HD-MTX, we stopped

supplemental prophylactic folic acid for SCD. Delayed clearance of MTX occurred four times

after HD-MTX courses and after the 2nd course, the patient developed self-limiting trouble

of conscience suspicious for seizure. We performed femur amputation due to N. peroneus

involvement with free transfer of the left distal tibia, fibula and calcaneus to the proximal

femur. Imaging and histology revealed a poor treatment response and pulmonary

metastases showed progression. In consequence, we added the immunomodulatory drug

mifamurtide to chemotherapy and pulmonary metastases were resected by bilateral

thoracotomy. One year after diagnosis, he is in complete remission and monthly apheresis is

still ongoing.

Discussion & Conclusion: This case highlights several challenges for treating a SCD patient

with osteosarcoma:

1.) diagnosis might be delayed due to skeletal pain mistaken for pain crisis and patients

coping with chronical pain

2.) regular erythrocyte apheresis can prevent sickle cell crisis during treatment.

Gastrin releasing peptide receptor is overexpressed in Wilms tumour patients and

cell lines

Elizaveta Fasler-Kan, Sabrina Ruggiero, Milan Milošević, Dietmar Cholewa and

Steffen M. Berger

Department of Paediatric Surgery, Children’s University Hospital, Bern

Gastrin.releasing peptide (GRP) regulates numerous functions of the gastrointestinal and

central nervous system, including release of gastrointestinal hormones, smooth muscle cell

contraction and epithelial cell proliferation. It is a potent mitogen for neoplastic tissues. The

effects of GRP are mediated through the gastrin-releasing peptide receptor (GRPR). This

receptor is glycosylated and belongs to the family of 7-transmembrane G-protein coupled

receptors that activate the phospholipase C signalling pathway. Recent studies have shown

that the GRPR is aberrantly expressed in numerous cancers such as those of the lung, colon,

prostate, breast and kidney. Interestingly, the GRPR was present only in tumour tissue, but

not in the normal kidney tissue of patients. Wilms tumour (WT) is one of the most common

paediatric solid tumours, occurring with the incidence of 1 in 10,000 births. WT are

complex childhood neoplasms and are frequently composed of various histological cell

types. With current therapy the prognosis is good. With protocols developed by the

National Wilms Tumor Study Group, the survival rate of children with WT reached 85-90%

by the end of the twentieth century. However, some of WT with a specific aggressive

behaviour have adverse clinical outcome in about 10-15% of the cases. The aim of the

study is to investigate the expression of GRPR in WT patients and in paediatric kidney

tumour cell lines. Our RT-PCR data showed that all analysed paediatric kidney tumour cell

lines (WT- 3 ab, WT-CLS 1. SK-NEP-1, G401) overexpress the GRPR.

Association of lung clearance index with survival in patients with cystic fibrosis

J. M. Kurz1, K. Ramsey1, R. Kraemer2, B. Spycher3, R. Fischer Biner4, P. Latzin1, F. Singer1

1 Division of Respiratory Medicine, Department of Paediatrics, University Hospital of Bern

2 Department of Clinical Research, University of Berne

3 Swiss Pediatric Respiratory Research Group, Institute of Social and Preventive Medicine,

University of Bern

4 Department of Pulmonology, Quartier Bleu, Lindenhofspital - Bern

Background: The lung clearance index (LCI), derived by the multiple-breath washout

(MBW) using nitrogen (N2) as tracer gas, is a sensitive marker to quantify ventilation

inhomogeneity in patients with cystic fibrosis (CF). So far, the association between LCI and

survival has not been investigated.

Aim: Examine the association between LCI and risk of death (mortality) or lung transplanta-

tion (LTX, morbidity) in children with CF.

Methods: Retrospective longitudinal study in a clinical cohort of 147 CF patients aged ≥ 3

years including 1585 LCI measurements between 01.01.1980 and 31.12.2005. First, we

performed cross-sectional analyses at baseline, i.e. when patients entered the study. We

averaged LCI (n = 438) across the first three visits per patient to avoid possible confounding

from indication to measure LCI. Primary outcome was the association of baseline LCI and

the compound outcome (death or LTX) determined in 2018. We fitted logistic regression

models adjusting for age, sex, and date of lung function measurement.

