Identification and characterization of nanobodies targeting the EphA4 receptor

The ephrin receptor A4 (EphA4) is one of the receptors in the ephrin system that plays a pivotal role in a variety of cell-cell interactions, mostly studied during development. In addition, EphA4 has been found to play a role in cancer biology as well as in the pathogenesis of several neurological disorders such as stroke, spinal cord injury, multiple sclerosis, amyotrophic lateral sclerosis (ALS), and Alzheimer's disease. Pharmacological blocking of EphA4 has been suggested to be a therapeutic strategy for these disorders. Therefore, the aim of our study was to generate potent and selective Nanobodies against the ligand-binding domain of the human EphA4 receptor. Weidentified two Nanobodies, Nb 39 and Nb 53, that bind EphA4 with affinities in the nanomolar range. These Nanobodies were most selective for EphA4, with residual binding to EphA7 only. Using Alphascreen technology, we found that both Nanobodies displaced all known EphA4-binding ephrins from the receptor. Furthermore, Nb39 andNb53 inhibited ephrin-induced phosphorylationoftheEphA4proteininacell-basedassay. Finally, in a cortical neuron primary culture, both Nanobodies were able to inhibit endogenous EphA4-mediated growth-cone collapse induced by ephrin-B3. Our results demonstrate the potential of Nanobodies to target the ligand-binding domain of EphA4. These Nanobodiesmaydeservefurtherevaluationaspotentialtherapeutics in disorders in which EphA4-mediated signaling plays a role

Onderzoek naar het mechanisme en therapeutisch potentieel van het efrinesysteem in ALS

Amyotrophic Lateral Sclerosis (ALS) is a dramatic neurodegenerative disease due to its progressive character, the short survival of the patient and the enormous impact on his/her quality of life and that of his/her caretaker. Interestingly, ALS patients, even carrying the same mutation in SOD1, have great variations in clinical characteristics as age of onset, disease duration and severity of the disease. This indicates that ALS is a multifactorial disease influenced by modifying genes and environmental factors. Epha4 is such a genetic factor that modifies ALS both in animal models as well as in patients. We investigated the mechanism in which Epha4 is modifying the disease pathogenesis in ALS. We found that vulnerable motor neurons have high Epha4 expression and that Epha4 reduced the reinnervating capacity of motor neurons, possibly contributing to the higher vulnerability of Epha4 expressing motor neurons in ALS. Furthermore, deleting the Epha4 cytoplasmic domain did not affect disease onset and survival in the SOD1G93A mouse model indicating that Epha4 forward signaling in the Eph-bearing cell does not play a role in the modifying effect of Epha4 in ALS. We therefore suggest that the reverse pathway in the ephrin-bearing cell should be modified in order to alter ALS pathology. We studied ephrinb2 as a possible ligand for the Epha4-mediated effect in ALS. It was highly expressed in motor neurons and in reactive astrocytes. However, deletion of ephrinb2 enhanced disease rather than attenuating it. Interestingly, deleting ephrinb2 from the astrocytes impaired the blood brain barrier integrity which possibly contributed to the reduced disease duration and survival in the SOD1G93A mouse model. To inhibit Epha4 reverse signaling we developed Nanobodies targeting the EphA4 ligandbinding domain. Nb 39 and Nb 53 were selective for EphA4, and had KD and IC50 values in the nanomolar range. Both Nanobodies were able to block the interaction of EphA4 with all ephrin ligands and they inhibited ephrin-induced phosphorylation. In conclusion, blocking the interaction of EphA4 and its ephrin ligands will be the most efficient way to block EphA4 reverse signaling and to develop a specific EphA4 antagonist.status: publishe

Acting with foresight in times of budget austerity

Horizon scanning is acknowledged to be one of the key components of a demand-driven healthcare system. We propose it to be a dynamic collaborative process driven by national unmet need matched with continued insight into the innovative pharmaceutical industry medicinal pipeline. Summer 2019 an international agreement was reached between payers to set up and commission an international horizon scanning initiative (IHSI), which is the international front-end of such a system. In this Policy Paper we propose a two-stage structure and organisation of the back-end national part of the horizon scanning process still to be implemented. This will lead to healthcare budgets managed with better foresight, a necessity in the face of breakthrough, some potentially curing therapies coming at a high cost. Taking the Belgian national component of the proposed horizon scanning system to implementation will require a pilot to be carried out. This to test the internal and external validity of the proposed design

Flexibel werken in ziekenhuizen: Over werklast, draagkracht en verandercultuur

Met schaarse middelen toch optimaal beantwoorden aan de toenemende zorgvraag, zonder dat het zorgpersoneel eronderdoor gaat: hoe doe je dat? Flexibiliteit kan alvast helpen, stellen prof. Brecht Cardoen, prof. Paul Gemmel en onderzoekster Lies Schoonaert, mede-auteurs van het MINOZ-rapport Flexibel werken in ziekenhuizen

