First Detection of sub-PeV Diffuse Gamma Rays from the Galactic Disk: Evidence for Ubiquitous Galactic Cosmic Rays beyond PeV Energies.

We report, for the first time, the long-awaited detection of diffuse gamma rays with energies between 100 TeV and 1 PeV in the Galactic disk. Particularly, all gamma rays above 398 TeV are observed apart from known TeV gamma-ray sources and compatible with expectations from the hadronic emission scenario in which gamma rays originate from the decay of π^{0}'s produced through the interaction of protons with the interstellar medium in the Galaxy. This is strong evidence that cosmic rays are accelerated beyond PeV energies in our Galaxy and spread over the Galactic disk.

First Detection of Photons with Energy beyond 100 TeV from an Astrophysical Source.

We report on the highest energy photons from the Crab Nebula observed by the Tibet air shower array with the underground water-Cherenkov-type muon detector array. Based on the criterion of a muon number measured in an air shower, we successfully suppress 99.92% of the cosmic-ray background events with energies E>100 TeV. As a result, we observed 24 photonlike events with E>100 TeV against 5.5 background events, which corresponds to a 5.6σ statistical significance. This is the first detection of photons with E>100 TeV from an astrophysical source.

Functional allele and genotype frequencies of CYP1A2, CYP2B6 and iNOS among mainland Chinese Tibetan, Mongolian, Uygur and Han populations.

WHAT IS KNOWN AND OBJECTIVE: Cytochrome P450 1A2 (CYP1A2), CYP2B6 and inducible nitric oxide synthase (iNOS) are involved in the metabolism and action of many important therapeutic drugs, and genetic variants have been associated with interethnic differences in response to treatment, including chemotherapy. METHODS: Eight hundred and forty-two unrelated Chinese healthy subjects (323 Tibetan, 134 Mongolian, 162 Uygur and 223 Han) were recruited for genotyping. Frequencies of CYP1A2 -163C>A, CYP2B6 516G>T and iNOS -954G>C were determined by the polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) methods. RESULTS AND DISCUSSION: The frequency of CYP1A2-163A was higher in Chinese Mongolian (0·698) than in Chinese Tibetan (0·633), Uygur (0·633) and Han populations (0·608, P < 0·05), respectively. The frequency of CYP1A2-163A in the Chinese population (total, 0·636) was intermediate between those reported in Caucasians (0·682, P < 0·05) and Africans (0·549, P < 0·01). The frequency of CYP2B6 516T in Chinese Uygur (0·287) was significantly higher than those in Chinese Tibetan (0·147, P < 0·01) and Mongolian (0·179, P < 0·01), respectively, but was similar to the frequency in Chinese Han (0·226). The frequencies of CYP2B6 516T were in the order of Africans (0·500) > Caucasians (0·286) > Chinese (0·200). The variant iNOS-954C was rare in Chinese Tibetan (0·005), Mongolian (0·004), Uygur (0·000) and Han (0·007), respectively, but showed higher frequencies in African ethnic groups. The frequencies of iNOS-954C were in the order of Africans (0·098) > Chinese (0·004) > Caucasians (0·000). WHAT IS NEW AND CONCLUSION: This is the first report of the distribution frequencies of functional CYP1A2, CYP2B6 and iNOS genes among mainland Chinese Tibetan, Mongolian, Uygur and Han populations. These results should help inform studies of interethnic differences in disease susceptibility or drug responses.

Differences in genotype and allele frequency distributions of polymorphic drug metabolizing enzymes CYP2C19 and CYP2D6 in mainland Chinese Mongolian, Hui and Han populations.

