A Hierarchical Approach for Traffic Sign Recognition Based on Shape Detection and Image Classification.

In recent years, the development of self-driving cars and their inclusion in our daily life has rapidly transformed from an idea into a reality. One of the main issues that autonomous vehicles must face is the problem of traffic sign detection and recognition. Most works focusing on this problem utilize a two-phase approach. However, a fast-moving car has to quickly detect the sign as seen by humans and recognize the image it contains. In this paper, we chose to utilize two different solutions to solve tasks of detection and classification separately and compare the results of our method with a novel state-of-the-art detector, YOLOv5. Our approach utilizes the Mask R-CNN deep learning model in the first phase, which aims to detect traffic signs based on their shapes. The second phase uses the Xception model for the task of traffic sign classification. The dataset used in this work is a manually collected dataset of 11,074 Taiwanese traffic signs collected using mobile phone cameras and a GoPro camera mounted inside a car. It consists of 23 classes divided into 3 subclasses based on their shape. The conducted experiments utilized both versions of the dataset, class-based and shape-based. The experimental result shows that the precision, recall and mAP can be significantly improved for our proposed approach.

Stem cell mobilizers targeting chemokine receptor CXCR4: renoprotective application in acute kidney injury.

We have discovered a novel series of quinazoline-based CXCR4 antagonists. Of these, compound 19 mobilized CXCR4(+) cell types, including hematopoietic stem cells and endothelial progenitor cells, more efficiently than the marketed 1 (AMD3100) with subcutaneous administration at the same dose (6 mg/kg) in mice. This series of compounds thus provides a set of valuable tools to study diseases mediated by the CXCR4/SDF-1 axis, including myocardial infarction, ischemic stroke, and cancer metastasis. More importantly, treatment with compound 19 significantly lowered levels of blood urea nitrogen and serum creatinine in rats with renal ischemia-reperfusion injury, providing evidence for its therapeutic potential in preventing ischemic acute kidney injury. CXCR4 antagonists such as 19 might also be useful to increase circulating levels of adult stem cells, thereby exerting beneficial effects on damaged and/or inflamed tissues in diseases that currently are not treated by standard approaches.

Discovery of 1-(2,4-dichlorophenyl)-N-(piperidin-1-yl)-4-((pyrrolidine-1-sulfonamido)methyl)-5-(5-((4-(trifluoromethyl)phenyl)ethynyl)thiophene-2-yl)-1H-pyrazole-3-carboxamide as a novel peripherally restricted cannabinoid-1 receptor antagonist with significant weight-loss efficacy in diet-induced obese mice.

After extensive synthetic efforts, we found that many structurally diverse bioisosteres could be generated via derivatizing the C-4 alkyl chain on the pyrazole ring of compound 3 (B/P = 1/33) with different electronegative groups. Especially when a sulfonamide or sulfamide moiety was added, resulting compounds exhibited not only potent CB1R activity but also a desired tPSA value over 90 Å(2), a threshold considered to possess a low probability to cross BBB, leading to the identification of compound 4 (B/P = 1/64) as a peripherally restricted CB1R antagonist. Apart from its significant weight-loss efficacy in DIO mice, compound 4 also displays 163 clean off-target profiles and is currently under development for treating obesity and the related metabolic syndrome.

Deregulated Cdk5 triggers aberrant activation of cell cycle kinases and phosphatases inducing neuronal death.

