The Genetic Background of Abnormalities in Metabolic Pathways of Phosphoinositides and Their Linkage with the Myotubular Myopathies, Neurodegenerative Disorders, and Carcinogenesis

Myo-inositol belongs to one of the sugar alcohol groups known as cyclitols. Phosphatidylinositols are one of the derivatives of Myo-inositol, and constitute important mediators in many intracellular processes such as cell growth, cell differentiation, receptor recycling, cytoskeletal organization, and membrane fusion. They also have even more functions that are essential for cell survival. Mutations in genes encoding phosphatidylinositols and their derivatives can lead to many disorders. This review aims to perform an in-depth analysis of these connections. Many authors emphasize the significant influence of phosphatidylinositols and phosphatidylinositols’ phosphates in the pathogenesis of myotubular myopathies, neurodegenerative disorders, carcinogenesis, and other less frequently observed diseases. In our review, we have focused on three of the most often mentioned groups of disorders. Inositols are the topic of many studies, and yet, there are no clear results of successful clinical trials. Analysis of the available literature gives promising results and shows that further research is still needed.


The Family of Phosphoinositol and Phosphoinositides
Phosphatidylinositol (PtdIns), the starting point of PIP metabolism, is a ubiquitous phospholipid in eukaryotic cells present in various proportions according to the type of membrane.PIPs are all metabolized directly or sequentially from PIs [8].The structural formulas of phosphoinositol and phosphoinositides are shown in Table 1.
Table 1.Structural formulas of phosphoinositol and phosphoinositides.ulating hormone (TSH), luteinizing hormone (LH), follicle-stimulating hormone (FSH), and insulin [1,4,6,7].The interconversions between this group of molecules are conducted by crucial enzymes, whose dysfunction can lead to severe abnormalities, disorders, and illnesses [4,6].This work aims to present and analyze the documented data concerning the association of cyclitol with pathological processes such as carcinogenesis, myotubular myopathies, and neurodegenerative disorders.

The Family of Phosphoinositol and Phosphoinositides
Phosphatidylinositol (PtdIns), the starting point of PIP metabolism, is a ubiquitous phospholipid in eukaryotic cells present in various proportions according to the type of membrane.PIPs are all metabolized directly or sequentially from PIs [8].The structural formulas of phosphoinositol and phosphoinositides are shown in Table 1.

Routes and Interconversions of PIs
As previously mentioned, PI is a key compound and precursor of PIPs, which are all metabolized directly or sequentially from PI [8].The scheme below presents detailed metabolic routes and interconversions of the PIPs family.The detailed analysis of genes and encoded enzymes is described in the table below Figure 1.

Routes and Interconversions of PIs
As previously mentioned, PI is a key compound and precursor of PIPs, which are all metabolized directly or sequentially from PI [8].The scheme below presents detailed metabolic routes and interconversions of the PIPs family.The detailed analysis of genes and encoded enzymes is described in the table below Figure 1.PI itself is a product of the synthesis of cytidine diphosphate diacylglycerol (CPD-DAG) and MI.The reaction is conducted by PI synthase, also called phosphatidylinositol synthase 1 (PIS1) [9,10].Then, PI phosphorylates are converted into phosphatidylinositol 3-phosphate (PI3P/PtdIns3P) [11].The conversion is catalyzed by phospha- PI itself is a product of the synthesis of cytidine diphosphate diacylglycerol (CPD-DAG) and MI.The reaction is conducted by PI synthase, also called phosphatidylinositol synthase 1 (PIS1) [9,10].Then, PI phosphorylates are converted into phosphatidylinositol 3-phosphate (PI3P/PtdIns3P) [11].The conversion is catalyzed by phosphatidylinositol 3 kinase (PI3K) and class III PI 3-kinase-vacuolar protein sorting 34 (Vps34) [12].
As previously mentioned in the figure above, every phosphorylation or dephosphorylation reaction is conducted with enzymes such as phosphatidylinositol kinases and phosphatases.Every gene has a specific gene ID.The list of genes, encoded proteins, and their functions are presented in Table 2.As we can see in the table above, dysfunction of genes or encoded enzymes can lead to various defects and disorders.The range of illnesses varies widely.In this paper, we will focus on MTM and MTMR gene dysfunctions, neurodegenerative diseases, and carcinogenesis.

