NEURAL REGENERATION RESEARCH www.nrronline.org REVIEW Network-centric medicine for peripheral nerve injury: treating the whole to boost endogenous mechanisms of neuroprotection and regeneration David Romeo-Guitart, Caty Casas* Institut de Neurociències (INc) and Department of Cell Biology, Physiology and Immunology, Universitat Autònoma de Barcelona (UAB) & Centro de Investigación Biomédica en Red sobre Enfermedades Neurodegenerativas (CIBERNED), Bellaterra, Barcelona, Spain Funding: This work was mainly supported by the Ministerio de Economía y Competitividad of Spain (#SAF 2014-59701) (to CC). Downloaded from http://journals.lww.com/nrronline by BhDMf5ePHKav1zEoum1tQfN4a+kJLhEZgbsIHo4XMi0hCywCX1AWnYQp/IlQrHD3i3D0OdRyi7TvSFl4Cf3VC1y0abggQZXdgGj2MwlZLeI= on 03/22/2022 Abstract *Correspondence to: Caty Casas, PhD, Peripheral nerve injuries caused by accidents may lead to paralysis, sensory disturbances, anaesthesia, and Caty.Casas@uab.cat. lack of autonomic functions. Functional recovery after disconnection of the motoneuronal soma from target tissue with proximal rupture of axons is determined by several factors: motoneuronal soma viability, orcid: proper axonal sprouting across inhibitory zones and elongation toward specific muscle, effective synapse 0000-0002-2273-6321 contact rebuilding, and prevention of muscle atrophy. Therapies, such as adjuvant drugs with pleiotropic (Caty Casas) effects, that promote functional recovery after peripheral nerve injury are needed. Toward this aim, we de- signed a drug discovery workflow based on a network-centric molecular vision using unbiased proteomic doi: 10.4103/1673-5374.251187 data and neural artificial computational tools. Our focus is on boosting intrinsic capabilities of neurons for neuroprotection; this is in contrast to the common approach based on suppression of a pathobiological Received: March 12, 2018 pathway known to be associated with disease condition. Using our workflow, we discovered neuroheal, a Accepted: December 28, 2018 combination of two repurposed drugs that promotes motoneuronal soma neuroprotection, is anti-inflam- matory, enhances axonal regeneration after axotomy, and reduces muscle atrophy. This drug discovery workflow has thus yielded a therapy that is close to its clinical application. Key Words: motoneuron; neurodegeneration; axonal regeneration; muscle atrophy; systems biology; neuroheal; PI3K/AKT; sirtuin 1; motor function recovery Introduction et al., 2008; Castillo-Galván et al., 2014) and surgical repair Recent advances in systems-oriented approaches and ‘omics is the best therapeutical option depending on the PNI de- technologies have led to an assortment of new computation- gree. This, together with sick leave spending time, increases al tools that promise to enable a more informed and suc- substantially the economic and social burden being the cessful implementation of the drug discovery workflow than estimated annual cost in USA health care range from of the reductionist “one drug for one target for one disease” USD 5.13 Billion in 2016 and the compound annual growth approach (Westerhoff and Palsson, 2004). Computational rate of 12.3% (https://www.marketsandmarkets.com) . The tools for the analysis of biological data can help decipher the prognosis and functional recovery after PNI depends on mechanism or mechanisms that underlie a disease state or the severity of the injury, the gap distance between disrupt- the mechanism of action of a drug by offering a holistic view ed nerve stumps, the length of the nerve to be regenerated of the system. These tools enable identification of promising and the extend of target reconnection. Moreover, axonal pathways for modulation and selection of potential inhibi- integrity disruption or axonal retraction may be provoked tors (Fotis et al., 2018). In this review, we focus on the recent by non-traumatic disorders such as peripheral neuropathies discovery of neuroheal (NH) through a systems biology acquired during lifetime such as those induced by diabetes approach and the use of artificial intelligence-based tools. mellitus, antineoplastic agents or infiltrating tumours, or NH is neuroprotective and pro-regenerative and pre-clinical those relates with genetical disorders such as Charcot-Ma- data indicate that it holds promise as a co-adjuvant treat- rie-tooth or amyotrophic lateral sclerosis disease (England ment to facilitate recovery of nervous system function after and Asbury, 2004; Taylor et al., 2011; Laing, 2012; Rob- traumatic injury to the peripheral nerves. berecht and Philips, 2013; Ale et al., 2014). There is a medical need for function recovery after a dis- Injuries affecting the nerve roots, close