To watch the companion video to this series, click here.
To read the previous article in this series, click here.
THE PROBLEMS WITH EPO
While EPO exerts desirable effects on the brain, there are two problems with EPO treatment. First, the full EPO molecule crosses the blood-brain barrier poorly due to its size, necessitating high systemic dosages to treat the brain. Second, EPO’s erythropoietic and angiogenic effects have dangerous implications for oncogenesis and cardiovascular health, particularly at the high systemic dosages needed to produce an effect on the brain.
The major concern with EPO treatment is the risk for tumor progression and metastasis[1][2], likely due to its antagonistic effect on TNF-a, through its effect on STAT5, and through angiogenesis. First, though TNF-a’s role in tumorigenesis is bidirectional and beyond the scope of this article series[3], it can certainly inhibit the growth of tumors. TNF-a has been shown to suppress erythropoiesis, while EPO administration suppresses TNF-a[4]. Second, much of the effects of EPO exerted via EPOR are dependent on activation of the STAT5 pathway, and STAT5 has been considered an oncogene because STAT5 proteins are overexpressed in cancers and promote their survival[5]. Third, agonism of the EPOR increases expression of vascular endothelial growth factor (VEGF), which in turn promotes tumorigenesis[6] via angiogenesis[7]. It has also been documented that EPORs are expressed by tumors[8][9], though EPO may not be growing tumors preferentially, as it exhibits angiogenesis of both tumors and surrounding tissue[10][11].
In addition to EPO’s oncogenic nature there are cardiovascular concerns, including the serious potential for venous thrombosis[12][13] and hypertension[14]. When administering EPO for its protective effect at high doses, the procoagulant and hemodynamic effects become potentially life-threatening for patients who produce excess inflammatory cytokines.
Consequent to these serious concerns, the FDA has mandated a warning that recombinant EPO and darbepoetin treatment is associated with ‘increased mortality, serious cardiovascular and thromboembolic events, and tumor progression.’
Remember that splice variants of EPO that protect neurons from apoptosis are found naturally in the human body, including EV-3, the exon 3 deletion variant, which lacks erythropoietic activity[15]. Inspired by this observation, academics have produced eleven analogues and derivatives of the EPO molecule that lack an erythropoietic effect.
NEUROEPO
It was demonstrated that EPO produced in the brain and retina differ from EPO produced in the kidney, as the former lacks sialic acid. When the sialic acid level of EPO is between 4-7 mol/mol protein, the resultant molecule, called NeuroEPO, resembles mammalian brain EPO, which is smaller and more active in vitro at low ligand concentrations[16]. When sialic acid is removed, the resulting molecule is rapidly degraded by the liver, limiting the viability of systemic delivery[17].
Intranasal delivery of NeuroEPO has been determined to be as efficacious as EPO in gerbils[18] and rats[19], with a therapeutic window of up to 12 hours and a lower dosing requirement than systemic EPO. In gerbils and monkeys, intranasal administration of NeuroEPO reaches the brain 5 minutes after dosing[20]. Intranasal NeuroEPO has improved neurobehavioral outcomes in rodent models of cerebral ischemia[21] without producing a hematopoietic effect[22], likely due to its low sialic acid content[23].
Intranasal NeuroEPO also lowered oxidative stress, inflammatory cytokines (including TNF-a), reduced amyloid b deposits and attenuated memory impairment in a transgenic mouse model of Alzheimer’s disease[24]. Earlier, in a non-transgenic mouse model of Alzheimer’s disease, both recombinant EPO and NeuroEPO also prevented amyloid b induced learning deficits, reduced inflammatory cytokines IL-1b and TNF-a, and prevented amyloid b-induced cell loss[25]. NeuroEPO has also been shown to be neuroprotective from glutamate-induced excitotoxicity[26], likely mediated by the anti-apoptotic protein BCL-2’s effect on oxidative damage that results from the excitotoxicity[27].
Intranasal NeuroEPO has been suggested as a therapeutic method for the treatment of stroke in humans[28][29]. In 2017, a randomized trial of up to 1 mg of NeuroEPO delivered intranasally to humans determined that it was well-tolerated and did not stimulate erythropoiesis[30].
EPOL
In 2018, researchers tested the tissue protective effect of another low sialic acid naturally occurring EPO variant, termed EPOL, that is also devoid of hematopoietic effect due to its short half-life. EPOL was originally discovered in the mammary gland of animals[31], and it was shown in vitro to provide a greater neuroprotective effect against oxidative stress than EPO, through its agonism of EPOR, which had a resulting effect on JAK/STAT intracellular signaling and an upregulation of the BCL2 gene[32].
