Review 
Diabetic Kinome Inhibitors—A New Opportunity for β-Cells Restoration 
Barbara Pucelik 1 , Agata Barzowska 1, JanuszM.D˛abrowski2,* and Anna Czarna 1,* 




Citation: Pucelik, B.; Barzowska, A.; D˛abrowski, J.M.; Czarna, A. Diabetic Kinome Inhibitors—A New Opportunity for β-Cells Restoration. Int. J. Mol. Sci. 2021, 22, 9083. https://doi.org/10.3390/ ijms22169083 
Academic Editors: Maria Ruzzene and Christian Borgo 
Received: 19 July 2021 Accepted: 18 August 2021 Published: 23 August 2021 
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Copyright: © 2021 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https:// creativecommons.org/licenses/by/ 4.0/). 
1  Malopolska Centre of Biotechnology, Jagiellonian University, Gronostajowa 7A, 30-387 Krakow, Poland;  
barbara.pucelik@uj.edu.pl (B.P.); agata.barzowska@doctoral.uj.edu.pl (A.B.)  
2  Faculty of Chemistry, Jagiellonian University, Gronostajowa 2, 30-387 Krakow, Poland  

* Correspondence: jdabrows@chemia.uj.edu.pl (J.M.D.); anna1.czarna@uj.edu.pl (A.C.) 
Abstract: Diabetes, and several diseasesrelated to diabetes, including cancer, cardiovascular diseases and neurological disorders, represent one of the major ongoing threats to human life, becoming a true pandemic of the 21st century. Current treatment strategies for diabetes mainly involve promoting β-cell differentiation, and one of the most widely studied targets for β-cell regeneration is DYRK1A kinase,a memberoftheDYRKfamily. DYRK1Ahasbeen characterizedasakeyregulator of cell growth, differentiation, and signal transduction in various organisms, while further roles and substrates are the subjects of extensive investigation. The targets of interest in this review are implicated in the regulation of β-cells through DYRK1A inhibition—through driving their transition from highly ineffcient and death-prone populations into effcient and suffcient precursors of islet regeneration. Increasing evidence for the role of DYRK1A in diabetes progression and β-cell proliferation expands the potential for pharmaceutical applications of DYRK1A inhibitors. The variety of new compounds and binding modes, determinedbycrystal structure and in vitro studies, may lead to new strategies for diabetes treatment. This review provides recent insights into the initial self-activationof DYRK1Abytyrosine autophosphorylation.Moreover,the importance of developing novel DYRK1A inhibitors and their implications for the treatment of diabetes are thoroughly discussed. The evolving understanding of DYRK kinase structure and function and emerging high-throughput screening technologies have been described. As a fnal point of this work, we intend to promote the term “diabetic kinome” as part of scientifc terminology to emphasize the role of the synergistic action of multiple kinases in governing the molecular processes that underlie this particular group of diseases. 
Keywords: diabetic kinome; protein kinases; DYRK1A; diabetes; beta-cells 
1. Introduction 
The diabetic kinome consists of protein kinases that control and regulate protein functions involvedin diabetes.Arangeof experimental evidence indicates that pharmacological modulations of the diabetic kinome are inextricably linked to changes in metabolic homeostasis. Numerous cell types and signaling pathways in diabetes have been identifed (i.e., PI3K-AKT/PKB, ERK/MAPK,growth factor, and hormone signaling pathways)[1–3]. Among others, the selected proteins’ kinase activity, i.e., hexokinase, pyruvate kinase M2 ketohexokinase isoform A, phosphoglycerate kinase 1, and nucleoside diphosphate kinase 1and2(NME1/2), contribute to altered metabolic homeostasis[4,5]. 
Insulinregulates glucose homeostasisby modulatingprotein kinases’ activityin target tissues. The impairment of the kinome response to insulin leads to insulin resistance. Thus, many kinases, including (i) Jun N-terminal kinase (JNK), (ii)Ikappa beta kinase (IKK), (iii) protein kinaseC(PKC) theta, (iv) glycogen synthase kinase3(GSK3), (v) S6 kinase-1 (S6K1), and (vi) 5’AMP-activated protein kinase (AMPK), are critical factors that regulate the insulin-dependent processes. Moreover, many of them are also related tothe pathogenesisof diabetes[6]. Morerecently,theroleof dual-specifcitytyrosine 
Int. J. Mol. Sci. 2021, 22, 9083. https://doi.org/10.3390/ijms22169083 https://www.mdpi.com/journal/ijms 
phosphorylation-regulated kinaseA(DYRK1A) was identifedin β-cells’ function. Due to the large amount of data related to mutations, overexpression, and dysregulation of protein kinases in the pathogenesis of many diseases, this family of enzymes has become oneof the most importantdrugtargets during the past20 years[7,8].Amilestone was the FDA approval (in 2001) of the frst kinase inhibitor, imatinib, also known under the trade name Gleevec®(Novartis, Basel, Switzerland). It is an oral chemotherapy drug used to treat leukemia and gastrointestinal stromal tumors whose mechanism of action involves potent inhibitionof the constitutively active BCR-ABL fusionprotein[7–9]. Imatinibis also being studiedinphaseII clinical trialstotreattypeIdiabetes(T1D) (NCT01781975)[10].This study aimed to investigate the possibility of short-term therapy with imatinib to induce tolerance and long-termremissionof T1D[10]. The developmentof small-molecule kinase inhibitors has emerged as one of the most extensively pursued areas of drug discovery. In recent years, signifcant progress has been made in the battle against diabetes mellitus (DM) in understanding its biological mechanisms. However, despite a large amount of data, the search for diabetes-relevant kinases inhibitors with their binding modes and structural featuresisrequired. Especially,therearestill unexploredgapsinthe knowledge of how protein kinases from the DYRKs family affect apoptosis, cell cycle regulation, cellular proliferation, and insulin resistance in diabetes. DYRK1A has been confrmed as a regulator of regenerative pathways essential for proper pancreatic β-cells function in humans. Inhibitors of this kinase have been extensively studied to treat various types of diabetes[11,12]. Harmine and its derivatives are oneof the mostfrequently studied—and still the most potent therapeutic of this group of compounds[13–16]. 
Recently published review papers on diabetes and CNS disorders highlight the im-portanceof DYRK inhibitorsin the therapyof cancer and neurological disorders[17–19] and suggest the directions in the design and development of small-molecule inhibitors (Figure 1)[12,20–23]. Thisreview describes recentreports on the initial self-activation of DYRK1A by tyrosine autophosphorylation, the development of DYRK1A inhibitors, and their importance in the treatment of diabetes mellitus (DM). In addition, advances in understanding the structure and function of DYRK kinases and functions and emerging HTS technologies are described. 

Figure 1. DYRK1A role in human diseases and the potential use of its inhibitors. 
Modulating the activity of DYRK1A kinase, which is signifcantly involved in diabetes, with small-molecule inhibitors could be an attractive therapeutic strategy to tackle diabetes. 
The global diabetes population is estimated to be 9.3% (463 million people) in 2019, risingto 10.2%(578 million)by2030and 10.9%(700 million)by2045[24]. Thus diabeteshas becomea challenging healthproblemaffecting the global population, and theprevalenceis higherindeveloping countries[24]. Diabetes mellitus (DM)is causedby chronic hyper<BR>glycemia due to impaired β-cells from the islets of Langerhans, distributed throughout the endocrinepancreastoproduceappropriate insulin levelsorineffective insulinusage[25]. It is also associated with vascular complications, mainly diabetic neuropathy (DN), with an incidenceof about 50%[26]. TheDNprogresses with decreasing nerve functionality withahighriskofpain,trophic changes,and autonomic dysfunction. Diabetesmayalso lead to ketoacidosis, retinopathy, nephropathy, and skin complications. Moreover, diabetes dramatically increases the risk of various cardiovascular problems, including coronary artery disease with chest pain (angina), heart attack, stroke, and narrowing of arteries (atherosclerosis)[27,28]. 
Accordingtothelatest classifcation,thereareseveraltypesofdiabetes:type1diabetes designated as T1D, type2diabetes marked as T2D, gestational diabetes, and other variants listed inTable 1. It should be emphasized that the pathogenesis of each form differs signifcantly.T1Disan autoimmune disorder causedbytheT-cell-mediated destruction of the insulin-producing pancreatic β-cells. T2D is a consequence of impaired glucose tolerance and insulin resistance with the prominent risk factors: obesity and physical inactivity. In addition to these most common, thereis also monogenic diabetes (for example, MODY, neonatal diabetes), extrinsic pancreatic diseases (for example, cystic fbrosis-related diabetes, pancreatic diabetes [or type 3c], and drug-induced diabetes[29]. In type1 diabetes,a signifcantreductionin pancreatic β-cells, resulting in insulin insuffciency and hyperglycemia,is observed.Type2diabetesis associated with insulinresistance, which causes the compensatory expansion of pancreatic β-cells and increases plasma insulin levels[30,31]. Finally, insuffcient β-cell mass and insulin secretion also cause mature onset diabetes of the young and gestational diabetes[32]. Therefore, modern antidiabetic therapies are based on increasing functional pancreatic β-cell mass. This review briefy discusses only the most common forms of diabetes. According to the American Diabetes Association (ADA) position statement, “Diagnosis and Classifcation of Diabetes Mellitus” (Table 1)providesa detailed classifcationof diabetesby etiology[33,34]. 
Table 1.Classifcationof diabetes mellitusby etiology[33]. 
Type Etiology 
Immunologically-mediated 
typeIdiabetes 
Idiopathic 
Genetic predisposition, type II diabetes Insulin resistance (aging, physical inactivity, and overweight) Relative insulin defciency and decreased β-cell function 
gestational 
β-cell dysfunction on a background of chronic insulin resistance during pregnancy 
diabetes Genetic defects of β-cell function: 
• 
MODY3, MODY2, MODY1and others 

• 
transient and permanent neonatal diabetes 

• 
mitochondrial DNA and others 

• 
Genetic defects in insulin action: 


• typeAinsulinresistance 

• 
leprechaunism, Rabson-Mendenhall syndrome 


• Lipoathropic diabetes and others The disease of the exocrine pancreas: 
• pancreatitis, trauma/pancreatectomy, neoplasia, cystic fbrosis, hemochromatosis, 
fbrocalculous pancreatopathy, and others Endocrinopathies: 
• acromegaly, Cushing’s syndrome, glucagonoma, pheochromocytoma, hyperthyroidism, other types somatostatinoma, aldosteronoma of diabetes Drug or chemical-induced: 
• e.g., vacor, pentamidine, nicotinic acid, glucocorticoids, thyroid hormone, diazoxide, 
β-adrenergic agonists, thiazides, Dilantin, γ-IFN Infections: 
• congenitalrubella 
• cytomegalovirus and others Uncommon forms of immune-mediated diabetes: 
• Stiff-man syndrome 
• anti-insulinreceptor antibodies and others Other genetic syndromes sometimes associated with diabetes 
• Down syndrome, Klinefelter syndrome,Wolfram syndrome, Friedreich ataxia, 
Huntington chorea, Laurence-Mood-Biedl syndrome, myotonic dystrophy, porphyria, Prader-Willi syndrome and others 

2. Insulin Homeostasis and Diabetes 
Insulin plays a crucial role in many metabolic processes, including (i) facilitation of cellularuptakeofglucose,(ii)preventionoftheglucosereleasebytheliver,(iii) activationof themusclecellstotakeupaminoacids,and(iv)reductionthebreakdown, conversion,and releaseoffats(Figure2)[35].In severaltissues,suchastheliver,muscle,andadiposetissue, insulin participatesin glucose metabolismby stimulating glucose uptake and infuencing both glycolysis and gluconeogenesis. 

Figure 2. Thefowchart illustratingtheroleofthepancreasinregulatingbloodglucose concentration. Localizedinthe isletsof Langerhans, pancreatic β-cells respond to blood glucose levels, resulting in the release of the proper amounts of insulin. Insulin affects the liver, muscles, brain, erythrocytes, and adipocytes. The loss of β-cells leads to insuffcient insulin production, resulting in increased blood glucose levels and eventually causes diabetes or insulin resistance. Adopted, modifed, and re-drawn from[36]. 
Insulin stimulates glycogen synthesis by inhibiting glycogen synthetase kinase, en-hances its production through mTOR activation, promotes fatty acid synthesis, and inhibits lipolysis by activating Acetyl-CoA Carboxylase. It also inhibits hormone-sensitive lipase and modulates gene transcription through the MAPK pathway or Akt-mediated phosphorylationof FOXO transcription factors[37]. The secretionof insulinfrom the β-cells can be triggered eitherby somatotropin orby glucagon. The most important stimulant for insulin release is glucose, and when blood glucose levels rise, insulin is released to balance this process. The defciencyin insulinregulatory functionmaybe causedby inadequate insulin secretion and/orreduced tissueresponse.It alsoresultsfroma complete inabilityof islet cellstoproduce insulin(T1D)orthe failuretoproduceenough insulin(Figure 3)[38–40]. 
Diabetesisa heterogeneous disease,but most cases correspondingtotype1and type2diabetes. Nevertheless,a considerableproportionof patients does not ft into this classifcationandareknowntohavehyperglycemiacausedby a mutationinasinglegene. Despite the rapid evolution of molecular diagnosis methods, many MODY cases may be misdiagnosed as type1 or type2diabetes. Thus, in the following sections, we briefy characterized only these most common types of diabetes. 