to the interface ruption of the neuron-target connection resulting from between central and peripheral nervous systems, are the trauma-associated peripheral nerve injury (PNI) due to most devastating types of PNI. Nerve roots can be transect- traffic, laboral or sportive accidents. These injuries usually ed or separated by mechanical traction, which is commonly occur at working ages and its consequences range from mild termed root avulsion (RA). RA is often caused by high-ener- discomfort to life-long impairment accompanied of paral- gy traffic accidents (mainly motor vehicle) or sport-related ysis, sensory disturbances, anaesthesia, lack of autonomic incidents (e.g., American football), by iatrogenic injuries, functions in the affected body areas and neuropathic pain. and during obstetric interventions (Pondaag et al., 2004; PNI has a prevalence of 1.3–2.8% (Noble et al., 1998; Taylor Buitenhuis et al., 2012; Daly et al., 2016; Kachramanoglou 1122 Romeo-Guitart D, Casas C (2019) Network-centric medicine for peripheral nerve injury: treating the whole to boost endogenous mechanisms of neuroprotection and regeneration. Neural Regen Res 14(7):1122-1128. doi:10.4103/1673-5374.251187 et al., 2017). RA can affect single roots or nerve plexus from which process of cell death affects disconnected MNs. After brachial, lumbar, and facial zones (Htut et al., 2007). Adult axotomy, a neuronal retrograde response is initiated whose brachial plexus injury is present in 1.2% of the multi-trauma intensity and time course is influenced by maturational admission patients in Canada, and 450–500 supraclavicular stage and MN subset as well as by the site and type of axo- injuries with root affectation occur every year in UK (Goldie nal injury and the age and species of animal. In neonates, and Coates, 1992). Neonatal brachial plexus injury affects axotomized MNs undergo apoptosis as a result of the lack 2 or 3 of 1000 births (Pondaag et al., 2004). Currently, the of muscle-derived trophic supports (Snider et al., 1992). The only treatment for brachial plexus injury is surgical nerve aggressiveness of the process after RA is characterized by root reimplantation. The timing for surgical repairing is crit- a drastic reduction of MN survival, which results in death ical mainly due to the progressive motoneuronal soma (MN) of around 80% of MNs by 4 weeks post injury (Penas et al., death triggered after injury (Htut et al., 2007). 2009). Models obtained either through sciatic root or facial Complete motor functional recovery after PNI, and par- nerve avulsion result in similar losses of MNs but with some ticularly after RA, requires five main steps: i) sustained sur- mechanistic differences (Watabe et al., 2005). vival of axotomized MNs; ii) promotion of axonal regrowth We have recently shed light on the mechanisms of MN across the glial scar formed at the ventral-root intermediate death after RA in adult rats using unbiased comparative pro- zone; iii) axonal elongation through the degenerated distal teomic analysis combined with a systems biology approach nerve stump to the target muscle; iv) establishment of func- for data mining (Casas et al., 2015). Our study showed that tional connections to denervated muscle, and v) prevention several programs, such as apoptosis and anti-apoptosis path- of muscle atrophy. Whether or not these steps occur de- ways, run in parallel and counterbalance to result in an over- pends on factors such as the patient’s age, the type of lesion, all non-apoptotic degenerative process (Li et al., 1998; Penas the distance from the injury to the target organ, and the et al., 2011a) and anoikis. In contrast, when nerve avulsion timing and type of surgical repair (Navarro et al., 2007). We is milder, not performed at the root but distally, the process have performed a PubMed literature search of articles pub- may shift towards an apoptotic process, which upon sciatic lished with the search term “neuroheal”, “nerve root avul- nerve avulsion leads rapid loss of about 50% of MNs by two sion”, and “peripheral nerve injury”. weeks post injury ((Martin et al., 1999, 2005) and personal observations). These observations reveal that to be clinically Experimental Models to Study Peripheral effective a neuroprotective agent should be plastic enough to Nerve Lesions protect against both forms of neuronal death. Successful discovery of drugs that can be used as adjuvants Although these models of RA allow study of the mecha- nisms of degeneration and target discovery, more sophisti- to surgical intervention after PNI requires first the existence cated models have been also characterized that better mimic of good models. In vivo models are widely