Cannabidiol disrupts conditioned fear expression and cannabidiolic acid reduces trauma-induced anxiety-related behaviour in mice.

The major phytocannabinoid cannabidiol (CBD) has anxiolytic properties and lacks tetrahydrocannabinol-like psychoactivity. Cannabidiolic acid (CBDA) is the acidic precursor to CBD, and this compound appears more potent than CBD in animal models of emesis, pain and epilepsy. In this short report, we aimed to examine whether CBDA is more potent than CBD in disrupting expression of conditioned fear and generalised anxiety-related behaviour induced by Pavlovian fear conditioning. Mice underwent fear conditioning and 24 h later were administered CBD and CBDA before testing for fear expression and generalized anxiety-like behaviour. We found that CBD and CBDA had dissociable effects; while CBD but not CBDA disrupted cued fear memory expression, CBDA but not CBD normalized trauma-induced generalized anxiety-related behaviour. Neither phytocannabinoid affected contextual fear expression. Our findings form the basis for future experiments examining whether phytocannabinoids, alone and in combination, are effective in these mouse models of fear and anxiety.

A latent green fluorescent styrylcoumarin probe for the selective growth and detection of Gram negative bacteria.

A novel, green fluorescent ß-alanylstyrylcoumarin derivative was synthesized and evaluated for its performance as a fluorogenic enzyme substrate on a range of clinically relevant microorganisms. The substrate was selectively hydrolysed by ß-alanyl aminopeptidase producing P. aeruginosa resulting in an on-to-off fluorescent signal. Growth inhibitory effect of the substrate was observed on Gram positive bacteria and yeasts. Meanwhile, Gram negative species, despite their extremely protective cell envelope, showed ready uptake and accumulation of the substrate within their healthy growing colonies displaying intense green fluorescence.

Methods for the detection and identification of pathogenic bacteria: past, present, and future.

In order to retard the rate of development of antibacterial resistance, the causative agent must be identified as rapidly as possible, so that directed patient treatment and/or contact precautions can be initiated. This review highlights the challenges associated with the detection and identification of pathogenic bacteria, by providing an introduction to the techniques currently used, as well as newer techniques that are in development. Focusing on the chemical basis for these techniques, the review also provides a comparison of their advantages and disadvantages.

Prospective phase II trial of nimotuzumab in combination with radiotherapy and concurrent capecitabine in locally advanced rectal cancer.

PURPOSE: The aim of the study was to evaluate the safety and efficacy of adding concurrent nimotuzumab to preoperative radiotherapy with concurrent capecitabine in locally advanced rectal cancer. METHODS AND MATERIALS: Patients with rectal cancer (clinical stage T3/4 or N+) were scheduled to receive weekly nimotuzumab (400 mg; days -6, 1, 8, 15, 22, and 29). Capecitabine (825 mg/m(2)) was delivered orally twice daily for the duration of radiotherapy. Radiotherapy was administered at 50.4 Gy (45 + 5.4 Gy). The main endpoint was the pathologic complete response (pCR) rate. RESULTS: Twenty-one patients with T3 or T4 disease were enrolled; 66.7 % were nodal-positive; the median distance from the anal verge was 5.5 cm. A pCR was achieved in four patients (19.0 %); 71.4 % patients obtained moderate or good tumor regression (Grade 2 and 3). Downstaging occurred in 15/21 (71.4 %) patients by T stage and 11/14 (78.6 %) by N stage. The actual dose intensities (median/mean, %) were nimotuzumab (100, 100) and capecitabine (100, 99.5). The most frequent Grade 1/2 toxicities were radiation dermatitis (57.1 %), nausea/vomiting (52.4 %), leukocytopenia (47.6 %), diarrhea (47.6 %), and proctitis (38.1 %). Grade 3 diarrhea was observed in 9.5 % of patients and Grade 3 leukocytopenia in 4.8 %. CONCLUSION: These preliminary results indicate that nimotuzumab can be safely combined with radiotherapy plus concurrent capecitabine. The efficacy of this regimen (pCR = 19.0 %) was significantly higher than that observed in previous phase II trials of preoperative radiotherapy with concurrent capecitabine and cetuximab in rectal cancer. Further investigation of concurrent nimotuzumab with radiotherapy plus capecitabine is warranted.