Aberrant activation of cell cycle proteins is believed to play a critical role in Alzheimer's disease (AD) pathogenesis; although, the molecular mechanisms leading to their activation in diseased neurons remain elusive. The goal of this study was to investigate the mechanistic link between Cdk5 deregulation and cell cycle re-activation in ß-amyloid(1-42) (Aß(1-42))-induced neurotoxicity. Using a chemical genetic approach, we identified Cdc25A, Cdc25B and Cdc25C as direct Cdk5 substrates in mouse brain lysates. We show that deregulated Cdk5 directly phosphorylates Cdc25A, Cdc25B and Cdc25C at multiple sites, which not only increases their phosphatase activities but also facilitates their release from 14-3-3 inhibitory binding. Cdc25A, Cdc25B and Cdc25C in turn activate Cdk1, Cdk2 and Cdk4 kinases causing neuronal death. Selective inhibition of Cdk5 abrogates Cdc25 and Cdk activations in Aß(1-42)-treated neurons. Similarly, phosphorylation-resistant mutants of Cdc25 isoforms at Cdk5 sites are defective in activating Cdk1, Cdk2 and Cdk4 in Aß(1-42)-treated primary cortical neurons, emphasizing a major role of Cdk5 in the activation of Cdc25 isoforms and Cdks in AD pathogenesis. These results were further confirmed in human AD clinical samples, which had higher Cdc25A, Cdc25B and Cdc25C activities that were coincident with increased Cdk5 activity, as compared to age-matched controls. Inhibition of Cdk5 confers the highest neuroprotection against Aß(1-42) toxicity, whereas inhibition of Cdc25 isoforms was partially neuroprotective, further emphasizing a decisive role of Cdk5 deregulation in cell-cycle-driven AD neuronal death.

LIMK2 is a crucial regulator and effector of Aurora-A-kinase-mediated malignancy.

Aurora A is overexpressed in majority of breast carcinomas. With the exception of BRCA1 and PHLDA1, no oncogenic Aurora A substrates are known in breast cancer. In this study, a chemical genetic approach was used to identify malignant targets of Aurora A, which revealed LIMK2 as a novel Aurora A substrate. Aurora A regulates LIMK2 kinase activity, subcellular localization and protein levels by direct phosphorylation at S283, T494 and T505. In response, LIMK2 also positively regulates the level of Aurora A, thereby engaging in a positive-feedback loop, promoting Aurora-A-mediated oncogenic pathways. Most importantly, LIMK2 ablation fully abrogates Aurora-A-mediated tumorigenesis in nude mice, suggesting that LIMK2 is a key oncogenic effector of Aurora A. Furthermore, LIMK2 ablation acts synergistically with inhibition of Aurora A in promoting cell death. Finally, Aurora-A-mediated upregulation of LIMK2 appears to be a common mechanism in many cancers. LIMK2 inhibition or ablation is therefore an alternative approach for modulating Aurora A deregulation in cancer.

Direct effects of HIV-1 Tat on excitability and survival of primary dorsal root ganglion neurons: possible contribution to HIV-1-associated pain.

The vast majority of people living with human immunodeficiency virus type 1 (HIV-1) have pain syndrome, which has a significant impact on their quality of life. The underlying causes of HIV-1-associated pain are not likely attributable to direct viral infection of the nervous system due to the lack of evidence of neuronal infection by HIV-1. However, HIV-1 proteins are possibly involved as they have been implicated in neuronal damage and death. The current study assesses the direct effects of HIV-1 Tat, one of potent neurotoxic viral proteins released from HIV-1-infected cells, on the excitability and survival of rat primary dorsal root ganglion (DRG) neurons. We demonstrated that HIV-1 Tat triggered rapid and sustained enhancement of the excitability of small-diameter rat primary DRG neurons, which was accompanied by marked reductions in the rheobase and resting membrane potential (RMP), and an increase in the resistance at threshold (R(Th)). Such Tat-induced DRG hyperexcitability may be a consequence of the inhibition of cyclin-dependent kinase 5 (Cdk5) activity. Tat rapidly inhibited Cdk5 kinase activity and mRNA production, and roscovitine, a well-known Cdk5 inhibitor, induced a very similar pattern of DRG hyperexcitability. Indeed, pre-application of Tat prevented roscovitine from having additional effects on the RMP and action potentials (APs) of DRGs. However, Tat-mediated actions on the rheobase and R(Th) were accelerated by roscovitine. These results suggest that Tat-mediated changes in DRG excitability are partly facilitated by Cdk5 inhibition. In addition, Cdk5 is most abundant in DRG neurons and participates in the regulation of pain signaling. We also demonstrated that HIV-1 Tat markedly induced apoptosis of primary DRG neurons after exposure for longer than 48 h. Together, this work indicates that HIV-1 proteins are capable of producing pain signaling through direct actions on excitability and survival of sensory neurons.