Myotubular Myopathy
Centronuclear myopathy (CNM) is one of the disorders affecting the nervous and muscular systems characterized by two main criteria: symptoms of clinically congenital myopathy and multiple centrally located nuclei in muscle cells confirmed by biopsy [100].
Myotubularin 1 (MTM1) is an enzyme involved in the regulation of phosphoinositides, which are important molecules in cell signaling and membrane trafficking.Mutations in the myotubularin gene MTM1 which cause human myotubular myopathy dramatically reduce the phosphatase's ability to dephosphorylate PI3P, affecting the levels of inositol lipid PI3P in myogenesis.In addition, it inhibits the transport of EGFRs to lysosomes, causing the formation of large endosomal vacuoles through the effects of myotubular phosphatase and its interaction with PI(3,5)P 2 .In most patients, mutations in the MTM1 gene are associated with the recessive X chromosome form (Xq28) [40,41,43,45], while autosomal dominant and recessive forms primarily involve mutations in the dynamin 2 (DNM2) gene on chromosome 19p13.2and the amphisin 2 gene (BIN1) on chromosome 2q14 [100].
XLMTM is a specific subtype of myotubular myopathy, which is a rare genetic neuromuscular disorder characterized by severe muscle weakness and hypotonia (low muscle tone) caused by mutations in the MTM1 gene.As a result, the condition is X-linked, and therefore mainly affects males (2/100,000 male births) [44,45,100].The symptoms of XLMTM are generally similar to those of other myotubular myopathy subtypes, but they tend to be more severe in males.Newborn males with XLMTM often have profound muscle weakness, leading to severe respiratory difficulties and respiratory failure.The weakened muscles can affect an individual's ability to breathe, swallow, and move, which makes it essential for affected infants to receive immediate medical attention and respiratory support.Patients with XLMTM can be classified into mild, intermediate, or severe phenotypes based on the amount of ventilation support required.Long-term patients with a severe phenotype are often unable to move and require ventilatory support.Patients with intermediate and mild phenotypes breathe independently for at least a few hours a day and can walk.Moreover, such patients often have coexisting liver disorders [45].
Ross et al. used tissues from human patients and animal models, including dogs with XLMTM, which received increasing doses of recombinant AAV8 vector restoring MTM1 expression (rAAV8-cMTM1).They found that administration of rAAV8-cMTM1 at doses higher than 2.5 × 1013 vg kg −1 allowed full recovery of all these cellular defects in dogs with XLMTM [101].They came to a similar conclusion in their work with an experiment in mice after administration of rAAV8 [102], short-term replacement of myotubularin with a prototypical targeted protein replacement agent (3E10Fv-MTM1) [103], or used the selective inactivation of PI3KC2β kinase activity [104], which improved contractile function and muscle pathology, and thus is associated with a highly promising treatment potential for myotubular myopathy.
BIN1 modulation as well as DMN2 reduction may also be an effective treatment strategy for XLCNM, as it restores adequate myofibrillar integrity [105,106].
Normal cell function is regulated by myotubularins belonging to the phosphatidylinositol 3-phosphate phosphatase family PI3P, identified by positional cloning of the MTM1 gene in patients suffering from X-chromosome-associated myotubular myopathy (with reported specificity toward PI3P) and the myotubularin-related protein 2 (MTMR2) gene (hydrolyzes both PI3P and PI(3,5)P 2 together with myotubularin-related protein 3 (MTMR3) in patients suffering from CMT4B) [42].Myotubularin-related protein 1 (MTMR1) was shown to use PI3P and/or PI(3,5)P 2 as substrates.The overall structure was very similar to the previously described structure of MTMR2 [46,47].It turns out that MTMR2, whose abnormal function is revealed in the neurodegenerative Charcot-Marie-Tooth disease type 4B2, is also highly specific for PI3P as a substrate.In addition, the myotubularin-related phosphatases MTMR1, MTMR3, and myotubularin-related protein 6 (MTMR6) also dephosphorylate PI3P [47].MTMR3 shows great similarity to myotubularin, primarily due to the catalytic domain, in addition to also having an extension at the C-terminal end that contains the FYVE domain.It can hydrolyze PI3P and PI(3,5)P 2 and set the pathway for the production of PI5P in the cell.Overexpression of catalytically inactive MTMR3 in cells results in significant formation of vacuolar compartments [48,49].
MTMR3 and myotubularin-related protein 4 (MTMR4) are protein tyrosine phosphatases that dephosphorylate position 3 in PI and generate PI5P from PI(3,5)P 2; as well as PI from PI3P.These regulate the production of PI3P, which plays a key role in inhibiting the DNA immune response by regulating the transport of STING, which is an activator of the protein tank-binding kinase (TBK1) that catalyzes the phosphorylation of interferon regulatory factor 3 (IRF3) [51].Myotubularin-related protein 4 (MTMR4) present in endosomes and regulating their recirculation process has been identified as a novel factor interacting with the ubiquitin ligase neural precursor cell expression protein 4 (Nedd4), which is downregulated during development.MTMR4 expression decreased in atrophied muscles, while Nedd4 expression increased, and MTMR4 was ubiquitinated by Nedd4, indicating that this new relationship between MTMR4 and Nedd4 may underlie the biological process of muscle degradation [52][53][54].MTMR8/R9 complex controls a cellular pool of PI3P that has been proposed to be essential in autophagy, a conserved intracellular process for the degradation of cytoplasmic proteins or organelles.Overexpression of both MTMR8 and MTMR9 resulted in a significant increase in the level of p62, a protein that is degraded in autophagosomes and is used to monitor autophagy.Mutations in both active and inactive myotubularins (which may play a regulatory role) are associated with diseases such as myotubular myopathy, Charcot-Marie-Tooth (CMT), and others [56,63].
CMT4B is a large group of heterogeneous diseases that are inherited in an autosomal recessive manner and have a progressive sensorimotor neuropathy.The etiopathological basis of MTMR2 and MTMR13, which affect vesicular transport in Schwann cells, is one where the loss of these proteins can lead to uncontrolled myelin folding and, ultimately, to the development of CMT4B disease.Among them, CMT4B is distinguished as having three forms associated with myotubularin family genes: CMT4B1 (MTMR2 located on chromosome 11q22), CMT4B2 (MTMR13/SBF2 located on chromosome 11p15) and CMT4B3 (MTMR5/SBF1) [107,108].
The disease is childhood-onset and manifests itself mainly through cranial nerve involvement, including glaucoma, vision loss, and other severe disabilities [109].