Structure-Activity Relationships, Deuteration, and Fluorination of Synthetic Cannabinoid Receptor Agonists Related to AKB48, 5F-AKB-48, and AFUBIATA.

Synthetic cannabinoid receptor agonists (SCRAs) are a growing class of new psychoactive substances (NPS) commonly derived from an N-alkylated indole, indazole, or 7-azaindole scaffold. Diversification of this core (at the 3-position) with amide-linked pendant amino acid groups and modular N-alkylation (of the indole/indazole/7-azaindole core) ensures that novel SCRAs continue to enter the illicit drug market rapidly. In response to the large number of SCRAs that have been detected, pharmacological evaluation of this NPS class has become increasingly common. Adamantane-derived SCRAs have consistently appeared throughout the market since 2011, and as such, a systematic set of these derivatives was synthesized and pharmacologically evaluated. Deuterated and fluorinated adamantane derivatives were prepared to evaluate typical hydrogen bioisosteres, as well as evaluation of the newly detected AFUBIATA.

Evaluation of the Possible Anticonvulsant Effect of Δ9-Tetrahydrocannabinolic Acid in Murine Seizure Models.

Introduction: The cannabinoid Δ9-tetrahydrocannabinolic acid (Δ9-THCA) has long been suggested in review articles and anecdotal reports to be anticonvulsant; yet, there is scant evidence supporting this notion. The objective of this study was to interrogate the anticonvulsant potential of Δ9-THCA in various seizure models-the Scn1a+/- mouse model of Dravet syndrome, the 6-Hz model of psychomotor seizures and the maximal electroshock (MES) model of generalized tonic-clonic seizures. Materials and Methods: We examined the effect of acute Δ9-THCA treatment against hyperthermia-induced seizures, and subchronic treatment on spontaneous seizures and survival in the Scn1a+/- mice. We also studied the effect of acute Δ9-THCA treatment on the critical current thresholds in the 6-Hz and MES tests using outbred Swiss mice. Highly purified Δ9-THCA was used in the studies or a mixture of Δ9-THCA and Δ9-THC. Results: We observed mixed anticonvulsant and proconvulsant effects of Δ9-THCA across the seizure models. Highly pure Δ9-THCA did not affect hyperthermia-induced seizures in Scn1a+/- mice. A Δ9-THCA/Δ9-THC mixture was anticonvulsant in the 6-Hz threshold test, but purified Δ9-THCA and Δ9-THC had no effect. Conversely, both Δ9-THCA and Δ9-THC administered individually were proconvulsant in the MES threshold test but had no effect when administered as a Δ9-THCA/Δ9-THC mixture. The Δ9-THCA/Δ9-THC mixture, however, increased spontaneous seizure severity and increased mortality of Scn1a+/- mice. Discussion: The anticonvulsant profile of Δ9-THCA was variable depending on the seizure model used and presence of Δ9-THC. Because of the unstable nature of Δ9-THCA, further exploration of Δ9-THCA through formal anticonvulsant drug development is problematic without stabilization. Future studies may better focus on determining the mechanisms by which combined Δ9-THCA and Δ9-THC alters seizure thresholds, as this may uncover novel targets for the control of refractory partial seizures.

The anticonvulsant phytocannabinoids CBGVA and CBDVA inhibit recombinant T-type channels.

