h.o.p.e. Orchestra Proposal

PART 3

a new paradigm

for neurological regrowth

The h.o.p.e. Orchestra

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THE NONPROFIT RESEARCH FOUNDATION: THE H.O.P.E. ORCHESTRA PAGE 2

JUSTIN HILL, M.D.

Neurology, Neuroscience & Neurorehabilitation Founder - h.o.p.e. Orchestra Director of Brain Injury Unit Assistant Professor, Neurology & Neurorehabilitation Burke Rehabilitation Hospital 785 Mamaroneck Avenue White Plains, NY 10605 914-413-8526 jhill@hopeorchestra.org BROOKE ELLISON, PHD, MPP

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Ethics, Stem Cells, & Health Care Policy Co-director, Certificate in Health Care Management Associate Director, Center for Community Engagement & Leadership Development School of Health Technology and Management Director of Education and Ethics, Stem Cell Facility Stony Brook University Stony Brook, NY 11794 brooke.ellison@stonybrook.edu MAROM BIKSON, PHD

Biomedical Engineering, Image Analysis & 3D Printing Co-director of Neural Engineering Professor, Center for Biomedical Engineering The City College of New York of CUNY T-403B, 160 Covent Avenue New York, NY 10031 bikson@ccny.cuny.edu DAVID PUTRINO, PHD, PT THE H.O.P.E. ORCHESTRA FOUNDATION PROPOSAL

Computational Neuroscience & Virtual Rehabilitation Director of Rehabilitation Innovation Mount Sinai Health Network Assistant Professor, Department of Rehabilitation Medicine Icahn School of Medicine at Mount Sinai 1 Gustave L. Levy Place New York, NY 10029-5674 david.putrino@mountsinai.org IRINI VAZANELLIS

Brand Architecture, Graphic Design & Storytelling Engineering, Simulation Arbory Workshop www.arboryworkshop.com Barcelona, Spain 08002 irini@arboryworkshop.com arbory workshop creative arboryworkshop.com

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INTRODUCTION

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ORCHESTRA OVERVIEW

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SECTION I: ARTIFICIAL INTELLIGENCE PRO-GROWTH KNOWLEDGE BASE

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SECTION II: MISSING TISSUE HOLOGRAM

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SECTION III: COMPREHENSIVE CELL BIOLOGY

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SECTION IV: REGROWTH SIMULATION

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SECTION V: REGROWTH REGIMEN

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SECTION VI: LAB CLINICAL TRIAL

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THE NONPROFIT RESEARCH FOUNDATION: ORCHESTRA OPERATIONS

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SUMMARY

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HUMAN-SCALE. ORGAN-REGROWTH. PERSONALISED. EXPERIMENTS.

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PART 3

IN THIS PROPOSAL

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INTRODUCTION

MISSION STATEMENT

Neurological injury results in missing tissue and consequent permanent disability such as the inability to see, speak, hear, or move. The h.o.p.e.

Orchestra seeks to regrow the individual’s missing tissue and restore neurological function through the development of personalized humanscale models and comprehensive treatment protocols. Our legacy will be to build the algorithms and tools to solve this tragic puzzle first for one person – and then for everyone.

BACKGROUND

THE H.O.P.E. ORCHESTRA FOUNDATION PROPOSAL

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According to a 2007 report by the World Health Organization, approximately 1 billion people around the world live with a neurological injury or disorder. The magnitude of this number, whether looked at from an economic or humanitarian standpoint, is enormous.

Yet, there is no known treatment for severe neurological injury; despite decades of research, thousands of safe, FDA tested drugs and immeasurable need, injuries such as blindness and paralysis remain permanent. These catastrophic injuries can lead to despair, loss of hope, and susceptibility to false hope. This false hope may create an irrational belief that repairing a nervous system will arise out of rehabilitation, simplistic monotherapies, and the current clinical trial system. The h.o.p.e. Orchestra believes we can and must create a world in which there is a personalized and comprehensive regrowth treatment plan for each patient’s neurological injury. What the field absolutely requires is a platform to orchestrate all this knowledge into personalized treatment plans for the most severely injured individuals.

"We can and must create a world in which there is a personalized and comprehensive regrowth treatment plan for each patient’s neurological injury."

SCALE AND SCOPE OF NEUROLOGICAL INJURY AND REPAIR

For nearly a century, science and medicine have been structured along a population framework and not a personalized approach. Populationbased medicine has remained at the heart of medical practice, under the assumption medicine should be relevant to most people, if it cannot be relevant to all. However, through newfound realizations about neuroanatomy and new understandings of the relationship between disease and disability, it has become clear this long-standing approach might not always be the best one. And there is no clearer example of when this approach must be abandoned than in neurological injuries, in which each injury is unique, with different magnitudes of cell loss and, as a result, require unique treatments.

Medicine and science are intertwined, but their missions are not the same. Medicine’s mission is the elimination of pathology and the restoration of health, while science’s mission is the discovery of facts. Science serves all of our interests by establishing facts based on cause and effect. This is the essence of science: observe a phenomenon, make a prediction as to the cause (the hypothesis), and then test that single variable (experiment) to determine whether or not the prediction is correct. If this cause and effect is reproduced reliably, then we can determine it is a fact. This pursuit of facts is best served by producing a large amount of data, inexpensively, and as fast as possible. The simpler and smaller the model, the more elegantly a fact can be demonstrated in cell biology. Funding agencies support researchers with a degree of freedom to pursue their own scientific interests, helping diversify and promote the discovery of new facts (reproducible biologic cause and effect). Basic science does not – and often should not – use human-scale models because these models are too big, too costly, and often inappropriate for the discovery of facts.

In contrast to an elegant scientific experiment,our central nervous system (CNS) is the most complicated structure on this planet. We continuously marvel at our complexity yet still propose clinical trial protocols too simplistic in their nature and approach to repair the

masterpiece that is the human central nervous system. This has resulted in tens of thousands of failed clinical trials. Thus, while the need for basic science to determine cause and effect is paramount, each scientist should not be held responsible both for the novel discovery of a single variable scientific fact and for promoting that single fact as a cure for a severe neurological injury. Establishing scientific facts is the foundation of advancement, and has provided us with thousands of safe drugs with known biological effects, but the value structures of academia and industry has conflated single facts of basic science with complex cures on a human scale. The solution is to approach the medicine of repair in an expansive, comprehensive manner and not in a simple, reductionist model.

CENTRAL NERVOUS SYSTEM ORGAN REPAIR PARADIGM SHIFT

Severe neurological injuries result in a staggering number of cells (neurons, astrocytes, vasculature, immune cells, etc.) being killed and the necessity to regrow all absent tissue is absolute, as every function a person can perform is anatomically based in the central nervous system. Just as electricity is physical, so, too, are thoughts, dreams, language, cognition, motion, vision, and every other action performed by a person.

Without true physical tissue repair, functional deficits such as blindness and paralysis remain permanent. Restoring a patient’s neurological function requires the regrowth of his or her unique missing tissue, and is the most difficult challenge in medicine. This is not organ repair, given the tissue is absent, rather the organogenesis of new tissue.

Is it possible to regrow the central nervous system? Yes, but we need to think differently – think bigger – than we ever have before. Until now, research has used models that have not captured, and perhaps could not capture, the overwhelming complexity of the CNS. The CNS is a trillion-cell supercomputer, having carefully evolved over billions of years, consuming 20% of our energy every day. It runs degrees hotter than the rest of the body therefore needs to be continuously cooled with ever-flowing cerebrospinal fluid. It is built from thousands of different cell types, including billions of neurons, billions of glia, billions of immune cells, and hundreds of miles of blood vessels, all working synchronously with one another. This means that even an injury of 1% of the CNS still kills over 10 billion cells and miles of cellular networks.

After an injury, the adult CNS also faces unique translational barriers that other organs do not. For example, many CNS cellular network connections were established in utero and stretched – rather than migrated – over the extreme distances within the adult CNS. This means nature does not have the ability to regrow expansive cellular networks after an injury to the adult brain. Furthermore, an injury forms new barriers to overcome, such as disrupted myelin and scar tissue, both of which stop a new cell from migrating. The ability to answer these essential questions, to overcome these barriers, and to develop expertise in mastering this extremely complex system is the future of personalized neurological repair.

Our core value - the premise upon which this entire initiative is built - is regrowth must be comprehensive and personalized. You can't just put random neurons, stem cells or a single drug in the CNS and ignore the rest of the complexity.