AsialoEPO
In 2003, Japanese researchers removed sialic acid from EPO to develop the derivative AsialoEPO, which is neuroprotective without a hematopoietic effect[33], though it is still angiogenic[34]. It is thought that AsialoEPO does not promote hematopoiesis because of its short half-life, which is measured in minutes, though it has an even greater affinity at the EPOR than rEPO. AsialoEPO has been shown to protect animals from cerebral ischemia, spinal cord injury, and sciatic nerve trauma[35], and it has exhibited as protective an effect as EPO on hypoxic ischemia injury in 7-day old rats[36].
Interestingly, its neuroprotective effects lasted past a 5-minute exposure, showing that the long duration of rEPO was unnecessary for the neuroprotective benefits it produced[37]. Brines et al. have confirmed that pre-treatment with AsialoEPO before injury did not provide additional benefit for neuroprotection, nor did subsequent treatments – a single dose post-trauma was sufficient to yield a neuroprotective response. Because of its short half-life, it has proved ineffective in neurodegenerative models with repeated trauma, such as Huntington’s disease[38], though a two-week treatment in rodents promoted oligodendrogenesis – the development of myelin-forming oligodendrocytes[39]. Japanese researchers have patented the molecule heavily[40].
CARBAMYLATED EPO
To increase the half-life of EPO without the hematopoietic effect and to investigate whether agonism of the EPOR was necessary for its tissue-protective effect, in 2004 researchers produced carbamylated EPO (CEPO).[41]. In this method, lysine residues are carbamylated, which causes all lysines to be replaced by L-homocitrulline, leaving a molecule with a longer half-life and less affinity for the EPOR. Consequently, CEPO produced no hematopoiesis, did not produce a pro-coagulant effect, did not produce an angiogenic effect, did not produce a hypertensive effect, and did not stimulate platelet production[42], thereby indicating a potential use for sustained treatment.
CEPO was the first EPO variant devoid of an angiogenic effect, potentially diminishing the oncogenic effect. CEBO does not appear to act on EPOR homomeric units. Instead, it is thought that the carbamylated EPO exerts a neuroprotective effect by acting on heteromers that contain an EPOR and a beta common receptor (bCR)[43], presumably by which it exhibits its proliferation of adult neural progenitor cells[44]. It has been shown to influence the expression of 66 genes, many of which are strongly associated with neurotrophic processes[45].
CEPO has been shown to improve recovery after cerebral ischemia in rodents[46], to attenuate radiation induced injury[47], to attenuate NMDA-induced excitotoxicity[48], to enhance spatial learning and neuroplasticity as effectively as EPO in a rodent model of traumatic brain injury[49], to provide neuroprotection to rodents following spinal cord hemisection that is equivalent to EPO[50], to reduce depressive symptoms in rodents[51], and to reduce neurological impairment due to middle cerebral artery occlusion as well as EPO[52]. Interestingly, the tissue protective effect of CEPO was observed to occur even with delayed administration after trauma of up to 24 hours[53]. CEPO was also patented in 2006[54].
HBP, HBSP, and pHBSP
In an effort to determine which part of the CEPO molecule was tissue-protective, in 2008 researchers produced helix B 25-mer peptide (HBP) from the helix B (amino acids 58-82) that faces into the aqueous environment and away from the cell membrane in the CEPO molecule. HBP was shown to be tissue protective but not erythropoietic, reducing infarct volume by 30% in a rodent model of stroke and attenuating the development of retinal edema in a rodent model of diabetes[55].
In a further effort to identify the tissue-protective element of CEPO, the same researchers synthesized the spatially adjacent amino acids of the aqueous surface of helix B into a linear peptide, named helix B surface peptide (HBSP). Due to spontaneous cyclization of the N-terminal glutamine of HBSP, pyroglutamate HBSP (pHBSP, also called ARA 290) was produced with the same tissue-protective potency as HBSP.
Due to the size of the peptides, HBSP and pHBSP have a similar half-life as AsialoEPO, and yet they have exhibit wide-ranging neuroprotective and tissue-protective effects. The peptides have been shown to improve memory in rats after traumatic brain injury[56][57] and to be protective of insulin sensitivity[58][59], against diabetic retinopathy[60] and neuropathy[61], against ischemic damage due to myocardial infarction[62] and cardiomyopathy subsequent to it[63], of renal function following injury[64], to improve the pathology of emphysema[65], and the damage sustained from burns[66].
pHBSP is being studied for its effect on neuropathy involving chronic pain[67], likely exerted via its antagonism of the capsaicin receptor[68]. (Readers will recall that cannabidiol’s analgesic effect also involves antagonism of the capsaicin receptor). The compound may also have cardioprotective effects for the aging population, as aging rodent studies indicate[69]. Interestingly, short-term pHBSP administration has been shown to alter human emotional processing in a double-blind, randomized study[70]. Neither HBSP nor pHBSP appear to be patented in the US.
