scholarly journals Molecular Modeling and Functional Analysis of Exome Sequencing–Derived Variants of Unknown Significance Identify a Novel, Constitutively Active FGFR2 Mutant in Cholangiocarcinoma

2017 ◽  
pp. 1-13
Author(s):  
Jan B. Egan ◽  
David L. Marks ◽  
Tara L. Hogenson ◽  
Anne M. Vrabel ◽  
Ashley N. Sigafoos ◽  
...  
Keyword(s):  
Molecular Modeling ◽  
Tyrosine Kinase ◽  
Exome Sequencing ◽  
Receptor Tyrosine Kinase ◽  
In Silico ◽  
Point Mutations ◽  
Wild Type ◽  
Modeling Tools ◽  
Variants Of Unknown Significance ◽  
Unknown Significance

Purpose Genomic testing has increased the quantity of information available to oncologists. Unfortunately, many identified sequence alterations are variants of unknown significance (VUSs), which thus limit the clinician’s ability to use these findings to inform treatment. We applied a combination of in silico prediction and molecular modeling tools and laboratory techniques to rapidly define actionable VUSs. Materials and Methods Exome sequencing was conducted on 308 tumors from various origins. Most single nucleotide alterations within gene coding regions were VUSs. These VUSs were filtered to identify a subset of therapeutically targetable genes that were predicted with in silico tools to be altered in function by their variant sequence. A subset of receptor tyrosine kinase VUSs was characterized by laboratory comparison of each VUS versus its wild-type counterpart in terms of expression and signaling activity. Results The study identified 4,327 point mutations of which 3,833 were VUSs. Filtering for mutations in genes that were therapeutically targetable and predicted to affect protein function reduced these to 522 VUSs of interest, including a large number of kinases. Ten receptor tyrosine kinase VUSs were selected to explore in the laboratory. Of these, seven were found to be functionally altered. Three VUSs (FGFR2 F276C, FGFR4 R78H, and KDR G539R) showed increased basal or ligand-stimulated ERK phosphorylation compared with their wild-type counterparts, which suggests that they support transformation. Treatment of a patient who carried FGFR2 F276C with an FGFR inhibitor resulted in significant and sustained tumor response with clinical benefit. Conclusion The findings demonstrate the feasibility of rapid identification of the biologic relevance of somatic mutations, which thus advances clinicians’ ability to make informed treatment decisions.

Molecular Modeling of Novel Fluorophoric Thiazolo- [2, 3-B] Quinazolinones to Study Epidermal Growth Factor Receptor Tyrosine Kinase Inhibition Potency

2021 ◽  
Author(s):  
Showkat Mir ◽  
Ganesh Chandra Dash ◽  
Kumar Sambhav Chopdar ◽  
Prajna Mohanta ◽  
Pranab Kishor Mohapatra ◽  
...  
Keyword(s):  
Epidermal Growth Factor Receptor ◽  
Growth Factor ◽  
Tyrosine Kinase ◽  
Receptor Tyrosine Kinase ◽  
Active Site ◽  
In Silico ◽  
Growth Factor Receptor ◽  
Cancer Therapeutics ◽  
Tk Inhibitors ◽  
Epidermal Growth

Epidermal growth factor receptor tyrosine kinase (EGFR-TK) is one of the key regulators that exhibit pivotal role in proliferation of cancer cell. Quinazolinones are studiedly widely as effective EGFR-TK inhibitor because of their higher affinity to bind with adenosine triphosphate (ATP) binding site of receptor tyrosine kinases. However, their toxicity due to non-specific binding to tyrosine kinase in non-cancerous normally dividing cells of the body limits its applicability as a cancer therapeutics. In the present investigation a series of thirty-four novel synthesized thiazolo- [2, 3-b] quinazolinones were studied in silico as EGFR-TK inhibitors. All the thirty-four compounds were screened against EGFR-TK domain using multiple software’s (AutoDock Vina, Argus Lab, YASARA, and MOE). The interactions of the ligands with amino acid residues, namely, Lys721, Met769 and Asp831 of the active site were through the functional groups on aryl substituents at position 3 and 5 of the thiazolo- [2, 3-b] quinazolinone scaffold. The methyl substituents at position 8 of the ligands had prominent hydrophobic interactions in the active site cavity of EGFR-TK domain. The compounds 5ab, 5aq, and 5bq were predicted to be non-toxic and drug-like by in silico ADMET investigations. These compounds were considered for further investigation due to their non-toxicity and higher docking score ranking in different docking methods. The molecular dynamics (MD) simulation for 100 ns of docked complexes revealed the stability of these compounds. The binding free energy determined using Molecular Mechanics Generalized Born Model and Solvent Accessibility (MM-GBSA) method indicate that thiazolo- [2, 3-b] quinazolinone has high inhibitory efficacy similar to the standard drug, erlotinib (5ab - 22.45, 5aq -22.23, 5bq -20.76, and erlotinib -24.11 kcal/mol). In silico studies and MD simulations indicated that compounds (5ab, 5aq and 5bq) could be potential EGFR-TK inhibitors and require further validation as cancer therapeutics using carcinoma cell lines.<br><p></p>

scholarly journals Only the Substitution of Methionine 918 with a Threonine and Not with Other Residues Activates RET Transforming Potential*