used to mimic the clinical issues after surgical repair. Some authors have per- PNI and have been used to identify factors that modulate formed RA of ventral or brachial roots combined with an axonal regeneration and MN death. MN survival depends immediate reparative surgery (Eggers et al., 2010; Pintér et on the severity and type of the injury, the distance from the al., 2010; Torres-Espín et al., 2013). However, immediate neuronal soma, and the age of the animals. Rat spinal MNs repair cannot be carried out clinically because, among other are extremely susceptible to the transection of the nerve reasons, patients need to be stabilized after the accident. Gu by avulsion from the spinal cord affecting the roots and is et al. (2005) were the first to evaluate delayed repair of the defined as preganglionic (proximal to the dorsal root gan- avulsed roots, and recently we set up a new in vivo rat model glia). Usually affecting roots from brachial or lumbosacral that combines RA of L3–6 ventral roots with delayed surgical plexus (Penas et al., 2009). But adult MNs are resistant to repair (Romeo-Guitart et al., 2017). This model can be used postganglionic transection (Vanden Noven et al., 1993; to test new therapies against MN degeneration in the con- Valero-Cabre et al., 2004) with the exception of immature text of post-injury axonal regrowth. Importantly, this model MNs still in development (Snider et al., 1992; Lowrie et al., reproduces long-term denervated muscle atrophy, which is 1994; Sun and Oppenheim, 2003). There are also differences essential for functional recovery (Romeo-Guitart et al., 2017). among organisms since, for instance, cranial hypoglossal MNs are susceptible to the transection of the hypoglossal nerve at distal level in mice but not in rats (Kiryu-Seo et al., Network-Centric Discovery of a New 2005). Models based on nerve crush produced by compres- Neuroprotective Agent: Neuroheal sion that lead to the axonal continuity breakdown (axonot- Many observations indicate the existence of multiple en- mesis) allow a reasonably good recovery, whereas those in dogenous mechanisms of neuroprotection. Endogenous which neurotmesis is produced by complete transection of defence of the nervous system is a continuous process that the nerve leads to a poor functional recovery of the animals, simultaneously performs and integrates the neurobiological although the latter model better recapitulates the clinical processes of tropism, neuroprotection, neuroplasticity, and features of PNI (Lago and Navarro, 2006). neurogenesis. Clinical activation of endogenous protective The rational design of neuroprotective agents is often strategies dates back more than six decades when Noble and based on knowledge of the underlying mechanisms of neu- collaborators observed that short periods of global hypoxia rodegeneration. In this regard, it is important to determine can protect the entire mammalian organism and preserve 1123 Romeo-Guitart D, Casas C (2019) Network-centric medicine for peripheral nerve injury: treating the whole to boost endogenous mechanisms of neuroprotection and regeneration. Neural Regen Res 14(7):1122-1128. doi:10.4103/1673-5374.251187 brain energy metabolism when re-exposed to longer periods bases as templates to build associations between them. Re- of hypoxia (Noble, 1943). These early studies were the foun- cently developed computational tools allow unique virtual dation for subsequent intensive research on the mechanisms screening strategies for polypharmacology and prediction involved in this type of protective pre-conditioning. How- of adverse and therapeutic polypharmacological effects and ever, drug discovery efforts have focused on identification of supervised machine-learning algorithms that enable predic- agents that suppress pathobiological processes rather than tion of drug-target interactions (Lavecchia, 2015) have revo- on discovery of drugs that enhance intrinsic neuroprotective lutionized the workflow for drug discovery. Network models pathways. This reductionist vision has not led to many suc- enable analysis of complex interaction networks between cessful therapies for neurodegeneration. drugs and their targets aiding in elucidation of mechanisms To identify a neuroprotective agent capable of boosting of action (Cheng et al., 2012; Woo et al., 2015). Statistical the endogenous mechanisms normally triggered after trau- models that make use of a data integration framework have ma, we took advantage of the existence of solid molecular been effectively applied both to predict novel drug indica- knowledge on the mechanisms triggered after trauma and tions and to identify potent drug combination strategies