Introduction: Cannabidiol (CBD) has been clinically approved for intractable epilepsies, offering hope that novel anticonvulsants in the phytocannabinoid class might be developed. Looking beyond CBD, we have recently reported that a series of biosynthetic precursor molecules found in cannabis display anticonvulsant properties. However, information on the pharmacological activities of these compounds on CNS drug targets is limited. The current study aimed to fill this knowledge gap by investigating whether anticonvulsant phytocannabinoids affect T-type calcium channels, which are known to modulate neuronal excitability, and may be relevant to the anti-seizure effects of this class of compounds. Materials and methods: A fluorescence-based assay was used to screen the ability of the phytocannabinoids to inhibit human T-type calcium channels overexpressed in HEK-293 cells. A subset of compounds was further examined using patch-clamp electrophysiology. Alphascreen technology was used to characterise selected compounds against G-protein coupled-receptor 55 (GPR55) overexpressed in HEK-293 cells, as GPR55 is another target of the phytocannabinoids. Results: A single 10 µM concentration screen in the fluorescence-based assay showed that phytocannabinoids inhibited T-type channels with substantial effects on Cav3.1 and Cav3.2 channels compared to the Cav3.3 channel. The anticonvulsant phytocannabinoids cannabigerovarinic acid (CBGVA) and cannabidivarinic acid (CBDVA) had the greatest magnitudes of effect (≥80% inhibition against Cav3.1 and Cav3.2), so were fully characterized in concentration-response studies. CBGVA and CBDVA had IC50 values of 6 µM and 2 µM on Cav3.1 channels; 2 µM and 11 µM on Cav3.2 channels, respectively. Biophysical studies at Cav3.1 showed that CBGVA caused a hyperpolarisation shift of steady-state inhibition. Both CBGVA and CBDVA had a use-dependent effect and preferentially inhibited Cav3.1 current in a slow inactivated state. CBGVA and CBDVA were also shown to antagonise GPR55. Conclusion and implications: These findings show that CBGVA and CBDVA inhibit T-type calcium channels and GPR55. These compounds should be further investigated to develop novel therapeutics for treating diseases associated with dysfunctional T-type channel activity.

Putative Synthetic Cannabinoids MEPIRAPIM, 5F-BEPIRAPIM (NNL-2), and Their Analogues Are T-Type Calcium Channel (CaV3) Inhibitors.

Synthetic cannabinoid receptor agonists (SCRAs) are a large and growing class of new psychoactive substances (NPSs). Two recently identified compounds, MEPIRAPIM and 5F-BEPIRAPIM (NNL-2), have not been confirmed as agonists of either cannabinoid receptor subtype but share structural similarities with both SCRAs and a class of T-type calcium channel (CaV3) inhibitors under development as new treatments for epilepsy and pain. In this study, MEPIRAPIM and 5F-BEPIRAPIM and 10 systematic analogues were synthesized, analytically characterized, and pharmacologically evaluated using in vitro cannabinoid receptor and CaV3 assays. Several compounds showed micromolar affinities for CB1 and/or CB2, with several functioning as low potency agonists of CB1 and CB2 in a membrane potential assay. 5F-BEPIRAPIM and four other derivatives were identified as potential CaV3 inhibitors through a functional calcium flux assay (>70% inhibition), which was further confirmed using whole-cell patch-clamp electrophysiology. Additionally, MEPIRAPIM and 5F-BEPIRAPIM were evaluated in vivo using a cannabimimetic mouse model. Despite detections of MEPIRAPIM and 5F-BEPIRAPIM in the NPS market, only the highest MEPIRAPIM dose (30 mg/kg) elicited a mild hypothermic response in mice, with no hypothermia observed for 5F-BEPIRAPIM, suggesting minimal central CB1 receptor activity.

Synthesis and pharmacological evaluation of newly detected synthetic cannabinoid receptor agonists AB-4CN-BUTICA, MMB-4CN-BUTINACA, MDMB-4F-BUTICA, MDMB-4F-BUTINACA and their analogs.

