7-Hydroxy-2,3,4,5-tetrahydro-1H-benzofuro[2,3-c]azepin-1-one

IC50: Selective protein kinase D (PKD) antagonist with the IC50 of 0.182, 0.280, 0.227, >10, 15.3, 20.3, 40.5 and >50 μM for PKD1, PKD2, PKD3, PKC, CAK, PLK1, CAMKIIα and Akt respectively.

CID755673, benzoxoloazepinolone, is the first identified cell-active small molecule PKD antagonist. It inhibits the activity of PKD1 with an IC50 of 182 nM and demonstrates highest selectivity to PKD1 when compared with AKT, PLK1, CAK, CAMKIIα, PKD2 and PKD3. Moreover, it was not competitive with ATP for enzyme inhibition. [1]

In vitro: In cell based assays, CID755673 dose-dependently suppressed PKD1 activation induced by phorbol ester endogenous in LNCaP cells. It was also reported to reverse biological actions of PKD1 including class IIa histone deacetylase 5 nuclear exclusion, vesicular stomatitis, virus glycoprotein delivery from the Golgi to the plasma membrane as well as the ilimaquinone-induced Golgi fragmentation. Moreover, CID755673 suppressed prostate cancer cell proliferation, cell migration, and invasion. [1]

In vivo: Experimental models of acute pancreatitis were developed to study the effect of CID755673 on acute pancreatitis in vivo. Results demonstrated that this compound suppressed PKD1/2 and therefore significantly offset necrosis and severity of pancreatitis. [2]

Clinical trial: So far, no clinical trial has been conducted.

References:
[1]Sharlow ER, Giridhar KV, LaValle CR, Chen J, Leimgruber S, Barrett R, Altamirano KB, Wipf P, Lazo JS and Wang QJ.  Potent and selective disruption of protein kinase D functionality by a benzoxoloazepinolone. J Biol Chem. 2008 Nov. 283(48): 3351246.
[2]Yuan JZ, Liu YN, Tan TY, Guha S, Gukovsky I, Gukovskaya A and Pandol SJ.  Protein kinase D regulates cell death pathways in experimental pancreatitis. Front Physiol. 2012 Mar. 3: DOI: 10.3389/fphys.2012.00060.

Profiling of hydroxycinnamoylquinic acids in plant extracts using in-source CID fragmentation.[Pubmed:27591562]

J Mass Spectrom. 2016 Dec;51(12):1130-1145.

Hydroxycinnamoylquinic acids (HCQAs) are a major class of phenolic plant secondary metabolites, belonging to the chlorogenic acid family. Various health-beneficial properties of HCQAs have been shown, which has drawn interest for HCQA profiling in plants of human consumption. However, this task remains challenging, because several isomeric HCQAs can be present in the sample with identical molecular formulae and the limited availability of reference standards poses additional challenges to their identification. In the present work, a high performance liquid chromatography-electrospray ionization-quadrupole time-of-flight-mass spectrometry (HPLC-ESI-Q/TOF-MS) method accompanied with an effective data filtering protocol is presented, which is shown to be suitable for the identification of HCQAs in plant materials in a non-targeted manner. Both collision-induced dissociation (CID) fragmentation performed in a collision cell and in-source (CID) fragmentation were used to produce accurate mass fragments. It was shown that fragmentation characteristics required for identification of regio-isomers of HCQAs can be achieved with in-source CID fragmentation, enabling the use of a single-stage MS system with in-source fragmentation for convincing identification of HCQAs. Based on a thorough validation of identified HCQA compounds using coffee bean extracts as reference samples, comprehensive profiling of HCQAs in two apricot (Prunus armeniaca L.) genotypes ('Preventa' and 'Gonci magyarkajszi') was carried out for the first time and the following 10 HCQAs were shown to be present in apricot fruit: 3-caffeoylquinic acid (CQA), cis-3-CQA, 4-CQA, 5-CQA, cis-5-CQA, 3,5-diCQA, 3-p-coumaroylquinic acid (pCoQA), 4-pCoQA, 3-feruloylquinic acid (FQA) and cis-3-FQA. Copyright (c) 2016 John Wiley & Sons, Ltd. HIGHLIGHTS: An HPLC-ESI-Q/TOF-MS method suitable for the identification of hydroxycinnamoyilquinic acids (HCQAs) in plant material in a non-targeted manner was developed. Single-stage, high-resolution MS system with in-source fragmentation was shown to be suitable for convincing identification of HCQAs. Comprehensive profiling of HCQAs in two apricot (Prunus armeniaca L.) genotypes was carried out for the first time. Copyright (c) 2016 John Wiley & Sons, Ltd.

Analysis of matrix-assisted laser desorption/ionization quadrupole time-of-flight collision-induced dissociation spectra of simple precursor ions and isobaric oligosaccharide ion mixtures based on product ion intensities and pattern recognition.[Pubmed:28299859]

Rapid Commun Mass Spectrom. 2017 May 30;31(10):873-885.