Neurodegenerative Diseases
Neurodegenerative diseases are still a significant problem in our aging society [110].Their occurrence is associated with many factors, such as genetic defects and disturbances in immunological processes.Many neurodegenerative diseases have their cause in the accumulation of extra-and intracellular deposits in the nervous system [111].Among these disturbances, we can distinguish amyloidoses, tauopathies, α-synucleinopathies, and TDP-43 proteinopathies [110].
Abnormalities in PIP metabolism and their linkage to neurodegenerative disorders has, to date, been mentioned by many authors.Researchers have described the influence of PIPs on the incidence of central nervous system (CNS) diseases through changes in the levels of selected PIPs as well as the enzymes catalyzing their interconversions.The vacuole 14 protein homolog (Vac14) is a frequently mentioned protein, also known as the Associated Regulator of PIKfyve (ArPIKfyve) [112,113].
Zhang et al. postulate that PI(3,5)P 2 is critical to neuronal health.Vac14 protein is a regulator of the signaling lipid PI(3,5)P 2 synthesis.The loss of Vac14 results in neurodegeneration processes in the midbrain and peripheral sensory neurons of mice models [114].Other proteins, such as Fab1/PIKfyve and Fig4/Sac3, also have a regulatory role in the PI(3,5)P 2 biosynthesis and its relationship with PI5P.Mutations in the genes encoding these proteins lead to the occurrence of neurological diseases, including amyotrophic lateral sclerosis (ALS) and CMT syndrome [115].For example, Fab1 binds the PI3P and allows it to be converted to PI(3,5)P 2 , and consequently, the appropriate levels of PI(3,5)P 2 affect the proper functioning of cells in the nervous system.The complex responsible for the Fig1 activity consists of Vac14, Vac7, Fig4, and Atg18 proteins.
Other authors have suggested that changes in the regulation of the phosphatidylinositol-3 kinase (PI3-K) are also linked to many neurodegenerative diseases.One of them is a Nieman-Pick type C disease (NPC), caused by mutations in the NPC1 and NPC2 genes.This disorder results in the deposition of neurofibrillary tangles in the CNS, the occurrence of which has been suggested to be associated with increased levels of specific kinases such as phosphatidylinositol 3-kinase (PI3K), glycogen synthase kinase (GSK-3β), and protein kinase B (Akt/PKB).The PI3K cascade leads to the activation of Akt and the inactivation of GSK-3β.Research on the NPC1-deficient mice model has shown major disruptions in the PI3K cascade.The inactivated GSK-3 and phosphorylated Akt were elevated in the neuronal cells, which indicated an unusual level of activity of PI3K in the NPC1-deficient mice brains [116].Cathepsin D (CD) is one of the key lysosomal proteases.A lack of CD results in a neurodegenerative pediatric disease known as neuronal ceroid lipofuscinosis (NCL/Batten disease).In the research on CD-deficient brains in mice by Walls et al., a decrease in PI3K was observed [117].
The phosphatidylinositol 5-phosphate 4-kinases (PI5P4Ks) have been studied for possible therapeutic effects in many diseases, including neurodegenerative ones.PI5P4Ks regulate the cellular level of PI5K and generate a specific pool of PI(4,5)P 2 products.PI5P4K has free isoforms: α, β, and γ.All of them have different abilities concerning receptor recycling, gene expression regulation, insulin signaling, and cell stress responses [118].PI5P4Kγ overactivity has a linkage to carcinogenesis and neurodegenerative diseases; thus, it is suggested as a potential therapeutic target.The novel proposed substances in the treatment of neurodegenerative diseases are specific PI5P4Kγ inhibitors and degraders-NIH-12848 and JWZ-1-80 [118,119].