Endocrinology ◽  
1997 ◽  
Vol 138 (4) ◽  
pp. 1450-1455 ◽  
Author(s):  
Anna Maria Cirafici ◽  
Giuliana Salvatore ◽  
Gabriella De Vita ◽  
Francesca Carlomagno ◽  
Nina A. Dathan ◽  
...  
Keyword(s):  
Tyrosine Kinase ◽  
Receptor Tyrosine Kinase ◽  
Multiple Endocrine Neoplasia ◽  
Multiple Endocrine Neoplasia Type ◽  
Point Mutations ◽  
Early Response ◽  
Specific Point ◽  
Medullary Thyroid ◽  
Transforming Activity ◽  
Early Response Genes

Abstract Specific point-mutations of the RET receptor tyrosine kinase protooncogene are responsible for the inheritance of multiple endocrine neoplasia type 2A (MEN2A) and 2B (MEN2B), and familial medullary thyroid carcinoma (FMTC). MEN2B is caused by the substitution of methionine 918 by a threonine in the tyrosine kinase (TK) domain of RET. This mutation converts RET into a dominant transforming oncogene. We have substituted Met918 with four different residues and found that RET acquired transforming activity only when Met918 was substituted with a threonine. However, also when serine and valine, but not leucine or phenylalanine, were inserted in position 918, the RET TK function was activated and induced, especially in the case of the RET(918Ser), immmediate-early response genes. We conclude that the preservation of Met918 is critical for the control of RET kinase. However, only when a threonine residue is present in position 918, does RET efficiently couple with a transforming pathway.

Whole‐Exome Sequencing of a Saudi Epilepsy Cohort Reveals Association Signals in Known and Potentially Novel Loci

2021 ◽  
Author(s):  
Amein Kadhem AlAli ◽  
Abdulrahman Al-Enazi ◽  
Ahmed Ammar ◽  
Mahmoud Hajj ◽  
Cyril Cyrus ◽  
...  
Keyword(s):  
Exome Sequencing ◽  
Whole Exome Sequencing ◽  
Rare Variants ◽  
High Rate ◽  
Variants Of Unknown Significance ◽  
Pathogenic Variants ◽  
Whole Exome ◽  
Novel Variants ◽  
Unknown Significance ◽  
Rare Genetic Variants

Abstract Background Epilepsy, a serious chronic neurological condition effecting up to 100 million people globally, has clear genetic underpinnings including common and rare variants. In Saudi Arabia the prevalence of epilepsy is high and caused mainly by perinatal and genetic factors. No whole-exome sequencing (WES) studies have been performed to date in Saudi Arabian Epilepsy cohorts. This offers a unique opportunity for the discovery of rare genetic variants impacting this disease as there is a high rate of consanguinity amongst large tribal pedigrees. Results We performed WES on 144 individuals diagnosed with epilepsy, to interrogate known Epilepsy related genes for known and functional novel variants. We also used an American College of Medical Genetics (ACMG) guideline based variant prioritization approach in an attempt to discover putative causative variants. We identified a 32 potentially causative pathogenic variants across 30 different genes in 44/144 (30%) of these Saudi Epilepsy individuals. We also identified 232 variants of unknown significance (VUS) across 101 different genes in 133/144 (92%) subjects. Strong enrichment of variants of likely pathogenicity were observed in previously described epilepsy-associated loci, and a number of putative pathogenic variants in novel loci are also observed. Conclusion Several putative pathogenic variants known to be epilepsy-related loci were identified for the first time in our population, in addition to several potential new loci have been identified which may be prioritized for further investigation.

scholarly journals Two Cases of Recessive Intellectual Disability Caused by NDST1 and METTL23 Variants

Genes ◽  
2020 ◽  
Vol 11 (9) ◽  
pp. 1021
Author(s):  
Amjad Khan ◽  
Zhichao Miao ◽  
Muhammad Umair ◽  
Amir Ullah ◽  
Mohammad A. Alshabeeb ◽  
...  
Keyword(s):  
Intellectual Disability ◽  
Secondary Structure ◽  
Homology Modeling ◽  
Exome Sequencing ◽  
In Silico ◽  
Brain Function ◽  
Sanger Sequencing ◽  
Autosomal Recessive ◽  
Wild Type ◽  
Genetic Condition