Results: At baseline, patients were mean (SD) 8.7 (3.0) years old and 48% were females.

Follow-up was on average 16.9 (7.6) years. Until 12/2018, 20% (29/147) died or received

LTX. Age and sex distribution was comparable between patients who survived and those

who died or received LTX. The latter group entered the study earlier. The crude odds ratio

(OR, 95%CI) for death or LTX by one LCI unit increase was 1.09 (1.03 to 1.15), p = 0.004,

the adjusted OR was 1.16 (1.08 to 1.25), p < 0.001.

Conclusion: This unique data suggests that elevated LCI is associated with mortality in

school-aged children with CF born 19 to 39 years ago. We acknowledge the limitations of

retrospective analysis of observational, routinely collected health data and historical MBW

measurements. Completion of the database and longitudinal analyses is ongoing.

Improving opportunities for physical activity and participation in inpatient settings

for adolescent cancer patients

Sophie Wist1, Rebecca Näff1, Sonja Lüer2, Eva Brack2, Jochen Rössler2, Ruth Stauffer

Lacorcia1

1 Department of Physiotherapy, Inselspital, Bern University Hospital, University of Bern,

Bern, Switzerland

2 Division of Pediatric Hematology/Oncology, Department of Pediatrics, Inselspital, Bern

University Hospital, University of Bern, Switzerland

Background: Physical activity has shown good results reducing cancer related fatigue and

negative side effects related to chemotherapy that induce a reduction of participation in

daily living and lead to a low quality of life in paediatric cancer patients. Clinical experience

indicates that in particularly adolescent cancer patients reduce their activity level during

intensive chemotherapy. Therefore, an investigation was started to clarify the requirements

and desires of our patients in order to improve their physical activity level.

Methods: Based on literature search, semi-structured interviews were developed and

conducted with four patients, four parents and a multidisciplinary team working with

adolescent cancer patients. The answers were analysed for inhibiting and promoting factors

for physical activity. Combining literature and interview results, recommendations and

potential measures were structured and discussed in an interprofessional setting.

Results: An improvement in the opportunities for physical activity and participation for

adolescent cancer patients meets a need and leads to greater quality of life. The interviewed

persons think that Physiotherapists should take a role in prevention, patient education and

as a motivator. Discussion: Although the investigation included only a small number of

participants, the chosen approach has allowed gathering different aspects for improvement

with relatively few resources. Physiotherapy should be involved earlier in the process and

aim to create an environment beneficial to physical activity.

Conclusion: The first interprofessional new measures to enhance physical activity in

inpatient settings have been recently implemented. The process is to be continuously

evaluated regarding its clinical feasibility.

Systematic differences in analysis software versions hamper infant lung

function testing

Marc-Alexander Oestreich, Florian Wyler, Philipp Latzin, MD, PhD, Kathryn Ramsey, PhD

Pediatric Respiratory Medicine, Inselspital, University Children›s Hospital of Bern, Universi-

ty of Bern, Switzerland

Background: Multiple-breath inert gas washout (MBW) is a non-invasive, feasible, and sen-

sitive technique to assess lung volumes and ventilation inhomogeneity in infancy. Poor

agreement among commercially available setups and software versions currently limits

widespread application as a surveillance tool in early cystic fibrosis (CF) lung disease.

Aim: We investigated differences in algorithms between two versions of an analysis

software and their impact on MBW outcomes functional residual capacity (FRC) and lung

clearance index (LCI).

Methods: Measurements were collected in healthy infants and infants with CF. Lung

function was performed within the first weeks of life and at one year of age. MBW was

performed using 4% sulphur hexafluoride (Exhalyzer® D, Eco Medics AG, Switzerland).

Data were analyzed using WBreath® versions 3.28.0 and 3.51.2 (ndd AG, Switzerland).

Results: Preliminary analysis included data from 14 infants (7 in CF). Using the older

software version (v3.28.0), mean (SD) FRC for healthy infants was 25.8 ml/kg (2.67) and

25.1 ml/kg (2.27) for CF patients. Using the newer version (v3.51.2), mean FRC was

systematically 10% lower (23.2 ml/kg (2.53) for healthy and 22.3 ml/kg (2.56) for CF

infants; p=0.002). In addition, LCI was approximately 14% higher using the newer software

with a mean of difference of 1.03 (0.57; p=0.003) for healthy infants and 0.97 (0.42;

p=0.0009) for CF infants.