Figure 3. Scheme illustrating possible effects of insulin and glucagon on body function. Adopted, modifed, and re-drawn from[41]. 
2.1. Type I Diabetes (T1D) 
T1D, referred to as insulin-dependent diabetes or juvenile-onset diabetes, concerns ca. 5–10% of cases. As already mentioned, it results from autoimmune destruction of the β-cells, accompaniedby cellular invasionbybothCD4+andCD8+Tcells,leadingto a decrease in β-cell mass[42,43]. Several markersof β-cell immune destruction include autoantibodies to islet cells, insulin, glutamic acid decarboxylase (GAD65), and tyrosine phosphatases IA-2 and IA-2β that are usually present in ca. 90% of patients[34]. T1D has genetic predispositions; the human leukocyte antigen (HLA) complex linked to the DQAandDQBgenes constitutesthemostrelevant susceptibilityregion[44,45].Itisalso relatedto poorly defned environmental factors.AsmallpercentageofT1D patients (<10%) display no autoimmune response evidence and are categorized as type 1B diabetic or idiopathic population[43]. 

2.2. Type 2 Diabetes (T2D) 
The most common form of diabetes is T2D (ca. 90–95% cases), also named non-insulin-dependent diabetes or adult-onset diabetes. Even though the etiologies of T2D are not fully explored. In this case (in contrast to T1D), autoimmune destruction of β-cells does not occur[25]. Lifestyle factors, including physical inactivity, sedentary lifestyle, smoking, andfrequent alcohol consumption,playanimportantrolein developingT2D[3,31].Itis characterized by increased hyperinsulinemia, insulin resistance, and β-cell dysfunction, with up to 50% cell loss at the time of diagnosis. T2D leads to a decrease in glucose transport into the liver, muscle cells, and fat cells. Recently, the involvement of impaired α-cell functionhas beenrecognizedinthe pathophysiologyofT2D[46]. Consequently, glucagon and hepatic glucose levels that rise during fasting are not suppressed witha meal due to inadequate insulin concentration, increased insulin resistance, and increased fat breakdown with hyperglycemia[32]. 
T2D contributes to a substantial increase in the risk of cardiovascular disease. Other mechanisms for developing hyperglycemia-induced micro and macrovascular complications include endothelial dysfunction, advanced glycation end-product formation, hypercoagulability, increased platelet reactivity, and overexpression of sodium-glucose cotransporter-2 (SGLT-2)[47–49]. Fibrinolysis and platelet aggregation canberemarkably improvedby metformin therapy. Glucagon-like peptide-1 (GLP-1)receptor agonists have been confrmed to have protective effects on the endothelium, which may help to reduce infammation (Figure 4.)[50–52]. 

Figure 4. Schematic illustration of the main types of diabetes in which the pancreas does not produce enough insulin, or the body’scellsdonotrespondappropriatelytothe insulinproduced. Adoptedandchangedfromastockimage. 

2.3. Maturity-Onset Diabetes of the Young (MODY) 
Other forms of diabetes are associated with monogenetic defects in β-cell function. In maturity-onset diabetes of the young (MODY), the onset of hyperglycemia occurs early (generally before age 25). This typeofDMis characterizedbyimpaired insulin secretion and minimal or no defects in insulin action[53]. The most common form of MODY is associated with mutations in the hepatic transcription nuclear factor HNF-1α [54–57]. It can alsoberelatedto mutationsinthe glucokinase gene, whichservesasa “glucose sensor”for the β-cell[58,59]. Due to the impairments in the glucokinase gene, higher plasma glucose levelsarerequiredto elicit normal insulin secretion[60].The critical factor distinguishing MODYfrom type1diabetesis the autoantibody negativity. Although GADA wasreported in 1% of individuals withMODY, the undetectable C-peptide concentration and lower HbA1c (GCK-MODY) may be determined[61,62]. Besides the genetic background, the features distinguishing MODY diabetes from T2D are the onset of the disease usually in the second or thirddecade of life, most often the absence of obesity, lower BMI, andthe predominance of insulin secretion defect in the absence of insulin resistance or even high insulin sensitivity[63]. 

2.4. Alzheimer’s Disease—Diabetes Mellitus (AD-DM) Relation and Type 3 Diabetes (T3D) 
Inrecent years,a signifcant increasein the incidenceof Alzheimer’s Diseaserelated to T2D, is observed. Patients with T2D are almost twice as likely to develop AD than patients who only have insulinresistance. T2D and Alzheimer’s Disease patients have similar β-amyloid depositsinthepancreasasinthebrain[64–66]. Severalresearchershave suggestedthatthisnew pathologyisreferredtoastype3diabetes(T3D) (Figure 5)[67]. Someof the targetreceptorsin T2D, e.g., IGF-1R,[68–70]PPAR,[71]IDE,[72,73], are also crucial regulators of tau protein expression and phosphorylation. For instance, it was reported that both hyperinsulinemia and IDE might be risk factors for Alzheimer’s disease[72,74]. 
The function of glucose transporter (GLUT) protein is controlled by the insulin-like growth factor (IGF) family, consisting of three ligands (insulin, IGF-1, and IGF-2), six IRs, and up to seven IGF-binding proteins (IGFBP1-7). IGF-1 and insulin can regulate neuronal excitability, metabolism, and survival through the insulin/IGF-1 signaling pathway. Few evidence on Alzheimer’s Disease patients’ brains showed a defcit ratio of insulin and resistancein IGF-1, suggesting thatAD mightbe diabetes type3[74]. Nevertheless, several studies also suggest theprotectiveroleofinsulin against apoptosis through various signaling pathways that suppress intracellular oxidative stress. For instance, the insulin/IGF/Akt pathway is considered to promote β-cell survival.[74]. 
The DYRK1A kinase is involved in molecular pathways relevant to human pancreatic β-cellproliferation,therebyprovidingapotential therapeutictargetfor β-cellsregeneration in T1D and T2D[75,76]. A further target of DYRK1A has been identifed as insulin receptor substrate-2 (IRS2)[77,78]. Moreover, hyperphosphorylation causedbyDYRK1A overexpression has been implicated in many pathogenetic changes attributed to brain diseases, particularlyin Down Syndrome and Alzheimer’s Disease[79,80]. 

Figure 5. Diagram illustrating the hallmarks of type III diabetes. Adopted and modifed from[80]. 


3. Molecular Basis of Diabetes 
3.1. Diabetic Kinome 
Protein kinasesarekeyregulatorsofsignal transduction pathwaysinmany physiolog
ical processes. Protein phosphorylation catalyzed by them is one of the major intracellular 
mechanisms of structural and enzymatic protein regulation. Reversible phosphoryla
tion/dephosphorylationisinvolvedin all physiological events, and its disruption can lead 
tomany pathological cases[81,82].Akinomeisasetofgenesforprotein kinasesinits 
genome. Serineandthreonine kinases contributeto insulinresistanceandthe development 
of diabetes (T2D)[83–85]. Kinases such as AMP-activated protein kinase (AMPK), Ikβ 
kinase(IKK),protein kinaseC(PKC),and mitogen-activatedproteinkinases(MAPKs)play 
importantrolesin the developmentof insulin sensitivity and insulinresistance[6,86–88]. 
Rho-associated coiled-coil-containing protein kinase (ROCK) and RNA-activated protein 
kinase(PKR)arealso involvedinthe pathogenesisof insulinresistance[89,90]. AMPK regulates lipid and glucose metabolism, therefore, this enzyme appears to be one of the main factorsresponsiblefor maintaining energy homeostasisinthebody. Activationof this protein leads to inhibition of anabolic pathways, and its dysregulation is one of the mech-anismsresponsible for insulinresistance-induced diabetes[91,92]. Thus, understanding the interplay between diabetes and protein kinome may help develop the targeted drug therapiesto minimize insulinresistance.Moreover,itmaybe criticalforthepreventionof diabetes. Therefore, the most crucial approach in discovering new drugs against diabetes is searching for pharmacological inhibitors of specifc kinases. Initially, the focus was mainly on tyrosine kinase inhibitors and cancer indications, but the feld is rapidly expanding towards serine/threonine kinases. 
Infammation, Diabetes, and Kinase Inhibition Infammation of pancreatic islets has emerged as a key contributor to the loss of functional β-cell mass in both T1D and T2D. In T1D, β-cells arethe target of an autoimmune assault. Chronic low-grade infammation and activation of the immune system are major factorsin obesity-induced insulinresistance and T2D[93,94]. Obesity is a strong antecedent of T2D, and both diseases are associated with adverse cardiovascular risk profles. Infammatory pathways have been suggested as the underlying unifying pathogenic mediators for excess weight, diabetes mellitus, and cardiovascular diseases. Chronic infammation is a common feature in the natural course of diabetes, and levels of infammatory biomarkers (secreted mainly by adipocytes) correlate with prevalent and incident diabetes and major complications and cardiovascular diseases in particular[93,94]. The developmentof insulinresistanceis also associated with low-grade tissue-specifc infammatoryresponses inducedby variouspro-infammatory and/or oxidative stress mediators, notably pro-infammatory cytokines such as IL-1β, IL-6, TNF-α, several chemokines, and adipocytokines. Chronic exposure of pro-infammatory mediators stimulates cytokine-signalingproteins, which ultimately blocksinsulin signalingreceptors’ activation in β-cells of pancreatic islets[95]. Some of the protein kinases are directly involved in these infammatory processes that underlie and accompany theprogressionofDM and its complications[95]. For instance, Iκβ kinase β (IKKβ),a central coordinatorof infammatoryresponses through activation of NF-κB, has been implicated as a critical molecular link between infammation and metabolic disorders[96]. Phosphorylationby IKKβ targetsIκBα to degrade proteasome that liberates NF-κB for translocation from the cytoplasm into the nucleus to promote expressionof numeroustarget genesand consequently induce insulinresistance[74].Xuet al. identifed inhibitorsof noncanonicalIκβ kinases (IKKs),TANK-binding kinase1(TBK1), andIκBkinaseε (IKKε), as enhancers of β-cellregeneration[97].In theprogressionof T1D andT2D,acommon featureisdecreasing β-cell massbycytokine-and/or glucolipotoxicityinduced apoptosis. Thus, prevention of β-cell loss by diabetic kinome inhibition can be an alternative approach for increasing β-cell mass in diabetes[97]. 


4. DYRK Family of Protein Kinases 
Among the 518 human kinases, dual-specifcity tyrosine phosphorylation-regulated kinase 1A (DYRK1A) is a conserved eukaryotic serine/threonine protein kinase. Other kinases belonging to the DYRKs family areDYRK1B, DYRK2, DYRK3, and DYRK4. DYRKs are from the CMGC group, which also includes other kinases: CDKs (cyclin-dependent kinases), CDKLs (CDK-like kinases), CK2 (casein kinase 2), CLKs (CDC-like kinases), GSKs (glycogen synthase kinases), and MAPKs (mitogen-activated protein kinases). Among them, CDKs, CKs, and MAPK have been well investigated in their functions in transcription, DNA damagerepair,protein degradation, and neurogenesis[98,99]. However, DYRKs and CLKs in the signaling pathways remain not completely understood. DYRKs isoforms are subdivided into two classes based on their subcellular localization. DYRK1A/B belonging toclass1arefoundinnuclei,whereasthoseofclass2prefer cytoplasmic localization.They all possess a kinase domain[100]. 
4.1. DYRKs Activity and Regulation 
Many protein kinases can adopt an active and inactive conformation. The transition between these conformations is regulated by the reversible phosphorylation of discrete serine,threonine,ortyrosineresiduesinthe ‘activationloop’[101].The DYRKs activation is dependent on the phosphorylation of a conserved tyrosine residue in the activation loop. The phosphorylatedtyrosine formssalt bridgeswithtwoargininesintheP +1 loop[102,103]. In DYRK1A, pY321 is important in the same interactions with two arginines (R325, R328) (Figure 6)[98,99,102–104]. 