EPOTRIS, EPOBIS, and NL100
In 2010, a group of European researchers built a peptide from the helix C motif of EPO, named Epotris. Epotris was shown to bind to the EPOR binding site 2 with about 10x less affinity than EPO (4.5 nM), potentially only partially agonizing it. It was shown to grow neurites and protect neurons from cell death without producing erythropoiesis when injected systemically[71]. It was determined through blocking the EPOR that both the observed neuritogenic effect and neuroprotective effect of Epotris were dependent on activation of the receptor. Recall that EPOR signaling involves a Jak2-Stat5 pathway[72], and in blocking Jak2 and Stat5 independently, researchers further confirmed that the neuritogenic effect was mediated by the Jak2-Stat5 signaling pathway. Epotris was also shown to decrease neurodegeneration, attenuate seizures, and triple the survival rate in a rodent model of kainic acid-induced brain injury.
In 2012, the developers of Epotris produced a second peptide, Epobis, derived of an 18-mer sequence motif from the C-terminal of the AB loop of EPO. The peptide Epobis was found to bind to the EPOR at binding site 1 (with a KD of about 60 nM,) with 60x less potency than EPO, producing no hematopoietic effect. It was shown to induce dose-dependent neurite growth mediated by activation of the EPOR that induced Stat5 activation in hippocampal neurons and to protect neurons in a kainic acid-induced model of brain injury[73]. Moreover, Epobis was shown to reduce TNF-a production similarly to EPO, to improve long-term but not acute working memory in rodents, and to cross the blood-brain barrier when delivered systemically (at a ratio of 20:1 in plasma and CSF)[74].
In 2019[75], the same team sought to produce a smaller EPO-mimetic peptide because smaller particles can more readily cross the blood-brain barrier[76]. The result, NL100, is 12-mer and as such has 3x lower molecular weight than Epotris. It is derived of the C-terminal side of the EPO helix C, including four of the sixteen amino acids involved in the second, low-affinity binding site of EPO with the EPOR. NL100 bound to the EPOR with a KD of 4.2 nM, crossed the blood-brain barrier, and improved working memory, long-term memory, and social recognition deficits in an animal model of amyloid b induced toxicity without inducing hematopoiesis. The authors speculated that the low affinity of NL100 for the EPOR may limit its cognitive effects, despite its ready crossing of the blood-brain barrier. Epotris, Epobis, and NL100 do not appear to be patented in the US.
JM-4
In 2015[77], a third team of researchers developed the non-hematopoietic 19-mer peptide JM-4 from the EPO AB loop with 2 cysteine molecules. JM-4 originally displayed immunomodulatory, neuroprotective, and tissue-protective effects in an animal model of multiple sclerosis, including against demyelination and axonal damage. JM-4 has been shown to attenuate damage from traumatic brain injury[78] and delay neurodegeneration due to taupathy[79] (a hallmark of Alzheimer’s disease). Between 2016 to 2018, the researchers developed at least four patents on JM-4, including its use in inflammatory diseases[80], to attenuate damage from traumatic brain injury[81], and to treat neurodegenerative diseases[82].
GENETIC ALTERATIONS: EPO MUTANTS
In 2012, researchers in Tennessee developed two new mutants of the EPO gene devoid of erythropoietic effects and delivered them via virus to mice[83]. EpoR76E was produced by converting an arginine to a glutamate at position 76 and EpoS71E was produced by converting a serine to a glutamate at position 71, both based from an EPO molecule taken from a rhesus monkey.
The two molecules were studied in an MPTP model of Parkinson’s in which the neurotoxin MPTP destroyed over 40% of dopaminergic neurons in control mice, while an intramuscular injection of EpoR76E delivered 3 weeks before the neurotoxin reduced dopaminergic neuron damage by half and fully protected the mice from neurodegenerative-induced tremor. EpoR71E failed to protect dopaminergic neurons from the neurotoxin in the model, though it improved TH-positive fibers on surviving neurons 5 weeks after the experiment and inhibited Parkinsonian tremor. The authors speculated that neuroprotection may have partially been due to EpoR76E’s modulation of the inflammatory cytokine response that occurs with MPTP delivery[84]. Interestingly, this team was the first to posit that EPO exerted an axonal regeneration effect that is independent of both its neuroprotection and erythropoiesis (in the case of EpoR71E).
To read the fifth article in the series, click here.