Conclusion: Different versions of the same analysis software revealed significant and

systematic differences in MBW outcomes. The newer software version (v3.51.2) calculates

FRC and LCI according to recent consensus guidelines and therefore we recommend using

this for future analysis.

Joint effects of prenatal smoke exposure and polymorphisms in the MBL2 gene on

cord blood levels of mannose-binding lectin

A. Soti1, O. Gorlanova2, L. Müller1, J. Usemann2, L.J Schlapbach1,3, M. Kabesch4, U. Frey2,

P. Latzin1, O. Fuchs1

1 Department of Paediatrics, Inselspital, University of Bern, Bern, Switzerland

2 University Children’s Hospital (UKBB), University of Basel, Basel, Switzerland

3 Paediatric Critical Care Research Group, Mater Children`s Hospital, Brisbane, Australia

4 Department of Paediatric Pulmonology, Allergy and Neonatology, Hannover Medical

School, Hannover, Germany

Background: Prenatal smoke exposure is associated with increased risk for respiratory

disease in offspring and impacts on the developing immune system. In addition, levels of

mannose-binding lectin (MBL), a soluble pattern-recognizing molecule of innate immunity,

are also related to respiratory morbidity early in life. So far, joint effects of prenatal smoke

exposure and single-nucleotide polymorphisms (SNPs) in the MBL2 gene and secondly of

MBL levels in umbilical cord blood (UCB) are unknown.

Objective: To investigate whether prenatal smoke exposure influences MBL levels in UCB in

concert with SNPs in the MBL2 gene region.

Methods: We measured MBL levels in UCB in 221 study participants of a birth cohort of

unselected, healthy, term-born infants. We assessed prenatal smoke exposure by questi-

onnaire, validated by cotinine levels in the first urine. After genome-wide chip-based

genotyping (Illumina HumanOmniExpress) we assessed the association of MBL levels with

MBL2 SNPs in a genome-wide association study (GWAS). In regression analyses, we

analysed effect modification of SNPs by smoking, adjusting for known and possible

confounders. For a preliminary assessment of gene-environment interaction (GxE), we used

logistic regression of MBL levels dichotomised at 700 ng/ml or 300 ng/ml levels.

Results: Prenatal smoke exposure significantly decreased MBL levels in UCB (p=0.006).

MBL2 SNPs were significantly associated with MBL levels. Except for a subgroup of SNPs

(n=6/24, rs920727, rs2506, rs2083771, rs2099903, rs1800450, rs11003134), smoking

significantly lowered MBL levels in addition to MBL2 SNPs. Preliminary data so far suggests

that such interaction exists for the downstream SNP rs10824787 and for rs3829168 in the

promoter region.

Conclusions: Prenatal smoke exposure seems to lower MBL levels in cord blood indepen-

dently as well as in a joint effect with MBL2 SNPs. Preliminary data furthermore suggests a

link between genetic and environmental effects on respiratory morbidity early in life.

Lack of Aquaporin 9 Expression in iPSC-Derived Hepatocytes Impairs Urea

Secretion

Laemmle A, Borsuk M, Robinson J, Gallagher RC, Nuoffer JM, Häberle J, Willenbring H

Reprogramming of patient-derived skin fibroblasts into induced pluripotent stem cells

(iPSCs) followed by directed differentiation into hepatocytes (iPSC-Heps) allows modeling

of genetic liver diseases in vitro. We aim to develop an iPSC-Hep-based model of the urea

cycle disorder ornithine transcarbamoylase (OTC) deficiency (OTCD) that can be used to

screen for pharmacological chaperones (PCs) to treat OTCD.

We generated iPSCs from fibroblasts of OTCD patients and controls and differentiated

them into iPSC-Heps. We compared urea cycle enzyme expression and ammonia metabo-

lism between patient-derived and normal iPSC-Heps and primary human hepatocytes

(PHH). We included human fetal and adult liver tissue as additional controls.