Synthetic cannabinoid receptor agonists (SCRAs) continue to make up a significant portion new psychoactive substances (NPS) detected and seized worldwide. Due to their often potent activation of central cannabinoid receptors in vivo, use of SCRAs can result in severe intoxication, in addition to other adverse health effects. Recent detections of AB-4CN-BUTICA, MMB-4CN-BUTINACA, MDMB-4F-BUTICA and MDMB-4F-BUTINACA mark a continuation in the appearance of SCRAs bearing novel tail substituents. The proactive characterization campaign described here has facilitated the detection of several new SCRAs in toxicological case work. Here we detail the synthesis, characterization, and pharmacological evaluation of recently detected SCRAs, as well as a systematic library of 32 compounds bearing head, tail, and core group combinations likely to appear in future. In vitro radioligand binding assays revealed most compounds showed moderate to high affinity at both CB1 (pK i = < 5 to 8.89 ± 0.09 M) and CB2 (pK i = 5.49 ± 0.03 to 9.92 ± 0.09 M) receptors. In vitro functional evaluation using a fluorescence-based membrane potential assay showed that most compounds were sub-micromolar to sub-nanomolar agonists at CB1 (pEC50 = < 5 to 9.48 ± 0.14 M) and CB2 (pEC50 = 5.92 ± 0.16 to 8.64 ± 0.15 M) receptors. An in silico receptor-ligand docking approach was utilized to rationalize binding trends for CB2 with respect to the tail substituent, and indicated that rigidity in this region (i.e., 4-cyanobutyl) was detrimental to affinity.

Systematic evaluation of a panel of 30 synthetic cannabinoid receptor agonists structurally related to MMB-4en-PICA, MDMB-4en-PINACA, ADB-4en-PINACA, and MMB-4CN-BUTINACA using a combination of binding and different CB1 receptor activation assays: Part I-Synthesis, analytical characterization, and binding affinity for human CB1 receptors.

Synthetic cannabinoid receptor agonists (SCRAs) are one of the largest and most structurally diverse classes of new psychoactive substances (NPS). Despite this, pharmacological data are often lacking following the identification of a new SCRA in drug markets. In this first of a three-part series, we describe the synthesis, analytical characterization, and binding affinity of a proactively generated, systematic library of 30 indole, indazole, and 7-azaindole SCRAs related to MMB-4en-PICA, MDMB-4en-PINACA, ADB-4en-PINACA, and MMB-4CN-BUTINACA featuring a 4-pentenyl (4en-P), butyl (B/BUT), or 4-cyanobutyl (4CN-B/BUT) tail and a methyl l-valinate (MMB), methyl l-tert-leucinate (MDMB), methyl l-phenylalaninate (MPP), l-valinamide (AB), l-tert-leucinamide (ADB), l-phenylalaninamide (APP), adamantyl (A), or cumyl head group. Competitive radioligand binding assays demonstrated that the indazole core conferred the highest CB1 binding affinity (Ki = 0.17-39 nM), followed by indole- (Ki = 0.95-160 nM) and then 7-azaindole-derived SCRAs (Ki = 5.4-271 nM). Variation of the head group had the greatest effect on binding, with tert-leucine amides and methyl esters (Ki = 0.17-14 nM) generally showing the greatest affinities, followed by valine derivatives (Ki = 0.72-180 nM), and then phenylalanine derivatives (Ki = 2.5-271 nM). Adamantyl head groups (Ki = 8.8-59 nM) were suboptimal for binding, whereas the cumyl analogues consistently conferred high affinity (Ki = 0.62-36 nM). Finally, both butyl (Ki = 3.1-163 nM) and 4-cyanobutyl (Ki = 5.5-44 nM) tail groups were less favorable for CB1 binding than their corresponding 4-pentenyl counterparts (Ki = 0.72-25 nM).

Straw decreased N2O emissions from flooded paddy soils via altering denitrifying bacterial community compositions and soil organic carbon fractions.

