Medulloblastomas are the most frequent encephalon tumours in kids and remained a major curative challenge. Clinical and histopathological characteristics are used for disease categorization and patient omen. Presently, several molecular surveies utilizing transcriptomic and genomic attacks suggested the being of four molecular subtypes, increasing the complexness and cognition of medulloblastoma biological science. Despite these important progresss, the molecular footing of medulloblastomas is non to the full understood. MicroRNAs ( miRNAs ) are a group of little non-protein cryptography RNA molecules that mark cistrons by bring oning mRNA debasement or translational repression. They represent an evolutionary conserved mechanism that controls cardinal cellular procedures, such as development, distinction, metamorphosis, proliferation and programmed cell death. Aberrant look of miRNAs correlatives with assorted malignant neoplastic diseases. This altered look can originate from mutant, methylation, omission, and addition of miRNA-encoding parts. We here review the cognition of miRNAs in medulloblastomas. The look forms of miRNAs in medulloblastomas were comprehensively evaluated and their diagnostic, predictive and curative biomarker function assessed. miRNAs are of import participants in medulloblastoma tumorigenesis and their curative development can represent an alternate attack to this annihilating disease.
Keywords: miRNAs, medulloblastoma, kids, diagnostic, predictive.
Childhood malignant neoplastic disease is comparatively uncommon, nevertheless about 15,000 kids ( & A ; lt ; 19 old ages old ) are diagnosed with malignant neoplastic disease each twelvemonth and [ American Cancer Society, 2009 # 6 ] it remains the taking cause of disease-related mortality in kids [ 1 ] . Central Nervous System ( CNS ) tumours, chiefly happening in the encephalon, are the 2nd most frequent malignance ( first is leukemia ) and is the most common solid tumour of the childhood, stand foring at least 30 % of all malignant neoplastic diseases in kids and striplings in the United States [ 2 ] . In Brazil, it is the 3rd most common tumour in kids [ 3 ] . Among childhood CNS tumours, medulloblastoma ( MB ) is the most frequent stand foring ~20 % of all encephalon tumours in kids [ 4 ] .
Medulloblastoma is a extremely aggressive tumour that arises from altered ( mutated ) staying crude neuroectoderm cells in the ventricle and grows in the cerebellar vermis, frequently occupying through the ependyma and brain-stem [ 5 ] . Around 30 % of kids present grounds of disseminated disease at diagnosing [ 6 ] . The classical neurological symptoms in kids are associated with an increasing in intracranial force per unit area and include sickness, concern, puke, crossness and ataxy [ 5 ] . Harmonizing to the WHO ( World Health Organization ) categorization, medulloblastomas are grade IV malignances and can be divided into five subtypes: classical, anaplastic, big cell, nodular desmoplastic and medulloblastoma with extended nodularity [ 7 ] .
The pick of intervention in medulloblastoma is based on the hazard stratification ( high hazard or standard hazard ) of the disease, and the current intervention modes range from surgery, craniospinal radiation therapy and standard chemotherapy intervention [ 8 ] . The hazard stratification is established harmonizing to the possibility of tumour resection, presence of metastasis and histology. The overall five-year endurance stands about 75 % , runing from 55 % to 80 % in high hazard and standard hazard patients, severally [ 9 ] . Despite the diverseness of curative options, the major concern about the intervention is that it lacks specificity, frequently ensuing in long late-effects that include neurocognitive damage, neuropathy, endocrine changes, impaired bone growing and impaired motor map, hearing loss every bit good as secondary tumours [ 8 ] .
The bulk of medulloblastomas is non associated with a predisposing familial cause, nevertheless its happening may be associated with familial upsets such as Li-Fraumeni syndrome ( TP53 mutants ) , Turcot syndrome ( APC mutants ) [ 10 ] or Gorlin syndrome ( PTCH1, PTCH2 and SUFU mutants ) [ 11, 12 ] .
Recently, a collaborative attempt affecting groups that performed schemes uniting high throughput testing ( genomics and transcriptomics ) has resulted in a consensus that medulloblastomas can be grouped into four distinguishable classs ( WNT/?-catenin, Sonic Hedgehog ( SHH ) , group 3 and group 4 ) harmonizing to alone molecular features [ 13-17 ] . In this manner, it is possible to set up hazard stratification for patients with medulloblastoma based on the molecular characteristics of the tumour.
