Analysis of epidermal growth factor receptor gene mutations in patients with non-small cell lung cancer in Shiyan, Hubei / 肿瘤研究与临床

Objective:To investigate the epidermal growth factor receptor (EGFR) gene mutation status in patients with non-small cell lung cancer (NSCLC) in Shiyan, Hubei and its relationship with the clinicopathological characteristics of patients.Methods:The data of 173 NSCLC patients who were admitted to Affiliated Dongfeng Hospital of Hubei University of Medicine from November 2017 to January 2020 were retrospectively analyzed. EGFR gene mutations in NSCLC tissues were detected by amplification refractory mutation system (ARMS)-TaqMan probe method, and clinicopathological data of patients were also collected to analyze the relationship between EGFR gene mutation status and clinicopathological characteristics of patients.Results:EGFR gene mutations were found in 76 of 173 patients, and the total mutation rate was 44.5%. The mutation rate of exon 18 was 6.6% (5/76), all of which were G719X mutation; the mutation rate of exon 19 was 46.1% (35/76), all of which were Del mutation; the mutation rate of exon 20 was 1.3% (1/76), which were EGFR gene 20ins mutation; the mutation rate of exon 21 was 44.7% (34/76), of which 33 cases were EGFR gene L858R mutation, and 1 case was EGFR gene L861Q mutation; 1 case was a double mutation of exon 19 Del combined with exon 20 T790M. The differences in EGFR mutation rates were statistically significant among patients with different gender, smoking history and pathological staging (all P < 0.05), and EGFR mutation rate in female patients was higher than that in male patients [59.7% (46/77) vs. 31.3% (30/96), χ2 = 14.08, P < 0.001], it was lower in patients with smoking history than that in patients without smoking history [27.1% (13/48) vs. 52.9% (63/119), χ2 = 9.22, P < 0.001], and it was higher in patients with adenocarcinoma than that in patients with non-adenocarcinoma [50.7% (75/148) vs. 4.0% (1/25), χ2 = 18.92, P < 0.001]. EGFR mutation status had no relevance with patients' age. Conclusions:The main types of EGFR mutations are exon 19 Del and exon 21 L858R in NSCLC patients in Shiyan, Hubei. EGFR mutations are more likely to occur in adenocarcinoma, female and no-smoking patients.

Relationship between hepatitis B virus genotyping and primary hepatocellular carcinoma of 182 patients / 公共卫生与预防医学

Objective To analyze the relationship between hepatitis B virus genotyping and primary liver cancer (PHC) in Wuhan, and to provide a theoretical basis for the early prevention and diagnosis of PHC. Methods Patients with chronic hepatitis B (CHB) from Wuhan Sub-heart General Hospital for treatment from February 2020 to February 2021 were selected and divided into PHC group (182 cases) and control group (189 cases) according to whether they were complicated with primary liver cancer. 5ml of fasting elbow venous blood was taken from all subjects at admission. HBV genotyping was determined by real-time fluorescence quantitative PCR. The DNA of CHB virus was determined by fluorescence probe hybridization and PCR amplification, and genotyping and drug-resistant mutation points were detected according to the product sequencing analysis. Spearman linear correlation analysis was used to analyze the correlation between genotyping and mutation rate of PHC patients. Results The proportion of C genotype in PHC group was significantly higher than that in non-PHC group (P0.05). The proportion of HEPATITIS B virus mutation in PHC group (114/182) was significantly higher than that in control group (84/189) (χ2=12.331, P0.05). The proportion of HBV C mutant in PHC group was significantly higher than that in control group (P1=0.349, r2=0.305, P<0.05). Conclusion The HBV genotype of PHC patients is mainly TYPE C, and has a high mutation rate of C genotype. It can be used for diagnosis of PHC by detecting the genotyping of CHB and mutation rate of C genotype in clinic.