This is illustrated by asking the opposite question: What part of an individual's injured brain do we not want to regrow? Do we not want new cells to have a blood supply? Do we not want to remove the scar tissue, through which cells cannot migrate? Do we not want new neurons to reestablish these missing networks? The absence of any cell type in the brain is a known pathologic condition. For example, the presence of neurons and their axons without white matter (oligodendrocytes) is called leukodystrophy. The absence of blood flow to a region is called an ischemic stroke. This means treatment strategies that are not comprehensive literally introduce a new pathology on top of any existing injury.

Once a comprehensive, personalized approach is envisioned, one realizes a smaller, reductionist approach will not regrow the patient’s missing neurological tissue.

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Given the size and scope of the problem, it’s obvious why no comprehensive treatment plan has ever been developed for a patient suffering from a neurological injury. Comprehensive means the leading experts for every cell type in the CNS are assembled to calculate the number of missing cells for one individual patient and thereby personalize the resultant treatment protocol. This means experts on neurons (there are over 300 types of neurons), astrocytes, microglia, oligodendrocytes, the cerebrospinal system, ependymal, vasculature, lymphatic, and pericytes, are all assembled at a bare minimum, respecting that each cell category contains multiple types. Comprehensive treatment also means the calculations are made for what each cell type needs to both survive and integrate into

What if regrowth of the human brain indeed required the most complicated science ever conceived? That would actually make sense, and this is the scientific approach of the h.o.p.e.

Orchestra. We will assemble all of the necessary global scientific talent and perform the steps essential for comprehensive, personalized tissue regrowth. Each scientist will be asked to answer their specific part of the puzzle. We will test these calculations on a human-scale cell culture model, a replica of the patient’s missing tissue

The single biggest absolute requirement for our scientific success is to think on the scale of the human injury and not on current laboratory preclinical models. This immediately dwarfs the current size of any cell culture model on the planet, any government or industry research budget, the complexity currently allowed for a publication or regulatory application, and most people’s ambition. Therefore, we must be able to both advance this project with achievable steps, while ensuring every project is building toward human scale repair. This means allowing every member of the Orchestra to publish, to obtain grants, and to commercialize whatever he or she sees fit. We will accelerate more quickly toward our goal if every Orchestra member is thriving while improving the necessary tools and sophistication that we need to achieve functional regrowth.

The approach to neurological repair that will grow out of the h.o.p.e. Orchestra will be unlike anything ever before seen in the field.

Science requires integration, and integration, in turn, requires a new paradigm for research collaboration. The h.o.p.e Orchestra will

When it comes to neurological repair, the entire world of science and medicine needs to be harnessed around the individual. Nothing we do has value unless it is reduced to an individual or, as science says, an n = 1. Unfortunately, no government or industry research project would focus such intense resources on a single neurologically injured individual; yet this is the only way the necessary rigor will be applied to solve the translational barriers and for the decision points to be correctly made. Therefore, each personalized treatment protocol design needs to be sponsored by a patron with an unwavering interest in achieving functional neurological regrowth and allow h.o.p.e.

Orchestra to move beyond the normal barriers to success.

The expertise and technology that needs to exist, but currently does not, is the ability to 1. Know all existing pro-growth strategies; 2. Calculate the number of missing cells and cellular networks; 3. Understand the comprehensive cell biology necessary for complete regrowth of all cell types; 4. Calculate and simulate in silico personalized cellular regrowth requirements; 5. Choose specific cells/drugs for regrowth of the individual’s missing tissue; 6. Orchestrate the regrowth of the patient’s missing tissue in multiple, parallel, life- sized lab experiments using clinical delivery protocols.

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The strength of this team is we embrace the daunting nature of repair, and we seek to employ every good idea necessary to surmount all of the translational barriers. This global panel of experts will be equipped with the best knowledge graph ever to exist. They will be focused on what it would take – from their perspective – to regrow their piece of the puzzle. This will produce the most comprehensive repair strategy that has ever been attempted. Never before will so much computing power – both human and silicon – be applied to repairing an individual.

move beyond the barriers in academia and industry that have created intellectual silos focused on grants, patents, or publications at the expense of the teamwork required for comprehensive treatments. This historical approach has impeded meaningful progress for far too long. The immensity of this challenge demands we incorporate every necessary repair treatment into one comprehensive model, an unprecedented attempt to merge academic, pharmaceutical and biotech interests. If the goal is the restoration of function, the scar tissue must be removed, this injury cavity must be repopulated with the exact number of previously held cells, and the lost cellular networks must be regrown. Anything less will result in continued functional deficits. Previous research has skipped these essential steps, leading to innumerable failed clinical trials, a belief in the research community that brain repair is not realistic or achievable, and a sense of resignation among the neurologically injured that treatment will never come.

Neurological repair is never going to happen if any of these steps are missed or skipped because thousands and thousands of decision points need to be correctly made. The enormity of this challenge immediately requires a team approach to harness all required global technology, expertise and therapeutics.

It requires a scientific orchestra to regrow this masterpiece of biology. This new level of comprehension will create a more sophisticated approach to increase the speed and scale of tissue regrowth while reducing future costs so that everyone can benefit. Since brain repair will be the result of the algorithms and tools we propose, each step we master will create the foundation for the future. However, there is no time to waste.

PART 3

5NEW PARADIGM FOR ORGAN REGROWTH

with clinically relevant delivery methods.

Simple, yet staggeringly complex.

HUMAN-SCALE. ORGAN-REGROWTH. PERSONALISED. EXPERIMENTS.

functional, far-reaching cellular networks. This means, for each cell type, cell biology experts must calculate the absolute amount of growth factors, metabolites, apoptosis inhibitors, cell adhesion proteins, guidance cues, cell migration gradients, network integration factors – local, regional, and distant- as well as gap junctions, synapses, etc. These cellular requirements must then be reduced to clinical practice, which means each aforementioned cellular requirement needs a specific drug, dose, gradient, duration of treatment, sequence of administration, toxicity profile, delivery route, and excretion pathway.

This information is still not translatable until the actual drugs and cells are procured (a difficult task given the competing interests of each company and academic lab), ready for clinical delivery, and approved by regulatory agencies. Because most physicians and scientists do not believe repair is possible or are trapped in career metrics, no group has invested in the work necessary to make the above a reality. This is the mindset and barrier that we will have to overcome, ultimately positioning us for success. We do not believe neurological regrowth is impossible; it is simply the most difficult, most complex journey any scientific team has ever attempted.

THE H.O.P.E. ORCHESTRA FOUNDATION PROPOSAL

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ORCHESTRA OVERVIEW

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Where to start? Each neurological injury raises essential questions that must be answered in order to achieve success.

The immensity of the task we propose is by no means underestimated, nor is the magnitude of the challenge. The solution is a global orchestration of all required technology, expertise and therapeutics with each Orchestra member committed to the singular goal of regrowing the missing tissue of the individual patient. The ability to handle this complexity will be built into a novel, six-section scientific team designed to answer the essential questions and to fill the current technological gaps in performing human-scale science.

We will be able to handle extreme complexity, perform experiments on the human scale, and build algorithms to develop personalized regrowth strategies. And we will perform the calculations and experiments for as long as it will take to achieve functional repair. Given the career metrics in academia and industry, no one would undertake the immensity of this experiment except for those who work on h.o.p.e.

This will be the first human-scale laboratory organogenic model of the central nervous system, pioneering actual human regrowth on the scale of 1) human meters – not rodent millimeters, 2) human-scale billions of cells – not thousands of cells in cell culture dishes, 3)

nanometer resolution of human cell connections – not current micrometer resolution used in modeling and 4) months of treatment for cells to migrate human distances – not days used in current, arbitrary clinical trial timepoints. This also will be the first paradigm to choreograph basic science with the exact protocols to be used in the hospital, thereby addressing clinical translational barriers. Success awaits us at the intersection of the correct scale of sophistication with a personalized, comprehensive approach.

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ESSENTIAL QUESTIONS

h.o.p.e. ORCHESTRA

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The h.o.p.e. Orchestra is unique among science and medicine institutions. It will assemble the first comprehensive scientific orchestra to pioneer personalized functional regrowth of the CNS. In essence, members of this orchestra have been specifically selected because of their ability to help answer the essential questions of neurological repair, and because of their ability to synthesize knowledge into a complete, comprehensive, and personalized treatment protocol. This orchestra is assembled based on the specific requirements of a patient’s unique injury.

HUMAN-SCALE. ORGAN-REGROWTH. PERSONALISED. EXPERIMENTS.

1. Artificial Intelligence Pro-Growth Knowledge Base of all existing therapeutic options; 2. Missing Tissue Hologram to calculate the number of missing cells and networks; 3. Comprehensive Cell Biology for calculation of complete regrowth of all cell types; 4. Cellular Regrowth Simulation to organize

the sequence and duration of cellular requirements; 5. Regrowth Regimen Design to establish specific cells/drugs for personalized tissue regrowth; 6. Lab Clinical Trial section to orchestrate the regrowth of the patient’s missing tissue in multiple, parallel, life-sized experiments using clinically applicable delivery methods.

