Dissecting the impact of target-binding kinetics of protein binders on tumor localization.
Ontology highlight
ABSTRACT:
SUBMITTER: Song Y
PROVIDER: S-EPMC7881221 | biostudies-literature | 2021 Feb
REPOSITORIES: biostudies-literature
Publications
Dissecting the impact of target-binding kinetics of protein binders on tumor localization.
Song Yunjin Y Jeong Hoibin H Kim Song-Rae SR Ryu Yiseul Y Baek Jonghwi J Kwon Jinhak J Cho Hyeongjun H Kim Kil-Nam KN Lee Joong-Jae JJ
iScience 20210129 2
Systematic control of <i>in vivo</i> behavior of protein-based therapeutics is considered highly desirable for improving their clinical outcomes. Modulation of biochemical properties including molecular weight, surface charge, and binding affinity has thus been suggested to enhance their therapeutic effects. However, establishing a relationship between the binding affinity and tumor localization remains a debated issue. Here we investigate the influence of the binding affinity of proteins on tum ...[more]
Similar Datasets
Project description:Rapid recognition of DNA target sites involves facilitated diffusion through which alternative sites are searched on genomic DNA. A key mechanism facilitating the localization of the target by a DNA-binding protein (DBP) is one-dimensional diffusion (sliding) in which electrostatic forces attract the protein to the DNA. As the protein reaches its target DNA site, it switches from purely electrostatic binding to a specific set of interactions with the DNA bases that also involves hydrogen bonding and van der Waals forces. High overlap between the DBP patches used for nonspecific and specific interactions with DNA may enable an immediate transition between the two binding modes following target site localization. By contrast, an imperfect overlap may result in greater frustration between the two potentially competing binding modes and consequently slower switching between them. A structural analysis of 125 DBPs indicates frustration between the two binding modes that results in a large difference between the orientations of the protein to the DNA when it slides compared to when it specifically interacts with DNA. Coarse-grained molecular dynamics simulations of in silico designed peptides comprising the full range of frustrations between the two interfaces show slower transition from nonspecific to specific DNA binding as the overlap between the patches involved in the two binding modes decreases. The complex search kinetics may regulate the search by eliminating trapping of the protein in semispecific sites while sliding.
| S-EPMC3207685 | biostudies-literature
Project description:A growing consensus is emerging that optimizing the drug-target affinity alone under equilibrium conditions does not necessarily translate into higher potency in vivo and that instead binding kinetic parameters should be optimized to ensure better efficacy. Therefore, in silico methods are needed to predict the kinetic parameters and the mechanistic determinants of drug-protein binding. Here we demonstrate the application of COMparative BINding Energy (COMBINE) analysis to derive quantitative structure-kinetics relationships (QSKRs) for the dissociation rate constants (k off) of inhibitors of heat shock protein 90 (HSP90) and HIV-1 protease. We derived protein-specific scoring functions by correlating k off rate constants with a subset of weighted interaction energy components determined from the energy-minimized structures of drug-protein complexes. As the QSKRs derived for these sets of chemically diverse compounds have good predictive ability and provide insights into important drug-protein interactions for optimizing k off, COMBINE analysis offers a promising approach for binding kinetics-guided lead optimization.
| S-EPMC6231175 | biostudies-literature
Project description:Cheap and massively parallel methods to assess the DNA-binding specificity of transcription factors are actively sought, given their prominent regulatory role in cellular processes and diseases. Here we evaluated the use of protein-binding microarrays (PBM) to probe the association of the tumor suppressor AP2? with 6000 human genomic DNA regulatory sequences. We show that the PBM provides accurate relative binding affinities when compared to quantitative surface plasmon resonance assays. A PBM-based study of human healthy and breast tumor tissue extracts allowed the identification of previously unknown AP2? target genes and it revealed genes whose direct or indirect interactions with AP2? are affected in the diseased tissues. AP2? binding and regulation was confirmed experimentally in human carcinoma cells for novel target genes involved in tumor progression and resistance to chemotherapeutics, providing a molecular interpretation of AP2? role in cancer chemoresistance. Overall, we conclude that this approach provides quantitative and accurate assays of the specificity and activity of tumor suppressor and oncogenic proteins in clinical samples, interfacing genomic and proteomic assays.
| S-EPMC3158074 | biostudies-literature
Project description:Inducible dimers are powerful tools for controlling biological processes through colocalizing signaling molecules. To be effective, an inducible system should have a dissociation constant in the "off" state that is greater (i.e., weaker affinity) than the concentrations of the molecules that are being controlled, and in the "on" state a dissociation constant that is less (i.e., stronger affinity) than the relevant protein concentrations. Here, we reengineer the interaction between the light inducible dimer, iLID, and its binding partner SspB, to better control proteins present at high effective concentrations (5-100 ?M). iLID contains a light-oxygen-voltage (LOV) domain that undergoes a conformational change upon activation with blue light and exposes a peptide motif, ssrA, that binds to SspB. The new variant of the dimer system contains a single SspB point mutation (A58V), and displays a 42-fold change in binding affinity when activated with blue light (from 3 ± 2 ?M to 125 ± 40 ?M) and allows for light-activated colocalization of transmembrane proteins in neurons, where a higher affinity switch (0.8-47 ?M) was less effective because more colocalization was seen in the dark. Additionally, with a point mutation in the LOV domain (N414L), we lengthened the reversion half-life of iLID. This expanded suite of light induced dimers increases the variety of cellular pathways that can be targeted with light.