Figure 6. The DYRK1A kinase domain and DYRK homology box with the inhibitor DJM2005 bound in theATP binding site. Magenta—theDH box and CMGC, orange—the activation segment(left). Ribbonrepresentationof the kinase domain, with one orientation,rotated90◦ around the depicted axisrelativetothe other(PDBentry2VX3)(right)[104]. 
DYRK possesses dual specifcity, as it can autophosphorylate tyrosine Y321 in the acti-vation loop and phosphorylate its substrates on either serine or threonineresidues[103,105]. While dual MAPK phosphorylation is the primary process of upstream kinase regulation, tyrosine phosphorylationof DYRK and GSK3 occurs via autophosphorylation[106]. Ac-tivated DYRKs phosphorylate their substrates only on serine or threonine residues and cannot rephosphorylate on tyrosine. Therefore, a translational intermediate folding of DYRKs with different biochemical properties and tyrosine phosphorylation ability has been proposed[107].Ithasalsobeen suggestedthatthedual specifcityof DYRKsis associated with dual sensitivity to kinase inhibitors[108].Tyrosine phosphorylation during DYRKs activationisrequiredto switchthe conformation,butitdoesnot maintainthis state.For this purpose, the stabilizing effect of salt bridges formed between phosphotyrosine and the twoargininesin theP+1loop mayplaya crucialrole[102]. 

4.2. DYRK1A Expression and Its Role in Neurological Diseases 
DYRK1A is a dosage-sensitive gene, and the imbalance in its expression affects brain structure and function[109]. It was reported that DYRK1A defciency might lead to autosomal dominant mentalretardation[109]. Therefore, both low and high DYRK1A expression can participatein developing several disorders[110]. DYRK1A expressionis regulatedby transcriptional factors, tumor suppressors, neurogenic factors, andprotein-protein interactions. Reducedexpressionoftherepressor complexAP4resultsinpremature overexpressionof DYRK1Ainthefetalbrain[111].Itwasalsoshownthatthe β-amyloid peptide increases DYRK1A mRNA SH-SY5Y cells[112]. 
Overexpression of another transcription factor, E2F1, enhanced DYRK1A activity by increasing its mRNA level in phoenix cells. Thus, DYRK1A may also be involved in cell-cycleregulation[113,114].The DYRKsarekeyproteinsregulatingNFAT1 phosphorylation[115,116]. Overdosage of DYRK1A associated with the DSCR1 gene (resident of the “Downsyndrome candidateregion”andasa shockorstressgene) wasreportedto diminishNFATc activityin the immuneresponse[117]. Protein p53‚a 345 well-known tumor suppressor gene—has been identifed to reduce DYRK1A expression. This process is mediated through the induction of miR-1246, resulting in the nuclear retention of NFATc1 and inductionof apoptosis (overexpressionof miR-1246reduces DYRK1A levels)[118]. There is also evidence that upregulation of DYRK1A leads to changes in neuronal proliferation in 
Down Syndrome[119]. 
The WDR68protein (also called HAN11, DCAF7) may act asaregulatory subunitof DYRK1A and DYRK1B[120,121]. Its overexpression inhibited the DYRK1A stimulation of GLI1-dependentreportergene activity[122].Thecircadianchangesin DYRK1A levelshave been reported, and DYRK1A was identifed as a molecular clock component leading to CRY2 degradation[123]. Recently, SPRED1and SPRED2(sprouty-relatedproteinwith an EVH1domain)werefoundtointeractwiththe catalytic domainofDYRK1A,leadingtothe inhibitionof phosphorylationof tau andSTAT3[124]. Thus, the DYRK1A-STATpathway is involvedinDS development. Phosphorylationby DYRK1Aattau Thr212residue primes tau phosphorylation by GSK3 at the Ser208 residue, resulting in increased neurofbrillary accumulation tangles existsin the brainof Alzheimer’s Disease patients[125,126]. 
DYRK1A plays an important role in cytoplasm homeostasis by localizing in the nucleus, as evidencedby increased immunoreactivityin this area[125]. The importanceof DYRK1Ain several biologicalprocessesis summarizedin (Figure 7). 

Figure 7. DYRK1A phosphorylation targets. Dual specifcity tyrosine-phosphorylation-regulated kinase 1A (DYRK1A) is encoded by Hsa21 and phosphorylates multiple targets that play roles in various biological processes. Adopted and modifed from[127]. 

4.3. DYRK1A Expression Affects Mechanisms of Diabetes 
DYRK1A has been found to affect multiple signaling processes in DM context by acti-vating/inactivating signalsof transcriptional and translation factors (RNAPII CTD[128], Sprouty2[129], DREAM complex[130], CREB[131], FKHR[132]), splicing factors(regulation of Cyclin D1 turnover as well as miscellaneous proteins including caspase-9[109,133,134], Notch[135], as well as glycogen synthase. It was shown that DYRK1Ais involvedinGSK3 phosphorylationatthe Ser640residue[136]. This interac<BR>tion subsequently causes the activation of glycogen synthase, a key enzyme in glycogen synthesisregulatedby insulin (Figure 8)[136,137]. DYRK1Aisan important kinasefor β-cellgrowth[138]. Studies using DYRK1A haploinsuffcient mice have confrmed that they are burdened with severe glucose intolerance, reduced β-cell mass, and proliferation, leading to diabetes. Upregulation of DYRK1A in β-cells was found to enhance this phenomenon signifcantly[11,30]. DYRK1A emergedin thedrugdiscovery feld as oneof the most attractive therapeutic targets for developing selective inhibitors as new drugs. They may have a high therapeutic potential for diabetes. The involvement of DYRK1A in the molecular pathways of different diseases is well described (see above). Therefore, we have focused on the new DYRK1A inhibitors discovered or specifcally developed to provide the basis for the future development of these promising drugs. 

Figure 8. Scheme illustrating DYRK1A substrates and predicted roles in biological processes. Adopted and modifed from[139]. 


5. CurrentTreatmentsof Diabetes 
Pharmacological treatment of DMis based on the following strategies: (i) insulin infusion; (ii) administration of drugs that increase insulin secretion (sulfonylureas, meglitinides); (iii) enhancement of insulin sensitivity (metformin, thiazolidinediones); (iv) prevention of glucagon synthesis (DPP-4 inhibitors and GLP-1 receptor antagonists; and 
(v) application of substances that increase glucose excretion (SGLT-2 inhibitors)[140]. These strategiesofferreasonable controlof disease symptoms. However, thereis simply no therapy to help DM patients to return to euglycemia. Thus, the focus on DM treatment’s development shifted towardpopulation of functional, insulin-producing β-cells. It allows the patient to achieve insulin homeostasis and relieve hyperglycemia. One of the most promising advances in diabetes therapy is enabling β-cells to replicate. Several classes of drugs, hormones, or growth factors such as PPARg agonists, GLP-1 agonists, DPP-4 inhibitors, GSK3β inhibitors, prolactin, IGF-1, HGF, and PTHRP have been tested for their ability to stimulate β-cellproliferation[141,142]. However, all theproposed approaches failed in inducing β-cell proliferation in clinical conditions. Nevertheless, in the majority of treated diabetic patients, some β-cells were able to survive after the treatments listed above. In view of these considerations, it seems reasonable that modifcations of current drugs or new appropriately designed small molecules or molecular targets could lead to β-cells proliferation/restoration. Under physiological conditions, human β-cells replicate at a low rate, about 2% per day, and only in the frst few years of life. So far, attempts to stimulate adult human β-cells to replicate have failed pharmacologically. Nevertheless, it changed in 2015 with the discoveryof harmine and other DYRK1A inhibitors, discussed below[16]. 
Harmine,a well-known DYRK1A kinase inhibitor, was identifedbyWanget al., throughahighthroughputscreening(HTS) campaign.It inducesamildlevelofc-Mycpro-tein expression in rodent islets. The mechanism of action involves inhibiting the DYRK1A kinase (likely a primary target of harmine), which allows the NFATpathway to induce c-Mycexpression[16].Moreover,it servesasthe terminatorofNFATdephosphorylation byrephosphorylatingNFATandactsasabrakeonthecellcycle[142]. SeveralDYRK1A inhibitors were identifed among other known protein kinases inhibitors, and a few are used as tool compounds for β-cell regeneration[138]. 
5.1. DYRK1A Inhibitors for β-Cell Function Restoration 
5.1.1. Harmine and Its Analogues—SAR Approach Screening of a panel of 69 kinases identifed harmine as a potent DYRK1A inhibitor[143]. Comparative invitro assaysrevealed that harmineis moderately spe-cifc towards DYRK1A[144]. The IC50 for DYRK1A hasreached33 nM. The DYRK1B showed an IC50 of 166 nM, and the other distant members DYRK2, DYRK3, and DYRK4 indicate IC50 values as following: 1.9 µM, 0.8 µM, and 80 µM[144]. Additionally, cell culture assays confrmed the potency of harmine for DYRK1A and its lack of toxicity. The DYRK1A⁄harmine complex’s crystal structure showed harmine blocking theATP-binding pocket and interacting with the backbone NH of methionine on position 240 of the hinge-region, as well as with the conserved lysine on position 188, by forming two hydrogen bonds (Figure 9)[145]. Furthermore, the DYRK1A/harmine complex structure suggests thatthe accessible volumeoftheATP-binding pocket can accommodate substituentsat 
the β-carbolinestructure[102]. Consequently, harminecanlikelybe modifedintoan even more potent and selective DYRK1A inhibitor. 

Figure 9. DYRK1A/harmine complex. DYRK1AATP-binding pocket with harmine (PDB: 4YU2). 
Although several new DYRK1A inhibitors have been identifed and described so far, none meet the selectivity standardsrequired for kinase-targetedprobe molecules. Harmine has been recognized as a potent monoamine oxidase (MAO) inhibitor, which is associated with a number of side effects. Due to the limited selectivity of harmine, its derivative AnnH75 (Figure 10)was developed, which, unlike harmine, does not interact with MAO while maintaining DYRK1A inhibition. Epigallocatechin gallate (EGCG) from green tea has also been shown to be a DYRK1A inhibitor. 

In 2018 an integrated approach to investigate the structure-activity relationship of harmine derivatives for diabetes management (DYRK1A activity and β-cell proliferation) was developed[15].Structure-baseddrugdesignand developmentwereusedto identify kinome and CNS off-targets and harmine-like molecules for more specifc therapy. The crystal structure of DYRK1A withATP-binding inhibitor DJM2005, 1, 7, and 9-amino harmine analogs were synthesized and examined in terms of their effect on DYRK1A binding and β-cellproliferation (Figure 11andFigure 12)[15,147,148]. 


Harmine analogs with polar substituents, e.g., hydroxymethyl or -acetyl, at position 1-C (Figure 12),showed good DYRK1A inhibition (IC50 49-67 nM). However, the 1-hydroxy moiety negatively impacted DYRK1A inhibition. The presence of a halogen atom at the position 1-C signifcantly increased the inhibition potency, making the 1-chloro substituted analog the most potent DYRK1A inhibitor, with an IC50 of 8.8 nM. Among synthesized 8 compounds with IC50 < 250 nM against DYRK1A, only four affected human β-cell proliferation. In contrast, the 1-amino analogs indicated no effect on β-cell proliferation. Notably, 1-and 3-hydroxymethyl compounds were most effective in vitro, indicating that these modifcations improve the β-cell proliferation and potentially increase the selectivity towardDYRK1A[15]. 
In the subsequent paper, the same authors reported the set of harmine derivatives modifed in 7-position as DYRK1A inhibitors with activity on human β-cell proliferation and targeteddrug delivery[147]. The harmine backbone was substitutedby terminal methyl ester, carboxamide, carboxylic acid, and amino/substituted amino groups with various carbon (1-5) chain lengths. Biochemical assays allow the selection of two 7-O analogs with activity towardDYRK1A (>100 nM). These compounds indicated an increase in β-cell proliferation (3-fold less than harmine). The reduced (in comparison to harmine) effcacy of described 7-O derivatives may be caused by several structural and biological factors, including lower potency for DYRK1A inhibition, limited cell permeability, reduced DYRK1A targeting and/oroff-target kinase activity[147]. 
9-N-substituted analogues of harmine were also studied in order to eliminate the disadvantagesof kinase andoff-target[148]. The libraryof62 compounds was tested, and among them, 4-(7-methoxy-1-methyl-β-carbolin-9-yl)butanamide proved to be the most promising DYRK1A inhibitor. The compound was tested in vivo and was signifcantly more effective than bare harmine (Figure 13). After treatment with 4-(7-methoxy-1-methyl-βcarbolin-9-yl)butanamide, Ki67 expression was increased in C57 mouse and human β-cells (Figure 13). In the PPX model, faster regeneration of β-cells was observed at a 10-fold lowerdrug dose than harmine. Similarresults were also obtainedin the NOD-SCID mouse model with transplanted human islets. Furthermore, no CNS side effects were observed at the dose of 30 mg/kg. Thus, this compound was selected as the lead candidate with high in vivo effcacy. Noteworthily identifed inhibitor is also characterized by improved selectivity and CNS off-targets and superior activity in β-cells restoration—crucial for the treatmentof diabetes[148]. The studies described above, the successof which has been confrmed in several animal models, demonstrate the validity of the modifcations used within the harmine structure. This research direction is defnitely worth continuing, but large-scale clinical trialswillonlyprovide answersto whetherthemosteffective compound of this family will be equally effective in clinical treatment. 