[1] Bohlius, J., Wilson, J., Seidenfeld, J., Piper, M., Schwarzer, G., Sandercock, J., ... & Djulbegovic, B. (2006). Erythropoietin or darbepoetin for patients with cancer. Cochrane Database of Systematic Reviews, (3). [2] Henke, M., Laszig, R., Rübe, C., Schäfer, U., Haase, K. D., Schilcher, B., ... & Frommhold, H. (2003). Erythropoietin to treat head and neck cancer patients with anaemia undergoing radiotherapy: randomised, double-blind, placebo-controlled trial. The Lancet, 362(9392), 1255-1260. [3] Mocellin, S., & Nitti, D. (2008). TNF and cancer: the two sides of the coin. Front Biosci, 13(2774), 83. [4] Johnson, C. S., Cook, C. A., & Furmanski, P. (1990). In vivo suppression of erythropoiesis by tumor necrosis factor-alpha (TNF-alpha): reversal with exogenous erythropoietin (EPO). Experimental hematology, 18(2), 109-113. [5] Yu, H., & Jove, R. (2004). The STATs of cancer—new molecular targets come of age. Nature Reviews Cancer, 4(2), 97-105. [6] Carmeliet, P. (2005). VEGF as a key mediator of angiogenesis in cancer. Oncology, 69(Suppl. 3), 4-10. [7] Maiese, K., Li, F., & Chong, Z. Z. (2005). New avenues of exploration for erythropoietin. Jama, 293(1), 90-95. [8] Szenajch, J., Wcislo, G., Jeong, J. Y., Szczylik, C., & Feldman, L. (2010). The role of erythropoietin and its receptor in growth, survival and therapeutic response of human tumor cells: from clinic to bench—a critical review. Biochimica et Biophysica Acta (BBA)-Reviews on Cancer, 1806(1), 82-95. [9] Jelkmann, W., Bohlius, J., Hallek, M., & Sytkowski, A. J. (2008). The erythropoietin receptor in normal and cancer tissues. Critical reviews in oncology/hematology, 67(1), 39-61. [10] Hardee, M. E., Cao, Y., Fu, P., Jiang, X., Zhao, Y., Rabbani, Z. N., ... & Arcasoy, M. O. (2007). Erythropoietin blockade inhibits the induction of tumor angiogenesis and progression. PLoS One, 2(6). [11] Okazaki, T., Ebihara, S., Asada, M., Yamanda, S., Niu, K., & Arai, H. (2008). Erythropoietin promotes the growth of tumors lacking its receptor and decreases survival of tumor-bearing mice by enhancing angiogenesis. Neoplasia (New York, NY), 10(9), 932. [12] Rosenzweig, M. Q., Bender, C. M., Lucke, J. P., Yasko, J. M., & Brufsky, A. M. (2004). The decision to prematurely terminate a trial of R-HuEPO due to thrombotic events. Journal of pain and symptom management, 27(2), 185-190. [13] Bennett, C. L., Silver, S. M., Djulbegovic, B., Samaras, A. T., Blau, C. A., Gleason, K. J., ... & Edwards, B. J. (2008). Venous thromboembolism and mortality associated with recombinant erythropoietin and darbepoetin administration for the treatment of cancer-associated anemia. Jama, 299(8), 914-924. [14] Vaziri, N. D. (1999). Mechanism of erythropoietin-induced hypertension. American journal of kidney diseases, 33(5), 821-828. [15] Bonnas, C., Wüstefeld, L., Winkler, D., Kronstein-Wiedemann, R., Dere, E., Specht, K., ... & Sillaber, I. (2017). EV-3, an endogenous human erythropoietin isoform with distinct functional relevance. Scientific reports, 7(1), 1-15. [16] Calapai, G., Marciano, M. C., Corica, F., Allegra, A., Parisi, A., Frisina, N., ... & Buemi, M. (2000). Erythropoietin protects against brain ischemic injury by inhibition of nitric oxide formation. European journal of pharmacology, 401(3), 349-356. [17] García-Rodríguez, J. C., & Sosa Teste, I. (2009). The nasal route as a potential pathway for delivery of erythropoietin in the treatment of acute ischemic stroke in humans. The Scientific World Journal, 9, 970-981. [18] Sosa Testé, I., García Rodríguez, J. C., García Salman, J. D., Santana, J., Subirós Martínez, N., González Triana, C., ... & Cruz Rodríguez, J. (2006). Intranasal administration of recombinant human erythropoietin exerts neuroprotective effects on post-ischemic brain injury in Mongolian gerbils. Pharmacol Online, 1, 100-112. [19] Lagarto, A., Bueno, V., Guerra, I., Valdés, O., Couret, M., López, R., & Vega, Y. (2011). Absence of hematological side effects in acute and subacute nasal dosing of erythropoietin with a low content of sialic acid. Experimental and toxicologic pathology, 63(6), 563-567. [20] Sosa, I., Cruz, J., Santana, J., Mengana, Y., García-Salman, J. D., Muñoz, A., ... & García, J. C. (2008). Paso