RATIONALE: Qualitative analysis of glycomic tandem mass spectrometry (MS/MS) data based on m/z values of product ions alone is widely used, and often sufficient for analysis of single analytes. However, most complex glycomic mixtures contain multiple isobaric oligosaccharides, in which case this approach is often limited. Here we show how ion intensity information can be used in order to enhance MS/MS data analysis, and extract both qualitative and semiquantitative information from complex glycomic MS/MS datasets. METHODS: A matrix-assisted laser desorption/ionization quadrupole time-of-flight (MALDI QTOF) mass spectrometer was used in this study. We compared the intensities of product ions within a single product ion series, determined by their length, across the whole glycomic MS/MS dataset. In order to detect discernable patterns, the intensity data was normalized to the intensity of each product ion within the series. In most cases, normalized data yielded easily discernable patterns, relevant either for analysis of specific glycomic structure types, or mechanistic MS studies. RESULTS: We used our approach on a glycomic sample consisting of human milk oligosaccharides. The approach yielded useful results for both qualitative and semiquantitative analyses. All normalizations performed were not equally rich in information and the information content of generated tables was not possible to predict. These analyses were shown to be independent of instrument manufacturer. CONCLUSIONS: Our approach enabled more detailed qualitative analysis of MS/MS spectra of precursor ions containing isobaric oligosaccharide structures. While limited semiquantitative information could be extracted from the raw data as well, the accuracy of this method should be significantly enhanced when standard calibration mixtures can be prepared. Copyright (c) 2017 John Wiley & Sons, Ltd.

Gas-Phase Fragmentation Behavior of Oxidized Prenyl Peptides by CID and ETD Tandem Mass Spectrometry.[Pubmed:27785692]

J Am Soc Mass Spectrom. 2017 Apr;28(4):704-707.

Farnesylation and geranylgeranylation are the two types of prenyl modification of proteins. Prenylated peptides are highly hydrophobic and their abundances in biological samples are low. In this report, we studied the oxidized prenylated peptides by electrospray ionization mass spectrometry and identified them by collision-induced dissociation (CID) and electron-transfer dissociation (ETD) tandem mass spectrometry. Modified prenyl peptides were generated utilizing strong and low strength oxidizing agents to selectively oxidize and epoxidize cysteine sulfur and prenyl side chain. We selected three peptides with prenyl motifs and synthesized their prenylated versions. The detailed characteristic fragmentations of oxidized and epoxidized farnesylated and geranylgeranylated peptides were studied side by side with two popular fragmentation techniques. CID and ETD mass spectrometry clearly distinguished the modified version of these peptides. ETD mass spectrometry provided sequence information of the highly labile modified prenyl peptides and showed different characteristic fragmentations compared with CID. A detailed fragmentation of modified geranylgeranylated peptides was compared by CID and ETD mass spectrometry for the first time. Graphical Abstract .

Modeling collision energy transfer in APCI/CID mass spectra of PAHs using thermal-like post-collision internal energy distributions.[Pubmed:27802636]

J Chem Phys. 2016 Oct 28;145(16):164311.

The internal energy transferred when projectile molecular ions of naphthalene collide with argon gas atoms was extracted from the APCI-CID (atmospheric-pressure chemical ionization collision-induced dissociation) mass spectra acquired as a function of collision energy. Ion abundances were calculated by microcanonical integration of the differential rate equations using the Rice-Ramsperger-Kassel-Marcus rate constants derived from a UB3LYP/6-311G+(3df,2p)//UB3LYP/6-31G(d) fragmentation mechanism and thermal-like vibrational energy distributions pME,Tchar. The mean vibrational energy excess of the ions was characterized by the parameter Tchar ("characteristic temperature"), determined by fitting the theoretical ion abundances to the experimental breakdown graph (a plot of relative abundances of the ions as a function of kinetic energy) of activated naphthalene ions. According to these results, the APCI ion source produces species below Tchar = 1457 K, corresponding to 3.26 eV above the vibrational ground state. Subsequent collisions heat the ions up further, giving rise to a sigmoid curve of Tchar as a function of Ecom (center-of-mass-frame kinetic energy). The differential internal energy absorption per kinetic energy unit (dEvib/dEcom) changes with Ecom according to a symmetric bell-shaped function with a maximum at 6.38 +/- 0.32 eV (corresponding to 6.51 +/- 0.27 eV of vibrational energy excess), and a half-height full width of 6.30 +/- 1.15 eV. This function imposes restrictions on the amount of energy that can be transferred by collisions, such that a maximum is reached as kinetic energy is increased. This behavior suggests that the collisional energy transfer exhibits a pronounced increase around some specific value of energy. Finally, the model is tested against the CID mass spectra of anthracene and pyrene ions and the corresponding results are discussed.

Potent and selective disruption of protein kinase D functionality by a benzoxoloazepinolone.[Pubmed:18829454]

J Biol Chem. 2008 Nov 28;283(48):33516-26.

Protein kinase D (PKD) is a novel family of serine/threonine kinases targeted by the second messenger diacylglycerol. It has been implicated in many important cellular processes and pathological conditions. However, further analysis of PKD in these processes is severely hampered by the lack of a PKD-specific inhibitor that can be readily applied to cells and in animal models. We now report the discovery of the first potent and selective cell-active small molecule inhibitor for PKD, benzoxoloazepinolone (CID755673). This inhibitor was identified from the National Institutes of Health small molecule repository library of 196,173 compounds using a human PKD1 (PKCmu)-based fluorescence polarization high throughput screening assay. CID755673 suppressed half of the PKD1 enzyme activity at 182 nm and exhibited selective PKD1 inhibition when compared with AKT, polo-like kinase 1 (PLK1), CDK activating kinase (CAK), CAMKIIalpha, and three different PKC isoforms. Moreover, it was not competitive with ATP for enzyme inhibition. In cell-based assays, CID755673 blocked phorbol ester-induced endogenous PKD1 activation in LNCaP cells in a concentration-dependent manner. Functionally, CID755673 inhibited the known biological actions of PKD1 including phorbol ester-induced class IIa histone deacetylase 5 nuclear exclusion, vesicular stomatitis virus glycoprotein transport from the Golgi to the plasma membrane, and the ilimaquinone-induced Golgi fragmentation. Moreover, CID755673 inhibited prostate cancer cell proliferation, cell migration, and invasion. In summary, our findings indicate that CID755673 is a potent and selective PKD1 inhibitor with valuable pharmacological and cell biological potential.