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Objective:To screen m6A modification-related genes, and to establish a prognostic model in patients with FLT3 mutated acute myeloid leukemia(AML), especially in older patients and to evaluate the prognostic efficiency of the model.Methods:Gene expression omnibus(GEO)datasets were used to analyze abnormally expressed m6A enzymes and reading proteins in FLT3 mutated AML; Correlation analysis was used to screen m6A modified-related genes in expression profiles.By integrating TCGA and BEAT data, 83 FLT3 mutated AML patients were included, and 32 of them were older than 60 years.Univariate Cox analysis and Lasso regression were conducted to construct the risk model.Kaplan-Meier curve and time-dependent receiver operating characteristic curve(tROC)were used to evaluate the prognostic efficiency of the model; subgroup analysis was conducted in the older patients.The concordance index(C-index)and calibration curve were used to evaluate the discrimination and accuracy of the model.Results:14 m6A modification enzymes or reading proteins were abnormally expressed in patients with FLT3 mutated AML.Correlation analysis filtered out 2 476 m6A related genes in expression profile.In TCGA and BEAT integrated data, univariate Cox analysis identified 132 prognostic genes.Lasso regression selected seven candidate genes to establish the prognostic risk model, including AKAP9, AVEN, DMCA1, DPYD, FAR2, GPHN and SPECC1L.Kaplan-Meier curve showed that high-risk group of the model had significantly shorter overall survival with a hazard ratio( HR)of 5.08(95% CI: 2.54-10.14, P<0.001).The area under the curve(AUC)in tROC for 1-year survival was 0.83; the C-index of risk model was 0.737.In older patients, the hazard ratio( HR)of the risk model for 1-year overall survival was 3.40(95% CI: 1.25-9.24, P=0.017)with an AUC of 0.79. Conclusions:The risk model based on m6A modified-related genes has some predictive value in assessing the prognosis of patients with FLT3 mutated AML, especially indicative to prognosis prediction in the elderly.
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Objective:To identify pathogenic genes in 3 cases of piebaldism, and to explore the genotype-phenotype relationships in piebaldism.Methods:Clinical data were collected from 3 patients with piebaldism and their parents at the Department of Dermatology, Henan Provincial People′s Hospital from January 2019 to December 2021. Peripheral blood samples were obtained from them and 100 unrelated healthy controls, and DNA was extracted. Whole-exome sequencing technology was used to screen genetic variation sites, and then Sanger sequencing was performed for verification. The deleteriousness of genetic variants was evaluated by using pathogenicity analysis software tools.Results:Case 1: a 23-year-old male patient presented with white patches on the forehead, chest, and abdomen for 23 years, and his parents had no similar symptoms; case 2: a 1-year- and 5-month-old male infant presented with white patches on the forehead and abdomen for 1 year, and his parents had no similar symptoms; case 3: a 6-year-old male child presented with white patches on the forehead and limbs for 6 years, and his parents had no similar clinical manifestations. Genetic testing showed that a missense mutation c.2033T>C (p.L678P) in exon 14 of the KIT gene, a splice site mutation c.2485-1G>C in exon 18 of the KIT gene, and a heterozygous missense mutation c.2346C>G (p.F782L) in exon 16 of the KIT gene were identified in the case 1, 2, 3 respectively, but no above mutations were identified in the patients′ parents or 100 unrelated healthy controls. The 3 genetic variants were all novel pathogenic mutations, and all were deleterious mutations.Conclusions:Three novel pathogenic mutations in the KIT gene were identified in the 3 cases of piebaldism, namely c.2033T>C (p.L678P), c.2485-1G>C, and c.2346C>G (p.F782L). It was further verified that the severity of piebaldism was closely related to the type and location of KIT gene mutations.
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Objective:To analyze clinical characteristics of and causative genes in two families with dystrophic epidermolysis bullosa, and to reveal the pathogenesis of the disease and mechanisms underlying phenotypic differences between patients.Methods:DNA was extracted from peripheral blood samples of members from two families with dystrophic epidermolysis bullosa, and subjected to high-throughput sequencing and Sanger sequencing.Results:The clinical manifestations of the 2 probands in the 2 families were consistent with the diagnosis of dystrophic epidermolysis bullosa, and the symptoms of the proband in family 1 were more serious than those of other patients in the family. Genetic testing showed that all patients in family 1 carried a mutation c.6082G>C (p.G2028R) in the COL7A1 gene, and the proband and her phenotypically normal mother and uncle also carried a splice-site mutation c.7068+2 (IVS91) T>G in the COL7A1 gene, both of which were first reported. The proband in family 2 carried the mutations c.6081_6082 ins C (p.G2028Rfs*71) and c.1892G>A (p.W631X, first reported) in the COL7A1 gene, which were inherited from her father and mother, respectively.Conclusion:The two pathogenic mutations may be the molecular mechanism underlying the severe clinical phenotype in the proband in family 1; the first reported mutations enriched the mutation spectrum of the COL7A1 gene.