5.1.2. Perha Pharmaceutics Inhibitors Despitethe therapeutic potentialof DYRK1A inhibitors,onlyafewofthemhavebeen 
well-characterizedtodatein termsof selectivityand biologicaleffects[149]. These include a series of (i) pyrazolidine-3,5-dione derivatives, (ii) 6-arylquinazolin-5-amines, (iii) the βcarboline alkaloid harmine, (iv) the green tea polyphenol epigallocatechin-3-gallate, (v) the benzothiazole INDY, (vi) bauerineCderivatives, and (vii) leucettines[150]—agroupof aminoimidazolinones derived from the marine sponge natural product leucettamine L41 was shown to be a most promising kinase inhibitor (Figure 14). The molecular interac<BR>tions of leucettine L41 with its targets and its neuroprotective properties were extensively studied.[150]leucettineL41(anATP-competitive inhibitorof DYRKsandCLKs)mayalso interact withGSK3β and CK2. Moreover, it causes cellular effects, including pre-mRNA splicing, HT22 hippocampal cells protection cell death. Furthermore, it may induce au-tophagy and inhibit tau phosphorylation[151]. It wasrecentlyreported that leucettine L41 could prevent DYRK1A proteolysis, inhibit STAT3α phosphorylation, and reduce pro-infammatory cytokine secretion (IL1-β, TNF-α, and IL-12) in APP/PS1 mice model. These results confrm the role of DYRK1A proteolysis in Alzheimer’s disease (AD) and suggesta possible mechanismasa noveltargetto counteractthe disease[152]. 

Another approach for discovering DYRK1A inhibitors was screening a library of plant and fungal extracts.[153]. Several compounds were identifed, including harmine, anthraquinone emodine and several favonoids. These molecules were isolated and characterized as the active constituents from four plant extracts. However, due to the moderate activity of selected anthraquinone and favonoids, the potential for further development is limited.In particular, favonoidsare knowntobeverypromiscuous kinase inhibitors[153]. Lamellarins are natural marine products isolated from mollusks, ascidians, and sponges. LamellarinDdisplays broad-spectrum kinase inhibition (i.e., CDK1, CK5, GSK3, PIM1, and DYRK1A) in the sub-nanomolar range. It is also toxic to cancer cells due to strong topoisomeraseIinhibition. LamellarinsBandDdiffer onlyin theOH and OMegroups’ number and position on a common pyrrolo(2,1-a)isoquinoline scaffold (Figure15). 

The synthetic model for modulation of lamellarins’ activity has been developed[154]. Based on the natural structure’s fne-tuning, it was possible to eliminate topoisomerase affnity and cytotoxicity while retaining the kinase inhibition (Figure 16). The pyrrole moiety wasreplaced with an indole skeleton and designing new chromeno[3,4-b]indoles. The otherparts of lamellarinD are unchanged (C, B, A,) as are the substituents most strongly interactingwiththeAring,i.e.,OHand-OCH3groups.Asaresultofthepresence ofahydroxylgroupat positionC-2, topoisomerase inhibitionis lost. Interestingly, selective inhibitionof DYRK1A was observed simultaneously.Without any other substituent or the addition of a hydroxyl group in C-10, two derivatives (4-hydroxychromeno(3,4-b)indol6(7H)-one and 3-hydroxychromeno(3,4-b)indol-6(7H)-one) were selected with IC50 = 74 and76 nM,respectively[154]. 

Similarly,DYRK1Ainhibitorscomprising:(i)meridianines,(ii)indirubin50-carboxylates, (iii) thiazolo[5,4-f]quinazolines, (iv) pyrido[2,3-d]pyrimidines, (v) 3,5-diaryl-7-azaindoles (DANDYs), (vi) KH-CB19, (vii) 2,4-disubstituted thiophenes, and (viii) hydroxybenzothiophenes are tested not only towards DYRK1A selectivity but, additionally, towards structurally closelyrelated kinase isoforms[155]. The Meijergroup has been intensively in-vestigating the C, N, S-or C, N, O-containing heterocycles representing precursors of biologically important molecules able to alter the kinases activity. One of the most promising compounds from this group is 8H-thiazolo[5,4-f ]quinazolin-9-ones, with micromolar DYRK1A inhibitory potency (Figure 17). 

Benzo-, pyrido-and pyrazinothieno[3,2-d]pyrimidines derivatives as DYRK1A in-hibitors were also investigated. Thiazolo[5,4-f]quinazoline scaffolds also indicate the potential for DYRK1A inhibition. Among the compounds of this library, methyl 9-(4-methoxyphenylamino)thiazolo[5,4-f]quinazoline-2-carbimidate, methyl 9(benzo[d][1,3]dioxol-5-ylamino)thiazolo[5,4-f]quinazoline-2-carbimidate, and methyl 9-(4bromo-2-fuorophenylamino)thiazolo[5,4-f]quinazoline-2-carbimidate inhibited DYRK1A withIC50at nanomolar values(40,47and50nM,respectively)(Figure 18)[156,157]. 

It was also reported that the other derivative, the methyl 9-anilinothiazolo[5,4f ]quinazoline-2-carbimidate (EHT 5372), inhibits DYRK1A and DYRK1B at subnanomolar concentrations with IC50 = 0.22 nM for DYRK1A and 0.28 nMfor DYRK1B, respectively). EHT 5372 and its derivatives are one most potent DYRK1A inhibitors reported so far, with high selectivity towardDYRK1A compared to other kinases of the CMGC group (Figure 19). EHT 5372 also inhibits cellular DYRK1A-mediatedtau phosphorylation andAβ production. However, it indicates signifcantly lower potency with IC50 1.06–1.17 µM[156,157]. 

Another compound belonging to the DYRK1A inhibitor class and characterized by nanomolar IC50 values is 8-cyclopropyl-2(pyridin-3-yl)thiazolo[5,4-f ]quinazolin-9(8H)-one (also called FC162,Figure 20)[158]. FC162has emergedasthe mostpromising candidate based on in vitro cell studies than well-characterized DYRK1A inhibitors (e.g., leucettine 41 and EHT1610). It was reported that FC162 could cross the BBB and is effective in Thr212 phosphorylation[158]. 

In further studies, the activity of FC162 on tau-4R cells (SH-SY5Y cells which overexpressing the four-repeat human tau isoform) was examined. The results indicated a dose-dependent inhibition of tau phosphorylation at Thr212. Moreover, the decrease in cyclin D3 phosphorylation at Thr283 was observed in murine pre-β-cells. After long-term treatment of FC162, a decreased G0 cell population was observed. Thus, these data re-veal that FC162 phenocopies the effect of Dyrk1a genetic deletion[158]. Moreover, the following compound from this group—10-iodo-11H-indolo[3,2-c]quinoline-6-carboxylic acid (KuFal194)[155]also indicated anin vitro activity against DYRK1A withIC50 =6nM and considerable selectivity in comparison to DYRK1B and CLK1. Nevertheless, due to the low water solubility ofKuFal194, further in vitro and in vivo studies should be performed with caution. It seems reasonable to use appropriate formulations that not only solve the problem of lipophilicity but, perhaps, by increasing stability, also improve other important parameters. 
The SAR evaluation of KuFal194 derivatives and kinome selectivity analysis including DYRK1A and CMGC protein kinases: CDK1/cyclin B, CDK2/cyclin A, CDK5/p25, CK1, GSK-3, and ERK2 were performed. Substituents in the 8-position eliminated the DYRK1A inhibitory activity, suggesting steric exclusionfrom theATP-binding pocket(Figure 21). Moderate, selective inhibitorsof GSK3 werealso obtainedbyadding polar H-bond acceptor substituents in 8-position. These showed no activity against DYRK1A. Strikingly, the 10chloroderivative (10-chloro-11H-indolo[3,2-c]quinoline-6-carboxylic acid) showed two-fold higher DYRK1A inhibitory potency (IC50 = 31 nM) than 11H-indolo[3,2-c]quinoline-6carboxylic acid without inhibiting other kinases[155]. 

Inorderto enhancethe physicochemicalpropertiesof KuFal194,asetof [b]-annulated chloro-substituted indoles were designed and developed. Compared to the iodine atom, the main rationale was that chlorine decreases the molar mass and lipophilicity and diminishes the overall toxicity[159]. Theresultsof kinase inhibition studies performed using proper bioassays have revealed that most of the tested compounds, except Mannich base, act as DYRK1A inhibitors with micromolar or even sub-micromolar concentrations applied. Compared to KuFal194, these novel compounds were less active and non-selective towards CLK1[158]. 4-chlorocyclohepta[b]indol-10(5H)-one was identifed asa novel dual DYRK1A/CLK1 inhibitor with slightly better solubility. X-ray structure analysis confrmed the binding mode of this compound to DYRK1A, exploiting mainly shape complementarity for tight-binding (Figure 22)[158]. 

In summary, inhibitors such as harmine, INDY, and leucine L41 have shown some promise in cellular assays due to their signifcant DYRK1A inhibitory activity. Results obtained againstrelated kinases were no longer aspromising,indicating their low selectivity. On the other hand, KuFal194 and EHT 5372 are characterized by proper selectivity against DYRK1A, but their use in in vivo studies is still limited due to high lipophilicity (Figure 23)[154]. Therefore, furtherdesignofimproved water-soluble derivativesorthe use of appropriate formulations is required. 

Ahalogen-substituted indole group was chosen to develop morehydrophilic DYRK1A inhibitors with reduced molecular weight. The received fragment served as a template to design and develop a series of substituted indole-3-carbonitriles with inhibitory properties against CMGC kinases[160]. Computational studies indicated that the halogen substituents,ata7positionoftheindolering,aremostlikelyto interactwiththehinge regionbya water-mediatedhalogen bond[160].Ata2positionofthe indole core,only aromatic or lipophilic residues were tolerated. The 2-phenyl-substituted derivative (7Iodo-2-phenyl-1H-indole-3-carbonitrile) was the most potent inhibitor of the series (IC50 against DYRK1A at 10 nM) and DYRK1A-mediated phosphorylation of SF3B1 in HeLa cells (IC50 =320nM) (Figure 24)[160]. However,itresultedinonlylow selectivitytorelated kinasesof the CMGCgroup and poor aqueous solubility.To increase the solubilityof the compounds,hydrophilicor aliphaticresiduesata2position were introduced.Byreplacing the 2-phenyl substituent with pyridin-3-yl or cyclopentyl residues, the reduction of logP value and increased solubility were obtained, while the DYRK1A activity was only slightly affected. Further modifcations of the 7-halogenindole-3-carbonitrile parent structure are underway to develop potent, highly selective, and water-soluble DYRK1A inhibitors[160]. 

The tetracyclicV-shaped pyridine-, pyrazine-or indole-containing compoundsrepresent the next set of molecules that target DYRK1A in the nanomolar range[12,161]. Inspired by the 6,5,6-fused tricyclic skeleton of harmine molecule, Meijer and Besson synthesized and characterized a series of N-aryl-7-methoxybenzo[b]furo[3,2-d]pyrimidin-4-amines and their N-arylbenzo[b]thieno [3,2-d]pyrimidine analogs substituted in position 4of the pyrimidine ring by an aromatic amine (Figure25). The kinase inhibition evalu<BR>ation was carried out on Ser/Thr kinases, including CDK5, GSK3, DYRK1A, CLK1, and CK1. The benzothieno[3,2-d]pyrimidines derivatives-N-(2,3-dihydrobenzo[b][1,4]dioxin-6yl)-7-methoxybenzothieno[3,2-d]pyrimidin-4-amine and N1-(7-methoxybenzothieno[3,2d]pyrimidin-4-yl)-N4,N4-dimethylbenzene-1,3-diamine were found as a most active in-hibitors submicromolar IC50 value and selectivity towards DYRK1A and CLK1 (0.5 and 
0.68 nM, 0.7 and 0.66 for each inhibitor)[162]. 

Another study by Dehbi et al. reported the synthesis and biological evaluation of 4,7-disubstituted pyrido[3,2-d]pyrimidines. Using the SAR approach, the authors indicated that some of these compounds could selectively inhibit DYRK1A and CDK5 without affecting GSK3. The most active compound was 4-[7-(5-methyl-thiophene-3-yl)-pyrido[3,2d]pyrimidin-2-yl]-phenol, which exhibited IC50: 110 nM for CDK5, 24 nM for DYRK1A and only 1.2 µMfor GSK3. In the C-7 amino derivatives, the best was indubitablycompound 1-[2-(4-hydroxyphenyl)pyrido[3,2-d]pyrimidin-7-yl]piperidin-2-one with IC50 = 60 nM against DYRK1A (Figure 26)[163]. These molecules have been tailored as dual selective DYRK1A and CDK5 kinase inhibitors involved in regulation processes, including CNS-related disorders like diabetic neuropathy[164]. 