computational tools for handling and extracting meaning (Keiser et al., 2007). Computational/experimental frame- from biological data. We realized that dissimilar reactions works have also been implemented for the systematic design occur in MNs after proximal and distal axotomy that lead of multi-targeted drugs (Bottegoni et al., 2012). to opposite readouts: MN degeneration vs. MN survival, In the discovery of NH, we used the technology known as respectively. We wondered which mechanisms triggered the therapeutic performance mapping system, which allows by distally axotomized MNs allowed them to recover from generation of disease-specific mathematical models for drug damage and even regenerate. We envisaged that mimicking screening using neural artificial intelligence-based methods these endogenous mechanisms might be neuroprotective for for machine learning (Pujol et al., 2010; Herrando-Grabulo- proximal axotomized degenerating MNs as well. In support sa et al., 2016; Romeo-Guitart et al., 2017, 2018). Therapeu- of this hypothesis, we observed that although similar mo- tic performance mapping system is based on the building of lecular mechanisms were initially triggered by both distal the molecular networks characteristic of an specific health and proximal RA axotomy, there are differences in extent condition (either normal or pathological) using network and of neurodegeneration at seven days post injury (Penas et al., systems biology approaches (Pujol et al., 2010). Therapeutic 2011a; Casas et al., 2015). Hence, we had identified the pre- performance mapping system facilitates the screening of cise time to investigate and dissect out pro-survival mecha- drugs for their capacity to shift the profile of topological mo- nisms from pro-degenerative ones. We used proteomics and lecular maps from pathological to beneficial or normal (Her- a systems biology approach to perform unbiased analysis of rando-Grabulosa et al., 2016). For this purpose, maps are both models. We discovered that MN degeneration after RA converted into mathematical models using artificial artificial was characterized by concurrent programs such as apoptosis neural networks and sampling methods as computational and anti-apoptosis, anoikis and cytoskeletal rearrangements, tools, and machine learning allow the screening and choice among others, and that distally axotomized MNs survive be- of thousands of drug combinations with a desirable effect cause these MNs differently engage the anti-anoikis program (e.g., neuroprotection) (Badiola et al., 2013). In our work, we and selective autophagy (Casas et al., 2015). Although not ful- sought to identify a combination of clinically used drugs that ly understood, this divergence in the molecular mechanisms would be capable of converting the degeneration-associated between MN degeneration or regeneration may be due to the map that describes the RA condition into the neuroprotec- proximity of the injury to the soma. This provokes a more tive and regenerative-associated map described by the distal severe disturbance of the cell organelles after preganglionic axotomy condition. Our therapeutic performance mapping injury and a harmful response to stress which leads to the op- system analysis resulted in a list of putative neuroprotective posite molecular responses (Penas et al., 2011b). combinations of US Food and Drug Administration-ap- These descriptions were not enough to identify specific proved drugs. Thus, computational tools allowed us to focus targets for a neuroprotective therapeutical strategy. We did on the network and to select pharmaceutical combinations not want to implement the reductionist vision – one drug putatively capable of boosting endogenous neuroprotective for one target for one disease – approach. Instead, we want- mechanisms. Experimental investigations led us to the com- ed to take advantage of breakthroughs in systems biology bination now known as NH. and pharmacology approaches as well as in computational We validated that NH increases MN survival after RA, re- tools for big data analysis to design a network-centric-based duces the inflammatory response, protects against endoplas- drug discovery process. Network analysis can be used to mic reticulum stress in vitro, and induces pro-regenerative elucidate the intricate connections that form the biological profiles in MN tracks in spinal cords; the NH components systems and how they are disturbed in pathological condi- act synergistically (Romeo-Guitart et al., 2018). In addition, tions (Barabási, 2007). The increasing amount of biological we demonstrated that NH treatment accelerates long-term data from a variety of ‘omics platforms (genomics, tran- motor