Synthesis and in-vitro anticancer activity of 3,5-bis(indolyl)-1,2,4-thiadiazoles.

A series of 3,5-bis(indolyl)-1,2,4-thiadiazoles were synthesized and evaluated for their cytotoxicity against selected human cancer cell lines. The reaction of indole-3-thiocarboxamide 3 with iodobenzene diacetate underwent oxidative dimerization to give 3,5-bis(indolyl)-1,2,4-thiadiazoles 4a-n. Among the synthesized bis(indoly)-1,2,4-thiadiazoles, the compound 4h with 4-chlorobenzyl and methoxy substituents showed the most potent activity.

PHLDA1 is a crucial negative regulator and effector of Aurora A kinase in breast cancer.

Aurora A kinase is overexpressed in the majority of breast carcinomas. A chemical genetic approach was used to identify the malignant targets of Aurora A, which revealed pleckstrin-homology-like domain protein PHLDA1 as an Aurora A substrate. PHLDA1 downregulation is a powerful prognostic predictor for breast carcinoma, which was confirmed in our study. We further show that downregulation of PHLDA1 is associated with estrogen receptor (ER) expression in breast carcinoma. Aurora A directly phosphorylates PHLDA1 leading to its degradation. PHLDA1 also negatively regulates Aurora A, thereby triggering a feedback loop. We demonstrate the underlying mechanisms by which PHLDA1 upregulation strongly antagonizes Aurora-A-mediated oncogenic pathways, thereby revealing PHLDA1 degradation as a key mechanism by which Aurora A promotes breast malignancy. Thus, not surprisingly, PHLDA1 upregulation acts synergistically with Aurora A inhibition in promoting cell death. PHLDA1 overexpression might therefore be an alternative method to modulate Aurora A deregulation in breast carcinoma. Finally, this study led to the discovery of a mutation in the Aurora A active site that renders it amenable to the chemical genetic approach. Similar mutations are required for Aurora B, suggesting that this modified approach can be extended to other kinases that have hitherto not been amenable to this methodology.

Glutathione-S-transferase P1 is a critical regulator of Cdk5 kinase activity.

Cyclin dependent kinase-5 (Cdk5) activity is deregulated in Alzheimer's disease (AD) and contributes to all three hallmarks: neurotoxic ß-amyloid formation, neurofibrillary tangles, and neuronal death. However, the mechanism leading to Cdk5 deregulation remains controversial. Cdk5 deregulation in AD is usually linked to the formation of p25, a proteolysis product of Cdk5 activator p35, which leads to Cdk5 mislocalization and hyperactivation. A few studies have indeed shown increased p25 levels in AD brains; however, others have refuted this observation. These contradictory findings suggest that additional factors contribute to Cdk5 deregulation. This study identified glutathione-S-transferase pi 1 (GSTP1) as a novel Cdk5 regulatory protein. We demonstrate that it is a critical determinant of Cdk5 activity in human AD brains and various cancer and neuronal cells. Increased GSTP1 levels were consistently associated with reduced Cdk5 activity. GSTP1 directly inhibits Cdk5 by dislodging p25/p35, and indirectly by eliminating oxidative stress. Cdk5 promotes and is activated by oxidative stress, thereby engaging a feedback loop which ultimately leads to cell death. Not surprisingly, GSTP1 transduction conferred a high degree of neuroprotection under neurotoxic conditions. Given the critical role of oxidative stress in AD pathogenesis, an increase in GSTP1 level may be an alternative way to modulate Cdk5 signaling, eliminate oxidative stress, and prevent neurodegeneration.

Synthesis of novel 1,2,4-oxadiazoles and analogues as potential anticancer agents.