Despite the great advancement made in the apprehension of medulloblastoma biological science, it is still a heterogenous disease with a different molecular behaviour. Therefore, more attempts needed to be driven to specify the most relevant predictive stratification factors and besides to place biological changes that could be marks for molecular specific therapies lending to the decreasing of hurtful and long-run effects of the current intervention schemes.
Emerging in this scenario is the increasing figure of little RNA molecules, peculiarly microRNAs ( miRNAs ) and the established grounds sing the cardinal functions of these molecules in human diseases, chiefly malignant neoplastic disease. Despite the fact that miRNAs are involved in the tumorigenesis of a scope of different tumours [ 18 ] , the cognition about the predictive, diagnostic and/or curative mark potency of these molecules in encephalon malignant neoplastic disease, particularly medulloblastomas, is still in its beginning.
Biosynthesis and map
miRNAs are short ( 19 to 25 bases ) and evolutionary conserved RNA constructions that can adhere to the courier RNA ( messenger RNA ) of protein coding cistrons [ 19 ] . Normally, miRNAs bind within the 3 ‘ untranslated part ( UTR ) of the cistrons and they are responsible for cistron look hushing by quashing interlingual rendition or directing the sequence-specific debasement of mark messenger RNA [ 20 ] .
These little RNAs are encoded within the genome and are ab initio transcribed as primary transcripts that can be several kilobases ( kilobit ) in length. The primary transcripts are ab initio cleaved by a RNase III enzyme called Drosha in the nucleus resulting in a precursor miRNAs ( pre-miRNAs ) with ?70 bases ( National Trust ) . The pre-miRNAs are rapidly transported to the cytol through the action of exportin-5 ( Exp5 ) , a atomic exportation protein that makes usage of Ran-GTP as a cofactor. Once in the cytol the pre-miRNA is cleaved once more by another RNase III enzyme called Dicer, that produces a mature miRNAs with 22 National Trusts. Finally, the mature miRNAs are incorporated to the RNA-induced silencing composite ( RISC ) and can modulate the cistron look post-transcriptionally by partner offing with their mark messenger RNA, taking to the suppression of interlingual rendition or debasement of mark messenger RNA ( Figure 1 ) [ 21 ] .
In general, to exercise its map the perfect biding of miRNA is merely necessary at the seed part ( 2-8 National Trust in the 5 ‘ part of miRNA ) frequently found in the 3 ‘ UTR of the mark messenger RNA. Therefore, their little size and their combined capacity to the progressive fiting acknowledgment site allow miRNAs to modulate an inordinate figure of messenger RNA. It is estimated that a individual miRNA might be involved in the ordinance of more than 200 different messenger RNA. Even though, they represent less than 2 % of human cistrons, miRNAs are responsible for the ordinance of the look of ~ 30 % of the human cistrons [ 22 ] . Although, we are at the beginning of understanding the biological science of miRNAs and their action mechanism, a turning figure of surveies have revealed the importance of these little RNAs in diverse biological procedures. Furthermore, through the planetary ordinance of cistron look of cells associated with different maps, it is sensible that miRNAs can be involved in the patterned advance of several diseases, including malignant neoplastic disease [ 18, 23, 24 ] .
miRNAs in malignant neoplastic disease
Recently, miRNAs have been proposed to be involved in the pathogenesis of malignant neoplastic disease and altered miRNA look profiles are associated with forecast in several human malignant neoplastic diseases [ 24, 25 ] . The look of miRNAs is deregulated in human malignant neoplastic disease and some of them are systematically up or downregulated in more than one type of malignances. miRNA look profiles in human solid and haematological malignances have shown their possible value as tumour markers in malignant neoplastic disease patient direction [ 26 ] . The miRNAs may hold a double function in malignant neoplastic disease and can move as transforming genes ( upregulated ) or tumour suppresser cistrons ( downregulated ) [ 27 ] .
The first known coverage of deregulated miRNAs in malignant neoplastic disease showed a omission of miR-15a and miR-16-1 in chronic lymphocytic leukaemia ( CLL ) , which suggested their function as tumour suppresser cistrons in CLL [ 28 ] . Members of the let-7 miRNA household besides act as tumour suppresser cistrons and are often downregulated in human malignant neoplastic disease, taking to the upregulation of several proto-oncogenes [ 29 ] . In add-on, increased look of miRNAs may besides move as transforming genes ( oncomiRs ) , as the illustration of the miR-17~92 bunch that is overexpressed in several tumours [ 30 ] . The deregulating of miRNAs look is the chief mechanism of loss or addition of map of these molecules in malignant neoplastic disease cells. In fact, the activation of oncogenic written text factors, such as MYC, represents an of import mechanism for altered miRNA look [ 31 ] . These informations strongly suggest that miRNAs play an of import function in human malignant neoplastic disease.