function recovery after PNI and root reimplantation scriptomics, proteomics, and metabolomics) have assisted in (Romeo-Guitart et al., 2017). the development of various in silico approaches that connect It is important to highlight that NH is a combination of molecular data to their biological functions using knowledge two repurposed drugs: acamprosate and ribavirin. Drug re- 1124 Romeo-Guitart D, Casas C (2019) Network-centric medicine for peripheral nerve injury: treating the whole to boost endogenous mechanisms of neuroprotection and regeneration. Neural Regen Res 14(7):1122-1128. doi:10.4103/1673-5374.251187 purposing facilitates clinical translation because it drastically using an adeno-associated viral vector increased MN survival reduces the need for preclinical toxicity and efficacy assays after RA. Although SIRT1 activation is essential for NH neu- and for safety profiling in human volunteers (Ashburn and roprotection, SIRT1 activation alone is not responsible for the Thor, 2004). Some successful examples of drug repurposing beneficial spectrum of neuroprotection that NH exerts reveal- include a glycogen synthase kinase- 3β-targeted therapy for ing the importance of its multi-target nature (Romeo-Guitart the treatment of glioblastoma (Furuta et al., 2017) and the et al., 2018). Moreover, NH normalized the location or abun- promising combination of two anti-diabetic drugs that exert dance of cytoskeletal motor proteins KIF5C and DCNT1, two neuroprotection in a mouse model of Alzheimer´s disease transport proteins related with neurodegeneration or axonal and in patients with Parkinson disease (Aviles-Olmos et al., disturbances. The kinesin family is formed by the kinesin-1 2013; Hettich et al., 2014). (historically named kif5c) and the kinesin-3 (KIF1A, KIF- 1Bα and KIF1Bβ) members (De Vos and Hafezparast, 2017). Neuroheal’s Mechanism of Action in KIF5C is enriched in MNs (Kanai et al., 2000) and its genetic Neuroprotection and Nerve Regeneration ablation or spontaneous mutation are linked with MN dis- As evidenced by the increasing number of publications in eases and paralysis (Xia et al., 2003; De Vos and Hafezparast, the recent years, drug discovery is undergoing a steady para- 2017). Besides, a dysfunctional dynein/DCTN1 has been used digm shift from the ideology of selective agents (‘magic bul- as an ALS mice model. Its mutation causes a defective axonal lets’) toward selectively promiscuous multi-targeting drugs transport that leads to an amyotrophic lateral sclerosis-like (‘magic shotguns’) (Roth et al., 2004; Strebhardt and Ullrich, phenotype in mice and to MN degeneration (LaMonte et al., 2008). The polypharmacology-based “one drug, multiple 2002; Heiman-Patterson et al., 2015). Finally, the increase on targets” paradigm has benefits relative to the conventional β-1 integrin can be yielding anti-anoikis effects to the MNs. de novo approaches to drug discovery (DiMasi, 2001; Dick- In fact, the concomitant activation of AKT axis with an in- son and Gagnon, 2004). In the case of nerve regeneration, crease of β-1 integrin may be inhibiting anoikis in our model multi-targeting gives the opportunity to act through several (Bouchard et al., 2007). pathways in parallel to support the neuroprotective and Although maintenance of MNs can be achieved after RA pro-regenerative phenotype. Analysis with computational experimentally by different pharmacological treatments, tools allowed us to identify putative proteins (nodes) that translation to the human healthcare has never been per- mediate the synergistic mechanism of action of NH. Im- formed. For example, the Pre084 compound, an agonist portant nodes belong to the phosphatidylinositol-4,5-bis- of the sigma receptor 1, is a good neuroprotectant but use phosphate 3-kinase (PI3K)/protein kinase B (AKT) axis and systemically causes neuropathic pain, precluding clinical anti-apoptotic pathways (p53, the apoptosis blocker X-linked application (Penas et al., 2011b; Roh et al., 2011; Pyun et al., inhibitor of apoptosis protein, cytoskeletal motor protein 2014). Other compounds such as N-acetyl-cysteine, GM-1 kinesin heavy chain isoform 5 (KIF5), and dynactin subunit ganglioside, and valproic acid maintain MN pool, but their 1 (DCNT1)), and the Src-integrin axis (the integrin subunit ability to enhance nerve regeneration has not been reported integrin beta-1 and the nicotinamide adenine dinucleo- (Oliveira and Langone, 2000; Zhang et al., 2005; Wu et al., tide-dependent deacetylase sirtuin 1 (SIRT1)) (Romeo-Gui- 2013). Riluzole, lithium, and an inhibitor of the proteogly- tart et al., 2018) (Figure 1). can receptor are neuroprotective in a model of RA with the We have shown