Proper assembly and disassembly of both immature and mature HIV-1 hexameric lattices are critical for successful viral replication. These processes are facilitated by several host-cell factors, one of which is myo-inositol hexaphosphate (IP6). IP6 participates in the proper assembly of Gag into immature hexameric lattices and is incorporated into HIV-1 particles. Following maturation, IP6 is also likely to participate in stabilizing capsid protein-mediated mature hexameric lattices. Although a structural-functional analysis of the importance of IP6 in the HIV-1 life cycle has been reported, the effect of IP6 has not yet been quantified. Using two in vitro methods, we quantified the effect of IP6 on the assembly of immature-like HIV-1 particles, as well as its stabilizing effect during disassembly of mature-like particles connected with uncoating. We analyzed a broad range of molar ratios of protein hexamers to IP6 molecules during assembly and disassembly. The specificity of the IP6-facilitated effect on HIV-1 particle assembly and stability was verified by K290A, K359A, and R18A mutants. In addition to IP6, we also tested other polyanions as potential assembly cofactors or stabilizers of viral particles.IMPORTANCE Various host cell factors facilitate critical steps in the HIV-1 replication cycle. One of these factors is myo-inositol hexaphosphate (IP6), which contributes to assembly of HIV-1 immature particles and helps maintain the well-balanced metastability of the core in the mature infectious virus. Using a combination of two in vitro methods to monitor assembly of immature HIV-1 particles and disassembly of the mature core-like structure, we quantified the contribution of IP6 and other small polyanion molecules to these essential steps in the viral life cycle. Our data showed that IP6 contributes substantially to increasing the assembly of HIV-1 immature particles. Additionally, our analysis confirmed the important role of two HIV-1 capsid lysine residues involved in interactions with IP6. We found that myo-inositol hexasulphate also stabilized the HIV-1 mature particles in a concentration-dependent manner, indicating that targeting this group of small molecules may have therapeutic potential.
- MeSH
- genové produkty gag - virus lidské imunodeficience chemie genetika metabolismus MeSH
- HIV-1 chemie genetika MeSH
- missense mutace MeSH
- polymery chemie MeSH
- sestavení viru * MeSH
- substituce aminokyselin MeSH
- vztahy mezi strukturou a aktivitou MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
As compounds of natural origin enter human body, it is necessary to investigate their possible interactions with the metabolism of drugs and xenobiotics in general, namely with the cytochrome P450 (CYP) system. Phytic acid (myo-inositol hexaphosphoric acid, IP6) is mainly present in plants but is also an endogenous compound present in mammalian cells and tissues. It has been shown to exhibit protective effect in many pathological conditions. For this paper, its interaction with CYPs was studied using human liver microsomes, primary human hepatocytes, the HepG2 cell line, and molecular docking. Docking experiments and absorption spectra demonstrated the weak ability of IP6 to interact in the heme active site of CYP1A. Molecular docking suggested that IP6 preferentially binds to the protein surface, whereas binding to the active site of CYP1A2 was found to be less probable. Subsequently, we investigated the ability of IP6 to modulate the metabolism of xenobiotics for both the mRNA expression and enzymatic activity of CYP1A enzymes. Our findings revealed that IP6 can slightly modulate the mRNA levels and enzyme activity of CYP1A. However, thanks to the relatively weak interactions of IP6 with CYPs, the chances of the mechanisms of clinically important drug-drug interactions involving IP6 are low.
- MeSH
- kyselina fytová * MeSH
- lidé MeSH
- messenger RNA MeSH
- savci MeSH
- simulace molekulového dockingu MeSH
- systém (enzymů) cytochromů P-450 MeSH
- xenobiotika * MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
Kyselina fytová (IP6) je nejhojněji se vyskytující inositolfosfát v přírodě, přítomný především v rostlinách, ale také v savčích buňkách. V současnosti je široce studována pro své rozmanité vlastnosti. Má schopnost vázat polyvalentní kationty, což bylo dříve považováno za nevýhodu ve spojitosti se sníženou dostupností minerálů z potravy. Na druhé straně jí tato struktura umožňuje funkci přírodního antioxidantu. Protektivní účinky kyseliny fytové byly popsány u patologických stavů včetně neurodegenerativních onemocnění, urolithiázy a rovněž zhoubného bujení.