WHAT IS KNOWN AND OBJECTIVE: Cytochrome P450 2C19 (CYP2C19) and CYP2D6 are important xenobiotic metabolic enzymes and both show considerable genetic variability between Orientals and Caucasians. There are known marked heterogeneity in susceptibility to various cancers and hypertension among Chinese Mongolian, Hui and Han ethnic groups, but the molecular mechanisms are unknown. Our objective was to investigate the patterns of distribution of CYP2C19 and CYP2D6 polymorphisms among healthy Chinese subjects to determine whether any observed inter-ethnic variability might be worth further investigation as possible contributors to the known differences in disease prevalence. METHODS: Blood samples were collected from 454 unrelated Chinese healthy subjects (214 Han, 111 Hui, 129 Mongolian) for genotyping analysis. The single nucleotide polymorphisms (SNPs) CYP2C19*2 (681G>A in exon 5), CYP2C19*3 (636G>A in exon 4) and CYP2D6*10 (188C>T in exon 1) were determined by the polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method. RESULTS AND DISCUSSION: Significantly higher frequencies of the CYP2C19 poor metabolic genotypes were observed in Chinese Han (18·7%), Chinese Hui (25·0%) and Chinese Mongolian (10·9%) subjects than has been reported for Caucasians (1·7-3·0%, P < 0·01). The prevalent defective allele CYP2C19*2 occurred more frequently in both Chinese Hui (32·4%) and Han (29·7%) than in Chinese Mongolian (18·2%, P < 0·01) subjects. The CYP2C19*2 and CYP2C19*3 defective alleles were significantly more frequent in Chinese Han and Chinese Hui ethnic groups than have been reported for Caucasians (11·1-16·3% and 0-0·2%, P < 0·01). CYP2D6*1/*10 heterozygotes and CYP2D6*10/*10 homozygotes were observed more frequently in Chinese Han (43·1% and 27·2%), Hui (40·6% and 30·7%) and Mongolian subjects (31·3% and 9·6%, both P < 0·01) than have been reported for Caucasians (5·5% and 0·3%, P < 0·01). In Chinese Mongolians, the CYP2D6*10 allele occurred at a frequency (25·2%, P < 0·01) intermediate between those reported for Caucasians and the other two Chinese ethnic populations. WHAT IS NEW AND CONCLUSIONS: This is first report of interethnic differences in frequencies of functional CYP2C19 and CYP2D6 genes among Chinese Mongolian, Hui and Han populations. These differences may be important in explaining reported inter-ethnic differences in disease prevalence and response to drugs.

Chemotherapy plus Erlotinib versus Chemotherapy Alone for Treating Advanced Non-Small Cell Lung Cancer: A Meta-Analysis.

BACKGROUND: Whether a combination of chemotherapy and erlotinib is beneficial for advanced non-small cell lung cancer (NSCLC) remains controversial. This study aimed to summarize the currently available evidence and compare the efficacy and safety of chemotherapy plus erlotinib versus chemotherapy alone for treating advanced NSCLC. METHODS: EMBASE, PubMed, and the Cochrane Central Register of Controlled Trials were searched for relevant studies. Our protocol was registered in PROSPERO (CRD42014015015). RESULTS: Nine randomized controlled trials with a total of 3599 patients were included. Compared to chemotherapy alone, chemotherapy plus erlotinib was superior in PFS (HR = 0.76 [95% CI 0.62, 0.92], P = 0.006), and no statistically significant difference was observed in OS (HR = 0.94 [95% CI 0.86, 1.03], P = 0.16). Intercalated erlotinib plus chemotherapy demonstrated improvements in PFS (HR = 0.67 [95% CI 0.50, 0.91], P = 0.009) and OS (HR = 0.82 [95% CI 0.69, 0.98], P = 0.03). Continuous erlotinib plus chemotherapy treatment failed to demonstrate improvements in PFS (HR = 0.91 [95% CI 0.80, 1.04], P = 0.16) and OS (HR = 0.98 [95% CI 0.89, 1.09], P = 0.75). The association of chemotherapy plus erlotinib with improvement in PFS was significant in never smoking patients (HR = 0.46 [95% CI 0.37, 0.56], P<0.00001) but not in smoking patients (HR = 0.70 [95% CI 0.49, 1.00], P = 0.05). Among patients with EGFR mutant tumors, chemotherapy plus erlotinib demonstrated significant improvements in PFS (HR = 0.31 [95% CI 0.17, 0.58], P = 0.0002) and OS (HR = 0.52 [95% CI 0.30, 0.88], P = 0.01). Among patients with EGFR wild-type tumors, no statistically significant difference was observed with respect to PFS (HR = 0.87 [95% CI 0.70, 1.08], P = 0.21) and OS (HR = 0.78 [95% CI 0.59, 1.01], P = 0.06). CONCLUSION: Combination of chemotherapy and erlotinib is a viable treatment option for patients with NSCLC, especially for patients who never smoked and patients with EGFR mutation-positive disease. In addition, intercalated administration is an effective combinatorial strategy.