de la molécula de eritropoyetina humana recombinante con bajo contenido de ácido siálico al Sistema nervioso central por la vía intranasal en los modelos del meriones unguiculatus y el primate no humano Macaca fascicularis. Revista de Salud Animal, 30(1), 39-44. [21] Rodriguez Cruz, Y., Yuneidys Mengana, T., Munoz Cernuda, A., Subirós Martines, N., González-Quevedo, A., Sosa Teste, I., & García Rodríguez, J. C. (2010). Treatment with nasal neuro-EPO improves the neurological, cognitive, and histological state in a gerbil model of focal ischemia. The Scientific World Journal, 10, 2288-2300. [22] Lagarto, A., Bueno, V., Guerra, I., Valdés, O., Couret, M., López, R., & Vega, Y. (2011). Absence of hematological side effects in acute and subacute nasal dosing of erythropoietin with a low content of sialic acid. Experimental and toxicologic pathology, 63(6), 563-567. [23] Hermann, D. M. (2010). Nonhematopoietic variants of erythropoietin in ischemic stroke: need for step-wise proof-of-concept studies. The Scientific World Journal, 10, 2285-2287. [24] Rodriguez Cruz, Y., Strehaiano, M., Rodriguez Obaya, T., Garcia Rodriguez, J. C., & Maurice, T. (2017). An intranasal formulation of erythropoietin (Neuro-EPO) prevents memory deficits and amyloid toxicity in the APP Swe transgenic mouse model of Alzheimer’s disease. Journal of Alzheimer's Disease, 55(1), 231-248. [25] Maurice, T., Mustafa, M. H., Desrumaux, C., Keller, E., Naert, G., Garcia-Barcelo, M. D. L. C., ... & Garcia Rodríguez, J. C. (2013). Intranasal formulation of erythropoietin (EPO) showed potent protective activity against amyloid toxicity in the Aβ25-35 non-transgenic mouse model of Alzheimer’s disease. Journal of Psychopharmacology, 27(11), 1044-1057. [26] Garzon, F., Coimbra, D., Parcerisas, A., Rodriguez, Y., García, J. C., Soriano, E., & Rama, R. (2018). NeuroEPO preserves neurons from glutamate-induced excitotoxicity. Journal of Alzheimer's Disease, 65(4), 1469-1483. [27] Rama, R., Garzón, F., Rodríguez-Cruz, Y., Maurice, T., & García-Rodríguez, J. C. (2019). Neuroprotective effect of Neuro-EPO in neurodegenerative diseases:“Alea jacta est”. Neural regeneration research, 14(9), 1519. [28] García-Rodríguez, J. C., & Sosa Teste, I. (2009). The nasal route as a potential pathway for delivery of erythropoietin in the treatment of acute ischemic stroke in humans. The Scientific World Journal, 9, 970-981. [29] Lagarto Parra, A., & Cesar Garcia Rodriguez, J. (2012). Nasal neuro EPO could be a reliable choice for neuroprotective stroke treatment. Central Nervous System Agents in Medicinal Chemistry (Formerly Current Medicinal Chemistry-Central Nervous System Agents), 12(1), 60-68. [30] Santos-Morales, O., Díaz-Machado, A., Jiménez-Rodríguez, D., Pomares-Iturralde, Y., Festary-Casanovas, T., González-Delgado, C. A., ... & Garcia-Garcia, I. (2017). Nasal administration of the neuroprotective candidate NeuroEPO to healthy volunteers: a randomized, parallel, open-label safety study. BMC neurology, 17(1), 129. [31] Toledo, J. R., Sánchez, O., Seguí, R. M., García, G., Montañez, M., Zamora, P. A., ... & Cremata, J. A. (2006). High expression level of recombinant human erythropoietin in the milk of non-transgenic goats. Journal of biotechnology, 123(2), 225-235. [32] Castillo, C., Zaror, S., Gonzalez, M., Hidalgo, A., Burgos, C. F., Cabezas, O. I., ... & Gavilán, J. (2018). Neuroprotective effect of a new variant of Epo nonhematopoietic against oxidative stress. Redox biology, 14, 285-294. [33] Erbayraktar, S., Grasso, G., Sfacteria, A., Xie, Q. W., Coleman, T., Kreilgaard, M., ... & Yilmaz, O. (2003). Asialoerythropoietin is a nonerythropoietic cytokine with broad neuroprotective activity in vivo. Proceedings of the National Academy of Sciences, 100(11), 6741-6746. [34] Mori, S., Sawada, T., & Kubota, K. (2007). Asialoerythropoietin is a strong modulator of angiogenesis by bone-marrow cells. Journal of Investigative Surgery, 20(6), 357-362. [35] Erbayraktar, S., Grasso, G., Sfacteria, A., Xie, Q. W., Coleman, T., Kreilgaard, M., ... & Yilmaz, O. (2003). Asialoerythropoietin is a nonerythropoietic