Phytic acid (IP6, inositol hexaphosphate) is the most abundant inositol phosphate in nature. It is present mostly in plants but it has ben found also in mammalian cells. IP6 is extensively studied because of enormous variety of its properties. It binds polyvalent cations which was formerly considered as disadvantage responsible for lowered bioavailability of minerals in food. On the other hand, its stucture allows it to act as a natural antioxidant. Protective effect of IP6 has been found in varoius pathologies incl. neuorodegenerative disorders, urolithiasis as well as in malignancies.
- MeSH
- kyselina fytová * chemie škodlivé účinky terapeutické užití MeSH
- lidé MeSH
- protinádorové látky MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- práce podpořená grantem MeSH
We characterized the effect of phytic acid (inositol hexaphosphate, IP6) as a potential adjuvant in treatment of colorectal carcinoma and evaluated the optimal concentration and treatment time to produe the maximal therapeutic effect. There is some evidence that myoinositol (Ins) can potentiate anti-cancer effects of IP6. Therefore, we tested both IP6 and Ins individually and in combination on human cell lines HT-29, SW-480 and SW-620 derived from colorectal carcinoma in different stages of malignancy. The effect of tested chemicals on the cells was measured using metabolic activity assay (WST-1), DNA synthesis assay (BrdU), protein synthesis assay (Brilliant Blue) and apoptosis (caspase-3 activity). We tested IP6 and Ins at three concentrations: 0.2, 1 and 5 mM for 24, 48 and 72 h. The concentrations and incubation periods were chosen according to low toxicity of the tested substance that was observed in a long-term clinical study. We found that all employed concentrations of IP6 or IP6/Ins decreased proliferation of the cell lines, with the maximum decrease being observed in HT-29 cells. Metabolic activity of treated cells differed in response to IP6 and IP6/Ins treatment; in HT-29 and SW-620 significant decrease was observed only at the highest concentration, whereas in SW-480 cells metabolic activity was lower at each concentration except 0.2 and 1 mM IP6 or IP6/Ins in 24-h incubation. The results from protein content assay corresponded to the results obtained from WST assay. In addition, we found maximum increase in caspase-3 activity at concentration 5 mM IP6 or IP6/Ins in HT-29 cells and with IP6 at concentration of 0.2 mM or IP6/Ins in SW-480 cells with clear indication of Ins enhancing the proapoptotic effect of IP6 in all the cell lines studied.
- MeSH
- apoptóza účinky léků MeSH
- inositol farmakologie MeSH
- kaspasa 3 metabolismus MeSH
- kolorektální nádory farmakoterapie patologie MeSH
- kyselina fytová farmakologie MeSH
- lidé MeSH
- nádorové buněčné linie MeSH
- nádorové proteiny analýza MeSH
- proliferace buněk účinky léků MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Inositol hexaphosphate (IP6), also known as phytic acid, has been shown to exhibit anticancer effects in a number of preclinical tumor models. IP6 decreases proliferation by arresting cells in the G0/G1 phase, inhibits iron-mediated oxidative reactions, enhances differentiation and stimulates apoptosis. The present study attempted to characterize the effect of IP6 on the migration and adhesion of colon cancer SW620 cells. IP6 was assessed at concentrations of 0.2 and 1 mM during 12, 24 and 48 h of exposure. Migration ability was measured with the real-time xCELLigence Real-Time Cell Analyzer Dual Purpose system. The expression of mRNA and proteins involved in migration and cancer progression [epithelial cell adhesion molecule, intercellular adhesion molecule-1, β-catenin, N-cadherin, E-cadherin, matrix metalloproteinase (MMP)-2 and MMP-9] was determined by reverse transcription-quantitative polymerase chain reaction and western blot analysis. The changes in the expression and subcellular localization of E-cadherin were determined by indirect immunofluorescence. IP6 induced a decrease in the migration ability of the tested SW620 cell line. IP6-treated cells also showed decreased expression of N-cadherin, increased levels of E-cadherin and decreased expression of MMP-2 and MMP-9. These results indicated that IP6 has potential to modulate the migration ability and expression of markers associated with invasion in SW620 cells; however, further analysis is necessary to obtain a detailed understanding of the mechanism of action.