Diazepam metabolism in native Chinese poor and extensive hydroxylators of S-mephenytoin: interethnic differences in comparison with white subjects.

A single oral 5 mg dose of diazepam was given to 16 healthy native Chinese Han volunteers. Eight volunteers were extensive metabolizers of S-mephenytoin, and eight were poor metabolizers of S-mephenytoin. Plasma levels of diazepam and its demethyl metabolite were determined by HPLC in blood samples drawn during 4 weeks. There was no difference in diazepam disposition between the two phenotypes. However, the plasma half-life of demethyldiazepam was longer in poor metabolizers than in extensive metabolizers of mephenytoin (mean +/- SD: 161 +/- 37 and 116 +/- 29 hours, respectively; p less than 0.02). The plasma concentrations of demethyldiazepam at 7, 14, and 21 days after intake of diazepam were significantly higher in poor metabolizers than in extensive metabolizers. We compared the pharmacokinetic parameters of diazepam in Chinese subjects with our previously reported data from white subjects. The mean plasma half-life values of diazepam in Chinese extensive metabolizers (85.1 hours) and poor metabolizers (88.3 hours) were very similar to those in white subjects who were poor metabolizers (88.3 hours), and more than twice those in white subjects who were extensive metabolizers (40.8 hours). In parallel, the mean clearance of diazepam in Chinese subjects (independent of phenotype) was similar to that in white subjects who were poor metabolizers, but half that in white subjects who were extensive metabolizers. Chinese subjects had a slightly larger volume of distribution of diazepam than white subjects.(ABSTRACT TRUNCATED AT 250 WORDS)

Pronounced differences between native Chinese and Swedish populations in the polymorphic hydroxylations of debrisoquin and S-mephenytoin.

The frequency of poor metabolizers of debrisoquin was low and similar in four different native Chinese nationalities. In a total sample of 695 Chinese subjects, only seven (1.01%) had a urinary ratio between debrisoquin and 4-hydroxydebrisoquin greater than 12.6, which is the antimode between poor metabolizers and extensive metabolizers in white populations. This is significantly lower than the 6.82% found in 1011 white Swedish healthy subjects (p less than 0.0001). Admixture analysis indicated the occurrence of two distributions within extensive metabolizers among both Chinese and white subjects. The mean of the distribution of metabolic ratios among Chinese extensive metabolizers was shifted toward higher values compared with Swedish extensive metabolizers (p less than 0.01). The frequency of poor metabolizers of S-mephenytoin was higher in 137 Chinese (14.6%) than in 488 Swedish (3.3%) subjects (p less than 0.0001). Our findings imply that drugs metabolized by these two polymorphic hydroxylases should be prescribed in different dosages to Chinese and white subjects.

[In vitro studies on omeprazole metabolism in rat liver microsomes].

The metabolism of omeprazole to its two major metabolites, hydroxyomeprazole (OH-OPZ) and omeprazole sulfone (OPZ-SFN) was studied in rat liver microsomes by a reversed phase HPLC assay. The formation of metabolites of OPZ depended on incubation time, substrate concentration, microsomal protein concentration, and was found to be optimal at pH 7.4. The Vmax and Km of OPZ hydroxylation in the rat liver microsomal preparation were 2033 nmol/(min.mg protein) and 46.8 mumol.L-1 respectively. The maximum rate of formation of OPZ-SFN (Vmax) was 187.9 nmol/(min.mg protein), with a Km value of 120.7 mumol.L-1 in rat liver microsome. Moreover, the effects of 7 drugs on OPZ metabolism were tested. The results showed that mephenytoin, benzodiazepines (DZ, NDZ, TMZ, FNZ, NZ) and papaverine caused inhibition of OPZ metabolism, among them papaverine was the only fairly strong inhibitor.