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This six-section orchestra is comprised of the following

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SECTION I: ARTIFICIAL INTELLIGENCE PRO-GROWTH KNOWLEDGE BASE

THE H.O.P.E. ORCHESTRA FOUNDATION PROPOSAL

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Machine learning allows curation and ingestion of the staggering amount of information now available. The computer is able to “learn” based on parameters it is instructed to do, such as determining which words appear most frequently and in what relationship to each other.

For example, including search terms such as “brain,” “growth factors,” “journal,” and “clinical trial” would allow the computer to produce a knowledge graph of which particular growth factors were studied in a clinical trial related to the brain. This narrowing of the field would allow the h.o.p.e. Orchestra experts on growth factors to review this knowledge graph and add their expertise to decide what information is of the highest quality with respect to each cell type.

This additional curation of the knowledge base would allow the computer to further narrow the literature based on even more specific search parameters, repeated over and over again.

There are more individual proteins (over 30,000) in a single cell than most people have days in their lives, making it impossible for a single person to “know” all of the complexities of a single cell. There are tens of millions of published papers on pro-growth strategies, tens of millions of unique chemical compounds from which to choose a therapeutic regimen, and a trillion-cell CNS supercomputer made up of a thousand different cell types to regrow. This complexity is beyond the understanding of a single person, and not knowing all of the relevant progrowth strategies makes designing a successful comprehensive treatment strategy impossible.

The old adage in medicine is immediately relevant: There are two types of treatments patients never receive - those the doctor does not know or those the doctor forgot. There is a third type of treatment a patient does not receive - a treatment that does not yet exist.

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A database of all pro-growth strategies needs to exist to identify which treatment options are the best and to identify where treatments do not yet exist. This will allow us to design our regrowth strategies with a realistic understanding of the current strengths and weaknesses of existing technologies.

TECHNOLOGY TO BE ORCHESTRATED

The field of neuroscience has produced an enormous amount of information over the last century. This has allowed an understanding of the complexities and nuances of neural repair never before conceivable. Just as importantly, rapid advances across equally vital fields have accelerated, including machine learning, brain atlases, informatics, engineering, cell biology, computer modeling, pharmacology, human physiology, etc. The amount of information generated in the last 50 years is remarkable and ready to be integrated into a comprehensive strategy.

Informatics The field of informatics has evolved to allow the processing of complex information in ever more simple schematics. This is incredibly important in order to narrow the complexity of the billions of data points into digestible parts for understanding and manipulation. Search parameters have continually improved, allowing for the user-friendly experience of these databases to thrive.

GOAL OF H.O.P.E.

We will use artificial intelligence to develop the first knowledge base of pro-growth strategies.

The integration of all brain atlases, drug library databases, all cellular pro-growth strategies, drug delivery options, toxicology, and the scientific literature base will ensure our scientific orchestra is always absolutely current. A comprehensive cell biology team, encompassing every cell type in the brain, will curate this knowledge base alongside machine learning experts. Bioinformatic experts will organize this database relative to the patient’s specific injury and translational disciplines (cell type, drugs, clinical delivery, etc.). This knowledge base section will assist the comprehensive cell biology section to increase the sophistication of the regrowth simulation of the individual’s injury, the design of the regrowth regimen, and the testing of this regimen in the lab

This knowledge base will be built as answers are sought to each question relevant to the patient.

Therefore, this knowledge base will be divided into the specific focuses of cell types to make progress realistic.

This database will by organized by 1. The individual’s neurological region of injury; 2. Cell type: neurons, astrocytes, microglia, oligodendrocytes, cerebrospinal system, ependymal, vasculature, lymphatic, and pericytes; 3. Technology arena: brain atlases, PubMed, image analysis techniques, human connectome project, stereology, drug libraries, Roche biochemical cell pathways, Ventner Institute mathematical modeling, cell migration kinetics, drug combo ranker, Food and Drug Administration database, biotechnology companies’ annual reports, pharmaceutical companies’ annual reports, world patent database, drug toxicology database, drug–drug interaction database, drug delivery literature, drug clearance database, bioreactors, tissue engineering, human brain physiology, electrophysiology, hydrodynamics, mechanotransduction, clinical delivery, material science, cell culture reagents, cell culture imaging, and 3D printing.

The machine learning team and comprehensive cell biology team will curate the knowledge base to select data and technology with the largest consensus regarding quality in the relevant areas necessary for pro-growth strategies for the patient’s unique anatomical injury. The informatics team will display the resultant knowledge graph in a searchable database easily queried by the key areas. We will maintain absolute rigor, so each promising therapy is considered and the context of each drug is known. We will make continual scheduled updates to ensure all literature and ideas are current. Given literature is delayed from idea creation to publication, continual check-in with the field is a must, and will be provided by each scientist. We will integrate vertical knowledge (e.g., information on a cell protein) for the first time with horizontal knowledge (e.g., information

on functional nervous tissue) in a translational approach around an individual injury. This will be the first knowledge base in a useable format for each orchestra member to integrate essential vertical and horizontal decision points around an individual injury.

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What are the best pro-growth ideas for each cell type in the brain? This information must be known to increase our odds for success.

clinical trial. We must ensure every good idea is known and considered, as any gap in knowledge decreases our chance for success.

IMPACT OF H.O.P.E.

The h.o.p.e. Orchestra will produce a pro-growth knowledge base that will be a resource for all future pro-growth steps, both for the h.o.p.e.

Orchestra and the entire biomedical research community. Artificial intelligence will allow more literature and data to be absorbed than previously possible by any academic or industry lab, as no academic lab or company has ever invested the resources to ensure they have the knowledge of every promising treatment option, as their incentives are different than ours.

The field will be dramatically accelerated forward, as will our chance for success, because it will be the first knowledge base to be 1.

2.

3.

4.

5.

Curated by each cell type by a comprehensive cell biology team; Focused on solving the problems of one patient’s unique anatomical injury at a time; Focused on pro-growth solutions; Pulled from all relevant databases; More expansive than any individual scientist physician could individually obtain.

"...a resource for all future pro-growth steps, both for the h.o.p.e.

Orchestra and for the entire biomedical research community. "

PART 3

Machine Learning

HUMAN-SCALE. ORGAN-REGROWTH. PERSONALISED. EXPERIMENTS.

PROBLEM TO BE SOLVED BY h.o.p.e.

THE H.O.P.E. ORCHESTRA FOUNDATION PROPOSAL

PART 3 PAGE 10

SECTION II: MISSING TISSUE HOLOGRAM

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TECHNOLOGY TO BE ORCHESTRATED

Central Nervous System Atlases Neuroanatomical atlases are ever increasing in their understanding of the exact cellular composition of the central nervous system.

The Automated Anatomical Labeling Atlas, Pubbrain, Brain Parser, Segmentation Validation Engine, Statistical Parametric Mapping, Human Connectome Project, Yale Neuro library, Allen Brain Atlas, and Montreal Neurological Institute all combined with a novel, global comprehensive cell biology team - will allow worthwhile calculations of the number of cell types missing from each region after the patient’s injury.

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Current MRI Image Analysis Technology Current medical imaging technology provides a gross structural view of the damaged brain, an accomplishment that revolutionized the field of diagnosing neurological pathology. To date, medical imaging of the central nervous system is similar to other organs. A typical MRI scan of the brain shows the injured area of the brain and shows evidence to help diagnose why the brain was injured.

Volumetric analysis is the ability to examine a region of the brain based on its size, and it has been a critical field of neuroscience for over a century. This involves regions of the brain being segmented out by researchers to further examine the interface among anatomy, function, and injury. The computing software, mathematics, and imaging technology are rapidly improving to further refine calculations of cell loss. The field is striving toward a goal of rapid, reproducible, and

Advanced imaging can now render a patient’s MRI, not just in 2D, but in more sophisticated 3D images called diffusion tensor images. This technology calculates the ease of water flow through a neuron. Given neurons extend great distances, this technology allows scientists to see the distant cellular network the brain relies on for rapid communication. This is especially important after a neurological injury because neuronal networks die back if their connections are not used in a process called Wallerian degeneration.

This die back is significant and must be quantified to understand what needs accurate regrowth on the network scale. Currently, no cellular network quantification is performed clinically.

However, a recent tool, called the NeMo Tool (network modification tool), quantifies losses in the brain connectivity network by mapping a specific patient’s lesions onto 116 regions of the brain, compared with 70 healthy, digitized MRI scans at the Montreal Neurological Institute.