| S-EPMC5332220 | biostudies-literature
Project description:PURPOSE:Deep learning is an emerging technique that allows us to capture imaging information beyond the visually recognizable level of a human being. Because of the anatomic characteristics and location, on-board target verification for radiation delivery to pancreatic tumors is a challenging task. Our goal was to use a deep neural network to localize the pancreatic tumor target on kV x-ray images acquired using an on-board imager for image guided radiation therapy. METHODS AND MATERIALS:The network is set up in such a way that the input is either a digitally reconstructed radiograph image or a monoscopic x-ray projection image acquired by the on-board imager from a given direction, and the output is the location of the planning target volume in the projection image. To produce a sufficient number of training x-ray images reflecting the vast number of possible clinical scenarios of anatomy distribution, a series of changes were introduced to the planning computed tomography images, including deformation, rotation, and translation, to simulate inter- and intrafractional variations. After model training, the accuracy of the model was evaluated by retrospectively studying patients who underwent pancreatic cancer radiation therapy. Statistical analysis using mean absolute differences (MADs) and Lin's concordance correlation coefficient were used to assess the accuracy of the predicted target positions. RESULTS:MADs between the model-predicted and the actual positions were found to be less than 2.60 mm in anteroposterior, lateral, and oblique directions for both axes in the detector plane. For comparison studies with and without fiducials, MADs are less than 2.49 mm. For all cases, Lin's concordance correlation coefficients between the predicted and actual positions were found to be better than 93%, demonstrating the success of the proposed deep learning for image guided radiation therapy. CONCLUSIONS:We demonstrated that markerless pancreatic tumor target localization is achievable with high accuracy by using a deep learning technique approach.
| S-EPMC6732032 | biostudies-literature
Project description:Abdurins are a novel antibody-like scaffold derived from the engineering of a single isolated CH2 domain of human IgG. Previous studies established the prolonged serum half-life of Abdurins, the result of a retained FcRn binding motif. Here we present data on the construction of large, diverse, phage-display and cell-free DNA display libraries and the isolation of high affinity binders to the cancer target, membrane-bound ephrin receptor tyrosine kinase class A2 (EphA2). Antigen binding regions were created by designing combinatorial libraries into the structural loops and Abdurins were selected using phage display methods. Initial binders were reformatted into new maturation libraries and low nanomolar binders were isolated using cell-free DNA display, CIS display. Further characterization confirmed binding of the Abdurins to both human and murine EphA2 proteins and exclusively to cell lines that expressed EphA2, followed by rapid internalization. Two different EphA2 binders were labeled with 64Cu, using a bifunctional MeCOSar chelator, and administered to mice bearing tumors from transplanted human prostate cancer cells, followed by PET/CT imaging. The anti-EphA2 Abdurins localized in the tumors as early as 4 hours after injection and continued to accumulate up to 48 hours when the imaging was completed. These data demonstrate the ability to isolate high affinity binders from the engineered Abdurin scaffold, which retain a long serum half-life, and specifically target tumors in a xenograft model.
| S-EPMC4552014 | biostudies-literature
Project description:Cobalamin uptake and transport in mammals are mediated by three cobalamin-binding proteins: haptocorrin, intrinsic factor, and transcobalamin. The nature of cobalamin-binding proteins in lower vertebrates remains to be elucidated. The aim of this study was to characterize the cobalamin-binding proteins of the rainbow trout (Oncorhynchus mykiss) and to compare their properties with those of the three human cobalamin-binding proteins. High cobalamin-binding capacity was found in trout stomach (210 pmol/g), roe (400 pmol/g), roe fluid (390 nmol/liter), and plasma (2500 nmol/liter). In all cases, it appeared to be the same protein based on analysis of partial sequences and immunological responses. The trout cobalamin-binding protein was purified from roe fluid, sequenced, and further characterized. Like haptocorrin, the trout cobalamin-binding protein was stable at low pH and had a high binding affinity for the cobalamin analog cobinamide. Like haptocorrin and transcobalamin, the trout cobalamin-binding protein was present in plasma and recognized ligands with altered nucleotide moiety. Like intrinsic factors, the trout cobalamin-binding protein was present in the stomach and resisted degradation by trypsin and chymotrypsin. It also resembled intrinsic factor in the composition of conserved residues in the primary cobalamin-binding site in the C terminus. The trout cobalamin-binding protein was glycosylated and displayed spectral properties comparable with those of haptocorrin and intrinsic factor. In conclusion, only one soluble cobalamin-binding protein was identified in the rainbow trout, a protein that structurally behaves like an intermediate between the three human cobalamin-binding proteins.