Flexibel werken in ziekenhuizen: OM en HRM perspectief

Iedereen spreekt over “flexibel werken”, maar al snel blijkt dat dit een term is die vele interpretaties kent. Flexibiliteit wordt zo een verzamelnaam van variabele werkomstandigheden telkens met een ander doel indachtig. Het kan bekeken worden vanuit het standpunt van de werkgever en vanuit het standpunt van de werknemer. Een werkgever wil zijn personeel zo flexibel mogelijk kunnen inzetten om zo goed mogelijk aan de variabele vraag van zorg te kunnen voldoen. Een werknemer bekijkt hoe de job zo flexibel mogelijk kan worden ingevuld (tijd, plaats, inhoud) om leuk en uitdagend te zijn, en een ideale balans tussen het werk en het privéleven te creëren. In deze tekst geven we een overzicht van de verschillende types flexibiliteit die relevant zijn voor een ziekenhuis. Hierbij geven we ook een aantal voorbeelden hoe flexibiliteit vorm kan krijgen in de dagelijkse realiteit. We focussen ons voornamelijk op verpleegkundigen, maar veel aangehaalde vormen van flexibel werken kunnen ook van toepassing zijn op andere werknemers

Access decision-making in the Belgian Commission for reimbursement of medicines 2010-2017: Investigating the readiness for value-based pricing

To balance the societal need for affordability of medicines with the industrial need for sustained innovation, the present pharmaceutical technology supply-driven system needs to become a societal demand-driven system. Value-based pricing is considered to be a key component of such a system, next to the conditional dialogue between payer and industry we proposed in previous work (Van Dyck, De Grève et al. 2016) in which it should be embedded. To find out how far Belgian pharmaceutical healthcare-related decision-making has evolved within this paradigm, we empirically investigated the access and reimbursement decision-making of the Belgian Commission for Reimbursement of Medicines (CRM) for the period 2010 – 2017. We combined this investigation with previous work in a meta-analysis in order to have the most complete picture possible of the present factors influencing decision-making in the Belgian system

Cochlear supporting cell transdifferentiation and integration into hair cell layers by inhibition of ephrin-B2 signalling

In mammals, cochlear sensory hair cells that are responsible for hearing are postmitotic and are not replaced after loss. One of the most promising strategies to regenerate hair cells is to identify and inhibit the factors preventing the conversion of adjacent non-sensory supporting cells into hair cells. Here we demonstrate that mammalian hair cells can be directly generated from supporting cells by inhibition of ephrin-B2 signalling. Using either ephrin-B2 conditional knockout mice, shRNA-mediated gene silencing or soluble inhibitors, we found that downregulation of ephrin-B2 signalling at embryonic stages results in supporting cell translocation into hair cell layers and subsequent switch in cell identity from supporting cell to hair cell fate. As transdifferentiation is here a result of displacement across boundary, this original finding presents the interest that newly generated hair cells directly integrate either hair cell layer, then would be likely more rapidly able to fit into functional circuitry

Cochlear supporting cell transdifferentiation and integration into hair cell layers by inhibition of ephrin-B2 signalling

In mammals, cochlear sensory hair cells that are responsible for hearing are postmitotic and are not replaced after loss. One of the most promising strategies to regenerate hair cells is to identify and inhibit the factors preventing the conversion of adjacent non-sensory supporting cells into hair cells. Here we demonstrate that mammalian hair cells can be directly generated from supporting cells by inhibition of ephrin-B2 signalling. Using either ephrin-B2 conditional knockout mice, shRNA-mediated gene silencing or soluble inhibitors, we found that downregulation of ephrin-B2 signalling at embryonic stages results in supporting cell translocation into hair cell layers and subsequent switch in cell identity from supporting cell to hair cell fate. As transdifferentiation is here a result of displacement across boundary, this original finding presents the interest that newly generated hair cells directly integrate either hair cell layer, then would be likely more rapidly able to fit into functional circuitry.status: publishe

Cochlear supporting cell transdifferentiation and integration into hair cell layers by inhibition of ephrin-B2 signalling

In mammals, cochlear sensory hair cells that are responsible for hearing are postmitotic and are not replaced after loss. One of the most promising strategies to regenerate hair cells is to identify and inhibit the factors preventing the conversion of adjacent non-sensory supporting cells into hair cells. Here we demonstrate that mammalian hair cells can be directly generated from supporting cells by inhibition of ephrin-B2 signalling. Using either ephrin-B2 conditional knockout mice, shRNA-mediated gene silencing or soluble inhibitors, we found that downregulation of ephrin-B2 signalling at embryonic stages results in supporting cell translocation into hair cell layers and subsequent switch in cell identity from supporting cell to hair cell fate. As transdifferentiation is here a result of displacement across boundary, this original finding presents the interest that newly generated hair cells directly integrate either hair cell layer, then would be likely more rapidly able to fit into functional circuitry