This will allow a more sophisticated approach for calculating the number of missing networks to be reconnected.

Stereology, the science of calculating particles in 3D space, has been applied to the field of neuroscience over previous decades to provide necessary rigor to our understanding of the cellular composition of each brain region.

Stereology, the quantification of objects in space, was initially developed to ensure that materials such as cement were correctly composed.

For example, if cement has the wrong ratio of water to limestone and other oxides, it will cause structures such as bridges and buildings to collapse. When this is applied to the brain, it allows new levels of rigor in the calculations of cell loss. Technologies, such as StereoTool, are beginning to perform computer-based simulations and are being tested against the rigor of the stereological vertical sections method, thereby calculating the patient’s missing amount of tissue.

Advanced imaging techniques - partial leastsquares regression, bootstrapping, change in connectivity, NetMo, network modification,

GOAL OF h.o.p.e.

The input for calcutating the 3D missing tissue hologram is the patient’s available medical imaging (X-ray, CT, MRI, angiogram). The image analysis team then determine the negative space of missing tissue, the injury cavity, the absent networks, and the amount of scar tissue.

Cell biology experts, equipped with the most sophisticated knowledge graphs, anatomy atlases, and stereology algorithms, will calculate how much scar tissue is present and how many cells and cellular networks are missing per specific cell type. This output will be transformed into a 3D hologram to then be imported into the regrowth simulation as the beginning of repair calculations, as well as lab clinical trials as the template for the regrowth experiments.

We will enhance current advanced imaging technologies in conjunction with the comprehensive cell biology section to include the calculations answering the following questions: 1. How big is the negative space of the acellular cavity per volumetric analysis?

2. How much scar tissue (chondroitin sulfate proteoglycans, reactive astrocytes, immune cells, etc.) is surrounding the patient’s injury cavity?

3. How many cells (neurons, astrocytes, oligodendrocytes, pericytes, vasculature, lymphatics, ependymal, cerebrospinal, extracellular matrix and microglia cells) were lost?

4. How extensive is the cellular network loss (neuronal, astrocytic, oligodendrocytic, pericyte, vasculature, lymphatic, ependymal, cerebrospinal, extracellular matrix, and

microglia networks) in meters both for efferent (exiting) and afferent (incoming) connections?

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similar lesion dysfunction mapping, voxel-based morphometry, tract-based spatial statistics, vascular tree influence on connectivity, goodness of fit, and Akaike Information Criterion - are all advanced imaging analysis tools integrating brain atlases with a patient’s MRI images to produce a render of the lost cellular network following an injury. The techniques can be performed on any region of the brain and allow an understanding of the network loss. All of these are academic pursuits, but none yet have clinical applications.

IMPACT OF h.o.p.e.

Applying comprehensive cell biology expertise will dramatically enhance the sophistication of every brain atlas, connectome project, and image analysis technique. It will demonstrate to the field the rigor necessary to calculate the unique injury of a single person. By solving one injury at a time, an algorithm will be built to integrate into all future imaging, allowing all future patients to know the extent, prognosis, and required interventions for their unique neurological injury.

For the first time, we will actually know the number of cells needed to replace the missing part of the individual’s damaged CNS. This will have a clarifying effect on the entire field of basic neuroscience by showing how overly simplistic monotherapy rationales are toward repairing the vast number of missing cells. For clinical trials, this new information will finally allow the correct dosing of therapeutics.

All neurological disorders need these calculations applied to them in the future. As funding agencies and advocacy groups have their particular disease of interest, these analyses will set an accurate benchmark for future grants, lab research, and clinical trials, so that their research questions will actually match the disease burden to be overcome.

"...this new information will finally allow the correct dosing of therapeutics."

PART 3

No medical imaging or research group can currently calculate the amount of cell loss, scar tissue burden, or cellular network loss after a neurological injury. Furthermore, no brain atlas, publication, or research group has achieved a comprehensive cell atlas capable of calculating the amount of cell loss for a specific injury.

Without this knowledge, it is impossible to accurately dose any therapeutic. A “damage report” needs to exist to quantify how much missing tissue needs to be regrown.

automated calculations of tissue loss, but has not yet incorporated comprehensive cell biology calculations.

HUMAN-SCALE. ORGAN-REGROWTH. PERSONALISED. EXPERIMENTS.

PROBLEMS TO BE SOLVED BY h.o.p.e.

THE H.O.P.E. ORCHESTRA FOUNDATION PROPOSAL

PART 3 PAGE 12

SECTION III: COMPREHENSIVE CELL BIOLOGY

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A new comprehensive biology team is necessary for a successful approach to tissue regrowth.

TECHNOLOGY TO BE ORCHESTRATED

Comprehensive Cell Biology

13The central nervous system is made of thousands of cell types. The major categories are the following:

1. Neurons: process and transmit information that underpin all functions; 2. Astrocytes: blood brain barrier, process information, glial scar formation, process metabolites; 3. Oligodendrocytes: increase conduction speed of information; 4. Glial scar: limits damage from spreading, barrier to new growth, influences immune system; 5. Pericytes: integral to blood supply and tissue remodeling; 6. Vasculature: supplies metabolites, stem cell migration pathway; 7. Lymphatic: clears extracellular fluid; 8. Ependymal: forms the cerebrospinal system compartments; 9. Cerebrospinal system and choroid plexus: cools the brain, establishes growth factor gradients; 10. Extracellular matrix: provides tertiary structure for brain, component of scar tissue; 11. Microglia: immune cells of brain, regulates new synapse formation.

It is a generational achievement that this level of cellular biology information is available in multiple databases such as PubMed (the national literature database supported by the National Institute of Health). Searching for the scientific literature on pro-growth strategies alone (cell types, growth factors, metabolism, apoptosis, cell adhesion, guidance cues, cell migration, nutrient migration, network integration – local, regional, distant, etc.) yields over 12 million papers. This level of complexity needs to be organized into translational steps, and no part ignored.

Every field of neuroscience will be greatly improved by adding a comprehensive biologic approach and a bold insistence to solve the problem of regrowing a single patient’s missing neurological tissue. This applies to knowledge bases, imaging, understanding neurological injury, improving computer models of repair, designing therapeutic repair strategies, and conducting laboratory cell culture experiments.

This is the paradigm shift that will permanently shape the field of personalized neurorepair and begin to close the translational gap between this enormous scientific expertise and individual patient treatment plans.

GOAL OF h.o.p.e.

Cell biologists spend decades developing sophisticated vertical and horizontal knowledge about their one cell of expertise. They have strong opinions on what the best papers are, the best growth factors, the best cell sources, etc.

For each of these decision points, the specific scientific talent will be assembled to orchestrate the best practices available to date. This is the vehicle by which each scientific soloist can imagine, decide, and implement exactly what

Artificial Intelligence Pro-Growth Knowledge Base Comprehensive cell biologists will work with the bioinformatics and artificial intelligence teams to curate the literature base by each cell type to improve all decision points regarding tissue regrowth.

Missing Tissue Hologram Comprehensive cell biologists will calculate 1) the total amount of the patient’s glial scar tissue; 2) the number of missing neurons, astrocytes, oligodendrocytes, pericytes, vasculature, lymphatics, ependymal, cerebrospinal system, extracellular matrix, and microglia resulting from the patient’s injury; and 3) the extent of the patient’s missing neurons, astrocytes, oligodendrocytes, pericytes, vasculature, lymphatics, ependymal, cerebrospinal system, extracellular matrix, and microglia cellular networks.

Regrowth Simulation Comprehensive cell biologists will calculate what is needed to 1) remove all the patient’s glial scar tissue; 2) replace all the patient’s missing cells; and 3) reconnect all the patient’s missing cellular networks.

Comprehensive Cell Biology Comprehensive cell biologists will calculate what is needed to 1) keep the injury cavity permeable; 2) ensure the survival and function of each cell type regarding their absolute requirements of growth factors, metabolites, blood supply, cell trafficking, cell communication, apoptosis, cell adhesion, cell-cell interactions, guidance cues, gap junctions, etc.; and 3) ensure the survival and function of each cellular network regarding the absolute requirements of growth factors, metabolism, apoptosis, cell adhesion, guidance cues, cell migration, nutrient migration, network integration – local, regional, and distant, gap junctions, synapses, etc.

Regrowth Regimen Comprehensive cell biologists will decide 1) what is the best cell replacement source per cell type; 2) which drugs, dose, duration, sequence,

delivery, toxicity, and excretion are needed to remove the glial scar; 3) which drugs, dose, duration, sequence, delivery, toxicity, and excretion are needed for each cell type survival and function; and 4) which drugs, dose, duration, sequence, delivery, toxicity, and excretion are needed for each cellular network survival and functional integration.