Técnicas de substituição de DNA mitocondrial para reduzir a transmissão da Síndrome de Leigh / Mitochondrial DNA replacement techniques to reduce Leigh syndrome transmission

A Síndrome de Leigh (SL) é uma doença neuro-metabólica congênita, que faz parte do grupo das encefalopatias fatais, com progressão e morte dentro de 2 anos, em média. A SL é causada por mutações no DNA que causam alterações na geração de ATP celular pelas mitocôndrias. As mitocôndrias contêm seu próprio DNA (mtDNA) e, ao contrário do DNA nuclear, o mtDNA é herdado somente da mãe. Mulheres portadores de mutações causadoras da SL podem vivenciar experiências muito tristes ao tentarem realizar o sonho da maternidade. As técnicas de substituição de mtDNA mutado com mtDNA saudável de doadora, oferecem a essas mulheres a possibilidade de terem uma criança geneticamente relacionada sem a SL. O desenvolvimento e a aplicação clínica de terapias de substituição de mtDNA já são uma realidade, tendo o primeiro bebê gerado a partir da técnica nascido em 2016. Mas será que essas técnicas são seguras? Neste trabalho, revisamos a SL e algumas técnicas de substituição de mtDNA já aplicadas em humanos, que envolvem a transferência de pronúcleos de zigotos ou de fuso acromático de oócitos. Concluímos que, apesar dos resultados promissores, ainda é cedo para assegurar a aplicabilidade clínica de técnicas de substituição de mtDNA em seres humanos. (AU)

Leigh syndrome (SL) is a congenital neurometabolic disease included in the group of fatal encephalopathies, with progression and death within 2 years on average. SL is caused by mutations in the DNA that cause changes in the generation of cellular ATP by mitochondria. Mitochondria contain their own DNA (mtDNA) and, unlike nuclear DNA, mtDNA is inherited only from the mother. Women with SL mutations may experience mournful situations when attempting to fulfill the dream of motherhood. Techniques for replacing mutant mtDNA with healthy donor mtDNA provide these women with the possibility of having a genetically related child without SL. The development and clinical application of mtDNA replacement therapies is a reality, and the first baby generated using the technique was born in 2016. However, are these techniques safe? In this article, we review SL and some mtDNA replacement techniques that have been used in humans, which involve zygote pronuclear transfer or oocyte spindle transfer. We conclude that, despite the promising results, it is too early to ensure that mtDNA replacement techniques are clinically applicable to humans. (AU)

Development of a 30 Y-STR Loci with middle or low mutations multiplex PCR system / 中国法医学杂志

Objective To deveplope construct and validate a novel multiplex PCR system comprised of 30 Y-STR markers only with low and moderate mutation rates. Methods 30 Y-STRs characterized by low/moderate mutation rate and middle/high polymorphic was amplified simultaneously in a multiplex PCR system using the six color labeling fluorescence. PCR product was analyzed in a ABI 3500XL Genetic Analyzer. The accuracy, specifity, sensitivity and stability of the system and its validation on the mixtures were evaluated. Results The validation studies demonstrated that the system is a stable, accurate, and sensitive multiplex PCR system. The sensitivity was 0.0625ng DNA. Y-STR could be detection in a male/female DNA mixture ratio of 1:4. Conclusion The primary study demonstrates that this multiplex PCR system is effective and reliable for forensic routine DNA analysis. It will be very helpful for constructing Chinese forensic Y-STR database and population genetic research.

Correlation analysis of reject reaction and corneal endothelial cells after penetrating keratoplasty from donation after cardiac death / 国际眼科杂志(Guoji Yanke Zazhi)

AIM:To study the relativity between reject reaction from donation after cardiac death (DCD) and corneal endothelial cell source of corneal graft after penetrating keratoplasty.METHODS:Totally 28 cases of corneal graft rejection after penetrating keratoplasty with cardiac death donor cornea were analyzed using corneal endothelial microscope at less than 1mo,2-3mo,4-6mo,7-12mo after operation.RESULTS:Coefficient variation of corneal endothelial cell of the 28 cases at less than 1 mo,2-3mo,4-6mo and 7-12mo were 38.23％,49.56％,57.18％,65.04％.Corneal endothelial cell density were 2071.15 ± 311.47,1771.33 ± 348.18,1626.59±353.92,1553.14±307.31.The coefficient variation of corneal endothelial cells was positively correlated with rejection (r =0.95,P ＜ 0.05).The postoperative corneal endothelial cell density was negatively correlated with rejection (r=-0.93,P＜0.05).CONCLUSION:The corneal endothelial cell coefficient variation increased gradually and the corneal endothelial cell density decreased gradually after DCD corneal allograft rejection.Corneal endothelial cell coefficient variation and corneal endothelial cell density can be used as indicators of early detection of postoperative rejection.

