A Biacore biosensor method for detailed kinetic binding analysis of small molecule inhibitors of p38α mitogen-activated protein kinase

Casper, David J.; Bukhtiyarova, Marina; Springman, Eric B.

doi:10.1016/j.ab.2003.10.025

Cited by 61 publications

(58 citation statements)

References 38 publications

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“…The use of calorimetry has, however, been restricted due to the requirement of relatively large protein concentration of the order of milligrams per milliliter (23). In many systems, such as new proteins being screened for kinase activity and͞or inhibition of ATP binding, only small or trace quantities of protein are available, thus precluding the use of calorimetry.…”

Section: Discussionmentioning

confidence: 99%

“…We have not achieved this level of quantitative analysis in our initial studies with SiNW FETs but note that implementation of background subtraction techniques used in SPR (23,24,26), which could be readily implemented with small arrays (27), should enable similar parameter analysis with our approach. We also believe that there are possible advantages to the SiNW devices compared with SPR, including (i) higher sensitivity, (ii) smaller quantity of protein required to make active sensor chips, and (iii) the potential for very large integrated arrays.…”

Section: Discussionmentioning

confidence: 99%

“…In this regard, it is important to compare our nanowirebased approach to other label-free methods used to study small-molecule-protein binding, including surface plasmon resonance (SPR) and calorimetry. In recent SPR studies of smallmolecule binding to kinases and other proteins (23)(24)(25)(26), the protein has been immobilized on the SPR sensor chip, and then the optical response was recorded as a function of time under different experimental conditions. These studies have shown time data recorded from Abl-modified SiNW devices by using solutions containing 100 nM ATP and 50 nM small molecules for Gleevec (red), A1 (blue), A2 (green), A3 (pink), and biotin (black).…”

Section: Discussionmentioning

confidence: 99%

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Label-free detection of small-molecule–protein interactions by using nanowire nanosensors

Wang

Chen

Lin

et al. 2005

Proc. Natl. Acad. Sci. U.S.A.

511

298

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Development of miniaturized devices that enable rapid and direct analysis of the specific binding of small molecules to proteins could be of substantial importance to the discovery of and screening for new drug molecules. Here, we report highly sensitive and labelfree direct electrical detection of small-molecule inhibitors of ATP binding to Abl by using silicon nanowire field-effect transistor devices. Abl, which is a protein tyrosine kinase whose constitutive activity is responsible for chronic myelogenous leukemia, was covalently linked to the surfaces of silicon nanowires within microfluidic channels to create active electrical devices. Concentration-dependent binding of ATP and concentration-dependent inhibition of ATP binding by the competitive small-molecule antagonist STI-571 (Gleevec) were assessed by monitoring the nanowire conductance. In addition, concentration-dependent inhibition of ATP binding was examined for four additional small molecules, including reported and previously unreported inhibitors. These studies demonstrate that the silicon nanowire devices can readily and rapidly distinguish the affinities of distinct smallmolecule inhibitors and, thus, could serve as a technology platform for drug discovery.inhibitors ͉ kinase ͉ Gleevec ͉ chronic myclogenous leukemia ͉ drug discovery I dentification of organic molecules that bind specifically to proteins is central to the discovery and development of new pharmaceuticals and to chemical genetic approaches for elucidating complex pathways in biological systems (1-4). Broadly representative of the importance of this concept for developing drugs to treat disease have been efforts focused on identifying inhibitors to protein tyrosine kinases (1, 5). Tyrosine kinases represent especially attractive targets because they are central elements in the networks that mediate signal transduction in mammalian cells. The regulatory function of tyrosine kinases occurs through phosphorylation of a tyrosine residue of a substrate protein by using ATP as a phosphate source (Fig. 1A) and subsequent transmission of this event through signal transduction cascade. Deregulation of phosphorylation through, for example, mutation or overexpression of protein tyrosine kinases has been linked to a number of diseases, including cancer (1, 5, 6).The identification of inhibitors to ATP or substrate protein binding thus can serve as a means of treating diseases linked to a tyrosine kinase. A successful example of this strategy has been the introduction of the small molecule STI-571, or Gleevec (Fig. 1B), which competitively inhibits ATP binding to the tyrosine kinase Abl and is a highly effective treatment for chronic myelogenous leukemia (1,5,7,8). This success and the recognition that Gleevec may be unable to cure late-stage chronic myelogenous leukemia because of mutations in the kinase (5,(8)(9)(10) suggest that the development of approaches that enable rapid, flexible, and quantitative comparison of small-molecule inhibitors of ATP or substrate protein binding to tyrosine ...

