Periodic Reporting for period 5 - GLADIATOR (Next-generation theranostics of brain pathologies with autonomous externally controllable nanonetworks: a trans-disciplinary approach with bio-nanodevice interfaces)
Reporting period: 2023-01-01 to 2023-09-30
Brain pathologies are highly complex. Despite recent progress, their prognosis is grim, defining a high societal challenge. Bridging life sciences, bio-nanotechnology, engineering and ICT, GLADIATOR promises a vanguard and comprehensive theranostic (therapeutic+diagnostic) solution for brain malignancies. GLADIATOR will provide, for the first time, a working prototype of a complete, autonomous and clinically applicable, nanonetwork-based, molecular communications system based on the conceptual framework of Externally Controllable Molecular Communications (ECMC). Using Glioblastoma Multiforme (GBM) tumors, the most detrimental of brain pathologies, as a proof-of-concept case, GLADIATOR will implement a platform of cell-based and electronic components, consisting of
1. Implantable autologous cell organoids, consisting of engineered induced neural stem cells (iNSCs), which will release specifically designed exosomal vesicles, acting as bio-nano-machines, delivering reprogramming (therapeutic) miRNAs and building nano-networks. Interfering with the underlying biological environment, they will provide a revolutionary intervention both killing the tumor cells but also reducing their aggressiveness and recurrence.
2. A hybrid bio-electronic interface, consisting of coupled external and implantable devices, which will enable communication channels with fluorescent bio-nano-machines, released by the modified cancer cells, via micro-optoelectronic sensors.
3. A wireless ECMC network integrating the cellular, sub-cellular and electronic components. This system will autonomously monitor the spatiotemporal tumor evolution and recurrence and generate, on demand, appropriate reprogramming interventions, by increasing iNSC renewal and multiplication via external radiofrequency stimulation.
GLADIATOR establishes the feasibility and innovation potential towards a far-reaching transformation in the investigation and management of complex malignancies and potentially other major central nervous system pathologies. It also promotes the emerging supra-disciplines of “bio-nano-machine diagnostics” and a profound shift towards “nano-network therapeutics” which lay the grounds for future autonomous, closed-loop, externally controllable, micro- or nano- scale devices for disease management.
1. Implantable autologous cell organoids, consisting of engineered induced neural stem cells (iNSCs), which will release specifically designed exosomal vesicles, acting as bio-nano-machines, delivering reprogramming (therapeutic) miRNAs and building nano-networks. Interfering with the underlying biological environment, they will provide a revolutionary intervention both killing the tumor cells but also reducing their aggressiveness and recurrence.
2. A hybrid bio-electronic interface, consisting of coupled external and implantable devices, which will enable communication channels with fluorescent bio-nano-machines, released by the modified cancer cells, via micro-optoelectronic sensors.
3. A wireless ECMC network integrating the cellular, sub-cellular and electronic components. This system will autonomously monitor the spatiotemporal tumor evolution and recurrence and generate, on demand, appropriate reprogramming interventions, by increasing iNSC renewal and multiplication via external radiofrequency stimulation.
GLADIATOR establishes the feasibility and innovation potential towards a far-reaching transformation in the investigation and management of complex malignancies and potentially other major central nervous system pathologies. It also promotes the emerging supra-disciplines of “bio-nano-machine diagnostics” and a profound shift towards “nano-network therapeutics” which lay the grounds for future autonomous, closed-loop, externally controllable, micro- or nano- scale devices for disease management.
Until the current reporting period, the consortium partners have worked towards the biological and technological goals of GLADIATOR:
1. Multiple constructs for Hsp70 promoter-dependent gene expression have been generated, which provide the possibility to monitor cell behaviour (Luciferase, EGFP), EV production by the cells (palmEGFP, palmtdTomato) and produce protein that is toxic for glioblastoma cells (TRAIL).
2. Other RF-responsive gene promoters are being investigated, including the Early Growth Response 1 (EGR-1) promoter and the Electromagnetic perceptive gene (EPG) promoter combined with miR-34a or/and NGF expressing vectors.
3. An in vitro platform for the combined evaluation of iR-NSCs, BBB, GBM constructs and iM-NSCs has been established including a triple culture with NSC spheroids, the BBB and U87 organoids as GBM construct.
4. A complete end-to-end simulator was created which includes: (i) molecular diffusion simulations in a 3D reconstructed volume, (ii) the link between the signal being transmitted and the data being received by the external sensors, and (iii) models of exocytosis and endocytosis.
5. A hybrid sensor prototype was constructed, including: (i) integration of the cellular components, (ii) miniaturization of a third generation implant PCB, and (iii) proof-of-concept of power transfer and backscatter modulation with redesigned (miniaturized) PCB
6. An orthotopic GBM in vivo model, in immunocompromised mice, was established with orthotopic implantation of tumour cells with high oncogenic potential recapitulating the human tumour evolution and clinical characteristics.
7. The tumourotropic and tumor homing effect of human iNSCs and exosomes was assessed in vivo.
8. Cellular and electronic components validated in vivo
1. Multiple constructs for Hsp70 promoter-dependent gene expression have been generated, which provide the possibility to monitor cell behaviour (Luciferase, EGFP), EV production by the cells (palmEGFP, palmtdTomato) and produce protein that is toxic for glioblastoma cells (TRAIL).