Assessment on Application of 24 Y-STR Loci in Forensic Science / 法医学杂志

OBJECTIVES@#To select a Y-STR marker system with strong haplotype identification ability, appropriate mutation rate and high compatibility and to assess its forensic application.@*METHODS@#The 24 Y-STR loci were tested by self-built fluorescent multiplex system, and the forensic assessment was conducted by 139 pairs of father-son samples collected in Jinan, Shandong province.@*RESULTS@#Totally 176 alleles were identified among the 24 Y-STR loci in the sample of 139 unrelated individuals labeled with father, and the gene diversity （GD） distributed between 0.083 7 （DYS645）-0.966 9 （DYS385a/b）. According to the 24 Y-STR loci, 139 different haplotypes were detected from 139 unrelated male individuals labeled with father in Han population of Shandong province and with no shared haplotype observed. The overall haplotype diversity （HD） was 1 and the discrimination capacity （DC） was 1. A total of 5 one-step mutations events were observed among the 24 Y-STR loci in 139 pairs of father-son. The average mutation rate was 0.001 5 [95% CI （0.000 5, 0.003 5）].@*CONCLUSIONS@#The system of 24 Y-STR loci shows a strong individual recognition ability and low mutation rate in the population in Jinan, Shandong province, and it has good application value in forensic science.

Statistical analysis of EGFR mutation and EML4-ALK gene fusion in non-small cell lung cancer patients of Zhongshan City / 国际检验医学杂志

Objective To detect the mutation of epidermal growth factor receptor(EGFR) gene,fusion of echinoderms microtubule associated protein sample-4 and gradual change of lymphoma kinase(EML4-ALK) gene,as well as describe their relationship with the clinicopathological features in patients with non-small cell lung cancer(NSCLC) from Zhongshan city of Guangdong province.Methods Mutations of EGFR gene and EML4-ALK fusion gene in 753 NSCLC patients from Zhongshan People's hospital were detected by ARMS real-time PCR.To study the relationship between the mutation and clinical features and explore the significance of EGFR gene mutation and EML4-ALK fusion in NSCLC.Results The EGFR mutation rate of 753 NSCLC patients is 43.16％(325/753),with highest mutation rate in 19 and 21 exons,43.08％(140/325) and 47.38％ (154/325) respectively,and the main mutation in 21 exon is L858R mutation.EGFR mutation is more common in female/non-smoking patients,or patients with adenocarcinoma/adenosquqmous carcinoma/adenocarcinoma metastasis(P＜0.05),but not relates with the age of patients(P＞0.05).The EML4-ALK fusion gene of 110 patients whose EGFR mutation were checked were simultaneously detected,showing a 9.09 ％ (10/110) mutation rate,and the mutation rate in type 1(80％) is significantly higher than type 2(10％) and 3(10％).Patients with EML4-ALK gene mutation tend to be younger(P＜0.05),but the EML4-ALK gene mutation rates show no significant differences in groups classified by gender,smoking history or pathological classification(P＞0.05).EGFR gene mutation and EML4-ALK fusion were detected in one patient simultaneously.Conclusion The EGFR mutation rate of patients with NSCLC in Zhongshan city is consistent with results reported in domestic and foreign literatures.Detections of EGFR gene mutation and EML4-ALK fusion are necessary test items,providing important evidence in molecular targeting therapy in NSCLS.

Statistical analysis of EGFR mutation and EML4-ALK gene fusion in non-small cell lung cancer patients of Zhongshan City / 国际检验医学杂志

Objective To detect the mutation of epidermal growth factor receptor(EGFR) gene,fusion of echinoderms microtubule associated protein sample-4 and gradual change of lymphoma kinase(EML4-ALK) gene,as well as describe their relationship with the clinicopathological features in patients with non-small cell lung cancer(NSCLC) from Zhongshan city of Guangdong province.Methods Mutations of EGFR gene and EML4-ALK fusion gene in 753 NSCLC patients from Zhongshan People's hospital were detected by ARMS real-time PCR.To study the relationship between the mutation and clinical features and explore the significance of EGFR gene mutation and EML4-ALK fusion in NSCLC.Results The EGFR mutation rate of 753 NSCLC patients is 43.16％(325/753),with highest mutation rate in 19 and 21 exons,43.08％(140/325) and 47.38％ (154/325) respectively,and the main mutation in 21 exon is L858R mutation.EGFR mutation is more common in female/non-smoking patients,or patients with adenocarcinoma/adenosquqmous carcinoma/adenocarcinoma metastasis(P＜0.05),but not relates with the age of patients(P＞0.05).The EML4-ALK fusion gene of 110 patients whose EGFR mutation were checked were simultaneously detected,showing a 9.09 ％ (10/110) mutation rate,and the mutation rate in type 1(80％) is significantly higher than type 2(10％) and 3(10％).Patients with EML4-ALK gene mutation tend to be younger(P＜0.05),but the EML4-ALK gene mutation rates show no significant differences in groups classified by gender,smoking history or pathological classification(P＞0.05).EGFR gene mutation and EML4-ALK fusion were detected in one patient simultaneously.Conclusion The EGFR mutation rate of patients with NSCLC in Zhongshan city is consistent with results reported in domestic and foreign literatures.Detections of EGFR gene mutation and EML4-ALK fusion are necessary test items,providing important evidence in molecular targeting therapy in NSCLS.