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is needed to regrow their part of a specific patient’s missing tissue.

Lab Clinical Trial Comprehensive cell biologists and physicians will decide what is required for 1) drug delivery in a clinically simulated lab trial to regrow the patient’s missing tissue; 2) clinical protocol orchestration and rehearsal; and 3) Food and Drug Administration and Hospital Institutional Review Board applications.

IMPACT OF h.o.p.e.

No academic or industry lab has a commitment to a comprehensive cell biology approach given the scale, scope, complexity, and current career incentives, thereby resulting in the oversimplification of treatment strategies. These diverse cell types in the central nervous system are irrefutable facts of nature. Ignoring them may be more expedient and simple, but this guarantees translational failure, as evidenced by the tens of thousands of failed clinical trials.

With this new commitment to comprehensive cell biology, the field of neuroscience will have a new standard to be met, and, finally, enhanced rigorous calculations will be applied on the human scale to an individual. For the first time, a single injury will have a repair strategy built around a complete understanding of all of the involved cells, the length of the missing cellular networks, and the individual requirements for the functioning of each cell.

"You can't just regrow random neurons."

PART 3

The central nervous system is made up of thousands of cell types. The major categories are neurons, astrocytes, oligodendrocytes, glial scar, pericytes, vasculature, lymphatics, ependymal, cerebrospinal system, extracellular matrix, and microglia. They all play an important role in a normal functioning brain. The absence of each and any cell type results in a known pathological disorder. Deciding to replace or treat only one cell type essentially overlaps a known pathologic condition onto the existing neurologic injury. Therefore, all must be replaced in order to restore function after a neurological injury. However, no industry or academic lab has undertaken a fully comprehensive cell biology approach to repair.

Human physiology is now better understood than ever before. We understand only a minority of cells are neurons, and they are actually part of a complex unit of different cell types. We understand the enormous influence that blood vessels exert in the brain, the cerebrospinal system’s ability to shape the structure of the brain, the effect of 3D growth on cell morphology, the effect of flowing hydrodynamic mechanical forces on cell function and form, and the other thousands of parts integrated into the trillion-cell supercomputer which is our central nervous system.

HUMAN-SCALE. ORGAN-REGROWTH. PERSONALISED. EXPERIMENTS.

PROBLEMS TO BE SOLVED BY h.o.p.e.

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SECTION IV: REGROWTH SIMULATION

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The ability to follow thousands of steps in an organized fashion has increased across multiple fields. Informatics continue to make great strides in developing the methods, algorithms, and tools necessary to unravel the complexities of enormous data. There are rigorous computational methods integrating medical and experimental observations with systems and chemical biology to explain drug effects and to further our understanding of basic biology and human disease. These fields are at the forefront of the use of high-throughput data-capture technologies - such as genome and transcriptome sequencing, metabolomics and proteomics.They use empirical medical observations to study the complex interplay among genetics, the environment, and disease. These technologies improve the modeling of an initial biochemical signal with the progression of a broad spectrum of molecular events. This progress also extends into 1) modeling the multiscale dimensions required in tissue engineering; 2) increasing the power of computing capabilities; and 3) the integration of predictive models among biology, materials science, and physical stimuli on cell phenotype.

All of this progress will be used to develop a patient-specific multiscale (both cellular and cell network) model.

TECHNOLOGY TO BE ORCHESTRATED

Cell Biology

15The global scientific community has spent

generations assembling a knowledge base of the 30,000 proteins in a single cell and the trillion cells in the CNS. Everything from DNA replication, RNA production of proteins, and the assembly of these proteins into the workhorses of the cell is described in exceptional detail. Institutes such as Roche Scientific and the Ventner Institute have impressively compiled this information into a single chart and mathematically modeled a single cell, respectively. There is an absolute wealth of information on guidance factors (proteins that control cell and cellular processes movement), growth factors (proteins responsible for cell survival and viability), metabolism (the processing of substrates for energy), blood supply (the complex flow of metabolites), cell communication (the regulation of information between cells), network integration (local, regional and distant), gap junctions (open pores between cells critical for cell regulation and communication), synapse formation (the unit of transmission of information between neurons), etc. This scientific exploration of biology produces daily insights and transforms how we understand life year after year.

Computing power and access to this processing power are also rapidly increasing. Use of the supercomputing grid allows the development of models with high enough complexity in their resolution and in the management of databases to make significant progress toward modeling a single patient.

Modeling extreme complexity can also make use of the experience and expertise offered by other fields. Of particular relevance on the micrometer and angstrom (cellular) scale is industrial engineering, which has expertise and operating procedural models to build computer chips in clean rooms requiring thousands of exacting steps to be perfectly performed in the correct sequence on microscopic scales.

On integrating multiple materials, architectural blueprints encapsulate the vast array of different materials and expertise that must be coordinated in the correct sequence and timeline to build towering skyscrapers. On a network scale, energy grids have the software to build, monitor, and manipulate the production and flow of

Data Visualization Informatics is ever improving in terms of the challenge of visualizing data and information.

The field is striving to create higher “data-toink ratios,” i.e., to layer more data into a figure with finer detail available to the user. Several companies can provide holographic visualization of scientific data with the most up-to-date 3D imaging techniques, or other complex engineering data, to produce digital holographic models for scientific and research display purposes.

GOAL OF h.o.p.e.

We will build a 3D spatial and temporal regrowth simulation designed to match treatment strategies for unique neurological injuries. We will master the complexity of cell biology by using machine learning, computational biology, and computer modeling in an evolving logic platform. This regrowth simulation will establish a treatment regimen that calculates on a cellby-cell basis the total amount and sequence of therapeutics required. This means, for the unique injury visualized by the 3D missing tissue hologram, the calculations will begin with The removal of the glial scar around the injury cavity: 1. How is the patient’s specific amount of glial scar removed (enzymes, small molecules, viral vector)?

2. How is the dissolved scar tissue cleared from the injury site?

3. How is the permeable environment maintained?

Cell replacement of all cell type: 1.

2.

What will the new cell sources be (stem cells, mature cells, subventricular zone, mitosis, migration)?

What are the cell survival requirements for each cell type (growth factors, metabolism, blood supply, cell trafficking,

cell communication, apoptosis, cell adhesion, cell-cell interactions, guidance cues, gap junctions)?

3. What is the sequence and timeline for each cell-type regrowth?

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The human central nervous system is composed of a trillion cells. Even a 1% loss of tissue is 10 billion cells extending over collective miles of cellular networks. No research group has the ability – or has even tried – to calculate precise, comprehensive cellular requirements for neurological regrowth on a rodent level, let alone a human scale. The CNS is too complex to be fixed with a single agent, and the complexity of repair must be calculated in terms of what is required to replace and reconnect the patient’s missing neurological tissue. Therefore, we must build the necessary logic engine, or “repair manual,” to match the thousands of treatment points onto the correct scale and scope of neurological damage, to be a repository for our evolving regrowth strategies, and to offer the means to track experiments and learn from them.

electricity from thousands of energy sources across expansive networks. The scales are enormous, the teams are large, and the budgets are significant. However, the ability to plan and execute thousands and thousands of decision points is not new and has been repeatedly performed, as witnessed by viewing any city skyline lit up at night.

Cellular network regrowth: 1. What are the migration cues and gradients (excitatory, inhibitory, rostral, caudal, ventral, dorsal)?

2. What are network survival requirements for each cell type (growth factors, metabolism, blood supply, cell communication, network integration – local, regional, and distant, gap junctions, synapse formation)?

3. What is the sequence and timeline for each cellular network regrowth?

These survival and functional calculations for every missing neuron, astrocyte, oligodendrocyte, pericyte, vasculature cell, lymphatic cell, ependymal cell, cerebrospinal system cell, extracellular matrix configuration, and microglia cell regarding the absolute number of required growth factors, metabolites, apoptosis cues, cell adhesions, migration guidance cues, nutrient gradients, network integration factors (local, regional, and distant), gap junctions, and synapses will be orchestrated regarding the dosing, gradient, sequence, and duration of treatment.

This calculation will be the template for the regrowth regimen design section and will be tested in the lab clinical trial to ensure tissue regrowth would unfold as predicted.

PART 3

Computer Modeling

HUMAN-SCALE. ORGAN-REGROWTH. PERSONALISED. EXPERIMENTS.

PROBLEM TO BE SOLVED BY h.o.p.e.

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IMPACT OF h.o.p.e.

This will be the first attempt to match the scale required for the regrowth of an individual’s unique neurological injury with all known progrowth cellular knowledge in a repair strategy built around a complete understanding of all the involved cells, their networks, and functional requirements.