Bruel et al. attempt to design new kinase inhibitors focused on a pyridazino[4,5b]indol-4-one scaffold and found an inhibitor with micromolar IC50 (5 µM) against DYRK1A (Figure 27)[165]. Due to its structural analogy with harmine, further optimization was performed. The pyridazino[4,5-b]indol-4-one series, the furan-2-yl-substituted derivative, was selected as a compound with submicromolar IC50 (0.22 µM) against DYRK1A. It was only four-fold less active than harmine (0.06 µM) and indicated no activity towards the other kinases. The mechanism of its activity was explained theoretically. Based on the presented docking model, the authors suggested differences in its affnity towards harmine. Harmine interacts with the Leu241 residue via hydrogen bonding and the presented inhibitor probablybind to the pyridazinone ring (backbone atoms of Glu239 and Leu241). Furthermore, selectivity to CDK5 and GSK3 kinases may be due to an additional hydrogen bonding interaction between a methoxyl group and an aspartate residue (Asn244) located in the kinase pocket (Asp86in CDK5; Thr138in GSK3)[165]. 

A systematic in vitro evaluation of 2500 plant extracts from New Caledonia and fromFrench Guyana was performed in another work. Aristolactams and lignan derivatives were purifed from Oxandra asbeckii and Goniothalamus dumontetii. Porphine alkaloids were isolated from Siparuna pachyantha, S. decipiens, S. guianensis, and S. poeppigii. Among these compounds, velutinam, aristolactam AIIIA, and medioresinol showed submicromolar IC50 values on DYRK1A (Figure 28)[166]. 

5.1.3. Azaindoles Azaindoles are structurally related to indoles, widely present in natural products and pharmaceuticals (Figure 29). Azaindole molecules appear to inhibit kinases than other 
targets preferentially. Moreover, their biological/pharmacological features are benefcial to treat many diseases, includingDM[167]. 

With some azaindoles being successfully developed as antidiabetic drugs, the 6azaindole and 7-azaindole derivatives have also been tested as DYRK1A inhibitors. For instance, the 3,5-diaryl-7-azaindole derivative, also called DANDY, represents one of the most potent inhibitors(IC50 =3nM)of DYRK1A. Besides DANDY, numerous DYRK1A inhibitor scaffolds have beenreported (Figure 30)[168]. 
Basedon moleculardockingstudyattheATP-bindingsite,itwas demonstratedthat there were multiple H-bond interactions with the peptide backbone (Glu239, Leu241) for the 7-azaindolecore, and the hydroxyl substituents interacted withLys188 and Ileu165. The hydroxyl derivatives showed moreremarkable activitythan their methoxy derivatives[168]. Moreover, it was indicated that 6-azaindole derivatives were considerably less active than the 7-azaindole ones. Interestingly, when these derivatives were tested against a representative kinase panel, a relative selectivity appeared with compounds acting mainly on the DYRK1A family[169]. 
The SAR study of the azaindoles shows that the nitrogen at the 7-position is indispensable, asreplacing the azaindole ring with the indole ring led to an inactive compound[170]. Methylation of the nitrogen at the N1-position of 7-azaindole indicates a similar effect. Thus, it suggests that the azaindole’s NH belongs to the pharmacophore. Furthermore, the additionofnitrogentothe azaindoleringledtoaless active molecule.Moreover,additional substitutionatthe2positionresultsinasignifcantdecreaseinits activity[170].It appeared that the 7-azaindole core was indeed critical for the strong protective effect effect of INS-1E cellsintheCK assay. Thus, 5-(3,4-difuorophenyl)-3-(pyrazole-4-yl)-7-azaindole (GNF3809) was selected for both, ex vivo and in vivo proof-of-concept effcacy studies (Figure 31)and demonstrated protective effects of β-cells. Future efforts directed at further optimization of GNF3809 and the elucidation of its molecular mechanism of action hold the substantial potentialtoaddressthe unmet medical needsofT1D patients[170]. 


5.1.4. Aminopyrazines The aminopyrazine scaffold was identifedfromaphenotypic high-throughput screening campaign measuring β-cell proliferation using mouse R7T1 β-cells. Lead optimization results in identifying a promising dual DYRK1A and GSK3β inhibitor aminopyrazine GNF4877 (Figure 32)[171]. Primingof GSK3β substrates by DYRK1A has linked the for-mer kinase and diabetes. The implication of these kinases in β-cell proliferation has been demonstrated via several screening tests and biological activity experiments. GSK3B action leads to NFATnuclear localization. Inhibition of GSK3β is required for β-cell proliferation. 
The inhibition of DYRK1A may stimulate the NFATsignaling, which infuences the β-cell proliferation. 

SAR studies on the aminopyrazine scaffold targeted the enzymatic inhibition of DYRK1Ausingastructure-directedapproach. GNF4877isan inhibitornotonlyof DYRK1A but also of GSK3β. It affects the proliferation of β-cells in both in vitro and in vivo con-ditions. Nevertheless, inhibition of GSK3β may also lead to the appearance of some side effects. For thisreason, GNF4877 was not selected for further clinical trials. Nevertheless, preclinical studies of this series of compounds have established a solid ground for the discovery of the next generation of selective DYRK1A inhibitors. 
Aminopyrazine compounds (GNF series) were designed and developed to increase β-cell proliferation in adult primary islets. Oral administration of these compounds to diabetic mice induced β-cell proliferation and increased insulin content and consequently improved glycemic control. Biochemical, genetic, and in vitro studies confrmed that DYRK1 affects β-cell proliferation induced by GNF7156. Furthermore, dual-inhibition of DYRK1A and GSK3β increased β-cellproliferation (Figure 33)[171]. 

However, GSK3β regulates various cellular processes, including behavior, immu-nity, and circadian rhythm. Its inhibition may also activate other pathways and lead to undesired side effects. Less than a year ago, research into optimizing the structure and function of aminopyrazine led to the discovery and development of GNF4877 as a dual-function DYRK1Aand GSK3β inhibitor of β-cell proliferation. Notably, compared to previously reported derivatives, this dual-mode agent was already active in nanomolar concentrations[171]. Another 6-azaindole derivative named GNF2133 has been developed as a DYRK1A inhibitor and has been shown to promote β-cell proliferation and restore its function (Figure 34). It was reported that the 6-azaindole was the most promising in DYRK1A inhibition and selectivity over GSK3β inhibition. It demonstrated signifcant dose-dependentglucose disposal functionand insulin secretioninresponseto glucosepotentiates arginine-induced insulin secretion (GPAIS) in rat insulin promoter and diphtheria toxinA(RIP-DTA) mice. Therefore,it shouldbe concluded thatitis an up-and-coming candidateforthetreatmentoftypeIdiabetes[172]. 

Three novel compounds, GNF-9228, GNF-4088, and GNF-1346 (Figure 35), effectively stimulated β-cellproliferation,but not the expressionof homeobox genes NKX6.1 or VGF, were described[173]. Subsequent studies demonstrated several salutary effects of the VGF prohormone and its encoded peptides, such as TLQP-21, on β-cell survival and function[173–175]. The mostpromising, GNF-9228, selectively activates human β-cell relative to α-cell proliferation and does not affect δ-cellreplication[173]. GNF-9228 stimulates proliferation by a mechanism distinct from DYRK1A inhibitors because DYRK1A overexpression does not infuence it and does not activate NFATtranslocation[173]. In conclusion, a small molecule with pleiotropic positive effects on islet biology was characterized, including stimulation of human β-cell proliferation and insulin secretion andprotection against multiple agentsof cytotoxic stress[173]. 

5.1.5. AC Inhibitors Asetof DYRK1A inhibitors were identifedby employing KINOMEscan[176]screening. These compounds, designated as AC, represent six different chemical scaffolds[177]. Compounds 12 and 15 share a 3-(3-pyridin-3-yl-1H-pyrrolo[2,3-b]pyridin-5yl)phenyl core with sulfonamide (para) or amine (meta) substituents on the terminal arene. Compounds 24, 25, and 28 are 4-[4-amino-2-[2-methoxy-4-(4-methylpiperazin1-yl)anilino]-1,3-thiazole-5-carbonyl]phenyl derivatives (Figure 36). Compound 24 constitutes the core scaffold, whereas25and28 include terminal acrylamidefunctionsaddedtothe terminalareneat para and meta positions. Compound 28 lacks a methoxy substitution on the central phenyl 
ring. Compound 20 represents the other core scaffolds with 7-azathiazole, compound 22 as a pyrazine,23 as an alkaloid, and27 asa substituted 1,6-phenanthroline[177]. 

Selected compounds compriseabroad spectrumofbiological activity towards DYRK1A kinase:from littletostrong inhibition, measuredbyremaining activity (70–100%upto<5%). The measured Ki of the inhibition of the phosphorylation of DYRKtide (peptide RRRFRPASPLRGPPK) shows the variation among the compounds, with preservation of the chemical differences between the scaffolds[177]. 
Compounds 23 and 27 showed the highest activityin cellular assays at concentrations signifcantly lower than harmine. Comparing the activity of these two inhibitors with harmine, a 5-fold and 50-fold increase in activity was observed for 23 and 27 at concentrations of1 µMand 0.1µM, respectively. Excessive increase in the dose of the inhibitor in the cells leadstoa decreasein activity, whichmay indicatea toxiceffect, while dose-dependent inhibitionis observedfor harmineat these concentrations[177]. 
Moreover, this diverse set of scaffolds revealed the ability to prevent tau phosphorylation. Some of the inhibitors were co-crystallized with DYRK1A (12, 15, 24, 25, 28, 22, 27). The obtained crystal structures show that, with one exception, the inhibitors are typical hinge binders[177]. The most promising of the reported compounds from the AC series, 27, has no hydrogen bond to the hinge. It is a unique feature. Hydrogen bonds with K188 and E203 are formed to its diazole group and N244 via its carbonyl. Additionally, the N292 side chain forms a hydrogen bond with the fuorinated arene. Bridging water between the hinge and the compound was found in onlyone of the chains of the tetramer. Itresidesatahydrogen-bonding distanceof2.7Åfromthe 1,6-phenanthrolinenitrogen, nearby(2.8Å)themain chainnitrogenofL241and2.6Å tothe carbonylof E239. The tri-fuoromethyl, fuorobenzyl ring is in perpendicular π-stacking with 1,6-phenanthroline and diazole rings. The trifuoromethyl group penetrates a hydrophobic pocket formed by G166ofthe glycine-rich loop, I165 and V173 side chains. Compounds EHT1610 and EHT5372 (the most selective DYRK inhibitors identifed so far) share remarkable similarities to compound 27. This suggests that the canonical hinge binding may be less critical for high affnity binding to DYRK2, as seen for this inhibitor. The benzyl rings of those scaffolds are roughly perpendicular to each other. While the overall orientation of the inhibitors differs, all three compounds interact with the P-loop. The trifuoromethyl moiety in 27 and the 2-fuoro-and 2-chlorobenzyl groups of EHT1610 and EHT5372 fll the same subpocket. The overall shape of these molecules can be described as “U” shaped. The opening of the “U” in AC27 is directed towardthe P-loop(F160). This arrangement is reversed for the EHT inhibitors (Figure 37)[177]. 