An analysis of mutations at 41 short tandem repeat loci in Han Chinese population / 中国法医学杂志

Objective The aim of this study was to investigate mutations of 41 STR loci. Methods 4546 bloodstain samples were typed from 1932 father–mother–child trios by using AGCU_21+1, AGCU_EX22 and GlobalFiler_ExpressTM amplification Kit. Calculate the mutation rates of STR loci. Results 154 mutations were identified at 32 of the 41 loci. The average mutation rate was 1.0×10-3per locus(95%CI: 0.8~1.1×10-3), and the mutations of SE33 was highest. 152(98.7%) mutation events were one-step mutation, 2(1.3%) events were two-steps. The mutation events occurred in 150 father–mother–child triplets. The mutations in 146(97.3%) triplets occurred at single locus, 8 mutations were observed at two loci in 4(2.7%) triplets simultaneously. 104 paternal and 22 maternal mutations could be determined under 79212 paternal and maternal allelictransfers. The ratio of paternal versus maternal mutations was 4.7:1, and 28 unassigned mutations were observed. Conclusion STR mutation are common in paternity testing, and we should pay more attention to it.

Analysis of mutation of 20 autosomal short tandem repeat loci in Henan Han population / 中国法医学杂志

Objective To investigate the mutation phenomenon of 20 autosomal STR loci in Henan Han population. Methods A total of 3011 parentage confirmed cases were collected to screen mutation events, ascertain the source of mutation, calculate mutation rate, analyze mutation rules and compare with the mutation condition of populations in different regions. Results 76 mutation events were observed in 19 STR loci, the average and accumulative mutation rate reached to 0.08% and 1.6629%, respectively. The ration of paternal versus maternal mutation was 8:1. Mutation rates of Penta E and D12S391 loci in Henan Han population were lower than the Han population of northern China(P<0.05); the mutation rate of CSF1PO locus were lower than Guangdong population and Yunan Han population(P<0.05); the mutation rates of D6S1043 and D12S391 loci were lower than Guangdong population(P<0.05). Conclusion STR mutation events were common in paternity testing. Region differences among mutation rate were significant.

Looking Back at Our 5-Year Experience of Paternity Testing: A Summary / 대한법의학회지

We have been testing familial relationships based on short tandem repeats (STRs) in families who requested it either voluntarily or by order of the court. Here, we present a summary of our 5-year experience of autosomal STR-based paternity tests. A total of 1,431 individuals from 588 cases were tested, including 878 pairs of either of the parent, and a child. Among these 588 cases, genetic information about the other parent was available only for 135 cases. Five hundred eighteen pairs were concluded to be parent-child relations, for which the median paternity index (PI) was 72,826, and the median decimal logarithm was 4.860. Autosomal mutation was observed in nine pairs (1.74%), and the pairs harbored only one mismatched locus among the 15 standard loci (D8S1179, D21S11, D7S820, CSF1PO, D3S1358, TH01, D13S317, D16S539, D2S1338, D19S433, vWA, TPOX, D18S51, D5S818, and FGA). The number of mismatched loci did not increase even after additional loci were included in the study. The observed mutation rates were D13S317 (0.193%), D18S51 (0.193%), D19S433 (0.193%), FGA (0.193%), vWA (0.386%), Penta D (0.387%), and Penta E (0.193%). There were 14 pairs with two mismatched loci, which we excluded through additional tests on either autosomal or X chromosomal STRs, and mitochondrial sequencing. Although PI is useful for determining parent-child relation, it provides indirect information; it is an interpretation of the test results that is based on probability. Additional genotyping on sex chromosome and mitochondrial DNA, or participation of other family members might be beneficial for a reliable conclusion.