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

See 1 more Smart Citation

Label-free detection of small-molecule–protein interactions by using nanowire nanosensors

Wang

Chen

Lin

et al. 2005

Proc. Natl. Acad. Sci. U.S.A.

511

298

View full text Add to dashboard Cite

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“…Detection of SSTR-2 in tissue sample with biomolecular interaction analysis system [17] The tissue samples of gastric cancer or no-cancer gastric mucosa were broken to pieces with scissors on ice before the addition of special buffer (4 µL/mg tissue) supplied by BIAcore AB (Amersham Biosciences, Uppsala, Sweden) for the extraction of proteins. Then the samples were homogenized in ice-cold phosphate buffered solution and centrifuged to pellets.…”

Section: Nag-1 Gene Detection By Rt-pcrmentioning

confidence: 99%

Preoperative growth inhibition of human gastric adenocarcinoma treated with a combination of celecoxib and octreotide

et al. 2007

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Aim: To gain insight into the histopathological responses and molecular targets in the inhibition of growth of human gastric cancer treated with celecoxib (a cyclooxygenase [COX]‐2 inhibitor) combined with octreotide. Methods: Seventy five patients with gastric cancer undergoing curative gastrectomy or extended resection were randomly divided into 3 groups. The apoptosis of tumor cells was measured by terminal deoxynucleotide transferase‐mediated dUTP nick end‐labeling (TUNEL) assay. Gastric cancer microvessel density (MVD) and the expression of COX‐2 were evaluated by immunohistochemical staining. The expression of somatostatin receptor (SSTR)‐2 was detected with the biomolecular interaction analysis system. The transcription of non‐steroidal anti‐inflammatory drug‐activated gene (NAG)‐1 was measured by RT‐PCR. Results: Compared with the control and celecoxib groups, more necrosis in the combination group was observed. The apoptotic rate in the combination group (7.06%±0.67%) was significantly higher than that in the control group (6.23%±1.29%, P<0.05). The MVD decreased considerably in the combination group. The upregulation of NAG‐1 was displayed both in the celecoxib and combination groups. The positive rate of SSTR‐2 in gastric cancers treated with celecoxib (48%) was significantly higher than that of control group (12%) after surgery (P<0.05). Conclusion: Celecoxib combined with octreotide significantly promoted necrosis in gastric adenocarci‐noma through the induction of apoptosis and the reduction of MVD. NAG‐1 and SSTR‐2 might be the molecular targets for celecoxib or octreotide.

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“…Since the SPR transducing mechanism relies on refractive index changes, SPR sensing has an intrinsic limitation in terms of the molecular weight of the molecule to be detected [4,5,6] . In order to be able to perform analytical biosensing measurements of low molecular weight molecules, typically below 30kDa, strategies have been developed which consist of either immobilising the low molecular weight target molecule non-specifically onto the sensor surface first and then binding specifically the corresponding antibodies to the target molecule [7] , using conformational changes of macromolecules, such as maltose binding protein and tissue transglutaminase upon analyte binding [8] or by using large particles that increase the local change of refractive index, therefore behaving like a plasmonic label [9] . In the later case, 1000 fold increase of the detection sensitivity has been observed.…”

Section: Introductionmentioning

confidence: 99%

Enzyme detection by surface plasmon resonance using specially engineered spacers and plasmonic labelling

et al. 2011

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A Biacore biosensor method for detailed kinetic binding analysis of small molecule inhibitors of p38α mitogen-activated protein kinase

Cited by 61 publications

References 38 publications

Label-free detection of small-molecule–protein interactions by using nanowire nanosensors

Label-free detection of small-molecule–protein interactions by using nanowire nanosensors

Preoperative growth inhibition of human gastric adenocarcinoma treated with a combination of celecoxib and octreotide

Enzyme detection by surface plasmon resonance using specially engineered spacers and plasmonic labelling

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