[Enzyme kinetic studies on the in vitro metabolism of omeprazole by Chinese human liver microsomes].

In the present study, a specific and reliable method was developed for the determination of omeprazole and its two major metabolites, hydroxyomeprazole (OH-OPZ) and omeprazole sulfone (OPZ-SFN) in Chinese adult human liver microsome by reversed phase HPLC assay. Formation of these metabolites is linear for at least 60 min and between 0.25 and 1 mg.ml-1 of microsomal protein. The enzyme kinetic analysis of the reaction revealed that the hydroxylation Vmax and K(m) obtained with the human liver microsomal preparation were 42.9 nmol.min-1.(mg protein)-1 and 6.49 mumol.L-1, respectively. The maximum formation rates of OPZ-SFN (Vmax) was 6.63 nmol.min-1.(mg protein)-1, with a K(m) value of 11.8 mumol.L-1. A number of compounds were tested for their ability to inhibit OPZ metabolism. Our results showed that mephenytoin, diazepam and nordiazepam are competitive inhibitors and papaverine is an uncompetitive inhibitor of OPZ hydroxylation. These studies suggest that the same isozyme metabolising MP, DZ and NDZ (may be P450 2C or P450 3A) may be involved in the hydroxylation of OPZ. Moreover, the compounds tested also have some effects on the formation of OPZ-SFN in vitro with Chinese human liver microsomes.

[HPLC determination of tetramethylpyrazine in human serum and its pharmacokinetic parameters].

An HPLC method for the determination of tetramethylpyrazine in serum and application of this method to tetramethylpyrazine deposition studies in human body were described. Tetramethylpyrazine was extracted from alkalinized serum with dichloromethane using methaqualone as internal standard. A RP mu-Bondapak-C18 (10 microns) column fitted with a variable-wavelength UV spectrophotometer operated at 280 nm was used. The mobile phase was methanol-water (58:42). The detection limit of the method was 0.0174 microgram/ml serum. Assay linearity was shown over the range of 0.0291-5.816 micrograms/ml serum with a regression coefficient of 0.9999. The extraction recovery was 99.84% and no interference was found from endogenous compounds, metabolites of parent drug or other commonly used drugs. For the serum concentration following oral administration of tetramethylpyrazine capsules to healthy volunteers (n = 6), the best fit was found to be with a two compartment open model. After administration of 174.5 mg dose, the pharmacokinetic parameters were as follows: Tp = 0.5102 h, Cmax = 3.114 micrograms/ml, AUC = 5.893 mg/L.h, T1/2 (Ka) = 0.1508 h, T1/2 (alpha) = 0.4855 h, T1/2 (beta) = 2.894 h, C1 = 15.7 L.h-1, Vc = 17.70 L, V = 66.77 L. The results imply that tetramethylpyrazine is absorbed rapidly, distributed widely in the body, and also eliminated at a fairly rapid rate.

[Methodological study on the determination of naftopidil concentration in biological samples by HPLC].

A method was developed for the determination of the novel antihypertensive drug naftopidil in biological samples by HPLC. Metoprolol was used as the internal standard. The analytical column was a stainless steel column filled with 10 micron C18 packing. The mobile phase was a mixture of methanol-acetonitrile-water-0.2 mol.L-1 HAc-0.2 mol.L-1 NaAc (50:45:5:0.9:0.1). Detection was performed at UV 232 nm. Biological samples can be well purified after two extractions with ether. The lowest detection limit was 5 ng.ml-1. The precision and accuracy within-day and day-to-day ranged from 3.17 to 10.88%. The mean recoveries were 79.35% to 95.72%. The results showed that this method was simple, sensitive and good enough to be used in pharmacokinetic study of naftopidil.