The mechanisms underlying miRNA cistron deregulating in malignant neoplastic disease are non good understood. However, the look of miRNAs can be modulated in response to assorted stimulations as oncogene activation or DNA harm [ 32 ] . Sing that more than one-half of the miRNAs have been aligned to genomic delicate sites or chromosomal parts associated with malignant neoplastic diseases [ 33 ] , genome transcript figure aberrances ( CNA ) is considered one of the major causes of miRNA deregulating in tumours [ 34 ] .
microRNAs IN MEDULLOBLASTOMAS
Despite the importance of miRNAs in tumorigenesis [ 35 ] merely late these molecules have been involved in the medulloblastoma pathogenesis ( Table I ) . In the following subdivision we will turn to the cognition of clinical and functional impact of miRNAs deregulating in medulloblastomas.
Diagnostic and predictive biomarkers
The first study depicting the engagement of a miRNA in medulloblastomas showed that miR-124, an enriched miRNA in normal encephalon, presented reduced look in MB samples. miR-124 modulates cell rhythm ordinance by aiming CDK6, a well-known inauspicious predictive marker in MB [ 36 ] . The low look of miR-124 in MB was corroborated in a subsequent study and it was suggested as a regulator of cell glycolysis ( bioenergetics trademark of malignant neoplastic disease ) by aiming SCL16A1 ( solute bearer household ) [ 37 ] . In a series of 61 MB samples, miR-199b-5p was described as upregulated in non-metastatic instances and its high look showed an association with a better overall endurance [ 38 ] . A good known miRNA associated with metastasis in several tumours, miR-21 is upregulated in MB samples. It was observed that miR-21 regulates the look of the metastasis suppresser PDCD4 and deficiency of this protein is indispensable for metastatic airing in MB [ 39 ] .
In a high throughput attack, MB of the SHH subgroup ( increased GLI1 look ) was associated with a notably downregulation of 28 miRNAs, proposing a tumour suppresser map moving through the ordinance of the SHH tract. In fact, miR-125b and miR-326 were identified as regulators of SMO ( smoothened ) and miR-324-5p of GLI1, that are cardinal activator constituents of the SHH tract. In the same work, miR-214 was demonstrated with an increased look in SHH subgroup tumours and its oncogenic map could be associated with the ordinance of SUFU, a SHH tract inhibitor [ 40 ] . Mutants or loss of map of SUFU are associated with medulloblastoma development [ 12 ] . In a 2nd attack, the same writers reported the look of 248 miRNAs in a representative aggregation of MB histopathological discrepancies with different clinical characteristics and molecular features. Consequently, the writers focused in 86 miRNAs already reported as expressed in neural tissues and/or associated with tumour. They observed that the bulk of the miRNAs were downregulated in MB samples [ 41 ] . The look of merely four miRNAs ( miR-let7g, miR-19a, miR-106b and miR-191 ) allowed the differentiation of MB histotypes ( authoritative, anaplastic and desmoplastic ) . Besides, the differential look of a group of six miRNAs ( miR-10b, miR-135a, miR-135b, miR-125b, miR-153, miR-199b ) was associated with tumours overexpressing ERBB2 and a group of three ( miR-181b, miR-128a, miR-128b ) with tumours overexpressing MYC. Furthermore, miR-31 and miR-153 were differentially expressed among high hazard and criterion hazard MBs and they were suggested as hapless predictive markers of the disease [ 41 ] . A recent study demonstrated the increased look of miR-182 and miR-183 in association with metastatic non-SHH MB subgroups [ 42 ] .