Differentiation of OTCD patient-derived iPSCs into iPSC-Heps revealed a disease-specific

phenotype with reduced OTC activity and urea secretion. However, urea secretion was also

low in control iPSC-Heps, which expressed all urea cycle enzymes at levels comparable to

PHH, even after ammonia challenge. iPSC-Heps are known to correspond to immature fetal

hepatocytes rather than mature adult hepatocytes. Therefore, we compared gene

expression profiles of human fetal and adult liver tissue to identify differentially expressed

genes that may be responsible for the low urea secretion in iPSC-Heps. We found that fetal

liver tissue was lacking expression of aquaporin 9 (AQP9), which is required for urea

secretion. Consistent with their fetal state of differentiation, AQP9 was absent in all of our

iPSC-Hep lines and its induction led to a significant increase in urea secretion.

These findings show that patient-derived iPSC-Heps replicate characteristic features of

OTCD. A limitation of this model is low urea secretion by iPSC-Heps because of their fetal

state of differentiation. We identify AQP9 as a target for overcoming this restriction and

realize the potential of iPSC-Heps.

Effect of metformin on ACTH receptor activation and downstream signaling

Shaheena Parween1, 2, Christa E Flück1, 2 and Amit V. Pandey1, 2

1 Department of Paediatric Endocrinology, Diabetology and Metabolism University

Children’s Hospital, Bern; 2Department of Biomedical Research (DBMR), University of

Bern, Bern, Switzerland

Background: The peptide hormone adrenocorticotropin (ACTH or Corticotropin) is a major

component of the stress response system in the Hypothalamus-Pituitary-Adrenal (HPA)

axis. Under stress, it is secreted from the anterior pituitary and stimulates cortisol producti-

on from the adrenal cortex. Changes in ACTH production or action are associated with

multiple disease conditions. In clinical situations like Cushing’s disease, ectopic ACTH

syndrome and congenital adrenal hyperplasia, there is excess ACTH production and

blocking the interaction of ACTH at its site of action would be a therapeutic option.

Currently, effective therapy to block the action of ACTH is unavailable. Insulin-sensitizing

treatment, such as metformin, has been used to ameliorate a few reported cases of adrenal

disorders. However, the exact mechanism of how these insulin-sensitizing drugs affect the

HPA axis is not known. Here we test whether an insulin-sensitizing drug, metformin have a

direct effect on the activity of ACTH.

Methods: Cell based in-vitro assays were performed to test the effect of metformin on

ACTH receptor activation and signaling. For assays, OS3 cells transfected with ACTH

receptor and luciferase reporter plasmids were used. Cyclic AMP (cAMP) generation upon

receptor activation was measured by dual luciferase assay (Promega). The potential to shift

the ACTH concentration-response curve (CRC) was evaluated to characterize the inhibitory

activity of metformin on ACTH receptor activation. Detailed characterization was done to

calculate the 50% inhibitory concentration (IC50) by varying concentration of metformin.

Results: Metformin was found to inhibit the activation of the ACTH receptor and downstre-

am signaling associated with ACTH response. Significant inhibition of ACTH induced

receptor activation upon treatment with 10 mM metformin was observed. Metformin

shifted the ACTH CRC towards the right by half log, indicating antagonism.

Conclusion: Treatment of an insulin-sensitizing drug, metformin reduces ACTH induced

receptor activation and signaling. This study could be useful in developing new strategies

for management of hyperandrogenic states especially associated with excess ACTH.

Expanding the phenotype of the p.R63W mutation of HNF4A – a case report

Mara Grassi

Masterstudent medicine, University of Bern

Background: The dominant mutation p.R63W (also known as p.R76W) in the hepatocyte

nuclear factor 4 alpha (HNF4A) leads to congenital hyperinsulinaemic hypoglycaemia (CHI),

macrosomia and Fanconi-type tubulopathy. It was only in 2014 that this mutation-specific

phenotype was recognized as such, and so far, 15 patients have been reported.

Case Report: We present a new case of a girl born in 2005 with severe neonatal hyperinsu-

linism, renal tubular disease and hepatopathy. Despite extensive investigations, the

diagnosis remained elusive for 12 years. A trio-whole exome sequencing finally showed this

specific mutation.

The case confirms the so far homogeneous phenotype of the p.R63W mutation, but in

addition, she presents some novel features such as liver cirrhosis, secondary mitochondrio-

pathy and several abnormal laboratory parameters (including low urate and FGF-23 levels).