cytokine with broad neuroprotective activity in vivo. Proceedings of the National Academy of Sciences, 100(11), 6741-6746. [36] Wang, X., Zhu, C., Wang, X., Gerwien, J. G., Schrattenholz, A., Sandberg, M., ... & Blomgren, K. (2004). The nonerythropoietic asialoerythropoietin protects against neonatal hypoxia‐ischemia as potently as erythropoietin. Journal of neurochemistry, 91(4), 900-910. [37] Morishita, E. M. M. Y. R., Masuda, S., Nagao, M., Yasuda, Y., & Sasaki, R. (1996). Erythropoietin receptor is expressed in rat hippocampal and cerebral cortical neurons, and erythropoietin prevents in vitro glutamate-induced neuronal death. Neuroscience, 76(1), 105-116. [38] Gil, J. M., Leist, M., Popovic, N., Brundin, P., & Petersén, Å. (2004). Asialoerythropoetin is not effective in the R6/2 line of Huntington's disease mice. BMC neuroscience, 5(1), 17. [39] Kaneko, N., Kako, E., & Sawamoto, K. (2013). Enhancement of ventricular-subventricular zone-derived neurogenesis and oligodendrogenesis by erythropoietin and its derivatives. Frontiers in cellular neuroscience, 7, 235. [40] Higuchi, Masato. "Method for removing sialic acid and method for producing asialoerythropoietin." U.S. Patent Application 12/087,945, filed 2009. [41] Leist, M., Ghezzi, P., Grasso, G., Bianchi, R., Villa, P., Fratelli, M., ... & Kallunki, P. (2004). Derivatives of erythropoietin that are tissue protective but not erythropoietic. Science, 305(5681), 239-242. [42] Coleman, T. R., Westenfelder, C., Tögel, F. E., Yang, Y., Hu, Z., Swenson, L., ... & Ghezzi, P. (2006). Cytoprotective doses of erythropoietin or carbamylated erythropoietin have markedly different procoagulant and vasoactive activities. Proceedings of the National Academy of Sciences, 103(15), 5965-5970. [43] Brines, M., Grasso, G., Fiordaliso, F., Sfacteria, A., Ghezzi, P., Fratelli, M., ... & Pobre, E. (2004). Erythropoietin mediates tissue protection through an erythropoietin and common β-subunit heteroreceptor. Proceedings of the National Academy of Sciences, 101(41), 14907-14912. [44] Montero, M., Poulsen, F. R., Noraberg, J., Kirkeby, A., van Beek, J., Leist, M., & Zimmer, J. (2007). Comparison of neuroprotective effects of erythropoietin (EPO) and carbamylerythropoietin (CEPO) against ischemia-like oxygen–glucose deprivation (OGD) and NMDA excitotoxicity in mouse hippocampal slice cultures. Experimental Neurology, 204(1), 106-117. [45] Tiwari, N. K., Sathyanesan, M., Schweinle, W., & Newton, S. S. (2019). Carbamoylated erythropoietin induces a neurotrophic gene profile in neuronal cells. Progress in Neuro-Psychopharmacology and Biological Psychiatry, 88, 132-141. [46] Wang, Y., Zhang, Z. G., Rhodes, K., Renzi, M., Zhang, R. L., Kapke, A., ... & Chopp, M. (2007). Post‐ischemic treatment with erythropoietin or carbamylated erythropoietin reduces infarction and improves neurological outcome in a rat model of focal cerebral ischemia. British journal of pharmacology, 151(8), 1377-1384. [47] Erbayraktar, S., De Lanerolle, N., De Lotbinière, A., Knisely, J. P., Erbayraktar, Z., Yilmaz, O., ... & Brines, M. (2006). Carbamylated erythropoietin reduces radiosurgically-induced brain injury. Molecular medicine, 12(4), 74-80. [48] Montero, M., Poulsen, F. R., Noraberg, J., Kirkeby, A., van Beek, J., Leist, M., & Zimmer, J. (2007). Comparison of neuroprotective effects of erythropoietin (EPO) and carbamylerythropoietin (CEPO) against ischemia-like oxygen–glucose deprivation (OGD) and NMDA excitotoxicity in mouse hippocampal slice cultures. Experimental Neurology, 204(1), 106-117. [49] Mahmood, Asim, et al. "Treatment of traumatic brain injury in rats with erythropoietin and carbamylated erythropoietin." Journal of neurosurgery 107.2 (2007): 392-397. [50] King, V. R., Averill, S. A., Hewazy, D., Priestley, J. V., Torup, L., & Michael‐Titus, A. T. (2007). Erythropoietin and carbamylated erythropoietin are neuroprotective following spinal cord hemisection in the rat. European Journal of Neuroscience, 26(1), 90-100. [51] Sampath, D., McWhirt, J., Sathyanesan, M., & Newton, S. S. (2020). Carbamoylated erythropoietin produces antidepressant-like effects in male and female