THE H.O.P.E. ORCHESTRA FOUNDATION PROPOSAL

PART 3

This is a comprehensive strategy to achieve central nervous system organogenesis, one that understands the complexity of the problem and tackles each challenge systematically.

If billions of cells are lost, how do we correctly replace those billions of cells? If a meter-long neural network is gone, how do we regrow that meter-long neural network? How much glial scar is present? How do we exactly bypass the inhibitory glial scar? Which cells need to be repopulated first? Second? Third? How far do all cells need to travel to integrate into their respective functional networks? How long for each axon to reach its target? How much growth factor is needed? Each of these questions, as well as many others, needs to be answered. A cell does not move without a combination of upstream guidance molecules cueing the internal cellular machinery to move. Therefore, if we want a cell to move 500 millimeters and then turn left, we have to provide it with the correct amount of drugs to travel 500 millimeters and turn left.

The current state of science does not have the capacity to answer these vital questions and falsely hopes that the cells have enough inherent knowledge to fix themselves. Unfortunately, when cells formed the correct neural networks, we were in utero, about the size of a grape, and our neural networks stretched rather than migrated. For a new neural network to migrate adult distances after an injury is unprecedented, and we must assist the cells in their attempts at regrowth. We must significantly “nurture” our “nature.”

This proposal is predicated on the belief that massive gaps in logic have caused these extreme failures of translational medicine. Easy examples are gathered from the fact that very few neuroscience experiments factor blood supply into their repair strategies. Without a blood supply, the attrition rate of every stem cell injection is catastrophic. The numbers from previous trials demonstrate injections of

millions of stem cells leads to the survival of only thousands, and those surviving were likely just luckily near a blood supply. Additionally, after a brain/spine injury, scar tissue forms around the damaged area, which prevents new cells from migrating to and through the area.

The combination of this scar tissue and the lack of blood supply leaves all stem cell trials with trapped cells that quickly die. This proposal’s new level of comprehension will help to create a more sophisticated approach to increase the speed, scale, and success of tissue regrowth.

It is imperative that these vital questions be addressed and orchestrated with the use of current best practices. A logical, organized computer model is vital toward the goals of not wasting time or resources, learning from failures, handling the staggering complexity (billions of cells at the nanometer and molecular level), being the repository of new knowledge, and creating the necessary clinical protocols, which will be used for the patient.

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"We must significantly 'nurture' our 'nature.'"

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THE H.O.P.E. ORCHESTRA FOUNDATION PROPOSAL

PART 3 PAGE 18

SECTION V: REGROWTH REGIMEN

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TECHNOLOGY TO ORCHESTRATE

Current Drug Libraries

19Science has discovered over 125 million

unique chemical and molecular compounds with tens of thousands of these compounds being drug candidates. These compounds are categorized in existing drug libraries and include information on FDA approval status, what cellular mechanisms are influenced, toxicology, drug– drug interactions, drug clearance and excretion, drug delivery options, etc. These drug libraries also can be examined per intellectual property, patents, cost, and supplier options. These massive resources will allow the selection of a comprehensive repair strategy, which has never before been possible.

Combination Drug Modeling The fields of pharmacology, informatics, and dynamic modeling are sophisticated in their analysis of millions of decision points within databases. Combination drug design is also making great progress in fields such as infectious disease and cancer, but has not yet been applied to neurorepair. Combinatorial drug design is a drug functional network analysis partitioned into numerous drug network communities, integrative informatics, and dynamic modeling.

Powerful computational tools can predict drug combinations, targeting multiple signaling modules of cellular molecular pathway networks

Safety Despite all of the FDA requirements and preclinical trials, drugs frequently have off-target effects, which are only detected once they are used in routine clinical practice. However, over 30 surveillance research organizations exist to address this problem. Each one is working to develop better algorithms to detect drug effects - both alone and in combination therapies - by using big data regarding molecular mechanisms, coupled with the electronic health record.

Predictive models are ever advancing and are critical tools to speeding up the discovery of safe, comprehensive therapies. The need to monitor for toxicity will always be vital.

A) Removal of scar tissue around patient’s injury cavity: 1.

2.

Which drugs for extracellular matrix removal, debris removal, and immune modulation?

What dose is required per number of each extracellular matrix protein, debris amount, and immune cells present?

3. What duration of drug delivery is needed?

4. In what sequence should drugs be delivered?

5. What is the delivery method for each drug?

6. What drug–drugs interactions will exist?

7. What toxicity may arise?

8. What excretion pathway will be present for each drug?

GOAL OF h.o.p.e.

B) Cell replacement survival (for each cell type):

Drug combinations are a hallmark of therapies for complex diseases such as cancer, HIV, and hypertension, but no one has applied this technology to brain tissue engineering. All of the aforementioned technologies need to be repurposed into pro-growth strategies for neural tissue regrowth. Also, because drug effect is dose-dependent, the effect of multiple doses of an individual drug needs to be examined, the result of which may yield a rapidly rising number of combinations and a challenging highdimensional statistical problem. The h.o.p.e.

Orchestra will make informed decisions about each possible drug combination and test these predictions in the lab, which is explained further in the following section about the lab clinical trial.

1. Which drugs for growth factors, metabolism, blood supply, cell trafficking, cell communication, apoptosis, cell adhesion, cell–cell interactions, guidance cues, gap junctions?

2. What dose and gradient are required for each cell?

3. What duration of drug delivery?

4. In what sequence should drugs be delivered?

5. What is the delivery method for each drug?

6. What drug–drug interactions will exist?

7. What toxicity may arise?

8. What excretion pathway will be present for each drug?

The regrowth regimen section begins with the calculated amount of cell damage from the 3D missing tissue hologram section and the biological requirements for regrowth calculated by the regrowth simulation section. The comprehensive cell biologists, computational modelers, drugcombo rankers, and informatics will develop the total amount of therapeutics needed to achieve cellular repair. “Comprehensive” remains the

C) Cellular network guidance and survival (for each cell network type): 1. Which drugs for growth factors, metabolism, blood supply, cell communication, network integration – local, regional and distant, gap junctions, synapse formation?

2. What dose and gradient are required for

each cellular network?

3. What duration of drug delivery?

4. In what sequence should drugs be delivered?

5. What is the delivery method for each drug?

6. What drug–drug interactions will exist?

7. What toxicity may arise?

8. What excretion pathway will be present for each drug?

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key term. This complex number of therapeutic options must be orchestrated to include dose, duration, sequence, cost, purchasing option, delivery route, toxicology, drug–drug interactions, pharmaceutical grade, and excretion per each cell type. The process begins with

The output of this phase will be a treatment strategy that is a detailed algorithm of the thousands of decision points performed in the lab clinical trial phase. Reproducible experiments will always remain the gold standard in science to validate the predictions made by these models.

IMPACT OF H.O.P.E.

As the team learns and the model develops simultaneously, there will be a feed-forward acceleration in sophistication to regrow a patient’s injured CNS. The scientific, medical, technical, legal, and financial challenges will be illuminated from this regrowth regimen, and identify additional translational barriers to be surmounted.

"No neural repair drugs are being used in clinical practice today."

PART 3

No neural repair drugs are being used in clinical practice today. One of the reasons is that neurological injuries are extremely complex and no academic or industry lab has applied the necessary skills and resources to solve the enormous challenge of how to regrow the missing tissue. Additionally, different incentives in industry and academic labs discourage tests on a drug for which they don’t own the patent or that doesn't present the possibility of a novel publication. This also means that the skill set to orchestrate thousands of therapeutic decision points into a safe, comprehensive, and translatable treatment plan does not currently exist. The field desperately needs a comprehensive approach to central nervous system tissue regrowth that overcomes these barriers.

through the analysis of proteomic pathways.

As drugs within overlapping communities share common mechanisms of action, scientists can build specific signaling networks based on targets.

The idea is to use as few drugs as possible to overlap with as many pro-growth pathways as necessary. A preference will be to use drugs with an established safety profile and have been used in prior clinical trials.

HUMAN-SCALE. ORGAN-REGROWTH. PERSONALISED. EXPERIMENTS.

PROBLEM TO BE SOLVED BY h.o.p.e.

PART 3

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SECTION VI: LAB CLINICAL TRIAL

THE H.O.P.E. ORCHESTRA FOUNDATION PROPOSAL

20

"We merge science and engineering to achieve our aims."

21

Clinical trials include test treatments under human physiologic conditions, but an individual cannot subject him or herself to every clinical trial, especially because being in previous clinical trials is an exclusion factor for others. This also makes it impossible to test multiple treatments in parallel with one another. This is even more relevant if multiple combinations of drugs need to be tested to determine which combination works best, a process not currently allowed.