Discussion: We will focus on the new features of this extensively investigated and

well-documented case. Hypotheses on the link between these features, as well as

hyperinsulinaemia, macrosomia and the impact of the p.R63W mutation on liver and

kidneys should result in a better understanding of this mutation-specific phenotype as well

as the function of HNF4A in general.

Conclusion: This case report helps us to gain insight into the role of HNF4A during

development and in metabolic regulation. It also underlines the importance of sharing case

histories.

New Mechanistic Insights into Androgen Production

Emanuele Pignatti1,2, Mihaela Zavolan3, Christa E. Flück1,2

1 Department of BioMedical Research, Inselspital, Bern University Hospital, University of

Bern, Bern, Switzerland

2 Division of Pediatric Endocrinology and Diabetology, Department of Pediatrics,

Inselspital, University Hospital of Bern, University of Bern, Bern, Switzerland

3 Biozentrum, University of Basel, 4056 Basel, Switzerland

Background: Androgens are essential for the development of sex features and for reproduc-

tion. Androgens peak around the age of 8 in both boys and girls during adrenarche, the

functional activation of the androgen-producing zona Reticularis in the adrenal glands.

However, what controls androgen production is poorly understood. At the same time, we

have little knowledge of what causes most androgen-related disorders, including Premature

Adrenarche (PA) and Polycystic Ovary Syndrome (PCOS).

Aim and Methods: To shed light on the mechanisms of androgen regulation, we used

established in vitro models of androgen activation and inhibition, based, respectively, on

serum starvation and metformin treatment of the steroidogenic H295R adrenal cell line. We

profiled mRNAs and miRNAs in parallel, to obtain simultaneous information on transcripti-

onal activity and suppression of translation. For the identification of regulatory gene

networks, gene-enrichment and pattern recognition algorithms will be used. Follow-up

experiments will allow the validation of our in vitro findings in human – these experiments

will include mRNA and miRNA profiling of the zona Reticularis from human adrenal

samples.

Relevance: Altogether, our results will define novel mechanisms of androgen regulation and

indicate candidate biomarkers for the preventive identification of PA and PCOS patients.

Level of glycemic control in pediatric patients with type 1 diabetes in Bern: a

cross-sectional study

Tanja Zingg, Michelle Dennig, Grit Sommer, Christa E. Flück

Department of Pediatrics (Division of Pediatric Endocrinology and Diabetology), Inselspi-

tal, Bern University Hospital, University of Bern, Switzerland

Background: Good glycemic control prevents long-term complications of microvascular and

macrovascular diseases in type 1 diabetes (T1DM). We aimed to investigate whether our

patients had A1c values <7.5% as recommended by ISPAD and how therapy modality,

duration of diabetes and pubertal status affected the metabolic control of our patients. We

also set out to compare our quality of care with our results of 2008 and with other

published data.

Methods: In 2017/18, we enrolled all patients with T1DM who were followed by the

outpatient clinic of the University Children’s Hospital Bern over a period of 6 months in an

observational cross-sectional study. Each patient was assessed once during the observatio-

nal period, including demographic and clinical data.

Results: 160 patients participated in the study, 41% (n=82) were boys and 49% (n=72)

were girls. Patients had a mean age (SD) at time of visit of 12.6 (3.5) years (range 2-17

years) and a mean duration (SD) of diabetes of 4.6 (3.6) years (range 1-16 years). Most

patients, 63% (n=100) received functional insulin treatment, 29% (n=47) used insulin

pump and 8% (n=13) injected insulin on multiple times per day (twice-daily/three-dose).

CGM devices were used by 43% (n=68) of patients in their diabetes management. Mean

A1c was 8% and 71% had A1c >7.5%. Compared to results from our hospital from 2008,

A1c was slightly higher (8% vs 7.6%), but more patients had diabetes for >2 years (80% vs

47%). Patients with T1DM duration >2 years had more often A1c levels above 7.5 % than

patients with duration <2 years (p<0.001). A1c values were significantly lower (p<0.05) in

patients using CGM devices (7.8% vs 8%).

Conclusion: The overall glycemic control was poorer 2017/2018 than in our study from

2008. This may be due to the higher percent of patients with diabetes duration >2 years,

thus with more patients out of the remission phase. Patients wearing CGM devices

performed better. Unfortunately, our patients in Bern did not reach the target A1c set by

ISPAD, similar to results of other diabetes centres in Europe and the United States. This

highlights the importance of regular consultations and extended use of CGM.