A library of 3,5-disubstituted-1,2,4-oxadiazoles 7-9 and their bioisosters, 1,3,4-oxadiazole 14 and 1,3,4-thiadiazole 16, were synthesized and evaluated in vitro for their anticancer potential against a panel of six human cancer cell lines. The key step in the synthesis of oxadiazoles 7-9 involve coupling of amidoxime 6 with an appropriate carboxylic acid followed by thermal cyclization. The bioisosteres, 1,3,4-oxadiazole 14 and 1,3,4-thiadiazole 16 were prepared from the reaction of a common precursor diacylhydrazine 13 with thionyl chloride and Lawesson's reagent, respectively. The anticancer studies on the synthesized compounds revealed that presence of a cyclopentyloxy or n-butyloxy on the C-3 aryl ring and piperdin-4-yl or trichloromethyl at the C-5 position of 1,2,4-oxadiazole is essential for good activity. In particular, 1,2,4-oxadiazole 7i and analogue 1,3,4-thiadiazole 16 exhibited significant activity against DU145 (IC(50): 9.3 µM) and MDA-MB-231 (IC(50): 9.2 µM) cell lines, respectively.

Nuclear envelope dispersion triggered by deregulated Cdk5 precedes neuronal death.

Nuclear fragmentation is a common feature in many neurodegenerative diseases, including Alzheimer's disease (AD). In this study, we show that nuclear lamina dispersion is an early and irreversible trigger for cell death initiated by deregulated Cdk5, rather than a consequence of apoptosis. Cyclin-dependent kinase 5 (Cdk5) activity is significantly increased in AD and contributes to all three hallmarks: neurotoxic amyloid-ß (Aß), neurofibrillary tangles (NFT), and extensive cell death. Using Aß and glutamate as the neurotoxic stimuli, we show that deregulated Cdk5 induces nuclear lamina dispersion by direct phosphorylation of lamin A and lamin B1 in neuronal cells and primary cortical neurons. Phosphorylation-resistant mutants of lamins confer resistance to nuclear dispersion and cell death on neurotoxic stimulation, highlighting this as a major mechanism for neuronal death. Rapid alteration of lamin localization pattern and nuclear membrane change are further supported by in vivo data using an AD mouse model. After p25 induction, the pattern of lamin localization was significantly altered, preceding neuronal death, suggesting that it is an early pathological event in p25-inducible transgenic mice. Importantly, lamin dispersion is coupled with Cdk5 nuclear localization, which is highly neurotoxic. Inhibition of nuclear dispersion rescues neuronal cells from cell death, underscoring the significance of this event to Cdk5-mediated neurotoxicity.

One-pot synthesis and anticancer studies of 2-arylamino-5-aryl-1,3,4-thiadiazoles.

A series of 2-arylamino-5-aryl-1,3,4-thiadiazoles 1a-j were synthesized and screened for their anticancer activity against various human cancer cell lines. The novel one-pot synthesis of 1,3,4-thiadiazoles was achieved by refluxing aryl aldehydes, hydrazine hydrate, and aryl isothiocyanates in methanol followed by oxidative cyclization with ferric ammonium sulfate. The compounds 1g-j with trimethoxyphenyl at the C-5 position displayed extremely potent anticancer activity with at least twofold selectivity (IC(50): 4.3-9.2 µM). The nature of substituent on the C-2 arylamino ring may be critical in opting for the selectivity towards a particular cancer cell.

Synthesis of novel indolyl-1,2,4-triazoles as potent and selective anticancer agents.

A diverse series of 22 indolyl-1,2,4-triazole congeners (6 and 7) have been synthesized from the reaction of indole-3-carbonitrile (4) or (5) with appropriate acid hydrazides in the presence of potassium carbonate. Synthesized compounds were evaluated for their cytotoxicity against six human cancer cell lines, and some of the compounds displayed promising activity. In particular, 3-(3',4',5'-trimethoxyphenyl)-5-(N-methyl-3'-indolyl)-1,2,4-triazole (7i) and 3-(4'-piperidinyl)-5-(N-methyl-3'-indolyl)-1,2,4-triazole (7n) were the most promising and broadly active compounds against the tested cell lines. It was interesting to note that the trimethoxyphenyl analog 7i showed twofold selective cytotoxicity against PaCa2 cell line (IC(50) 0.8 µm), whereas piperidinyl analog 7n was found to be selectively cytotoxic against MCF7 cell line (IC(50) 1.6 µm). Notably, the 4-fluorophenyl derivative 7c exhibited selective cytotoxicity against PC3 cell line (IC(50) 4 µm). The structure-activity relationship study revealed that substituents including 3,4,5-trimethoxyphenyl, 3,4-dimethoxyphenyl, 4-benzyloxy-3-methoxyphenyl, 4-piperidinyl, 4-fluorophenyl and N-methylindole are beneficial for the activity of indolyl-1,2,4-triazoles (6 and 7).