mice. Progress in Neuro-Psychopharmacology and Biological Psychiatry, 96, 109754. [52] Wang, Y., Zhang, Z. G., Rhodes, K., Renzi, M., Zhang, R. L., Kapke, A., ... & Chopp, M. (2007). Post‐ischemic treatment with erythropoietin or carbamylated erythropoietin reduces infarction and improves neurological outcome in a rat model of focal cerebral ischemia. British journal of pharmacology, 151(8), 1377-1384. [53] Lapchak, P. A., Kirkeby, A., Zivin, J. A., & Sager, T. N. (2008). Therapeutic window for nonerythropoietic carbamylated-erythropoietin to improve motor function following multiple infarct ischemic strokes in New Zealand white rabbits. Brain research, 1238, 208-214. [54] Christensen, S., Foldager, L., Valbjorn, J., Thuesen, M., Pedersen, A., & Munk, M. (2006). U.S. Patent Application No. 11/178,155. [55] Brines, M., Patel, N. S., Villa, P., Brines, C., Mennini, T., De Paola, M., ... & Ghezzi, P. (2008). Nonerythropoietic, tissue-protective peptides derived from the tertiary structure of erythropoietin. Proceedings of the National Academy of Sciences, 105(31), 10925-10930. [56] Robertson, C. S., Garcia, R., Gaddam, S. S. K., Grill, R. J., Cerami Hand, C., Tian, T. S., & Hannay, H. J. (2013). Treatment of mild traumatic brain injury with an erythropoietin-mimetic peptide. Journal of neurotrauma, 30(9), 765-774. [57] Robertson, C. S., Cherian, L., Shah, M., Garcia, R., Navarro, J. C., Grill, R. J., ... & Hannay, H. J. (2012). Neuroprotection with an erythropoietin mimetic peptide (pHBSP) in a model of mild traumatic brain injury complicated by hemorrhagic shock. Journal of neurotrauma, 29(6), 1156-1166. [58] Collino, M., Benetti, E., Rogazzo, M., Chiazza, F., Mastrocola, R., Nigro, D., ... & Thiemermann, C. (2014). A non‐erythropoietic peptide derivative of erythropoietin decreases susceptibility to diet‐induced insulin resistance in mice. British journal of pharmacology, 171(24), 5802-5815. [59] Brines, M., Dunne, A. N., van Velzen, M., Proto, P. L., Ostenson, C. G., Kirk, R. I., ... & Dahan, A. (2014). ARA 290, a nonerythropoietic peptide engineered from erythropoietin, improves metabolic control and neuropathic symptoms in patients with type 2 diabetes. Molecular medicine, 20(1), 658-666. [60] McVicar, C. M., Hamilton, R., Colhoun, L. M., Gardiner, T. A., Brines, M., Cerami, A., & Stitt, A. W. (2011). Intervention with an erythropoietin-derived peptide protects against neuroglial and vascular degeneration during diabetic retinopathy. Diabetes, 60(11), 2995-3005. [61] Schmidt, R. E., Feng, D., Wang, Q., Green, K. G., Snipes, L. L., Yamin, M., & Brines, M. (2011). Effect of insulin and an erythropoietin-derived peptide (ARA290) on established neuritic dystrophy and neuronopathy in Akita (Ins2Akita) diabetic mouse sympathetic ganglia. Experimental neurology, 232(2), 126-135. [62] Ahmet, I., Tae, H. J., Juhaszova, M., Riordon, D. R., Boheler, K. R., Sollott, S. J., ... & Talan, M. I. (2011). A small nonerythropoietic helix B surface peptide based upon erythropoietin structure is cardioprotective against ischemic myocardial damage. Molecular Medicine, 17(3-4), 194-200. [63] Ahmet, I., Tae, H. J., Brines, M., Cerami, A., Lakatta, E. G., & Talan, M. I. (2013). Chronic Administration of Small Nonerythropoietic Peptide Sequence of Erythropoietin Effectively Ameliorates the Progression of Postmyocardial Infarction–Dilated Cardiomyopathy. Journal of Pharmacology and Experimental Therapeutics, 345(3), 446-456. [64] van Rijt, W. G., Nieuwenhuijs-Moeke, G. J., van Goor, H., Ottens, P. J., Ploeg, R. J., & Leuvenink, H. G. (2013). Renoprotective capacities of non-erythropoietic EPO derivative, ARA290, following renal ischemia/reperfusion injury. Journal of translational medicine, 11(1), 286. [65] Howell, K., Whelan, S., Rochfort, K., & Brines, M. (2017). Specific ligand for the innate repair receptor, ARA290, attenuates pulmonary hypertension in an elastase-induced murine model of emphysema. [66] Bohr, S., Patel, S. J., Shen, K., Vitalo, A. G., Brines, M., Cerami, A., ... & Yarmush, M. L. (2013). Alternative erythropoietin-mediated signaling prevents secondary microvascular thrombosis and inflammation within cutaneous