Straw return is widely applied to increase soil fertility and soil organic carbon storage. However, its effect on N2O emissions from paddy soil and the associated microbial mechanisms are still unclear. In this study, wheat straw was amended to two paddy soils (2% w/w) from Taizhou (TZ) and Yixing (YX), China, which were flooded and incubated for 30 d. Real-time PCR and Illumina sequencing were used to characterize changes in denitrifying functional gene abundance and denitrifying bacterial communities. Compared to unamended controls, straw addition significantly decreased accumulated N2O emissions in both TZ (5071 to 96 mg kg-1) and YX (1501 to 112 mg kg-1). This was mainly due to reduced N2O production with decreased abundance of major genera of nirK and nirS-bacterial communities and reduced nirK and nirS gene abundances. Further analyses showed that nirK-, nirS- and nosZ-bacterial community composition shifted mainly along the easily oxidizable carbon (EOC) arrows following straw amendment among four different soil organic carbon fractions, suggesting that increased EOC was the main driver of alerted denitrifying bacterial community composition. This study revealed straw return suppressed N2O emission via altering denitrifying bacterial community compositions and highlighted the importance of EOC in controlling denitrifying bacterial communities.

Synthesis and in Vitro Cannabinoid Receptor 1 Activity of Recently Detected Synthetic Cannabinoids 4F-MDMB-BICA, 5F-MPP-PICA, MMB-4en-PICA, CUMYL-CBMICA, ADB-BINACA, APP-BINACA, 4F-MDMB-BINACA, MDMB-4en-PINACA, A-CHMINACA, 5F-AB-P7AICA, 5F-MDMB-P7AICA, and 5F-AP7AICA.

Synthetic cannabinoid receptor agonists (SCRAs) are an evolving class of new psychoactive substances (NPS) with structurally diverse compounds emerging each year. Due to the rapid pace at which these drugs enter the market, there is often little or nil information regarding the pharmacology of these substances despite widespread human use. In this study, 12 recently emerged SCRAs (reported between 2018 and 2020) were synthesized, analytically characterized, and pharmacologically evaluated using a live cell-based nanoluciferase complementation reporter assay that monitors in vitro cannabinoid receptor type 1 (CB1) activation via its interaction with ß-arrestin 2 (ßarr2). All synthesized SCRAs acted as agonists of CB1, although differences in potency (EC50 = 2.33-5475 nM) and efficacy (Emax = 37-378%) were noted, and several structure-activity relationships were identified. SCRAs featuring indazole cores (EC50 = 2.33-159 nM) were generally of equal or greater potency than indole analogues (EC50 = 32.9-330 nM) or 7-azaindole derivatives (EC50 = 64.0-5475 nM). Interestingly, with the exception of APP-BINACA (Emax = 75.7%) and 5F-A-P7AICA (Emax = 37.4%), all SCRAs showed greater efficacy than the historical SCRA JWH-018 to which responses were normalized (Emax = 142-378%). The most potent CB1 agonists in the study were ADB-BINACA (EC50 = 6.36 nM), 4F-MDMB-BINACA (EC50 = 7.39 nM), and MDMB-4en-PINACA (EC50 = 2.33 nM). Notably, all of these SCRAs featured an indazole core as well as a "bulky" tert-butyl moiety in the pendant amino acid side chain. This study confirms that recently detected SCRAs 4F-MDMB-BICA, 5F-MPP-PICA, MMB-4en-PICA, CUMYL-CBMICA, ADB-BINACA, APP-BINACA, 4F-MDMB-BINACA, MDMB-4en-PINACA, A-CHMINACA, 5F-AB-P7AICA, 5F-MDMB-P7AICA, and 5F-AP7AICA were all able to activate the CB1 receptor in vitro, albeit to different extents, and are potentially psychoactive in vivo. These results indicate that further evaluation of these widely used NPS is warranted to better understand the risks associated with human consumption of these drugs.

Inhibitory Kappa B Kinase α (IKKα) Inhibitors That Recapitulate Their Selectivity in Cells against Isoform-Related Biomarkers.