- MeSH
- buněčná adheze účinky léků MeSH
- invazivní růst nádoru prevence a kontrola MeSH
- kolorektální nádory farmakoterapie patologie MeSH
- kyselina fytová farmakologie terapeutické užití MeSH
- lidé MeSH
- nádorové biomarkery metabolismus MeSH
- nádorové buněčné linie MeSH
- pohyb buněk účinky léků MeSH
- progrese nemoci MeSH
- protinádorové látky farmakologie terapeutické užití MeSH
- screeningové testy protinádorových léčiv MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
The assembly of a hexameric lattice of retroviral immature particles requires the involvement of cell factors such as proteins and small molecules. A small, negatively charged polyanionic molecule, myo-inositol hexaphosphate (IP6), was identified to stimulate the assembly of immature particles of HIV-1 and other lentiviruses. Interestingly, cryo-electron tomography analysis of the immature particles of two lentiviruses, HIV-1 and equine infectious anemia virus (EIAV), revealed that the IP6 binding site is similar. Based on this amino acid conservation of the IP6 interacting site, it is presumed that the assembly of immature particles of all lentiviruses is stimulated by IP6. Although this specific region for IP6 binding may be unique for lentiviruses, it is plausible that other retroviral species also recruit some small polyanion to facilitate the assembly of their immature particles. To study whether the assembly of retroviruses other than lentiviruses can be stimulated by polyanionic molecules, we measured the effect of various polyanions on the assembly of immature virus-like particles of Rous sarcoma virus (RSV), a member of alpharetroviruses, Mason-Pfizer monkey virus (M-PMV) representative of betaretroviruses, and murine leukemia virus (MLV), a member of gammaretroviruses. RSV, M-PMV and MLV immature virus-like particles were assembled in vitro from truncated Gag molecules and the effect of selected polyanions, myo-inostol hexaphosphate, myo-inositol, glucose-1,6-bisphosphate, myo-inositol hexasulphate, and mellitic acid, on the particles assembly was quantified. Our results suggest that the assembly of immature particles of RSV and MLV was indeed stimulated by the presence of myo-inostol hexaphosphate and myo-inositol, respectively. In contrast, no effect on the assembly of M-PMV as a betaretrovirus member was observed.
- MeSH
- Alpharetrovirus fyziologie MeSH
- Betaretrovirus fyziologie MeSH
- buněčná membrána chemie metabolismus MeSH
- Gammaretrovirus fyziologie MeSH
- genové produkty gag chemie metabolismus MeSH
- interakce hostitele a patogenu * MeSH
- kultivované buňky MeSH
- polyelektrolyty chemie metabolismus MeSH
- Retroviridae fyziologie ultrastruktura MeSH
- sestavení viru * MeSH
- virion MeSH
- zvířata MeSH
- Check Tag
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
U některých typů onkologických onemocnění je suplementace glutaminu, citrulinu a argininu racionálnější, než doplňků např. s obsahem inositol hexafosfátu (IP6). Komplikované hledání příčinného vztahu mezi doplňky stravy a onkologickým onemocněním je citelně podněcováno komerčními zájmy. U většiny doplňků stravy chybí adekvátní důkazy a není možné marketingovou aktivitu kvalitně odborně podpořit.