Synthesis and anticancer activity of 5-(3-indolyl)-1,3,4-thiadiazoles.

A series of 5-(3-indolyl)-2-substituted-1,3,4-thiadiazoles 5a-m were synthesized and their cytotoxicity analyzed against six human cancer cell lines. The reaction of indole-3-carboxylic acid 3 with aryl or heteroaryl hydrazides afforded the N,N'-diacylhydrazines 4, which upon treatment with Lawesson's reagent resulted in the formation of indolyl-1,3,4-thiadiazoles 5a-m in good yields. Indolyl-1,3,4-thiadiazole 5m with 4-benzyloxy-3-methoxyphenyl and 5-bromo indolyl substituents is the most active in suppressing the growth of cancer cells (IC(50) 1.5 muM, PaCa2). The compounds 5b, 5e and 5h bearing C-2 substituent as benzyl, 3,4-dimethoxyphenyl and 4-benzyloxy-3-methoxyphenyl, respectively, have shown significant cytotoxicity against multiple cancer cell lines. Introduction of 4-dimethylamino (5d and 5k) and 3,4,5-trimethoxy (5l) groups in the C-2 phenyl ring induced selectivity against MCF7 and MDA-MB-231 cancer cell lines (compounds 5d, 5k and 5l).

Cdk5 is a major regulator of p38 cascade: relevance to neurotoxicity in Alzheimer's disease.

Cyclin-dependent kinase (Cdk) 5 and p38 activities are significantly increased in Alzheimer's Disease (AD). Both p38 and Cdk5 promote neurodegeneration upon deregulation. However, to date the mechanistic link between Cdk5 and p38 remains unclear. This study presents the first mechanism showing Cdk5 as a major regulator of p38 cascade in neurons and in transgenic mouse model of AD. Using beta-amyloid and glutamate as the neurotoxic stimuli, our results show that deregulated Cdk5 induces p38 activation by increasing reactive oxygen species (ROS) in neuronal cells and in primary cortical neurons. Elimination of ROS inhibits p38 activation, revealing ROS as major stimuli of the p38 cascade. Importantly, Cdk5-mediated p38 activation increases c-Jun expression, thereby revealing a mechanistic link between deregulated Cdk5 and c-Jun level in AD brains. c-Jun is over-expressed in AD, and is believed to contribute significantly to neurodegeneration. Based on the proposed mechanism, Cdk5 inhibition is more neuroprotective relative to p38 and c-Jun, suggesting that Cdk5 is an upstream regulator of neurodegenerative pathways triggered by p38 and a preferable therapeutic target for AD.

Stereoselective recognition of tripeptides guided by encoded library screening: construction of chiral macrocyclic tetraamide ruthenium receptor for peptide sensing.

[structure: see text] Molecule sensor 1 is devised by incorporating the reporting unit of ruthenium(II) complex and two recognition motifs of chiral cyclotetraamides on the sidearms. The target binding tripeptides for sensor 1 were readily identified by using an encoded library screening method. This solid-phase screening indicated a preferable binding of molecule 1 with d-alanine over the l-isomer. The optical and NMR studies for the binding events of 1 with tripeptides Ac-Ala-Gly-Ala-NHC(12)H(25) in the solution phase showed a consistent trend for the stereoselective recognition of the dd-isomer over the ld-, dl-, and ll-isomers.