burns. Proceedings of the National Academy of Sciences, 110(9), 3513-3518. [67] Swartjes, M., Morariu, A., Niesters, M., Brines, M., Cerami, A., Aarts, L., & Dahan, A. (2011). ARA290, a Peptide Derived from the Tertiary Structure of Erythropoietin, Produces Long-term Relief of Neuropathic PainAn Experimental Study in Rats and β-Common Receptor Knockout Mice. Anesthesiology: The Journal of the American Society of Anesthesiologists, 115(5), 1084-1092. [68] Zhang, W., Yu, G., & Zhang, M. (2016). ARA 290 relieves pathophysiological pain by targeting TRPV1 channel: Integration between immune system and nociception. Peptides, 76, 73-79. [69] Nanavati, A., Moen, J., Axsom, J., Krawczyk, M., Petrashevskaya, N., Beyman, M., ... & Morrell, C. (2018). ARA290, A Small Non-Hematopoietic Peptide Derived From Erythropoietin, Prolongs Healthspan And Attenuates Age-Associated Declines In The Heart’s Structure And Function. The FASEB Journal, 32(1_supplement), lb288-lb288. [70] Cerit, H., Veer, I. M., Dahan, A., Niesters, M., Harmer, C. J., Miskowiak, K. W., ... & Van der Does, W. (2015). Testing the antidepressant properties of the peptide ARA290 in a human neuropsychological model of drug action. European Neuropsychopharmacology, 25(12), 2289-2299. [71] Pankratova, S., Kiryushko, D., Sonn, K., Soroka, V., Køhler, L. B., Rathje, M., ... & Bock, E. (2010). Neuroprotective properties of a novel, non-haematopoietic agonist of the erythropoietin receptor. Brain, 133(8), 2281-2294. [72] Sirén, A. L., Faßhauer, T., Bartels, C., & Ehrenreich, H. (2009). Therapeutic potential of erythropoietin and its structural or functional variants in the nervous system. Neurotherapeutics, 6(1), 108-127. [73] Pankratova, S., Gu, B., Kiryushko, D., Korshunova, I., Køhler, L. B., Rathje, M., ... & Berezin, V. (2012). A new agonist of the erythropoietin receptor, Epobis, induces neurite outgrowth and promotes neuronal survival. Journal of neurochemistry, 121(6), 915-923. [74] Dmytriyeva, O., Pankratova, S., Korshunova, I., & Walmod, P. S. (2016). Epobis is a nonerythropoietic and neuroprotective agonist of the erythropoietin receptor with anti-inflammatory and memory enhancing effects. Mediators of inflammation, 2016. [75] Dmytriyeva, O., Belmeguenai, A., Bezin, L., Soud, K., Woldbye, D. P. D., Gøtzsche, C. R., & Pankratova, S. (2019). Short erythropoietin-derived peptide enhances memory, improves long-term potentiation, and counteracts amyloid beta–induced pathology. Neurobiology of aging, 81, 88-101. [76] Upadhyay, R. K. (2014). Drug delivery systems, CNS protection, and the blood brain barrier. BioMed research international, 2014. [77] Yuan, R., Wang, B., Lu, W., Maeda, Y., & Dowling, P. (2015). A distinct region in erythropoietin that induces immuno/inflammatory modulation and tissue protection. Neurotherapeutics, 12(4), 850-861. [78] Wang, B., Kang, M., Marchese, M., Rodriguez, E., Lu, W., Li, X., ... & Dowling, P. (2016). Beneficial effect of erythropoietin short peptide on acute traumatic brain injury. Neurotherapeutics, 13(2), 418-427. [79] Dmytriyeva, O., Belmeguenai, A., Bezin, L., Soud, K., Woldbye, D. P. D., Gøtzsche, C. R., & Pankratova, S. (2019). Short erythropoietin-derived peptide enhances memory, improves long-term potentiation, and counteracts amyloid beta–induced pathology. Neurobiology of aging, 81, 88-101. [80] Dowling, P. C., Yuan, R. R., & Lu, W. (2017). U.S. Patent Application No. 15/413,398. [81] Wang, B., Yuan, R. R., Lu, W., & Dowling, P. C. (2016). U.S. Patent No. 9,345,745. Washington, DC: U.S. Patent and Trademark Office. [82] Lu, W., Dowling, P. C., & Yuan, R. R. (2017). U.S. Patent No. 9,585,932. Washington, DC: U.S. Patent and Trademark Office. [83] Dhanushkodi, A., Akano, E. O., Roguski, E. E., Xue, Y., Rao, S. K., Matta, S. G., ... & McDonald, M. P. (2013). A single intramuscular injection of rAAV‐mediated mutant erythropoietin protects against MPTP‐induced parkinsonism. Genes, Brain and Behavior, 12(2), 224-233. [84] Sajja, V. S., Tenn, C., McLaws, L. J., & Vandevord, P. J. (2012). A temporal evaluation of cytokines in rats after blast exposure. Biomedical sciences instrumentation, 48, 374-379.
Comments