Trajectory of microbiota maturation in healthy Bern infants – a network approach

L. Cecchini, D. Marchukov, S. Ganal-Vonarburg, C. Sokollik, B. Misselwitz

Background: Intestinal microbiota composition is fundamental to human health and

undergoes critical changes within the first two years of life. Factors probably influencing the

microbiota are the maternal microbiota and the general environment in Switzerland.

However, the development of the intestinal microbiota is incompletely understood. Gaining

knowledge of the trajectory of microbiota maturation is likely key to the understanding of

the pathogenesis of many pathologies in childhood.

Aims: We aim for a deep understanding of the maturation of the healthy infant intestinal

microbiota regarding composition, diversity and metabolic activities. We aim for identifying

parameters affecting microbiota maturation and effects of the microbiota on infant

outcome.

Methods: We will recruit 120 pregnant mothers who will be followed as mother-baby pairs

until 10 years of age. Infants will be followed clinically to determine adequate growth and

development as well as pathology including abdominal pain. Epidemiological parameter

and infant nutrition will be assessed. We will collect biosamples such as stool, maternal milk

and skin swaps.

Species composition and diversity will be assessed by 16S sequencing. Metagenomic

shotgun sequencing and bacterial mRNA analysis will inform about metabolic potential and

metabolic activity of the microbiota. Mass spectrometry will assess the small molecule

content of stool and maternal milk samples. Network analysis will be used to assess the

complex relationships between bacteria metabolic activities and small molecular content.

Expected results: We expect an increase in complexity and metabolic potential and activity

with age. Microbiota parameters will differ according to nutrition and might predict infant

outcomes such as growth and abdominal pain. Systematic analysis of sequential maternal

and infant bacteria samples from stool, skin and maternal milk will help characterizing

bacterial transfer from mother to infant.

Conclusion: We propose an observational study of healthy Bern mother baby pairs with

clinical characterization and biosampling. Advanced analysis tools will be used to characte-

rize the microbiota and address mechanistic questions.

Human-induced steroidogenic cells from urine consist a new cell-based model

to study adrenal disorders in a personalised manner

Efstathios Katharopoulos

Background: Congenital adrenal hyperplasia (CAH) is an inherited recessive disorder

characterised by insufficient production of steroids by the adrenal cortex that can also lead

to disorders of sex development (DSDs). CAH is managed by life-long hormone replace-

ment, a suboptimal therapy which fails to mirror the physiological feedback loops of the

hypothalamic-pituitary axis. Recently, various mesoderm-derived cells, including urinary

stem cells (USCs), have been differentiated to adrenocortical-like cells. These innovative

differentiation protocols open the doors to new in vitro disease models and cell-based

treatments (possibly permanent) for CAH, and highlight the huge potential of USCs as

starting material.

Aim: We tried to reproduce and establish in our lab a previous protocol for the isolation of

urinary stem cells (USCs) and their characterisation.

Methods: Fresh urine samples were collected, cells were concentrated and expanded in

vitro using a common culture medium enriched with epinephrine, insulin, EGF, transferrin

and T3. Morphology was microscopically examined and expression of CD44+ was tested in

mRNA level.

Results: Urinary stem cell colonies were successfully (75% success rate) isolated after 2-3

weeks. Cells presented characteristic morphology and expressed the CD44+ mesenchymal

marker. Further experiments for the characterisation of USCs are ongoing.

Conclusions and perspectives: We are in the process of establishing a reproducible

protocol to isolate urinary stem cells in our laboratory. We anticipate that this protocol is

going to foster new research lines in our lab, since USC represent an easily-accessible source

of mesoderm-derived pluripotent cells that can be isolated rapidly and in a cost-effective

way. Future efforts will focus on further characterisation of the isolated USC using FACS

and immunohistochemistry with mesenchymal (CD29, CD166) and pluripotent (SSEA-4)

markers and steroid production.

2019

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MZ

Universitätsklinik für KinderheilkundeForschungssekretariat

Inselspital, Universitätsspital Bern

CH-3010 Bern

Tel.: +41 31 632 94 93