Web-based Y-STR Database for Haplotype Frequency Estimation and Kinship Index Calculation / 대한법의학회지

We developed a web-based Y chromosomal short tandem repeat (Y-STR) database (ySTRmanager, http://ystrmanager.yonsei.ac.kr) to facilitate calculation of Y-STR haplotype frequency estimates for random matches and kinship indices for various relationship levels. The ySTRmanager database provides 3 functions: (i) Y-STR haplotype search, (ii) kinship index calculation, and (iii) user database configuration. The Y-STR haplotype search function allows researchers to retrieve Y-STR haplotypes that meet queried Y-STR allele, Y-haplogroup affiliation, and/or sample information from a selected population in the open database, which consists of 12-17 Y-STR loci. The number of matches in a selected population, haplotype frequency estimator, and detailed results for matched and neighbor haplotypes are displayed as a set of search results. The kinship index calculation function provides kinship indices of 2 input Y-STR haplotypes for the relationship represented by the number of meioses, with consideration of target population and mutation rate of each Y-STR. In addition, ySTRmanager allows registered users to configure their own database to store and analyze Y-STR haplotype and/or mutation rate data. The stored Y-STR data can be used in the search function and in the analysis to obtain forensic statistical values. The ySTRmanager will be a useful system to analyze and manage Y-STR data in the practice of forensic genetics.

A simple and rapid quantitative method of detection of the common achondroplasia mutation: Analysis in mismatch repair deficient cells.

Achondroplasia is the most common form of dwarfism and has an incidence of approximately 1/7,500. In more than 97% of cases, it is caused by a recurrent point mutation, a G to A substitution at nucleotide position 1138 (G1138A) of the fibroblast growth factor receptor 3 gene. Although this is an autosomal dominant condition, more than 90% of all mutations occur sporadically making this one of the most mutagenic sites in the human genome. The reasons for the high spontaneous G1138A mutation rate are not known. This investigation was performed by developing a simple and rapid semi-quantitative allele specific PCR based assay capable of reliably detecting more than 25 mutant G1138A copies in a pool of 300,000 wild type molecules. Using this assay, the G1138A mutation frequency was measured in cell lines deficient in mismatch repair (LoVo, SW48) and comparing it with controls. No differences were found in the frequency of this point mutation between the mismatch repair deficient and wild type cell lines.

Allele frequency and its forensic application of STR Y27 in Korean males

The allele frequency and mutation rate in a Short Tandem Repeat locus, Y27 were studied in 247 unrelated Korean males using polymerase chain reaction followed by high-resolution polyacrylamide gel electrophoresis, a procedure called the amplification fragment length polymorphism technique. Six alleles were noted ranging from 190 bp to 210 bp. They existed as discrete bands with 4 bp discrepancy. Among which DY3(198 bp), DY4(202 bp) were common with the frequencies of 0.408, 0.356 respectively. Other alleles, DY1(190 bp, frequency 0.020), DY2(194 bp, frequency 0.121), DY5(206 bp, frequency 0.089), DY6(210 bp, frequency 0.004) were relatively uncommon. In a 78 subject father-son study with parenthood confirmed through other genetic studies, no case of mutation was noted. As the allele number was not as large as 6 and two alleles were dominant, the discrimination power in routine individual identification was thought to be low. But in selective cases such as father-son determination or sex determination, this locus could be a valuable genetic marker and we thought these results to be common for the Korean population. These results were also compared with that of other race.

A validation study of four single locus probes (MS1, MS31, MS43 and g3) in a Korean population: further evaluation for paternity testing

Hypervariable tandem repetitive regions in human DNA are proving to be increasingly useful for genetic analysis in humans. We chose four single locus probes (SLP; MS1, MS43, MS8 and g3) for a validation test among Koreans. The specimens were from 216 unrelated individuals and 33 paternity inclusion families. Extracted DNA from EDTA blood was restricted by Hinfl and electrophoresed in 0.7% agarose gel, transferred and hybridized with chemiluminescent probes. Heterozygosity was over 90% by all of the probes. Total numbers of unassignable mutant bands from 33 paternity inclusion cases were 5, and the highest mutation rate was determined in probe MS1(0.045). The probability of having the same DNA band between two unrelated individuals was 5.7 x 10(-10) when four SLPs were used at the same time. The data presented here on allele frequencies and mutation rates provide preliminary data supporting the validity of these probes in paternity analysis and forensic investigators in the Korean population.