Carcinogenesis
Carcinogenesis is a highly complex process involving environmental factors and gene mutations [123].Critical mutations involve proto-oncogenes, tumor-suppressor genes, and DNA-repair genes.Key principles of cancer are uncontrolled proliferation, metastasis, apoptotic loss, and angiogenesis [124].Here we present another gene reported to have a contribution to carcinogenesis.
Mutation of myotubularin-related protein 7 gene (MTMR7) has been described as contributing to colorectal cancer (CRC) development.MTMR7 itself participates in decreasing insulin-mediated activation of Akt and ERK1/2 signaling, resulting in proliferation reduction of human CRC cells.In human colorectal cancers, MTMR7 has been down-regulated, which has been related to a poor prognosis [62].
PI3K mis-activation has been widely reported in cancer diseases [69,125,126].Its activation can be signaled via various pathways including mTOR, JAK2/STAT5, Akt, or RTK [69], which gives a promising location for the development of target drugs, but can also be responsible for therapy resistance.Phosphatidylinositol 3-kinase catalytic subunit type 3 (PIK3C3), a subunit of the PI3K complex, takes part in the formation, initiation, and maturation of autophagosomes [127].It has been shown that, in the condition of oncogenic herpesvirus KSHV infection, the expression of PIK3C3 is upregulated, and takes part in tumor progression and metastasis [68].It is worth mentioning that various drugs have been developed to interfere with the PI3K/AKT/mTOR axis.So far, pan and isoform-specific PI3K inhibitors have been developed, and some have had promising results [128][129][130].The PI3K inhibitor copanlisib has been clinically approved for follicular lymphoma, and idelalisib for chronic lymphocytic leukemia, follicular lymphoma, and small lymphocytic lymphoma [128,131].There is also data suggesting the usage of copanlisib against solid tumors and other non-Hodgkin lymphomas may be warranted [129,132].
The PKR/PI4K2A axis, which takes part in the clearance of misfolded proteins in lysosomes, has been revealed to be a potential drug target-inhibiting tumor growth in the lung and breast [74].
The phosphatase and tensin homolog (PTEN) is a commonly known cancer suppressor that acts mainly via inhibiting PI3K/Akt activation [133].This molecule controls the cell cycle, driving apoptosis among pathological conditions [134].Mutations of PTEN are commonly known for the link to the PTEN hamartoma tumor syndrome (PHTS), which is characterized by a greater risk of cancer occurrence, includes Cowden syndrome (CS), Bannayan-Riley-Ruvalcaba syndrome (BRRS), and PTEN-related Proteus syndrome (PS) [135,136].To date, there has been some effort put into establishing PTEN as a potential drug target.For example, the research of inducing PTEN de-ubiquitination has resulted in the cessation of its degradation, which subsequently has driven PTEN activity in inhibiting the AKT signaling pathway and tumor growth [137,138].Other members of the PTEN family, TPTE and PTEN homologous inositol lipid phosphatase (TPIP), reported to occur in at least three forms (alpha, beta, and gamma), have been reported to act as tumor suppressors, as the overexpression of this molecule leads to cell proliferation inhibition and apoptosis induction [99,139].
Upregulation of Inositol polyphosphate 4-phosphatase type II (INPP4B) leads to colon cancer cell proliferation, growth, and survival, which is associated with increased activation of PI3K/Akt and SGK3.Of note, in this study, it turned out that PTEN inhibited INPP4B-mediated PI3K signaling activation [36].This finding is consistent with the study on pancreatic cancer using mice models, where INPP4B had additionally been found to inhibit apoptosis.In addition, INPP4B expression was correlated with a positive resection margin, poor overall survival, and disease-free survival.Moreover, INPP4B could be a potential diagnostic marker of use almost equal to or greater than that of other established diagnostic markers, such as CA125, AFP, and CEA [140].
Mutation of SACM1L, a gene for phosphatidylinositol-3-phosphatase SAC1, is common among cancers [95].In the breast cancer model, the loss of SAC1 function has led to their increased migration and invasion.Interestingly, the loss of SAC1 corresponded with increased levels of Golgi PI4P [141].
A protein and lipid kinase FYVE finger-containing phosphoinositide kinase (PIKfyve) has been described as taking a crucial role in regulating the endosomal system and transport in the cells, as well as to participate in downstream membrane signals, remodeling the cytoskeleton and even innate immune responses [142,143].Mutation in gene PIKFYVE for this kinase has been investigated as having a potential role in carcinogenesis, and there is also some evidence that this gene can be an oncogene [144].This makes PIKfyve a potential treatment target.Studies on PIKfyve inhibitors showed promising results in reducing multiple myeloma and non-Hodgkin lymphoma cell viability via inducing cell vacuolization, rupture of the plasma membrane, and non-apoptotic death [86,145,146].