A scheme utilizing deep sequencing of MB mouse theoretical accounts revealed the increased look of 26 miRNAs and reduced look of 24 miRNAs in MB tissue compared to normal encephalon [ 43 ] . In the group of downregulated miRNAs were included miR-124a and miR-128, antecedently identified by others [ 36, 37, 41 ] . Interestingly, among the upregulated miRNAs were nine members of the miR-17~92 bunch, a group of miRNAs implicated as transforming genes in several tumours [ 44 ] . The look of three of these miRNAs ( miR-19a, miR-20 and miR-92 ) was associated with the SHH subgroup in MB human samples [ 43 ] . These consequences were besides corroborated in another study utilizing microarray that identified miR-18a, miR-19b, miR-20a and the paralogous miR-106a ( miR-106a/363 bunch ) as upregulated in activated SHH tract MB samples. In this work, it was shown that the overexpression of these miRNAs occurred as a consequence of an elaboration on chromosome 13q31.3 ( miR-17~92 bunch parts ) in a subset of tumours. Furthermore, the high look of these miRNAs was associated with NMYC elaboration in these samples [ 45 ] . Novel perennial chromosome 8q24.22-q24.23 elaboration was besides described in MB and it was independent of MYC elaboration ( 8q24.1 ) . In this chromosome part, crossing 3 megabases ( Mb ) , are mapped two miRNAs ( miR-30b and miR-30d ) that presented higher look in 54 % and 12 % of the MB samples compared to normal cerebellum, severally. The look of these miRNAs was purely related to the presence of the chromosome elaboration [ 46 ] .
A miRNA profiling that evaluated 365 miRNAs was capable to place the four distinguishable molecular subgroups of MBs. Additionally, the writers identified a particular and robust miRNA signature ( 16 miRNAs ) associated with WNT subgroup and mutants of CTNNB1. Overexpression of miR-23b, miR-148a, miR-182, miR-193a, miR-224 and miR-365 was validated in WNT subgroup of MBs. The writers suggested that the look degree of these possible tumor/metastasis suppressive miRNAs is likely to find the response to intervention and would be an of import biomarker for hazard stratification in a WNT subgroup [ 47 ] . A similar analysis utilizing microarrays revealed a miRNA signature based on the differential look of 9 miRNAs ( 4 upregulated and 5 downregulated ) that distinguish MB samples from normal cerebellum specimens. Interestingly, four of these miRNAs ( miR-17, miR-100, miR-106b and miR-218 ) were non reported as differentially expressed in old surveies sing MBs [ 48 ] .
Since the development of high throughput techniques, the first studies sing integrative genomic analysis in medulloblastomas have begun to be published with really promising consequences. Recently, the look of 663 miRNAs was evaluated in MB primary samples, MB cell lines, nervous root cells ( CD133+ ) and nervous primogenitor cells ( CD133- ) . This analysis revealed 33 differentially expressed ( 21 upregulated and 12 downregulated ) miRNAs in MB primary samples compared to CD133+ cells. An integrative and functional analysis of negatively correlated predicted marks of both up and downregulated miRNAs revealed 106 altered important tracts in MB. Enrichment was observed in tracts modulating neural migration, nervous system development and cell proliferation [ 49 ] . Based on cistron look, other study proposed that MB nowadayss six molecular subgroups. The writers identified specific miRNA profiles that were correlated with the messenger RNA signatures. Integrating mRNA look, miRNA look and cistron copy figure analysis they showed that a specific group of MB characterized genetically by MYC transcript figure additions and transcriptionally by the enrichment of photoreceptor tracts and increased look of miR-183~96~182 bunch is associated with a hapless forecast [ 13 ] .
miRNA profiling has besides been used to measure differences among all paediatric CNS tumours. Actually, an attempt was driven in the analysis of 470 miRNAs in a assortment of paediatric CNS malignances including untypical teratoid/rhabdoid tumour ( AT/RT ) , glioblastoma ( GBM ) , MB, ependymoma ( EPN ) and pilocytic astrocytoma ( PA ) . Three miRNAs ( miR-25, miR-129 and miR-142-5p ) were found to be differentially expressed in every tumour type when compared to the normal controls. The analysis besides revealed important differences in miRNA look between the malignant neoplastic disease groups and allowed the assignment of specific miRNA signatures to each CNS tumour type. Performing unsupervised hierarchal bunch analysis utilizing miRNA look of the 20 % most variable miRNAs demonstrated that paediatric tumour samples formed tighter diagnostic groups, with the exclusion of GBMs that presented a widespread bunch [ 50 ] . The same bunch form was observed in a subsequent study utilizing a different proficient attack, which was interesting sing the heterogeneousness of encephalon tumours. These consequences could be explained by the greater grade of heterogeneousness of GBMs, the most aggressive and non-responsive paediatric encephalon tumour [ 51 ] .