Intellectual disability (ID) is a highly heterogeneous genetic condition with more than a thousand genes described so far. By exome sequencing of two consanguineous families presenting hallmark features of ID, we identified two homozygous variants in two genes previously associated with autosomal recessive ID: NDST1 (c.1966G>A; p.Asp656Asn) and METTL23 (c.310T>C; p.Phe104Leu). The segregation of the variants was validated by Sanger sequencing in all family members. In silico homology modeling of wild-type and mutated proteins revealed substantial changes in the secondary structure of both proteins, indicating a possible effect on function. The identification and validation of new pathogenic NDST1 and METTL23 variants in two cases of autosomal recessive ID further highlight the importance of these genes in proper brain function and development.

scholarly journals Molecular Modeling and Structural Stability of Wild-Type and Mutant CYP51 from Leishmania major: In Vitro and In Silico Analysis of a Laboratory Strain

Molecules ◽  
2018 ◽  
Vol 23 (3) ◽  
pp. 696 ◽  
Author(s):  
Masoud Keighobadi ◽  
Saeed Emami ◽  
Milad Lagzian ◽  
Mahdi Fakhar ◽  
Alireza Rafiei ◽  
...  
Keyword(s):  
Molecular Modeling ◽  
Structural Stability ◽  
In Silico ◽  
Leishmania Major ◽  
In Silico Analysis ◽  
Laboratory Strain ◽  
Wild Type ◽  
Silico Analysis

scholarly journals Novel frameshift variant in MYL2 reveals molecular differences between dominant and recessive forms of hypertrophic cardiomyopathy

2019 ◽  
Author(s):  
Sathiya N. Manivannan ◽  
Sihem Darouich ◽  
Aida Masmoudi ◽  
David Gordon ◽  
Gloria Zender ◽  
...  
Keyword(s):  
Hypertrophic Cardiomyopathy ◽  
Exome Sequencing ◽  
Rapid Screening ◽  
Functional Study ◽  
Ventricular Muscle ◽  
Loss Of Function ◽  
Variants Of Unknown Significance ◽  
Unknown Significance ◽  
The Impact

AbstractHypertrophic cardiomyopathy (HCM) is characterized by enlargement of the ventricular muscle without dilation and is often associated with dominant pathogenic variants in cardiac sarcomeric protein genes. Here, we report a family with two infants diagnosed with infantile-onset HCM and mitral valve dysplasia that led to death before one year of age. Using exome sequencing, we discovered that one of the affected children had a homozygous frameshift variant in Myosin light chain 2 (MYL2:NM_000432.3:c.431_432delCT: p.Pro144Argfs*57;MYL2-fs), which alters the last 20 amino acids of the protein and is predicted to impact the C-terminal EF-hand (CEF) domain. The parents are unaffected heterozygous carriers of the variant and the variant is absent in control cohorts from gnomAD. The absence of the phenotype in carriers and infantile presentation of severe HCM is in contrast to HCM associated with dominant MYL2 variants. Immunohistochemical analysis of the ventricular muscle of the deceased patient with the MYL2-fs variant showed marked reduction of MYL2 expression compared to an unaffected control. In vitro overexpression studies further indicate that the MYL2-fs variant is actively degraded. In contrast, an HCM-associated missense variant (MYL2:p.Gly162Arg) and three other MYL2 stopgain variants that lead to loss of the CEF domain are stably expressed. However, stopgain variants show impaired localization suggesting a functional role for the CEF domain. The degradation of the MYL2-fs can be rescued by inhibiting the cell’s proteasome function supporting a post-translational effect of the variant. In vivo rescue experiments with a Drosophila MYL2-homolog (Mlc2) knockdown model indicate that neither MYL2-fs nor MYL2:p.Gly162Arg supports regular cardiac function. The tools that we have generated provide a rapid screening platform for functional assessment of variants of unknown significance in MYL2. Our study supports an autosomal recessive model of inheritance for MYL2 loss-of-function variants and highlights the variant-specific molecular differences found in MYL2-associated cardiomyopathies.Author SummaryWe report a novel frameshift variant in MYL2 that is associated with a severe form of infantile-onset hypertrophic cardiomyopathy. The impact of the variant is only observed in the recessive form of the disease in the proband and not in the parents who are carriers of the variant. This is in contrast to other dominant variants in MYL2 that are associated with cardiomyopathies. We compared the stability of this variant to that of other cardiomyopathy associated MYL2 variants and found molecular differences in the disease pathology. We also show different protein domain requirement for stability and localization of MYL2 in cardiomyocytes. Further, we used a fly model to demonstrate functional deficits due to the variant in the developing heart. Overall, our study shows a molecular mechanism by which loss-of-function variants in MYL2 are recessive while missense variants are dominant. We highlight the use of exome sequencing and functional testing to assist in the diagnosis of rare forms of diseases where pathogenicity of the variant is not obvious. The new tools we developed for in vitro functional study and the fly fluorescent reporter analysis will permit rapid analysis of MYL2 variants of unknown significance.

Sign in / Sign up

Export Citation Format

Share Document