[Methodological study on the determination of oxiracetam concentration in serum and urine by HPLC].

A method was developed for the determination of oxiracetam concentration in serum and urine by HPLC. Acyclovir was used as an internal standard. The analytical column was a stainless-steel column (30 mm x 4.6 mm ID) filled with 10 microns Bondapak NH2. A mixture of acetonitrile and water (80: 20) as mobile phase was used at a flow rate of 1 ml.min-1. Detection was performed at 210 nm. The retention times were 6.3 min for oxiracetam and 8.1 min for the internal standard. The lower detection limits were 1 microgram.ml-1 for serum and 20 micrograms.ml-1 for urine. The precision and accuracy within-day and day-to-day for both serum and urine samples ranged from 5.0 to 10.7%. The mean recoveries were 99.7 +/- 5.9% and 99.0 +/- 5.6% for human serum and urine, respectively. The results showed that the method is simple, rapid, sensitive, reliable and good enough to be used in studying the clinical pharmacokinetics of oxiracetam.

[A chiral capillary gas chromatographic method for direct determination of enantiomers of mephenytoin in human urine].

A gas chromatographic method equipped with nitrogen-phosphorus detector was developed for the determination of the S- and R-enantiomers of the anticonvulsant, mephenytoin (MP) in human urine. Dichloromethane (4 ml) was added to 1 ml urine, the mixture was shaken and centrifuged. The organic phase was transferred to another tube and blown to dryness under nitrogen on water bath (37 degrees C). The residue was dissolved in 10 microliters ethylacetate and 1-2 microliters was injected into the GC. Our results showed that direct enantiomeric separation of mephenytoin was obtained by using a chiral capillary column, the retention times for S- and R-mephenytoin were 25.5 and 26.2 min respectively, with a detection limit less than 50 ng/ml of mephenytoin. Similar linear and reproducible standard curves were obtained over the concentration range of 53.2 to 2128.0 ng/ml (for S-MP, r = 0.9914 +/- 0.0070, n = 6; and for R-MP, r = 0.9939 +/- 0.0070, n = 6), and the mean recoveries of S- and R-MP were 95.4% and 95.8% respectively. The within-day relative standard deviations were less than 8.8% for both S- and R-MP, and that of between-days were less than 14.3%. There was a good reproducibility of the urine S/R mephenytoin determined in China and in Sweden by using similar method in 107 Chinese volunteers after a single oral dose of 100 mg racemic mephenytoin (r = 0.9091, P < 0.001).

[Study on the frequency distribution of debrisoquine hydroxylation deficiency in Chinese healthy Zang and Wei volunteers].

The metabolic ratio of debrisoquine hydroxylation (MR) determined as the ratio of debrisoquine over 4-OH-debrisoquine in 8 h urine after a single oral dose of DB (10 mg) in unrelated Chinese Zang and Wei volunteers was studied by using gas chromatography. The MR of 132 healthy Chinese Zang subjects ranged from 0.20 to 34.32, and that of 158 healthy Chinese Wei subjects ranged from 0.13 to 29.73. Two phenotypes were apparent in the frequency distribution histograms with an antimode at log MR = 1.10 (MR = 12.6). Two subjects were therefore identified as poor metabolizers (PMs) of DB in 132 Zang volunteers (the frequency of PMs was found to be 1.52%), and only one in 158 Wei volunteers (0.63%). Neither sex nor smoking habit affected the DB hydroxylation (P greater than 0.2). The recoveries of DB and 4-OH-DB in Zang volunteers were 19.83 +/- 10.99 and 11.57 +/- 7.04%, and in Wei volunteers they were 22.74 +/- 14.41 and 17.77 +/- 10.82%, respectively.

[Studies on the metabolism of antiepilepserine in isolated perfused rat liver].