Recently, in a different attack, miRNAs sequences were assayed for base brace omissions, elaborations and mutants. The writers evaluated nine specific miRNA ( miR-33b, miR-135a-1, miR-135a-2, miR-135b, miR-186, miR-200b, miR-512-2, miR-548d-1 and miR-548d-2 ) that were selected on the footing of the presence of possible mark sequences within the 3? UTR of the MYC messenger RNA. At least 73 % ( 35/48 ) of the analyzed MB samples presented an change in one of the nine miRNAs, bespeaking an alternate mechanism of MYC overexpression in MB [ 52 ] .
The information presented so far, sing paediatric medulloblastoma, indicate that miRNA profiling could be an of import tool to back up hazard stratification and molecular categorization of MBs. Meanwhile, due to the high heterogeneousness of this specific group of tumours, new attempts with greater cohorts and/or the usage of new engineerings ( particularly deep sequencing ) , which allow a deeper coverage of miRNAs, will be important to turn to this issue in the hereafter.
Potential curative marks: miRNA functional function in medulloblastomas
Current grounds demonstrates the engagement of miRNAs in the pathogenesis of MB. Under this fortunes, the transition of this RNA molecule points to the possibility of the usage of new curative schemes for MB intervention. Indeed, several groups around the universe evaluated the ectopic transition of miRNAs in MB. For curative intents distinguishable attacks can be used, depending on the purpose. Sing miRNAs downregulated in tumours and that map as tumour suppressers, the curative attack relies on the ability to reconstruct the look of the miRNA utilizing man-made RNA semidetached house that will be recognized by RISC composite and will mime miRNA maps [ 53 ] . For miRNAs that map as transforming genes, being upregulated in malignant neoplastic disease, the scheme should be the direct mark for silence or decrease of look through the usage of antisense oligonucleotides with specific alterations that bind specifically to the miRNA molecule impairing its maps [ 54 ] . Experimentally up or downregulated miRNAs and its effects in MB are shown in Table II.
The first grounds was demonstrated for the re-expression of miR-124 in MB cell lines. Transient look of a man-made miRNA that mimics miR-124 map resulted in a decrease in MB cell proliferation [ 36, 37 ] , non mediated by cell programmed cell death [ 36 ] . The re-expression of miR-124 was associated with downregulation of CDK6 [ 36 ] and SLC16A1 [ 37 ] , which could explicate the cell growing damage. Ectopic look of miR-129 was proposed to modulate CDK6 ensuing in cell growing apprehension by the same mechanism [ 55 ] . The re-expression of miR-199b-5p resulted in lessened MB tumour growing in vitro and in vivo. miR-199b-5p Restoration besides impairs the ingrafting potency of MB cell lines by diminishing the stem-cell-like ( CD133+ ) subpopulation of MB cells, as a effect of downregulation of HES1, a cardinal effecter of Notch signaling pathway [ 38 ] . The lessening in cell proliferation was besides achieved when miR-125b, miR-324-5p and miR-326 were ectopically expressed in vitro, by the targeting of SHH signaling pathway. The same molecular regulative circuitry could be observed in the development of cerebellar nervous primogenitors, in which the miRNAs look additions through distinction stairss thereby leting cell ripening and growing suppression [ 40 ] . In another study, the same writers demonstrated that the deliverance of look of miR-9 and miR-125 in MB cell lines, utilizing retinoic acid intervention, was associated with cell growing apprehension and programmed cell death and with a lessening in cell proliferation while aiming the abbreviated isoform of the neurotropin receptor TrkC ( t-TrkC ) [ 41 ] , a deregulated mechanism already described for neuroblastoma [ 56 ] . Enforced look of the full miR-17~92 bunch in a MB mouse theoretical account led to an early tumour development in the animate beings. miR-19a and miR-92 were expressed at elevated degrees in the tumours and look analysis revealed the loss of look of the wild type Ptch1 allelomorph, a bona fide tumour suppresser cistron. Besides, the tumours presented increased degrees of GLI1 bespeaking the constituent activation of SHH signaling pathway. Tumors showing this miRNA bunch were sensitive to cyclopamine, which decreased cell proliferation in add-on to the downregulation of GLI1 [ 43 ] . Other work demonstrated that the higher look of miR-17~92 bunch was driven by NMYC overexpression, which acts in synergism for the increased proliferation in a SHH subgroup of MB tumour cells. This manner, the writers suggested that miR-17~92 can supply a selective