that activation of SIRT1 mediated by NH immediate reimplantation of roots (Bergerot et al., 2004; Li promotes neuroprotection (Romeo-Guitart et al., 2017, 2018). et al., 2015; Fang et al., 2016). These compounds promote It has been extensively reported that activation of SIRT1 can long-term survival of MN, as does NH, after injury, but in exert neuroprotection in different neurodegenerative diseases tests of these compounds, reparative surgery was performed such as amyotrophic lateral sclerosis, Alzheimer and Hun- immediately after RA, so it is unclear if they also protect MNs tington diseases and in experimental autoimmune encepha- after a delayed root reimplantation. We showed that NH sus- lomyelitis induced by pharmacological or genetic activation tains MN survival for two weeks after RA in the absence of (Kim et al., 2007; Jiang et al., 2012; Nimmagadda et al., 2013; reparative surgery (Romeo-Guitart et al., 2017). Watanabe et al., 2014). In the context of trauma, SIRT1 acti- Although mammalian neurons have intrinsic capabilities to vation reduces functional impairments after spinal cord in- replace and repair axons by regeneration, in the best clinical jury (Chen et al., 2017). Although some reports indicate that scenario only 10% of axons regenerate to target organs after SIRT1 activation has beneficial effects and improves function- nerve transection (Janjic and Gorantla, 2017). The formation al recovery after traumatic brain injury (Wu et al., 2007), its of the inhibitory glial scar at the central nervous system-pe- role after stroke remains controversial (Petegnief and Planas, ripheral nervous system transition zone after root injury 2013). Mice that overexpress SIRT1 have ameliorated amy- make more difficult axon regrowth in this case than in distal otrophic lateral sclerosis pathophysiology (Watanabe et al., axotomy (Silver and Miller, 2004; Silver et al., 2014). Different 2014). We found that co-treatment of the RA-injured animals molecular pathways have been reported to mediate axonal re- with NH and the selective SIRT1-activity inhibitor Ex-527 growth within central nervous system and peripheral nervous blocked NH pro-survival effects (Romeo-Guitart et al., 2017, system (Tedeschi, 2012). Among them, the PI3K/AKT axis, 2018). Indeed, the overexpression of SIRT1 in spinal MNs which is activated by neurotrophic factors, is the best charac- 1125 Romeo-Guitart D, Casas C (2019) Network-centric medicine for peripheral nerve injury: treating the whole to boost endogenous mechanisms of neuroprotection and regeneration. Neural Regen Res 14(7):1122-1128. doi:10.4103/1673-5374.251187 mediated by neurotrophic factors (Tannemaat et al., 2008). Some compounds, including riluzole, lithium, and an inhib- itor of the proteoglycan receptor, have been demonstrated to increase axonal regeneration after RA with immediate root reimplantation (Bergerot et al., 2004; Li et al., 2015; Fang et al., 2016). However, in this model is difficult to distinguish whether functional recovery is mediated by an increase in MNs survival or by a specific effect on axonal regeneration. NH, as a combination of two repurposed drugs, has clear ad- vantages in terms of clinical translation. After traumatic injuries to the nervous system, such as PNI, neurodegenerative diseases, or intensive health-care periods, muscle is considerably atrophied (Powers et al., 2016). Skeletal muscle atrophy is correlated with a high mortality and morbidity risk (Calvani et al., 2013). The im- portance of muscle in recovery after PNI has recently been Figure 1 Neuroheal exerts neuroprotective and regenerative effects after peripheral nerve injury. acknowledged. Recovery depends on efficient reconnection Left, String protein network representation of minimal mechanism of action of axons with neuromuscular junctions (NMJs) from mus- of NeuroHeal. In red are highlighted key nodes experimentally proven to be cle, and this muscle must be minimally atrophied. We have modulated by neuroheal: an increased activity of nicotinamide adenine dinu- cleotide-dependent deacetylase sirtuin 1 (SIRT1) and serine/threonine-pro- observed that NH treatment reduces muscle-denervated tein kinase protein kinase B (AKT), and an augmented presence of integrin atrophy and increases the number of functional NMJs (Fig- subunit integrin beta-1 (ITGB1), kinesin-related protein KIF5 and dynactin subunit 1 (DCNT1). Right, pleiotropic effects promoted by neuroheal at ure 1). Hence, NH treatment shows clear positive combined different levels: the spinal cord motoneuronal soma (MNs) (blue), astrocytes effects on neuroprotection and axonal regeneration, and its (yellow), and microglia (green), the motor nerve at