2. Other RF-responsive gene promoters are being investigated, including the Early Growth Response 1 (EGR-1) promoter and the Electromagnetic perceptive gene (EPG) promoter combined with miR-34a or/and NGF expressing vectors.
3. An in vitro platform for the combined evaluation of iR-NSCs, BBB, GBM constructs and iM-NSCs has been established including a triple culture with NSC spheroids, the BBB and U87 organoids as GBM construct.
4. A complete end-to-end simulator was created which includes: (i) molecular diffusion simulations in a 3D reconstructed volume, (ii) the link between the signal being transmitted and the data being received by the external sensors, and (iii) models of exocytosis and endocytosis.
5. A hybrid sensor prototype was constructed, including: (i) integration of the cellular components, (ii) miniaturization of a third generation implant PCB, and (iii) proof-of-concept of power transfer and backscatter modulation with redesigned (miniaturized) PCB
6. An orthotopic GBM in vivo model, in immunocompromised mice, was established with orthotopic implantation of tumour cells with high oncogenic potential recapitulating the human tumour evolution and clinical characteristics.
7. The tumourotropic and tumor homing effect of human iNSCs and exosomes was assessed in vivo.
8. Cellular and electronic components validated in vivo
The GLADIATOR consortium is already producing original research results beyond the state of the art, demonstrating
1. Formation of the cellular components of the proposed platform with a complete characterization of transdifferentiated cells and protocol optimization, comparing the iNSC derived from iPSC and assessment of transfection efficacy in transdifferentiated iNSCs vs iPSC derived.
2. Experiments to validate the effects of RF exposure on cells, and to study RF effects on the expression of fluorescent proteins under the control of various promoters. Increases in proliferation as well as protein production have already been demonstrated.
3. A sustainable three-dimensional (3D) scaffold comprised of biocompatible hydrogel and of electrospun nanofibers, as the technological embodiment of 3D GBM tumor constructs and the proposed organoids.
4. The building blocks of a theoretical communication model, which include exosomal release distribution and binding, in an expanded molecular communications modelling platform.
5. The initial designs for novel technologies for ultrasound-based transcranial power transfer and passive communication schemes, which are currently being tested experimentally, as well as considering various designs of metasurfaces for transcranial RF transmission.
When the proposed platform is completed and becomes clinically available, it is expected to have a significant societal impact. The advances in cancer management, enabled by the innovations in GLADIATOR, will improve patient prognosis and prolong survival by minimizing recurrences and reducing drug toxicity. Improved health, extended life expectancy and productivity, reduced sick-leaves, shorter hospitalizations, reduced return visits, less personnel and caregiver involvement will also have a positive effect on the already overstrained Health Care Systems.
The ground-breaking biological and nanotechnology-based innovations of GLADIATOR, are also expected to have significant economic impact since they can enter into significant market segments. For example, the microelectronic medical implants and micro-sensors markets were valued at $35B in 2016 and expected to grow to $56B by 2021. The Point of Care (PoC) diagnosis market was valued at $10.3B in 2016 growing to $33.7B by 2025. GLADIATOR can lead to high-gain innovations that can have a long-lasting positive impact on Europe’s science and industry, with benefits proportional to the associated high-risks of the project.
1. Formation of the cellular components of the proposed platform with a complete characterization of transdifferentiated cells and protocol optimization, comparing the iNSC derived from iPSC and assessment of transfection efficacy in transdifferentiated iNSCs vs iPSC derived.
2. Experiments to validate the effects of RF exposure on cells, and to study RF effects on the expression of fluorescent proteins under the control of various promoters. Increases in proliferation as well as protein production have already been demonstrated.
3. A sustainable three-dimensional (3D) scaffold comprised of biocompatible hydrogel and of electrospun nanofibers, as the technological embodiment of 3D GBM tumor constructs and the proposed organoids.
4. The building blocks of a theoretical communication model, which include exosomal release distribution and binding, in an expanded molecular communications modelling platform.
5. The initial designs for novel technologies for ultrasound-based transcranial power transfer and passive communication schemes, which are currently being tested experimentally, as well as considering various designs of metasurfaces for transcranial RF transmission.
When the proposed platform is completed and becomes clinically available, it is expected to have a significant societal impact. The advances in cancer management, enabled by the innovations in GLADIATOR, will improve patient prognosis and prolong survival by minimizing recurrences and reducing drug toxicity. Improved health, extended life expectancy and productivity, reduced sick-leaves, shorter hospitalizations, reduced return visits, less personnel and caregiver involvement will also have a positive effect on the already overstrained Health Care Systems.
The ground-breaking biological and nanotechnology-based innovations of GLADIATOR, are also expected to have significant economic impact since they can enter into significant market segments. For example, the microelectronic medical implants and micro-sensors markets were valued at $35B in 2016 and expected to grow to $56B by 2021. The Point of Care (PoC) diagnosis market was valued at $10.3B in 2016 growing to $33.7B by 2025. GLADIATOR can lead to high-gain innovations that can have a long-lasting positive impact on Europe’s science and industry, with benefits proportional to the associated high-risks of the project.