Newly discovered binding features, such as CH-O interaction with Asn292 or binding water molecules that serve as catalytic lysine anchors, may provide valuable information for the optimization of these DYRK1A inhibitors and related kinases, which could be used in the future to treat not only diabetes but also neurodegenerative diseases, particularly Alzheimer’s Disease. The reported fndings, once again, confrm the importance of a multidirectional approach in the search and development of new DYRK1A-inhibitors[177]. 
Thereis emerging evidence demonstratingarole for DYRK1Ain diabetes and β-cell proliferation, which expands the potential for pharmaceutical applications of DYRK1A inhibitors. The diversity of the novel scaffolds and the binding modes determined by crystal structure and in vitro assays may lead to novel strategies for diabetestreatment. Small molecular inhibitors of DYRK1A developed in our group indicate specifc and strong binding affnity with promising therapeutical applications. One of our inhibitors is a potential regulatory agent for restoring pancreatic β-cell mass, secretory and regulatory functions to the organ. Hence, one of the aims is to further optimize the development of such inhibitors, depictingthe mechanisms involvedintheprogressionof diabetes.We have shownthattheAC inhibitors developedbyusareableto potentiatethe glucose-stimulated insulin secretion in cultured β-cells and isolated mouse islets of Langerhans. These results correlate with the inhibitory effcacy of the compounds against DYRK1A kinase selectivity and human β-cellproliferation.We assessed the AC27 inhibitor for its ex vivo activityin the freshly isolated pancreatic islets from mice. The results show that in both hiPSC-islets and isolated mouse islet models, AC27 signifcantly increased insulin secretion relative to untreated groups. Furthermore, this effect may be improved by co-addition of RepSox, a selective inhibitor of the TGF-β type1 receptor, orLY364947,a selectiveATP-competitive TGF-β receptor kinaseI inhibitor. Among others,the pathogenesisof impaired GSIS observed in T2D can be alleviated by these molecules. Controlled, stable regulation of cell functionatthe molecularlevelistakingitstollinregenerative medicine. Stimulation of functional cell growth will be more promising assuming that small molecule-induced human β-cell proliferation is reachable in clinical practice. This set of studies provides proof-of-concept that small-molecule-induced human β-cell proliferation is achievable and lends considerable promise to the goals of regenerative medicine for diabetes treatment. 
5.1.6. Miscellaneous Scaffolds and Drug Combinations In 2020, the novel DYRK1A inhibitor named KVN93 was identifed. This tau kinase inhibitor interacts with DYRK1Aby targeting theATP-binding sitein its active conformation when the activation loop is phosphorylated. It was investigated in Alzheimer’s Disease treatment asa compound able toregulate cognitive function, β-amyloid pathology, and neuroinfammation. The in vivo studies revealed that KVN93 improves long-term memory andreduces amyloid plaque levelsin 5XFAD miceby increasing theAβ degradation enzyme. KVN93 can modulate neuroinfammation in microglial cells by regulating TLR4/AKT/STAT3 signaling. The experiments carried out in wild-type mice injected with LPS confrmed that KVN93 treatment reduced microglial and astrocyte activation. These data suggest that KVN93 is a potential therapeutic DYRK1A inhibitor and is able to regulate (i) cognitive/synaptic function, (ii)Aβ plaque load, and (iii) neuroinfammatory reactions[178]. The studies describedby Allegretti and co-authorsrevealed that the anticancer kinase inhibitor OTS167 may act as a structurally novel, remarkably potent DYRK1A inhibitor to induce human β-cellreplication[179]. Despite the OTS167’s targetpromiscuity and cyto-toxicity,the multidimensional compound optimization was performed to tailor kinase selectivity towards DYRK1A and reduce its cytotoxicity. Indeed, the series of 1,5-naphthyridine derivative characterization yielded several leads with exceptional DYRK1A inhibition and human β-cell replication promoting potencies but substantially reduced cytotoxicity. The results suggest that these compounds are the most potent human β-cell replication pro-moting molecules described and exemplify the potential purposefully leverage off-target activitiesof advanced stage compounds for the desired application[179]. In order to elucidate the molecular pathways that control β-cell growth, Abdolazimi 
et al. screened about 2400 bioactive compounds for rat β-cell replication-modulating activity[180].Inthislibrary,theCC-401was identifedasasmall moleculethatpromoted human β-cell replication (Figure 38). CC-401 is known as an advanced clinical candidate previously characterized as a c-Jun-N-terminal kinase inhibitor. However, these studies re-vealedthat CC-401alsoactsvia DYRK1A/B inhibition[180].Moreover,itwasreportedthat DYRK1A/1B inhibition–dependent induction of β-cell replication is multifactorial. CC-401 treatmentledtorodent(invitro andinvivo)and human(invitro)β-cell replication via DYRK 1A/1Binhibition. In contrast to rat β-cells, which were broadly growth responsive to compound treatment(replication-inducing compounds like GSK3β or ALK5/TGFβ in-hibitors), human β-cellreplication was only consistently inducedbyDYRK1A/B inhibitors. In many reports, researchers identifed the DYRK1A/B inhibition–dependent activation of NFATas the primary mechanism of induction of β-cell–replication. Nevertheless, NFAT activity inhibition had a limited effect on CC-401–induced β-cell replication. Thus, the additional effects of CC-401–dependent DYRK1A/B inhibition were investigated. It has been found that CC-401 inhibited DYRK1A-dependent phosphorylation/stabilization of the β-cell–replication inhibitor p27Kip1. Additionally, CC-401 increased the expression of numerous replication-promoting genes generally suppressed by the dimerization partner, RB-like, E2F, and multi-vulval classB(DREAM) complex depends upon DYRK1A/B activity for integrity, including MYBL2 and FOXM1. These data demonstrate CC401 derivatives (abbreviated as STF compounds) and one of the commonly used DYRK1A inhibitors like harmine as a valuable resource for manipulating the signaling pathways that control β-cell replication and leverage DYRK1A/B inhibitors to expand understanding of the molecular pathways that control β-cell growth[180]. 

Additionally, the potential of combining small molecule inhibitors to augment the limited replication response of human β-cells was demonstrated. This effect was enhanced by simultaneous glycogen synthase kinase–3β (GSK3β)or activinAreceptor type II-like kinase/transforming growth factor-β (ALK5/TGFβ)inhibition[30]. 
It wasreported lately thatthe combinationof inhibition DYRK1A with transforming growth factor-beta superfamily (TGFβSF)/SMAD signaling leads to a synergistic increase in human β-cell proliferation and the number of β-cells in both mouse and human islets. This effect is related to the activation of cyclins and CDKs with the decreased levels in key cell-cycle inhibitors (including CDKN1C and CDKN1A) through altering their Trithorax-and SMAD-mediated transactivation. Additionally, this dual DYRK1A and TGFβ inhibition allow the preservation of β-cell functions. These effects were proved in healthy human-and stem cell-derived β-cells as well as patients with T2D both in in vitro andin vivo investigations[181]. Furthermore, therelationship between DYRK1A and insulin receptor substrate-2 (IRS2) has been thoroughly discussed. The loss of IRS2 expression in β-cells contributes to T2D. It was also indicated that IRS2 might be one of the DYRK1A targets. DYRK1A directly interacts with IRS2 through the N-terminal domain of DYRK1A. Moreover, DYRK1A promotes tyrosine(Y)-phosphorylation and K48linked poly-ubiquitination of IRS2 with the proteasomal degradation of IRS2. In vitro evaluationrevealed the expressionof DYRK1Ain MIN6cells and β-cells islets and pointed itsrolein inducing apoptosis. Furthermore, IRS2 expression was slightlyreducedin the hippocampus and islets of young APP/PS1 mice (3-month-old), while it was signifcantly suppressedinolder animals (6-month-oldmice).Itwas postulatedthatitmightberelatedto other mechanisms, e.g., activation of GSK3β and neuroinfammationintheearlystageofthe disease[182]. These fndingsalso complementthe current understandingoftherelationship between DM and AD[182]. Some evidence was also provided that the combination of any GLP1R agonist class member with any DYRK1A inhibitor class member induces a synergistic increase in human β-cell replication accompanied by an increase in human β-cells mass[50].Acombinationof small-molecule DYRK1A inhibitor (suchas harmine, INDY, leucettine, 5-IT, GNF4877) to any one of the antidiabetic drugs that directly (GLP-1 analogs) or indirectly (DPP4 inhibitors) activate the GLP1R and convert the mitogenically inactive GLP1R agonists into potent β-cell proliferative agents[50]. Combining these two agents boosted human pancreatic β-cell proliferation and expanded β-cell mass in human cadaveric islets ex vivo. For instance, cadaveric human islets were transplanted into immunodefcient mice with diabetes inducedby streptozocin, and the mice were then treated with the drug combination. The animals showed increased insulin production and improved glycemic control than mice treated with either compound alone or no treatment. Both phosphorylated NFATand cAMP-PKA mediated the synergistic effect of the two molecules on pancreatic β cell expansion–dependent activation of cell cycle genes such as cyclin-dependent kinases and β-cell-specifc genes (e.g., GLUT2, PDX1, and NKX6.1)[50,183]. Theresultingproliferation rates exceeded thoseof DYRK1A inhibitors alone and may be in a range that could restore β-cell mass in people with T2D and T1D[50,149,183]. 
Itiswellknownthatthenatureof diabetesisunlikelytobefullyaddressedbythemodulationofanysingletarget.The “paradigm shift” determinestheresearchwhen complexity prevails, and radical specifcity is no more the ultimate target. Simultaneous targeting of both DYRK1A kinase enhances the restoration of the β-cell population. Alternatively, the combinatory treatment with inhibitors, hypoglycaemic agents (glucagon-like peptide-1 (GLP-1)receptor agonists), and cell markers (e.g., TGF-β)improves the proliferation rate in human cadaveric β-cells. However, current inhibitors lack target specifcity, with risks of adverse effects. Thus, the need to identify drugs that provide an accelerated human β-cell proliferationof improved specifcityremains the priority. The developmentof inhibitorsis frequently compromised by suboptimal pharmacokinetics. Evidence has recently emerged that simultaneous targeting of both DYRK1A and GSK3β may further beneftinrestoring the insulin-producing β-cell population. Moreover, the recent studies on the DYRK family show the compensatory mechanism for DYK1A and DYRK1B synergistic effect on the proliferation of the β-cells in mammalian cell culture models. 


6. Biological Effects of DYRK1A Inhibitors 
6.1. Diabetes 
As mentioned above, several DYRK1A inhibitors are able to enhance β-cell proliferation and improve insulin secretion and glucose homeostasis[16]. The gold standard in thisresearch,harmine, may increase human β-cell proliferation in culture by ca. 2%. Nevertheless, DYRK1A inhibitors, including leucettine-L41 and INDY, indicate comparable proliferative potential, while 5-IT and GNF4877 were found to be 10-fold more potent. 
Therefore, inhibition of DYRK1A is an important mechanism underlying β-cell pro-liferation and emphasizes that the diabetic kinome is a key target that can increase the mitogenic activity of β-cells. Following DYRK1A inhibitor treatment, proliferation is en-hanced by induction and nuclear translocation of NFATtranscription factors that affect the cell cycle. Furthermore, it is suggested that DYRK1A inhibitors attract other targets involved in the stimulation of β-cell proliferation. Thus, the role of the diabetic kinome seemstobecrucialforthe future developmentof anti-diabetic strategies. Several studiesreveal that eachof these DYRK1A inhibitors alsoinhibits other kinases, particularly members of the CMGC family, including (i) cyclin-dependent kinase (CDK), (ii) mitogen-activated protein kinase (MAPK), (iii) glycogen synthase kinase-3 (GSK3), and (iv) CDC-like kinases notably: DYRKs, CLKs, GSKs[149]. Noteworthily,it canbe speculated that eachof them may be involved in human β-cell proliferation. Importantly, GSK3 (involved in insulin signaling and the replication of β-cells) may be recognized as the most prominent target of DYRK1A inhibitors because DYRK1A functions asapriming kinase for GSK3 signaling and playsasubstrateroleinpreparationforGSK3 phosphorylation.The interactionof DYRK1A inhibitors withGSK3β has been shown to lead to β-cellproliferationinrodents[3]. Fur-thermore, it has been suggested that dual-mode inhibition of DYRK1A and GSK3β may contributetotheeffcacyofthe aminoprazine derivative GNF4877[171]. 
GSK inhibitors (LiCl, 1-Akp) have also been shown to increase human β-cell proliferationfrom 0.17%to 0.71%[5]. However, DYRK1A inhibitors actina dose-dependent manner, with proliferation peaking after treatment with the optimal inhibitor concentration and decreasing at higher doses. These are results suggesting interactions with other kinases/targetsathigher doses[2,6,7]. Furthermore,off-targeteffectsarenot necessarily limited to protein kinases. 5-IT has also been found to be an adenosine kinase inhibitor, and its β-cell mitogeniccapacitymaybe attributedto adenosine kinase inhibition[8].Itisalso possible that DYRK1A inhibitors may affect targets other than kinases. Harmine not only inhibits DYRK1A in human β-cells but alsoreduces the abundanceof SMADproteins[6]. In addition,it actsasanMAO inhibitor[16,149]. 
Nevertheless, it can be stated that the mitogenic effects and enhanced proliferation mediatedby DYRK1A inhibitorsactas translocationofNFATtranscription factorstothe nucleus, with the consequent transactivation of cyclins (cyclin A, CDKs) and repression of CDK-inhibitors such as p15INK4, p21CIP1, and p57KIP2[149]. Other possible mechanisms involving DYRK1A, necessary for β-cell restoration are (i) phosphorylation and stabilization of p27KIP1, (ii) phosphorylation of D-cyclins and acceleration of their degradation; 
(iii) phosphorylation of the DREAM complex member, LIN52, enforcing cell cycle arrest; and (iv) phosphorylationof tauprotein crucial forAD[149]. 
All these data indicate that regulation of DYRK1A kinase activity is an important mechanism underlying human β-cell proliferation. Other potential kinases and therapeutic targets capable of enhancing β-cell mitogenic activityarealso indicated. Therefore,a better understanding of the diabetic kinome is crucial for the design and development of new and innovative, more potent and selective small molecules. 