the peripheral nerve sys- activity shows that mimicking endogenous mechanisms can tem (blue line), and the muscle (orange). NMJ: Neuromuscular junctions. lead to effective therapeutic designs. terized (Caporali et al., 2008). PI3K is very sensitive to PTEN Conclusion and Summary (gene of phosphate and tension homology deleted on chrom- An optimal therapeutic strategy after PNI must sustain some ten), which blocks PI3K and reduces the pro-regenera- long-lasting neuronal viability, enhance axonal regrowth, tive effect of the axis (Berry et al., 2016). Several approaches, favour functional reinnervation of NMJs, and reduce muscle including PTEN deletion (Park et al., 2008) and its pharma- atrophy. Covering all these aspects by influencing a single tar- cological inhibition, enhance axonal regrowth through the get is likely impossible, and, therefore, multi-targeting drugs glial scar after injury to the central nervous system (Ohtake et are desired. The nervous system, like other systems in the al., 2014). However, the suppression of PTEN activity cannot body, has the availability to engage endogenous mechanisms be translated to the clinic due to its tumour-suppressor role in the face of injury that allow neurons to survive, axons to (Tolkacheva and Chan, 2000). Thus, discovering new drug speedily regenerate, and healthy denervated muscle to be therapies that increase axonal regrowth by AKT activation sustained. We reasoned that potentiating these mechanisms without affecting PTEN has raised much interest (Tolkacheva might result in effective neuroprotection and repair of the and Chan, 2000). We demonstrated that NH reduces glial damaged nervous system. NH was discovered using a systems scar formation and enhances axonal regrowth after distal or biology and neural artificial intelligence-based computation- proximal injury (Romeo-Guitart et al., 2017, 2018). Accelera- al tools in a network-centric vision for drug discovery. In tion of nerve regeneration resulting from NH treatment is ac- animal models, NH boosts endogenous mechanisms and its companied by an increase in compound muscle action poten- multi-targeting nature activates multiple factors necessary for tial at the muscles innervated by the L4–6 MNs. NH probably functional recovery after PNI, resulting in neuronal survival, enhances nerve regeneration because it increases PI3K/AKT axonal regrowth, and limited denervated muscle atrophy. axis activity and growth associated protein 43 expression, a Author contributions: First draft of the manuscript: DRG; study guiding, pro-regenerative factor also called neuromodulin (Denny, critical revision and final draft of the manuscript: CC. 2006; Holahan, 2017). Conflicts of interest: The authors declare no competing interests. Neuroheal is currently under patent review. Very few drugs that enhance axonal regeneration after PNI Financial support: This work was mainly supported by the Ministerio de have been described in the literature (Al-Majed et al., 2000; Economía y Competitividad of Spain (#SAF 2014-59701) (to CC). Elzinga et al., 2015). These are FK506, geldanamycin, N-ace- Copyright license agreement: The Copyright License Agreement has been signed by both authors before publication. tylcysteine, acetyl-L-carnitine, IGF, chondroitinase ABC, Plagiarism check: Checked twice by iThenticate. and neurotrophic factors (Navarro et al., 2001; Welin et al., Peer review: Externally peer reviewed. 2009; Sun et al., 2012; Farahpour and Ghayour, 2014; Janjic Open access statement: This is an open access journal, and articles are distributed under the terms of the Creative Commons Attribution-NonCom- and Gorantla, 2017). All have undesired side effects such as mercial-ShareAlike 4.0 License, which allows others to remix, tweak, and immunosuppression in the case of FK-506 and tumorigenesis build upon the work non-commercially, as long as appropriate credit is given 1126 Romeo-Guitart D, Casas C (2019) Network-centric medicine for peripheral nerve injury: treating the whole to boost endogenous mechanisms of neuroprotection and regeneration. Neural Regen Res 14(7):1122-1128. doi:10.4103/1673-5374.251187 and the new creations are licensed under the identical terms. Elzinga K, Tyreman N, Ladak A, Savaryn B, Olson J, Gordon T (2015) Brief Open peer reviewers: Daoli Zhu, Nantong University, China; Shanji Nan, electrical stimulation improves nerve regeneration after delayed repair in The Second Hospital of Jilin University, China. Sprague Dawley rats. Exp Neurol 269:142-153. Additional file: Open peer review reports 1 and 2. England JD, Asbury AK (2004) Peripheral neuropathy. Lancet 363:2151- 2161. 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