The metabolism of antiepilepserine, 3,4-methylene dioxycinnamyl piperidine, was studied in isolated perfused rat liver. Two metabolites were separated and purified by means of HPLC. They were identified as 3,4-methylene dioxycinnamyl hydroxypiperidine and 4-hydroxy-3-methoxycinnamyl piperidine by UV and MS. The latter was further confirmed by chemical synthesis. The pharmacokinetics of antiepilepserine in isolated perfused liver was also studied. Parameters obtained include a first order elimination constant, k = 0.0157 +/- 0.0043 min-1, and a half-life, t 1/2 = 46.7 +/- 11.9 min, (n = 6). The amount of antiepilepserine lost during the perfusion could not be compensated by the increased amount of metabolites. Antiepilepserine was found to be tightly bound and stored in the liver. This might be one of the explanations of the first pass effect of antiepilepserine after oral administration.

[The molecular mechanism of polymorphism of S-mephenytoin hydroxylative metabolism].

The hydroxylation of S-mephenytoin exhibits a genetic polymorphism in humans and there are a large interethnic differences in the frequency of the poor metabolizer phenotype. A S-mephenytoin P450-hydroxylase termed P450 UK was purified and identified to be CYP 2C19 by amino-terminal amino acid analyses. The levels of P450 2C19 and the ability of the human liver samples to 4'-hydroxylate S-mephenytoin were found to be strongly correlative. Recent report showed that the principle defect in S-mephenytoin poor metabolizers is a single base pair (G-->A) mutation in exon 5 of CYP 2C19 resulting in an aberrantly spliced mRNA and a non-functional P450 2C19 protein in liver of S-mephenytoin PM. Further investigations demonstrate that this major defect is responsible for about 75% of poor metabolizer phenotype in both caucasians and orientals who are homozygous for S-mephenytoin hydroxylation defect. This genetic defect (CYP 2C19) also affects metabolism of several other widely clinical used drugs.

[Study of the molecular mechanism of human cytochrome P450 and genetic polymorphism of drug oxidative metabolizing enzyme].

Polymorphism has been detected in a variety of drug-metabolizing enzymes at both phenotypic and genotypic level. Human cytochrome P450 enzymes have been studied extensively in recent years, and the majority of mutations which give rise to a defective phenotype have now been identified(for example CYP2D6, CYP2C19, etc.). There are some groups of enzymes showing definite polymorphism at the phenotypic level but the exact genetic mechanisms are not yet clear(CYP1A1, CYP1A2, etc.). There are still other groups of enzymes, showing some indication of polymorphism at either the phenotypic or genotypic level yet have not been unambiguously demonstrated (for example CYP2A6, CYP2C9, CYP2E1, CYP3A4, etc.). The molecular mechanism of all these polymorphisms and possible polymorphisms is discussed, with particular reference to the effects of this variation on drug metabolism and on the susceptibility to chemically-induced diseases.

Role of leptin receptor gene polymorphisms in susceptibility to the development of essential hypertension: a case-control association study in a Northern Han Chinese population.

In order to explore the potential association between the leptin receptor (LEPR) gene polymorphisms and essential hypertension (EH) risk in the Northern Han Chinese population, we recruited 823 hypertensive subjects and 491 healthy control subjects from the Northern Han Chinese. Genotyping was performed to identify the Lys109Arg, Gln223Arg and Lys656Asn polymorphisms of the LEPR gene. Significant associations were found in a dominant genetic model ([GG+AG] vs. AA), P=0.007, odds ratio (OR)=3.697, 95% confidence interval (CI) 1.442-9.482), and in homozygote comparison (GG vs. AA, P=0.005, OR=3.890, 95% CI 1.501-10.077) for the Gln223Arg polymorphism. No significant association could be found between Lys109Arg or Lys656Asn polymorphism and EH risk. Linkage disequilibrium was detected between the Lys109Arg and Gln223Arg polymorphisms, and haplotype analyses identified that the G-A haplotype was a protective haplotype for EH. Our studies demonstrated that the LEPR Gln223Arg polymorphism had an important role in a patient's susceptibility to EH in the Northern Han Chinese population.