6.2. Other Diseases 
This review focuses on DYRK1A inhibitors developed for β-cell restoration and treatment of diabetes. However, it is worth noting that the development of DYRK1A inhibitors may be benefcial in the treatment of other diseases, including neurological disorders such as Alzheimer’s Disease(AD), Parkinson’s and Huntington’s diseases, Down Syndrome (DS)[79]and cancer[20]. 
6.2.1. Neurological Disorders DYRK1A has been implicated in neuronal development and many others related signaling pathways.InDS,thetriplicationofchromosome21resultsin ca.1.5-fold higher DYRK1A levels than the general euploid population. This DYRK1A overexpression has been linked to the cognitive defcits associated with Down Syndrome[184]. Moreover, through hyperphosphorylation of tau protein (Alzheimer’s Disease protein) and the for-mation of insoluble tau aggregates, DYRK1A is also involved in neurodegeneration and neuronal loss appearinginAD[185,186]. Therefore, a therapeutic strategy for cognitive defcits associated with DS, and ultimatelyAD, would involve controlled inhibitionof brain DYRK1A activity[187]. Overthe past few years, several DYRK1A inhibitors have been developed, most of which bind to the enzyme’s activeATP site. However, there are selected exceptions, such as epigallocatechin gallate (EGCG), an allosteric inhibitor of DYRK1A that improves cognition inTs65Dn mice(a well-established invivo modelforDS[185,188].ItwasalsoreportedthatT65Dn mice witha normalized DYK1A gene copy number (two copies) were characterizedby a decrease in (i) senescent cells population in the hippocampus and cortex, (ii) cholinergic 
neurodegeneration, as well as (iii) APP thatpromotes theproductionof pathogenicAβ, andtaulevels,in comparisontoDownSyndromemicewiththreecopiesof DYRK1A[189]. 
These data indicated that DYRK1A inhibition and normalizationof its level couldreduce or delayAD neuropathology[189]. 
6.2.2. Cancer Both overexpression and downregulation of DYRK1A areassociated with neurological defects, refecting the extreme gene-dosage sensitivity of this protein. It was reported that DYRK1A could act as both an oncogene anda tumor suppressor[190]. DYRK1A works as a negative regulator of the cell cycle, and its dosage can direct cells toward proliferation or exit from the cell cycle. It may also promote the survival of malignant cells by inhibiting pro-apoptotic pathways since the loss of DYRK1A can activate p53 (the increased degradation of DYRK1A caused by p53 activation is mediated by MDM2, which was found to interact with and ubiquitinate DYRK1A, ultimately leading to its proteasomal degradation)[191,192]. DYRK1A likely playsa tumor type-specifcrole, so whether DYRK1A inhibition would promote or inhibit tumor cell growth depends on the tissue type and tumor microenvironment.Although DYRK1A is most widely characterized for itsrolein brain development, DYRK1Ais overexpressedin various diseases,including many typesof cancers, such as leukemia[193,194], pancreatic adenocarcinoma[195–197], and gliomas[198,199]. ItwasalsoreportedthatDYRK1Acould positivelyregulatetheSTAT3/EGFR/Metsignaling pathway in human EGFR wild-type NSCLC cells. In addition, DYRK1A inhibition (by siRNA or an inhibitor) increased the anticancer activity of AZD9291 (EGFR inhibitor, Osimertinib) NSCLC cells[200]. Furthermore, it was reported that inhibition of DYRK1A destabilizesEGFRandreduces EGFR-dependent glioblastomagrowth[201].Itwasindi<BR>catedthatDYRK1AreducesthelevelofCyclinD1by phosphorylatingonThr286,inducing the proteasomal degradation of Cyclin D1 and cell cycle G1 phase arrest. Furthermore, DYRK1A suppression canpromote the degradationof EGFR andreduce the self-renewal capacity of glioblastoma cells[200,201]. Pozo et al. investigated the ability of harmine and INDY to inhibit GBM tumor growth and survival[201]. They suggested that DYRK1A functions upstream of SPRY2 to modulate EGFR lysosomal targeting. Phosphorylation of SPRY2 by DYRK1A decreases its inhibitory infuence on FGF-induced MAPK activation. In glioblastomas, several members of the SPRYfamily are included in a transcriptome module associated with the EGFR amplifcation status in GBMs, suggesting that they could act as oncogenes. Thus, destabilization of EGFR by DYRK1A inhibition may be a potential therapeutic target fora subsetof EGFR-dependent GBMs[201]. Another example is CX-4945 (silmitasertib),a casein kinase2inhibitor currentlyin clinical testing for various cancers[202].Itwas subsequently foundtoalso potently inhibit several membersofthe 
CLK and DYKRK families, including DYRK1A, and was able to block DYRK1A-related tau phosphorylation in a mouse model of Down Syndrome. 


7. Summary 
Diseases related to diabetes and obesity are one of the major threats to human life. According to WHO, approximately 300 million people will be obese in 2035[203]. This ever-increasing trend is diffcult to prevent due to changing lifestyles around the world and energy-rich diet availability. Only in 2015, more than 1.6 million human deaths were caused by hyperglycemia and diabetes. Type 2 diabetes is now treated with various pharmaceuticals, but in fact there is no effective treatment. Other types of diabetes rely solely on supplementing the body with external insulin. Despite signifcant advances in insulin-based and other therapies, patients with diabetes will continue to receive medication throughout their entire lives. This causes an enormous healthcare burden and limits the comfort of patient’s life. 
Two main types of diabetes—T1D and T2D—share similar mechanisms of β-cellfunction failurevia an insuffcient mass of the endocrine pancreatic cell fraction. In T1D, this phenomenonisdrivenbyautoimmuneassaultagainstowncells,whileT2Dis characterized by insulin resistance and subsequent β-cell mass decrease. In general, T1D and T2D are, by defnition, a blood hyperglycemia condition caused by total or relative defcits in β-cell mass. Existing therapiesimprove glycemic control butprovide onlya temporal relief, withlifetime dependency. Several earlyprevention measures and strategiesoffered for diabetic patients of T2D put this disease into reasonable control to delay the clinical onset. Such interventions do not exist for T1D. As stated by The Global Report of the World Health Organization (WHO), T1D cannot be prevented with current knowledge. Although effective approaches are available to prevent T2D, no cure for advanced disease is available.Theoptimalapproachshouldreverseits pathologicchangestoprovideacure rather thana lifetime pharmaceutical supplementation. Thus, fnding an accurate cure for diabetes is of critical importance. Restoring metabolic homeostasis would free the patient from constant reliance on pharmaceuticals and monitoring glucose level. Nevertheless, so far, all the possible therapies are only in very early preclinical stages. The treatment strategiesrely mainlyonpromoting β-cellsdifferentiation. Thispromising strategyrequires selective alteration of cellular differentiation to obtain a new, regenerated population of β-cells. Unfortunately, direct alteration of transcription factors is complicated, and there is no effcient strategy to affect the pancreas selectively. Therefore, more upstream biomolecular targets are sought. The importance of targeting protein kinases with small molecules is an irrefutable and great tool to establish therapeutical pathways to understand disease mechanisms. In particular, the fnding of DYRK1A, a crucial protein kinase that has been implicated as a potential regulator of β-cells, raises its potential application in diabetes. DYRK1A is involved in cellular processes related to the proliferation and differentiation of β-cells. Thus, DYRK1Ais oneofthe most extensively studied targets for β-cells regeneration. The β-cells differentiation observed when DYRK1A kinase activity is modulated points to a possibility of using “diabetic kinome” as a target for future DM therapies. Scientifc investigations and the pharmaceutical industry have confrmed the roleof DYRK1A kinasein various molecularprocesses. Thisreviewaimsto highlightthe knowledge and approaches taken under action within the past few years. These last fve years have brought progress and even more questions about the actual position of the approach for many scientifc felds.Wepresentrecent developmentsin diabetic kinome inhibitors, witha particular focus on DYRK1A. 
Wehavepaid particular attentioninthisreviewtothefactthatno DYRK1A inhibitors, to date, have met the selectivity standards needed for use as probe molecules. Harmine, oneof the most commonly usedinhibitorsin DYRK1A-relatedresearch, possesses strong cross inhibition of monoamine oxidase (MAO), which would cause some adverse effects. Thelow selectivityalsomakes harmine unsuitableasaprobetotest DYRK1A inhibition in cell lines. Efforts to eliminate the MAO inhibition while keeping DYRK1A inhibition led to the harmine derivative AnnH75. Another DYRK1A inhibitor, green tea favonol epigallocatechin-gallate (EGCG), was shown to correct cognitive defcits in Down Syn-drome mouse models and humans. However, it also potentially has multiple targets (and correspondingly is under consideration for use in a broad range of disorders) and cannot be considered a DYRK specifc inhibitor. Thus, structural modifcations may be introduced to achieve high selectivity. The so-called “gatekeeper” identity was identifed early as a principal determinant of inhibitor selectivity. This residue initiates the “hinge” segment thatlinksthetwofoldinglobesofprotein kinases,anditssidechainlies adjacenttoATP inhibitors that bind via hydrogen bonding to the hinge. DYRK protein kinase targets consist of phenylalanine, which simultaneously offers good opportunities for inhibitor design and polypharmacology. Another opportunity for selectivity andfavorable binding kinetics is covalent binding to sulfhydryl groups. The cysteine in the HCD (histidine cysteine aspartate)motifisthemostprominenttargetforthe DYRK1A.Athirdopportunity involves linkingATP-site inhibitorsto peptides correspondingto substraterecognition sequences. This allows for high potency and selectivity for research compounds. In order to briefy summarize all the DYRK1A inhibitors discussed in this review, their IC50 values, targets, biological activity with future directionof development are listedinTable 2. 
Table 2.IC50 value determined for DYRK1A inhibitors mentioned thorough thisreview. Abbrev. In order: ↑↑/↑↑↑ = moderate/most potent. 
IC50 
No. Compound Name OtherTargets InVitro Results Future Directions 
DYRK1A[nM] 

MAO-A 
increasing the selectivity 
1. 
harmine 33 CK1 ↑↑↑ 
of the compound 
PIM3 

the research direction is 4-(7-methoxy-1-methyl-β-defnitely worth 
2. 25 MAO-A ↑↑↑ 
carbolin-9-yl)butanamide pursuing, large-scale clinical trials are needed 

results will be use to the 
4-hydroxychromeno[3,4-CDK5 
3. 
0.074 not available chromeno[3,4-b]indole 
b]indol-6(7H)-one GSK3 
asa pharmacophore 

1-[2-(4-promising scaffolds for hydroxyphenyl)pyrido[3,2-CDK5 targetingprotein kinases 
4. 60 not available 
d]pyrimidin-7-yl]piperidin-2-GSK3 involved in the central one nervous system 

further modifcations are CLK1 underway, aiming at the 2-cyclopentyl-7-iodo-1H-CLK2 minimal cytotoxicity, development of potent, 
5. 
70 
indole-3-carbonitrile CLK4 more data not available highly selective and GSK3 water-soluble DYRK1A inhibitors 
Table 2.Cont. 
IC50 
No. Compound Name OtherTargets InVitro Results Future Directions 

DYRK1A[nM] 

results will be used to 3-hydroxychromeno[3,4-CDK5 the 
6. 500 not available 
b]indol-6(7H)-one) GSK3 chromeno[3,4-b]indole as a pharmacophore 

promising scaffolds for 
4-[7-(5-methyl-thiophen-3-yl)
CDK5 targetingprotein kinases 
7. 
pyrido[3,2-d]pyrimidin-2-yl]-24 not available 
GSK3 involved in the central 
phenol 
nervous system 

4-chlorocyclohepta[b]indol-biological data are 
8. 
200 CLK1 not available 
10(5H)-one needed 

CLK1 CLK2 use for the development 7-chloro-1H-indole-39. 
3300 CLK3 not available of new DYRK1A carbonitrile CLK4 inhibitors GSK3 

CLK1 
7-iodo-2-(pyridin-3-yl)-1H-biological data are 
10. 
80 CLK2 not available 
indole-3-carbonitrile needed 
CLK4 
Table 2.Cont. 
IC50 
No. Compound Name OtherTargets InVitro Results Future Directions 

DYRK1A[nM] 

CLK1 CLK2 7-iodo-2-phenyl-1H-indole-3-minimal cytotoxicity, biological data are 
11. 10 CLK3 
carbonitrile more data not available needed 
CLK4 GSK3 

CLK1 
10-chloro-11H-indolo[3,2
CLK2 biological data are 
12. 
c]quinoline-6-carboxylic 23 not available 
CLK3 needed 
acid 
CLK4 

new scaffolds offer novel 13. 
AC2 >16000 not available not available opportunities to design DYRK1A inhibitors 

new scaffolds offer novel 14. 
AC8 >8000 not available not available opportunities to design DYRK1A inhibitors 
Table 2.Cont. 
IC50 
No. Compound Name OtherTargets InVitro Results Future Directions 

DYRK1A[nM] 

new scaffolds offer novel 15. 
AC12 216 pan kinase not available opportunities to design DYRK1A inhibitors 

biological research in 
16. 
AC13 350 not available not available 
progress 

new scaffolds offer novel 17. 
AC14 221 not available not available opportunities to design DYRK1A inhibitors 

new scaffolds offer novel 18. 
AC15 329 pan kinase not available opportunities to design DYRK1A inhibitors 

new scaffolds offer novel 19. 
AC16 >6000 not available not available opportunities to design DYRK1A inhibitors 
Table 2.Cont. 
IC50 
No. Compound Name OtherTargets InVitro Results Future Directions 

DYRK1A[nM] 

new scaffolds offer novel 20. 
AC18 >4800 not available not available opportunities to design DYRK1A inhibitors 