growing advantage to MB cells [ 45 ] . Exogenous overexpression of miR-193 and miR-224, the most upregulated miRNAs in WNT subgroups, was found to suppress cell proliferation, anchorage-independent growing and increases radiation sensitiveness in MB cell lines. These features refer to the better response to intervention and first-class forecast in this subgroup of MB tumours [ 47 ] . Reintroduction of miR-128a in MB cell lines was associated with reduced cell growing through the downregulation of Bmi-1 and induced cell aging by the upregulation of reactive O species ( ROS ) degrees [ 57 ] . miR-34a impairs cell proliferation, invasion and endurance by aiming c-MET ( MB and gliomas ) and Notch-1, Notch-2, CDK6 ( in glioma ) [ 58, 59 ] . Recently, miR-34a look was shown to bring on programmed cell death, aging and chemotherapy sensitiveness of MB cell lines thought a positive feedback mechanism advancing MAGE-A downregulation with attendant increasing of the look of p53 and its marks ( p21/WAF1/CIP1 ) [ 60 ] . Besides, miR-34a look was related to cell proliferation suppression, initiation of programmed cell death and distinction in MB cell lines by the ordinance of Dll1, an of import effecter protein of the Notch signaling pathway [ 61 ] .
Because most of the studied miRNAs are downregulated in medulloblastomas, the huge bulk of the studies evaluated the re-expression of these miRNAs in MB cells. However, there are few illustrations in which suppression of miRNA resulted in alterations of biological procedures in MB cells. So, suppression of miR-21 impaired the cell ability of migration and invasion in MB cell lines due to the initiation of the look of PDCD4, which negatively regulated µPAR and integrin proteins ( indispensable for cell invasion ) and positively regulated E-cadherin and TIMP-2 proteins ( cell migration control ) [ 39 ] . Knockdown of miR-182 and miR-183 was related with decreased cell migration in MB cell lines. In heterograft experiments, MB cells overexpressing miR-182 were able to occupy normal tissue and capable to metastasise to the leptomeninges, this manner stand foring a really aggressive MB phenotype [ 42 ] .
So far, several attempts reported the importance and impact of miRNA transition in a series of cellular biological procedures. These surveies are of great value to clear up the biological science of medulloblastomas and more of import to specify fresh marks for curative development in future attacks.
The development of high throughput methodological analysiss and analysis of all biological molecules is taking to a wider and deeper apprehension of their engagement and cross-talk in several human diseases. We here highlighted the progresss and groundss demoing miRNAs as a cardinal molecular effecter in medulloblastoma pathogenesis and the possible usage of these molecules as biomarkers of diagnosing, forecast and therapeutics.
Despite the new penetration and enthusiasm in medulloblastoma sing miRNAs, the ordinance of these molecules as a curative attack is still in its babyhood and cautiousness most be taken during their design. Even more, one time the bulk of these findings were observed in in vitro surveies, more research is needed in vivo in MB theoretical accounts and in extended pre clinical attacks to clear up the cardinal function of miRNAs in the pathogenesis of MB and to set up its safety and existent importance as a curative mark for future intercessions. Furthermore, miRNAs present the characteristic to modulate 100s of messenger RNA, thereby the designation of the marks involved in the deregulated miRNA-mRNA web will be of polar importance to clarify fresh molecular tracts that could besides be used for new curative attacks in MB. A great challenge relies on the ability of the research workers in conveying the findings observed in big graduated table surveies to routine diagnostic. It is duty of these professionals to place a concise and smaller panel of relevant miRNAs that could be evaluated by everyday techniques for a faster appraisal of the related MB subtype ensuing in a better direction of the intervention for the patients.
Concluding, miRNAs are major regulators of mRNA look degrees and are often deregulated in medulloblastomas. Its word picture, biological and clinical impact in medulloblastomas is far from complete, but their unraveling is important in order to place fresh clinical biomarkers and development of new and more effectual hereafter curative options conveying hope for the patients affected by this blue disease.
Declaration of involvement statement
The writers have no struggle of involvement to unwrap. The writers entirely are responsible for the content and authorship of the paper.