Furthermore, a clinical trial (like animal research) cannot visualize cellular regrowth processes, let alone correct course midtrial based on the needs of the regrowth process. This is even more relevant given the regrowth of large cellular networks, at a rate of 1 millimeter per day, may require a thousand days to reach the necessary meter. No trial is conducted over this amount of time, which immediately halts any chance of success. Also, every injury has a different amount of missing tissue; yet the doses are circumscribed for an entire study cohort of patients.

Preclinical biological research must begin working on the human scale and use clinically realistic

The regrowth simulation and regrowth regimen must be able to successfully regrow the missing tissue hologram template before any clinical trial would be deemed both safe and worthwhile.

TECHNOLOGY TO BE ORCHESTRATED

3D Cell Cultures Scientific expertise now exists in regards to the importance of cells growing in three dimensions. It dramatically affects their size, shape, and function and even changes the genes and proteins they produce. 3D cell cultures have made significant progress in the technical requirements necessary for these experiments, which include incubators, fluid dynamics, cell media, scaffolding, etc. Furthermore, tissue engineering has expanded these 3D models to include multiple cell types representing a more physiologic environment in organs such as the lung and liver. However, no multi-cellular, 3D model currently exists for the human brain.

Human Physiology This personalized neurological repair model must be closer to human physiology than current lab cell culture models. There is already an enormous amount of intrinsic, unknown variables that make any model fraught with assumptions and translational failures. Therefore, this model will be incubated at the human brain temperature, and not at the current temperatures currently used in cell biology, which are degrees colder.

The body is in constant motion, so the direction of flow will be calculated for the patient’s injury proximity relative to his or her circulating cerebrospinal system and vascular system.

Current cell culture models have cells either lying stagnant in sitting media or sitting on a shaker, which provides bidirectional motion, but neither

Drug Delivery The central nervous system has a blood brain barrier, which separates the vascular compartment from the rest of the brain and prevents most pharmaceutical drugs from entering the brain. After an injury, this barrier can be destroyed, but typically direct access to the central nervous system remains limited. There are numerous approaches to drug delivery, including 1) direct injection; 2) vascular injection (arterial or venous); 3) cribriform plate entry (through the nose); and 4) a lumbar puncture (bottom of the spinal column). Drug delivery by direct injection requires an operating room and destruction of neighboring tissue and can only reach a small amount of the brain. Utilizing a lumbar puncture, a cribriform plate, or a vascular approach also face logistical challenges. The field does not have a gold-standard drug delivery method, which would allow any therapeutic option to be delivered in the right place and for the right amount of time. However, an astonishing array of promising approaches have been produced in the last ten years, including linking nanoparticles such as iron and nickel particles - to therapeutics,

allowing their manipulation with magnets. Each therapeutic option will need to be matched with the right mode of delivery, as stem cells, enzymes, and small molecules all pose distinctly different challenges.

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of these models actually exists in nature. Also, humans have lungs and a vasculature to deliver oxygen to cells, meaning cells do not live exposed to room air and atmospheric oxygen tension. For this reason, our model will exist at the correct exposure to oxygen and other metabolites.

Also, cells in the brain are born, live, and grow in complete darkness. They are never exposed to light, therefore requiring our model to employ infrared optics to capture cell migration and growth in real time. Drug deliveries to current cell culture and rodent models are also not what will be used in the clinical protocols. Therefore, a skull with accurate foramens, complete vertebral column, CSF, and vascular entry points will be incorporated into the model. The distance from the individual’s potential drug delivery entry points to their missing tissue will be measured from the individual’s own physical exam. A safe delivery mode is required as the duration of drug delivery will likely last months. These delivery points need to be tested in a life-size model of the individual in exactly the way we will later administer the treatment protocol in hospital.

Clinical Trials Clinical trials have rigorous documentation for the dosing, length of a trial, and monitoring of side effects and outcomes. For any trial to proceed, these protocols and documentation must be approved. The applications to regulatory bodies such as the FDA can cost millions of dollars and are another burden to translation, as the culture is risk-averse to future litigation.

Medical fields such as cancer treatment allow borderline lethal side effects, as death is the alternative outcome. But the FDA does have a regulatory pathway to expedite drug approval for severe, life-threatening illnesses. And although this proposal fits the Unmet Medical Need pathway, the regulatory bodies will have no appetite for any therapeutic regimen this complex for a patient who does not have death as an immediate consequence. Bioethics will usher this proposal through these riskaverse agencies.

PART 3

Current lab research does not perform experiments on the scale of humans. This means experiments are done on thousands of cells measuring distances in micrometers and not the billions of cells over the scale of meters as necessary in human brain functioning. Current lab research typically tests simple treatments on a single layer of neurons laying on a flat, stagnant dish, or small neurospheres, which do not approximate human physiology (hydrodynamics, temperature, oxygen concentration, 3D, gradients, etc.), let alone capture the thousands of treatments needed for 3D nervous system organogenesis. Furthermore, current preclinical animal research uses delivery methods for its experiments not allowed in humans and certainly do not use clinically relevant, FDA-approved delivery methods. Animal models are also too slow because each mouse, rat, or primate must be killed and dissected to examine the cellular growth produced in an experiment, meaning the experiment cannot make adjustments during the experimental phase, which is not how clinical medicine is practiced.

scenarios to achieve successful organogenesis of a unique, missing piece of an adult CNS. The ability to administer a comprehensive treatment this complicated must be practiced and rehearsed to ensure a safe, accurate procedure. Therefore, a life-sized, personalized model of the individual’s missing tissue must be created to perform multiple, parallel experiments with the goal of regrowing the individual’s specific missing tissue.

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PROBLEMS TO BE SOLVED BY H.O.P.E.

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GOAL OF h.o.p.e.

THE H.O.P.E. ORCHESTRA FOUNDATION PROPOSAL

PART 3

The lab clinical trial section will orchestrate the regrowth of the patient’s missing tissue in multiple parallel attempts, acceleraing progress, which is not possible in clinical trials. It will coordinate the skills of diverse researchers orchestrating the regrowth regimen to regrow the damaged tissue, as determined by the 3D missing tissue hologram template. The final therapeutic repair strategy will have well over 20,000 decision points, requiring rehearsal. We must ensure the performance is feasible, monitoring for cancer formation, drug toxicity, and cell death, before entering a clinical trial for an individual. Clinical delivery barriers also must be solved with the cell biology challenges, requiring this model to be as close to human physiology as technically possible and also to have testable, life-sized, and personalized clinical drug entry points.

Every experiment needs a positive control, a negative control, and a dynamic range to show a statistically significant separation between experimental and control groups.

This is less true for engineering, where the goal is visible and practical. This section is testing to see whether our tissue engineering perfectly overlaps with the negative space of the patient’s missing tissue. This is where we merge science and engineering to achieve our aims. Currently, science stops an experiment to measure its outcomes. In a clinical trial, a stop date is decided before the trial begins.

However, to save time and cost, we will use our Missing Tissue Hologram as our negative control as well as our positive control. Similar to building a bridge, we will visually inspect the personalized model in real time to see if cells are migrating and connecting according to the Regrowth Simulation and Missing Tissue Hologram. Mid-experiment corrections will be done, similarly to what is done clinically when doctors make rounds for patients in the hospital and adjust medication doses as needed. All data will be fed back into the Pro-Growth Knowledge Base, the Regrowth Simulation, and a new Regrowth Regimen will be generated based on the latest Lab Clinical Trial results to further refine the algorithms

until the right level of regrowth is achieved mirroring the Missing Tissue Hologram.

Functionality of the tissue will be measured by electroencephalography (EEG) and transcranial magnetic stimulation (TMS).

Clinical trials in the United States require an enormous amount of documentation to proceed. This applies both to the FDA and a hospital’s own institutional review board.

Documentation must be provided for the pharmaceutical grade of the drug, dosing, rationale, and purchase options as well as trial duration and numerous other categories.

The h.o.p.e. Orchestra is seeking to produce a therapeutic proposal dwarfing any previous application in size and complexity. However, these regulatory institutions have no inherent interest toward anything so complex. The burden will be on us to show compelling evidence with sufficient data and rigor to then proceed to clinical use.

IMPACT OF H.O.P.E.

Never before has cell biology been performed on the scale and scope of a human model; however, until we achieve the precision, comprehension, and complexity to the utmost perfection, our treatments will inevitably fail.

The ability to answer and orchestrate these essential questions, overcome these barriers, and develop expertise in mastering extreme complexity is the future of personalized neurological repair.