| S-EPMC3460486 | biostudies-literature
Project description:BackgroundSignaling through MEK-->ERK1/2 and PI3 kinases is implicated in many aspects of cell physiology, including the survival of oxidant exposure. Oxidants play a role in numerous physiological and pathophysiological processes, many of which rely on transport in and out of the nucleus. However, how oxidative stress impacts nuclear trafficking is not well defined.Methodology/principal findingsTo better understand the effect of stress on nucleocytoplasmic trafficking, we exposed cells to the oxidant diethyl maleate. This treatment activated MEK-->ERK1/2 as well as PI3 kinase-->Akt cascades and triggered the inhibition of classical nuclear import. To define the molecular mechanisms that regulate nuclear transport, we examined whether MEK and PI3 kinase signaling affected the localization of key transport factors. Using recently developed tools for image acquisition and analysis, the subcellular distributions of importin-alpha, CAS, and nucleoporins Nup153 and Nup88 were quantified in different cellular compartments. These studies identified specific profiles for the localization of transport factors in the nucleus and cytoplasm, and at the nuclear envelope. Our results demonstrate that MEK and PI3 kinase signaling as well as oxidative stress control nuclear trafficking and the localization of transport components. Furthermore, stress not only induced changes in transport factor distribution, but also upregulated post-translational modification of transport factors. Our results are consistent with the idea that the phosphorylation of importin-alpha, CAS, Nup153, and Nup88, and the O-GlcNAc modification of Nup153 increase when cells are exposed to oxidant.Conclusions/significanceOur studies defined the complex regulation of classical nuclear import and identified key transport factors that are targeted by stress, MEK, and PI3 kinase signaling.
| S-EPMC2793512 | biostudies-literature
Project description:Parafibromin, a tumor suppressor protein encoded by HRPT2/CDC73 and implicated in parathyroid cancer and the hyperparathyroidism-jaw tumor (HPT-JT) familial cancer syndrome, is part of the PAF1 transcriptional regulatory complex. Parafibromin has been implicated in apoptosis and growth arrest, but the mechanism by which its loss of function promotes neoplasia is poorly understood. In this study we report that a hypomorphic allele of hyrax (hyx), the Drosophila homolog of HRPT2/CDC73, rescues the loss-of-ventral-eye phenotype of lobe (Akt1s1). Such rescue is consistent with previous reports that hyx/parafibromin is required for the nuclear transduction of Wingless (Wg)/Wnt signals and that Wg signaling antagonizes lobe function. A screen using double hyx/lobe heterozygotes identified an additional interaction with orb and orb2, the homologs of mammalian cytoplasmic polyadenylation element binding protein (CPEB), a translational regulatory protein. Hyx and orb2 heterozygotes lived longer and were more resistant to starvation than controls. In mammalian cells, knockdown of parafibromin expression reduced levels of CPEB1. Chromatin immunoprecipitation (ChIP) showed occupancy of CPEB1 by endogenous parafibromin. Bioinformatic analysis revealed a significant overlap between human transcripts potentially regulated by parafibromin and CPEB. These results show that parafibromin may exert both transcriptional and, through CPEB, translational control over a subset of target genes and that loss of parafibromin (and CPEB) function may promote tumorigenesis in part by conferring resistance to nutritional stress.
| S-EPMC3109426 | biostudies-literature
Project description:The addition of expansive agents could overcome the main disadvantages of raw concrete including high brittleness and low tensile strength. Few studies have investigated the early hydration kinetics of expansive cementitious binders, though the findings from the early hydration kinetics are helpful for understanding their technical performances. In this study, mixtures of 3CaO•3Al2O3•CaSO4 and CaSO4 (i.e., ZY-type™ expansive agent) with different proportions of mineral admixtures (e.g., fly ash and slag) were added into cement pastes to investigate the early hydration kinetics mechanism of expansive cementitious binders. Early hydration heat evolution rate and cumulative hydration heat were measured by isothermal calorimeter. Kinetic parameters were estimated based on the Krstulovic-Dabic model and Knudsen equations. Mechanical performances of expansive cementitious binders were tested in order to evaluate if they met the basic requirements of shrinkage-compensating materials in technical use. The early hydration heat released from cementitious binders containing ZY-type™ expansive agent was much greater than that released by pure cement, supporting the idea that addition of the expansive agent would improve the reaction of cement. The early hydration kinetic rates were decreased due to the reactions of the mineral admixture (e.g., fly ash or slag) and the ZY-type™ expansive agent in the cement system. The hydration reaction of cementitious binders containing ZY-type™ expansive agent obeyed the Krstulovic-Dabic model well. Three processes are involved in the hydration reaction of cementitious binders containing ZY-type™ expansive agent. These are nucleation and crystal growth (NG), interactions at phase boundaries (I), and diffusion (D). The 14-day expansion rates of cementitious binders containing ZY-type™ expansive agent are in the range of 2.0 × 10-4 to 3.5 × 10-4, which could meet the basic requirements of anti-cracking performances in technical use according to Chinese industry standard JGJ/T 178-2009. This study could provide an insight into understanding the effects of expansive agents on the hydration and mechanical performances of cementitious binders.
| S-EPMC6631045 | biostudies-literature