CLK2 new scaffolds offer novel 21. 
AC20 3500 ABL not available opportunities to design PDGFR DYRK1A inhibitors 

GSK3β CLK2 biological research in 22. 
AC22 >6000 not available 
HIPK1/2 progress CDK7 

DRAK1/2 biological research in 
23. 
AC23 4200 not available 
ERK5 a progress 

new scaffolds offer novel 24. 
AC24 800 CLK2 not available opportunities to design DYRK1A inhibitors 
Table 2.Cont. 
IC50 
No. Compound Name OtherTargets InVitro Results Future Directions 

DYRK1A[nM] 

biological research in 
25. 
AC25 1200 CLK2 not available 
progress 

GSK3β 
biological research in 
26. 
AC27 532 PIK3CG not available 
progress 
PIK4CB 

biological research in 
AC28 -CLK2 not available 
progress 

possible use in viral infections, cancer and CDK1 neurodegenerative 
28. Aristolactam AIIIA 80000 CDK2 not available 
pathologies (e.g., 
CDK4 
Alzheimer’s and Parkinson’s diseases) 

c-Jun-N-terminal kinase  key issues are the  
PRKD2  development of  
CC-401  - PRKD3  ↑↑  strategies to target  
CSNK1G3  regenerative compounds  
MAPK9  selectively to the β cell  

Table 2.Cont. 
IC50 
No. Compound Name OtherTargets InVitro Results Future Directions 

DYRK1A[nM] 
Akt CDK2 CDK5 

CK1 3,5-di-(4-hydroxyphenyl)-1H-CLK1 Noncytotoxic, more data studies on appropriate 30. 
3 
pyrrolo[2,3-b]pyridine ERK2 not available animal models GSK3 JAK3 TRKA 
pim1kinase 
Akt CDK2 CDK5 

3-(4-hydroxyphenyl)-5-(2,4-CK1 dihydroxyphenyl)-1H-CLK1 Noncytotoxic, more data studies on appropriate 
31. 11.7 
pyrrolo[2,3-b]-ERK2 not available animal models 
pyridine GSK3 JAK3 TRKA 
pim1kinase 
Akt CDK2 CDK5 

CK1 3,5-di-(3,4-dihydroxyphenyl)-CLK1 Noncytotoxic, more data studies on appropriate 
32. 12.4 
1H-pyrrolo[2,3-b]pyridine ERK2 not available animal models GSK3 JAK3 TRKA 
pim1kinase 
Table 2.Cont. 
IC50 
No. Compound Name OtherTargets InVitro Results Future Directions 

DYRK1A[nM] 
Akt CDK2 CDK5 

3-(4-hydroxyphenyl)-5-(3,4-CK1 dihydroxyphenyl)-1H-CLK1 Noncytotoxic, more data studies on appropriate 
33. 23.1 
pyrrolo[2,3-b]-ERK2 not available animal models 
pyridine GSK3 JAK3 TRKA 
pim1kinase 

CLK1 
high-potential therapy 
CLK2 
34. 
EHT5372 0.22 not available for AD and other 
CLK4 
tauopathies 
GSK3 

CLK1 
high-potential therapy 
CLK2 
35. 
EHT1610 0.36 not available for AD and other 
CLK4 
tauopathies 
GSK3 

FC162 modifedTau 
development asa 
CLK1 phosphorylation and 
36. 
FC162 11 DYRK1A kinase 
GSK3 could alter cell cycle 
inhibitor 
progression of pre-B cells 
Table 2.Cont. 
IC50 
No. Compound Name OtherTargets InVitro Results Future Directions 

DYRK1A[nM] 

cytotoxic effects was 
CDK5/p25 GSK3α/β candidate for further 
37. 
furan-2-yl-substituted >10000 determined against six 
PI3Kα evaluations 
human cancer cell lines 

requires strategies to mitigate the 38. 
GNF2133 6 GSK3 ↑↑ observedhypertrophic effects in nonpancreatic tissues 
39. 

GNF1346 -undetermined ↑ for now, untested 
CDK  
40.  GNF3809  - CLK MAP4K4 GSK3 FLT HIPK  ↑↑  further optimization and elucidation of its molecular mechanism of action needed  
JAK3  

Table 2.Cont. 
IC50 
No. Compound Name OtherTargets InVitro Results Future Directions 

DYRK1A[nM] 
41. 

GNF4088 -undetermined ↑ for now, untested 

GNF4877 was 42. 
GNF4877 6 GSK3 ↑↑ notprogressed beyond preclinicalresearch 

may provide a path 43. 
GNF7156 100 GSK3 ↑↑ forwardto develop new drugs to treat diabetes 

development of chemically modifed versions of GNF9228 
44. GNF9228 -undetermined ↑↑↑ 
with enhanced bioavailability to allow in vivo testing 

promising starting point 
data available for 
45. 
Kufal194 6 CLK-1 for the development of 
zebrafsh 
therapeutics in DS 

biological research in progress, promising CLKs 
46. L41 170 not available compound forthe 
mTOR/PI3K development of novel AD therapeutic agents 
Table 2.Cont. 
IC50 
No. Compound Name OtherTargets InVitro Results Future Directions 

DYRK1A[nM] 
47. 

medioresinol 100 CDK1 not available for now, untested 

methyl 9-(4-bromo-2-CK1 a promising source for 48. 
fuorophenylamino)thiazolo[5,4-50 CDK5 not available the synthesis of novel f ]quinazoline-2-carbimidate GSK3 bioactive molecules 

methyl 9-(4-CK1 a promising source for 49. 
methoxyphenylamino)thiazolo[5,4-40 CDK5 not available the synthesis of novel f ]quinazoline-2-carbimidate GSK3 bioactive molecules 

methyl 9-(benzo[d][1,3]dioxol-CK1 a promising source for 50. 
5-ylamino)thiazolo[5,4-47 CDK5 not available the synthesis of novel f ]quinazoline-2-carbimidate GSK3 bioactive molecules 
Table 2.Cont. 
IC50 
No. Compound Name OtherTargets InVitro Results Future Directions 

DYRK1A[nM] 

N-(2,3dihydrobenzo[b][1,4]dioxin-6
TRPV1 starting point of a larger 
51. yl)-7-0.5 not available 
CLK1 program 
methoxybenzothieno[3,2d]pyrimidin-4-amine 

N1-(7methoxybenzothieno[3,2-starting point of a larger 52. 
0.68 CLK1 not available 
d]pyrimidin-4-yl)-N4,N4-program dimethylbenzene-1,3-diamine 

a model compound for 53. 
OTS167 -undetermined cytotoxic the design of less toxic compounds 
54. 

velutinam 600 CDK1 not available for now, untested 
Compared to the well-known and the best to date inhibitor for increasing human pancreatic β-cell replication, the advantages of the newly identifed fragments give us a privileged position in the race to the new therapeutics. Future studies should provide proof-of-concept that small-molecule–induced human β-cell proliferation is achievable with the use of regenerative medicine for diabetes therapy. The generation of the iPSCderived β-cells has been one of the most desired strategies, with several protocols being invented. Functional iPSC-derived β-cells bringreal hope for diabetic patients, who are not qualifed for transplantation, with severe glycemic lability, recurrent hypoglycemia, and a reduced ability to sense symptoms of hypoglycemia (reduced hypoglycemia awareness). Providing an unlimited source of autologous, engineered cells from the somatic pool could signifcantly shift the availability of transplants from very limited to plentiful. Therefore, every fnding and improvement in the prolonged intervention of diabetes is of the highest value. Current knowledge on the transplantation of the pancreatic islets tackles the severe problem of engraftment and stable implantation of the delivered cell mass into the organ. The importance of resolving this issue has been demonstrated broadly, and multiple methods are proposed to alleviate the problem. We would also like to propagate the term “diabetic kinome” within scientifc terminology to emphasize the role of multiple kinases’ synergistic action in directing molecular processes that underlie this particular set of diseases. The human kinome constitutes over 500 kinases, responsible for every biological function and regulation in the cell. Therefore, fnding the optimal selectivity profle for kinase inhibitors is of essential importance. 
Author Contributions: Conceptualization, A.C., B.P., J.M.D.; writing—original draft preparation, B.P., A.C., J.M.D., A.B.; writing—review and editing, B.P., J.M.D.; visualization, A.B., B.P.; supervision, A.C.; project administration, A.C., J.M.D.; funding acquisition, A.C., J.M.D. All authors have read and agreed to the published version of the manuscript. 
Funding: Thisresearch was fundedby National Science Center, within theprojects: Sonata Bis9 no 2019/34/E/NZ1/00467, Opus 19 no 2020/37/B/NZ7/04157, and NAWAPolish National Agency for Academic Exchange no PPN/PPO/2018/1/00046. 
Conficts of Interest: The authors declare no confict of interest. 
Abbreviations  
AD  Alzheimer ’s Disease  
Akt  Protein kinase B  
AMPK  50AMP-activated protein kinase  
BBB  Blood-brain barrier  
β-FGF  Fibroblast growth factor  
cAMP  30,50-cyclic adenosine monophosphate  
CDKLs  CDK-like kinases  
CDKN  Cyclin-dependent kinase inhibitor  
CDKs  Cyclin-dependent kinases  
CK2  Casein kinase 2  
CLKs  CDC-like kinases  
CMGC  Kinase family is named after the initials of its subfamily members, including  
cyclin-dependent kinase (CDK), mitogen-activated protein kinase (MAPK),  
glycogen synthase kinase (GSK) and CDC-like kinase (CLK)  
c-Myc  Family of regulator genes and proto-oncogenes that code for  
transcription factors  
CNS  Central nervous system  
CREB  cAMP response element-binding protein  
DM  Diabetes mellitus  
DN  Diabetic Neuropathy  
DPP-4  Dipeptidyl peptidase-4  
DREAM  Dimerization partner, RB-like, E2F, and multi-vulval class B  
DS  Down Syndrome  

DSCR1 gene Down Syndrome criticalregion gene1 
DYRK1A Dual-specifcity tyrosine phosphorylation-regulated kinaseA 
EGCG Epigallocatechin gallate 
ERK/MAPK Achain of proteins in the cell that communicates a signal from a receptor on the cell’s surface to the DNA in the nucleus. The pathway includes many proteins, including MAPK (mitogen-activated protein kinases, originally-called ERK, extracellular signal-regulated kinases) 
FDA Food and Drug Administration 
FKHR Forkhead protein 
FOX Forkhead box 
FOXM1 Forkhead Box M1 
GAD65 Glutamic acid decarboxylase 
GADA Glutamic acid decarboxylase antibodies 
GDM Gestational diabetes mellitus 
GLP-1 Glucagon-like peptide-1 
GLUT4 Glucose transporter type4 
GPAIS Glucose potentiates arginine-induced insulin secretion 
GSK3 Glycogen synthase kinase3 
GSKs Glycogen synthase kinases 
HbA1c Glycated hemoglobin 
HK Hexokinase 
HLA Human leukocyte antigen 
HNF-1α Hepatic transcription nuclear factor 
HTS High-throughput screening 
IDE Insulin-degrading Enzyme 
IGF-1R Insulin-likegrowth factor1 receptor 
IKKβ Ikappa beta kinase 
IL-1β Interleukin1beta 
IL-12 Interleukin 12 
IR Insulin receptor 
IRS1 Insulinreceptor substrate1 
IRS2 Insulin receptor substrate-2 
JNK Jun N-terminal kinase 
MAO Monoamine oxidase 
MAPK Mitogen-activated protein kinase 
MODY Maturity-onset diabetes of the young 
mTOR Mammalian target of rapamycin kinase 
MYBL2 Myb-relatedproteinB 
NFAT Nuclear factorof activatedT-cell 
NKX6.1 Protein that in humans is encoded by the NKX6-1 gene 
NME1/2 Nucleoside diphosphate kinase1and2 
p53 tumor protein P53 
PI3K Phosphatidylinositol 3-kinase 
PI3K-AKT/PKB Signal transduction pathway that promotes survival and growth in response to extracellular signals. Key proteins involved are PI3K (phosphatidylinositol 3-kinase) and Akt (protein kinase B) 
PKC Protein kinaseC 
PKR RNA-activated protein kinase 
PPAR Peroxisomeproliferator-activatedreceptors 
PPX Partial pancreatectomy model 
PTHRP Parathyroid hormone-related protein 
RNA Ribonucleic acid 
ROCK Rho-associated coiled-coil containing protein kinase 
S6K1 S6 kinase-1 SAR Structure-activity relationship 
SGLT-2 Sodium-glucose co-transporter-2 
SMAD Family of structurally similar proteins that are the primary signal 
transducers for receptors of the transforming growth factor-beta 
SPRED2 Sprouty-related protein with an EVH1 domain 
Sprouty2 Sprouty homolog2 
STAT3 Signal transducer and activatorof transcription3 
TGF-β Transforming growth factorβ 
TGFβSF Transforming growth factor-beta superfamily 
TNF-α Tumor necrosis factorα 
VGF Nerve growth factor 


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