IKKß plays a central role in the canonical NF-kB pathway, which has been extensively characterized. The role of IKKα in the noncanonical NF-kB pathway, and indeed in the canonical pathway as a complex with IKKß, is less well understood. One major reason for this is the absence of chemical tools designed as selective inhibitors for IKKα over IKKß. Herein, we report for the first time a series of novel, potent, and selective inhibitors of IKKα. We demonstrate effective target engagement and selectivity with IKKα in U2OS cells through inhibition of IKKα-driven p100 phosphorylation in the noncanonical NF-kB pathway without affecting IKKß-dependent IKappa-Bα loss in the canonical pathway. These compounds represent the first chemical tools that can be used to further characterize the role of IKKα in cellular signaling, to dissect this from IKKß and to validate it in its own right as a target in inflammatory diseases.

[Laser treatment of port wine stains].

OBJECTIVE: To assess the efficiency of the PDL in the treatment of PWS in Chinese patients with skin type III-IV. METHODS: 252 patients with PWS at different anatomical sites were retrospectively studied. A flashlamp-pumped pulsed dye laser was used. The differences in therapeutic effect of laser were analyzed in the following ways: (1) by dermatomal distribution of face and neck; (2) by anatomical subdivision of face; (3) by the size of the lesion. The therapeutic effect was graded as 0 < or = poor < 25%, 25% < or = fair < 50%, 50% < or = good < 75%, 75% < or = excellent < or = 100%. RESULTS: It revealed that the lesions at neck had the best response. The dermatome II zone showed the least response to PDL. Among the anatomical subdivision of face, the frontal area had the highest clearance, while the zygoma and cheek had the lowest clearance. Patients with PWS on the extremities respond less favorably with the fair lightening effect. Furthermore, the mean lightening was decreased as the size of PWS was increased. CONCLUSIONS: The response difference of PWS to PDL was not only related to the anatomical region, but also related to the lesion size. It is imperative to give the patients rational treatment suggestion and normal expectation.

[Application of fractional laser resurfacing in the treatment of superficial scar].

OBJECTIVE: To study the therapeutic effect, safety and risk of fractional resurfacing with ablative laser in the treatment of superficial scar. METHODS: 88 cases of superficial scar, including 66 cases of acne scar, 12 cases of burn scar and 10 cases of other scar, were treated. All the patients were treated with Pixel (Er:YAG 2940 nm, > or = 3 times), or Encore (Ultrapulse CO2 10600 nm, > or = 2 times), or a combination of Pixel and Encore( > or = 3 times). RESULTS: The therapeutic effect of treatment was graded as medium when the scar was improved > 25%, as good when the scar was improved > 50%. The effective result was achieved in 80% of the patients. Good effect was achieved in 50% of the patients. There were 244 treatments of Pixel and 86 treatments of Encore. Persistent hyperpigmentation was happened in one case with Encore treatment, which relieved four months later. No other complication was happened. CONCLUSIONS: The fractional resurfacing with ablative laser is an effective method for superficial scar, especially acne scar. It is mini-invasive with low risk. The key mechanism is the ability of enhancing the tissue remodeling induced by heat effect of laser.

Rhinoplasty in Chinese: management of lower dorsum and bulbous nasal tip.

BACKGROUND: Bulbous nasal tips and lower dorsa are common facial features in Chinese people, and surgery to reshape these is frequently requested. The use of silicone implants in rhinoplasty has been widely used in China for many years, but is not suitable for patients seeking Caucasian tip shapes. The creation of an excessively high tip supported only by a silicone implant inevitably leads to implant extrusion. Although many rhinoplasty techniques have been used in Caucasian patients, there is currently no suitable method for Chinese patients, whose anatomy differs from that of Caucasians. The present study was aimed to investigate the clinical outcome of a novel method of rhinoplasty in Chinese people. METHODS: Eighty patients underwent rhinoplasty using our method between 2002 and 2006. We classified the patients into three types, according to the distance between tip defining points, and used different techniques accordingly. Furthermore, an innovative cartilage carving method and a tip fibro-fatty tissue flap were designed and combined with traditional techniques, such as insertion of silicone implant, cartilage grafts, suture techniques and cephalic trimming to reshape the nasal contours. The followup period was 10 - 60 months (average, 21 months). RESULTS: Remarkable modifications in nasal contours were achieved. No complications developed in any of the 80 patients. Seventy-eight patients were satisfied with the results. The outcomes remained unchanged over time. CONCLUSION: Our method is effective and suitable for the treatment of Chinese patients with lower dorsa and bulbous nasal tips.