Testing combinations in a transparent replica of the patient’s injury will increase the speed of discovering therapies producing a plausible change in functional recovery. This transparency means we do not have to set a time-point for an experiment, which would result in losing time and resources; rather, we can course-correct the experiment until we achieve the desired cellular growth. This is not possible in a rodent, primate, or clinical trial approach. This has never been done before, and the novel insights produced will be extraordinary and immediately publishable.

As the team learns and the model develops, there will be a feed-forward acceleration

in the sophistication necessary to repair a patient’s injured CNS. No single patient can participate in every clinical trial, so this is a way to regrow the damaged part of the patient’s central nervous system by utilizing all prevailing options. Each technology used will be enhanced by having both comprehensive biology and the human scale applied to it, including bioreactors, tissue engineering, clinical delivery methods, 3D printing, etc. The individual patient will benefit from having every relevant facet of global technology applied to him or her to accelerate the goal of achieving functional regrowth of the damaged central nervous system.

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THE NONPROFIT RESEARCH FOUNDATION: ORCHESTRA OPERATIONS

THE H.O.P.E. ORCHESTRA FOUNDATION PROPOSAL

24

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We will write the score for the achievement of neurological regrowth.

NEW PARADIGM, NEW POSSIBILITIES

It is precisely because this is the only plausible path to patient recovery that everyone will benefit. The primary beneficiary of this new, orchestrated approach to neurological regrowth will be, deservingly, the patient. For as long as neurological injury has existed – which is to say for as long as science has existed – there has been no practical treatment. Patients arriving at hospitals or rehabilitation centers with paralysis, blindness, deafness, or the inability to communicate, are typically given little to no hope when it comes to treatment and recovery, and it is a social imperative to see this change.

Scientists and physicians have worked tirelessly on this precise question, but now we are in a unique age in which scientific and medical innovations of various kinds can collaborate to seriously pursue solutions requiring diverse arrays of knowledge working together. This is our most immediate and practical opportunity to treat neurological injuries, and it is our best opportunity to alleviate the suffering many of these people face every day.

Furthermore, each scientist will have access to

The number of Orchestra members, the timeline proposed and the budget, are all written as the starting minimum, and will continue to grow over time. The successful treatment protocols will eventually be on the scale of equal magnitude projects such as supercomputer construction ($3 billion) or a moon landing ($25 billion).

We will begin this process with small enough and achievable goals to demonstrate proof of concept, to successfully advance technology platforms, and to leverage this new standard of science into greater speed and sustainability. The timeline for each cycle of innovation will decrease as the size of the team and the budget increases.

The greatest cost will be the purchase of all the necessary drugs, cells, and reagents needed to create and test a comprehensive treatment protocol. We recognize the initial work on 1) the spinal injuries, 2) the brainstem injuries, 3) the subcortical injuries, and 4) the cortical injuries will be resource intensive, but the far-reaching goal of reducing the cost of treatment protocols for everyone will be achieved with significant forethought and the algorithms embodied in the repair simulation and repair regimen sections.

1. De-identified imaging relevant to their research interest; 2. An integrated network of image analysis, volumetric analysis, histology atlases, and stereology experts, which will help them become the first to answer foundational questions in their field; 3. A curated, up-to-date knowledge base, which is both horizontal and vertical in its sophistication; 4. The opportunity to repurpose their current scientific skills around personalized neurorepair; and 5. The chance to put their most ambitious scientific ideas to the test in a life-sized, patient-specific cell culture model.

The more everyone succeeds, the faster we will reach our common goal of translational success.

The Orchestra will establish goals for each progressive scientific innovation cycle. Each scientific soloist will be supplied with the financial and technical support to answer their individual piece of the equation. The h.o.p.e.

Orchestra has no intellectual or patent claims on technology or ideas created by scientists, as a result of participation in this project, but must always be allowed future use of the emergent technology. All members are encouraged to publish, seek grants, and commercialize any and all ideas arising from this collaboration. Financial support is provided through non-venture philanthropic donations to sponsor the above sections, similar to any musical orchestra. We want every scientist to give us his or her best

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ideas and effort, and we will respect the lifetime of work and study they have invested in their ideas and expertise. For technology partnerships and the acquisition of cells and drugs, we will ensure negative results reflect the complexity of the endeavor and do not single out a specific drug or cell which will prevent any company from allowing us to use their therapeutics. The incentive for all involved in the project is to achieve cures for devastating injuries, and we must continuously manage all the other metrics or motivations with potential to mitigate our progress.

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The integration of each scientific domain will offer the deepest understanding of brain injuries in history. With an 'n' of 1, we will define a new standard of therapeutic design, as this regrowth strategy will address what is demanded by nature. We cannot choose to be interested in certain parts of the problem and ignore others – nor will we be. No bias: just a rigorous treatment strategy.

The approach taken by the h.o.p.e. Orchestra will also have practical benefits for scientists.

Scientists will be able to publish their individual part of the equation and apply for grant funding, as each calculation and experiment will be on a small enough scale to be publishable and reductionist enough for specific aims for potential grants, yet the entire enterprise will simultaneously be engaging in science on the scale of the human CNS masterpiece. Each will be a foundational paper as the first approach to personalized neural repair medicine in each respective field.

“Our world is a better place when everyone is afforded the opportunity to be a part of it.”

- Brooke Ellison PhD, MPP

HUMAN-SCALE. ORGAN-REGROWTH. PERSONALISED. EXPERIMENTS.

The h.o.p.e. Orchestra is a nonprofit research foundation focused on solving the puzzle of organ regrowth for individuals with neurological injuries and permanent disability. This institute will be the center for the above sections, both for having dedicated infrastructure to maintain the lab clinical trial section but also for being a global hub, as experts are consulted to answer specific questions. As this institute solves the problems for each anatomical region – 1) spine, 2) brainstem, 3) subcortical, and 4) cortical - the entire world will benefit.

THE H.O.P.E. ORCHESTRA FOUNDATION PROPOSAL

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THE ORCHESTRA

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HUMAN-SCALE. ORGAN-REGROWTH. PERSONALISED. EXPERIMENTS.

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ORCHESTRA OPERATIONS PER INNOVATION CYCLE

THE H.O.P.E. ORCHESTRA FOUNDATION PROPOSAL

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BUDGET PER INNOVATION CYCLE

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HUMAN-SCALE. ORGAN-REGROWTH. PERSONALISED. EXPERIMENTS.

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TIMELINE FOR INITIAL INNOVATION CYCLES

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SUMMARY The overwhelming challenges are why this journey is both exciting and necessary. The overwhelming need is why we believe this groundbreaking and visionary undertaking will attract the correct patients, patrons and scientific Orchestra members: the best and brightest in their fields, the boldest thinkers, and the most revolutionary innovators – those resilient to temporary failure, full of grit, inspired by complexity, generous of spirit and undaunted by challenge. This proposal is written to clarify the vision of neural regrowth and to inspire a team of thinkers who are thrilled at the notion of accomplishing something great. Each Orchestra section will advance its field dramatically while, in parallel, also building toward our ultimate accomplishment of making functional brain and spine repair possible.

THE H.O.P.E. ORCHESTRA FOUNDATION PROPOSAL

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The more everyone involved thrives and innovates, the closer we are to achieving our goal.

Although naysayers have been present to oppose every advance in our culture and technology since time immemorial, we have luckily always been blessed through the millennia with a minority of talented, inspired souls who make the seemingly impossible a reality. Ultimately, everything is impossible until someone does it.

This is an opportunity unlike any other because personalized neurorepair is a puzzle more enormous than any other sought to be solved.

But it can be done. Indeed, it must be done.

Our legacy will be to build the algorithms and tools necessary to solve this tragic puzzle for one person – and then for everyone.

Regrowth of the missing tissue of an injured human central nervous system will be possible, and the achievement of this goal will be an unprecedented gift to all of humanity and will change the world as we know it.

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The h.o.p.e. Orchestra

Justin Hill, M.D. Neurology, Neuroscience & Neurorehabilitation Founder - h.o.p.e. Orchestra jhill@hopeorchestra.org Brooke Ellison, PhD, MPP Ethics, Stem Cells, & Health Care Policy Stony Brook University bellison@hopeorchestra.org

David Putrino, PhD, PT Computational Neuroscience & Virtual Rehabilitation Mount Sinai Health Network dputrino@hopeorchestra.org IRINI VAZANELLIS Brand Architecture, Graphic Design & Storytelling Engineering, Simulation Arbory Workshop irini@arboryworkshop.com

www.hopeorchestra.org

HUMAN-SCALE. ORGAN-REGROWTH. PERSONALISED. EXPERIMENTS.

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Biomedical Engineering, Image Analysis & 3D Printing The City College of New York of CUNY mbikson@hopeorchestra.org

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Marom Bikson, PhD