Conclusions
Summing up, MI is a forerunner of many derivatives, including PIs, PIPs, IPs, GPIs, and IPGs, which take part in transmitting various signals in the eukaryotic cells.That is why gene mutations for these molecules can play a pivotal role in the development of many widespread diseases, such as cancers or neurodegenerative disorders.Cancer diseases and neurodegenerative disorders, such as AD, are wide-spread problems among developing countries, and the number of patients has been increasing for many years.
Consequently, there is a growing need for studies to better explain the development of these diseases.In our review, we focused on phosphatydylinositols and the linkage between their gene mutations and connected disorders.These alterations contribute to, e.g., carcinogenesis, enabling cancer cells to get their key principles, such as uncontrolled proliferation, apoptosis inhibition, and increased migration.Other diseases whose pathogenesis is related to the improper functioning of PIPs are myotubular myopathies.Their impaired function of PIPs disrupts myogenesis, cellular compartmental organization, and the accumulation of improper metabolites, leading to macroscopic muscle defects.
In neurodegenerative disorders, it has been described that mutations of genes for PIPs lead to disruption of intracellular trafficking and, once again, the accumulation of defective metabolites.Subsequently, it disturbs the very complex and active functioning of the neural cells.All of the studies conducted on PIP gene mutations and mentioned in our review give a better insight into many common diseases.The connections described make PIPs and their genes potential drug targets and diagnostic markers in the disorders mentioned above.
Our review provides an expanded overview of the most important links between PIP mutations and the occurrence of diseases.To obtain more information, further studies focused on the particular mutations are needed.

Table 1 .
Structural formulas of phosphoinositol and phosphoinositides.

Table 1 .
Structural formulas of phosphoinositol and phosphoinositides.

Table 2 .
List of genes and proteins with their functions.