Effect of 7-hydroxystaurosporine on glioblastoma cell invasion and migration.

AIM: To investigate the effect of 7-hydroxystaurosporine (UCN-01), a selective protein kinase C (PKC) inhibitor, on cell growth, migration, and invasion in invasive human glioblastoma U-87MG cells. METHODS: PKC activity was determined based on the PKC-catalyzed transfer of the (32)P-phosphate group from [g-(32)P]ATP into a PKC-specific peptide substrate. Cell viability was measured by MTT assay. Cell invasion and migration were evaluated by a Boyden chamber assay and scratch wound assay, respectively. Protein expression was analyzed using Western blot assay. The formation of 3-dimensional cellular aggregates was examined by a cell-cell aggregation assay. RESULTS: UCN-01 treatment resulted in concentration- and time-dependent inhibition of U-87MG cell growth at higher doses (>100 nmol/L), and reduced cell invasion and migration capability at less cytotoxic doses (<100 nmol/L). UCN-01 significantly repressed PKC activity. Consistent with this result, UCN-01 blocked cell invasion stimulated by phorbel 12-myristate-13-acetate (PMA) and ethanol (EtOH), 2 PKC activators. Enforced expression of the tumor suppressor genes BRCA1 and PTEN increased the anti-invasion potential of UCN-01. Exposure to UCN-01 caused a dose-dependent increase in cell adhesion molecule E-cadherin. The effect of UCN-01 on the formation of cell-cell aggregation was significantly reduced by the addition of an anti-E-cadherin antibody. CONCLUSION: UCN-01 inhibits the invasion and migration of human glioma cells. Accordingly, UCN-01 can have potential clinical applications for the treatment of human glioma metastasis.

[Interaction between ATM and radiation-activated phosphorylation of P53 and P21].

BACKGROUND & OBJECTIVE: ATM gene is a member of PI-3K kinase family. ATM protein is capable of controlling DNA repair process and cell cycle checkpoint. In AT cells from ataxia-telangiectasia (AT) patients, ATM gene mutation leads to the deficiency of ionizing radiation-activated phosphorylation of P53 and P21. It shows ATM gene could mediate the phosphorylation of P53 and P21. This study was to explore the interaction between ATM and P53, and to observe whether ATM directly medicates the phosphorylation of P21 in a P53-independent way. METHODS: pEBS7-YZ5 vector containing ATM cDNA was transfected into AT cells by electroperforation. The cells expressing ATM protein stably were screened with hygromycin, and identified by reverse transcription-polymerase chain reaction (RT-PCR). The interaction between ATM and P53 in pEBS7-YZ5-AT cells was assessed by co-immunoprecipitation and Western blot. K562 cells served as a P53 mutation cell model to study whether ATM could interact with the phosphorylation of P21. RESULTS: pEBS7-YZ5 was transfected into AT cells successfully. RT-PCR detected fragment of ATM cDNA. After exposed to ionizing radiation, P53 of pEBS7-YZ5-AT cells was phosphorylated, and immunoprecipitation showed interaction between ATM and P53; P21 of K562 cells was phosphorylated, P21 protein was detected in the immunoprecipitation of ATM antibody-complex. CONCLUSION: Ionizing radiation-activated ATM kinase could interact with the phosphorylation of P53 and P21 in both P53 wild type and mutant type cells.