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Objective:To report a Chinese pedigree with autosomal dominant Waardenburg syndrome, and to identify causative gene mutations.Methods:Clinical data and peripheral blood samples were collected from the proband and her parents. Genomic DNA was extracted, gene mutations were detected through a next-generation skin-targeted sequencing panel, and Sanger sequencing was performed to verify causative mutations.Results:The proband clinically presented with irregular white patches on the abdomen and lower limbs, moderate to severe sensorineural deafness in the right ear, and iris heterochromia in both eyes. The proband′s mother presented with iris heterochromia in both eyes, epicanthus, early canities and thick eyebrows. In the family, both the proband and her mother were diagnosed with Waardenburg syndrome. A causative frameshift mutation c.976-977delinsT (p.Thr327Profs*54) was identified in both the proband and her mother, which caused the AG to TT base substitution at positions 976 - 977 in the coding region of exon 7 of the PAX3 gene, resulted in a frameshift from the amino acid position 327 to 54 in the PAX3 protein (threonine was substituted by proline at amino acid position 327). The proband′s father showed a normal phenotype, and his genetic test results were negative.Conclusion:The novel frameshift mutation c.976-977delinsT (p.Thr327Profs*54) in the PAX3 gene may contribute to the clinical phenotype of the patients with Waardenburg syndrome in the family.
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Objective:To detect gene mutations in 1 patient with Vohwinkel syndrome who presented with palmoplantar keratoderma, pseudo-ainhum and deafness.Methods:Clinical data were collected from the proband, and a genetic test was performed to identify mutation sites.Results:Clinical manifestations of the proband were consistent with classical Vohwinkel syndrome. The genetic test revealed a heterozygous mutation c.160A>C (p.N54H) in the GJB2 gene, which was not detected in her parents or healthy controls.Conclusion:The heterozygous mutation c.160A>C (p.N54H) in the GJB2 gene was first identified in a patient with classical Vohwinkel syndrome, and there were overlaps in mutation sites between classical Vohwinkel syndrome and palmoplantar keratoderma with deafness.
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Objective:To report a patient with congenital hypotrichosis 14 complicated by hypergonadotropic hypogonadism, and to analyze LSS gene mutations in his family.Methods:Peripheral blood samples were collected from the proband and his parents with normal phenotypes, and genomic DNA was extracted from these samples. Second-generation sequencing was performed to screen suspected mutations among hereditary hair disorder-associated genes. Possible causative genes were identified from the screened suspected variants based on clinical phenotypes, and verified using Sanger sequencing. The identified variants were also verified in healthy controls, and searched in the Human Gene Mutation Database, 1000 Genomes Project database, and ExAC database.Results:The patient harbored a homozygous missense mutation c.812T>C (p.Ile271Thr) in exon 8 of the LSS gene, and his parents were the mutation carriers. The variant was not present in healthy controls and databases.Conclusion:The homozygous mutation c.812T>C in the LSS gene may be the causative mutation for congenital hypotrichosis 14 in this family, which was a novel mutation that had not been reported before.
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Objective:To summarize the clinical characteristics and genetic etiology of infants with D-bifunctional protein deficiency (DBPD).Methods:This study involved two DBPD newborns who were hospitalized in the Second Affiliated Hospital of Wenzhou Medical University in August 2020 and November 2020. Clinical data including manifestations, radiographic findings and genetic testing results were retrospectively analyzed. Relevant articles up to November 2022 were retrieved from various databases including CNKI, Wanfang, CQVIP, Online Mendelian Inheritance in Man and PubMed using the terms of "D-bifunctional protein deficiency" and " HSD17B4" in both Chinese and English. Clinical data of the cases diagnosed with DBPD by genetic testing within two years of age were collected. Clinical features and genetic etiology of the children with DBPD were summarized by descriptive statistical analysis. Results:Both neonates in this report presented with dyspnea, hypotonia, intractable epilepsy, poor response, absence of primitive reflexes, and craniofacial malformation. Whole-exome sequencing revealed that patient 1 carried heterozygous variations of c.972+1G>T and c.727T>A (p.W243R) in the HSD17B4 gene, which were inherited from his father and mother, respectively. A homozygous variation of c.1333+4A>G in the HSD17B4 gene was identified in patient 2. All these mutations were pathogenic. Thirteen papers (12 in English and one in Chinese) involving 19 patients from 16 pedigrees were retrieved. Altogether 21 patients (eight males and 13 females) were included, and among them, four from two pedigrees were born to consanguineous parents. There were 21 patients with hypotonia, 20 with epileptic seizures (17 presenting with epileptic seizures within 5 d after birth) and 12 with craniofacial deformities including high forehead, long philtrum and high palatine arches. Genetic tests showed that 13 patients carried compound heterozygous variations in the HSD17B4 gene and eight patients had homozygous variations. Twenty-six variations in the HSD17B4 gene were detected, including 16 missense mutations, seven splicing mutations, two nonsense mutations and one frameshift mutation. Conclusion:DBPD should be considered and genetic tests should be given when newborns have dystonia and intractable epilepsy complicated by appearance deformity.
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Objective:To investigate the application value of noninvasive prenatal DNA screening combined with nuchal translucency thickness measurement in the diagnosis of fetal chromosome aneuploidy.Methods:A total of 5 730 pregnant women who were screened for fetal chromosomal diseases in the Quzhou Maternal and Child Health Hospital from January 2017 to March 2019 were included in this study. All of them underwent noninvasive prenatal DNA screening and nuchal translucency thickness measurement. The results of amniocentesis were used as the gold standard to evaluate the diagnostic efficacy of noninvasive prenatal DNA screening, nuchal translucency thickness measurement and their combination.Results:Noninvasive prenatal DNA screening revealed that 64 (1.12%) women out of 5 730 pregnant women had high risk of developing chromosomal abnormalities. Ultrasound examination results showed that nuchal translucency was thickened in 140 (2.44%) women. The outcome of adverse pregnancy increased with the increase of nuchal translucency thickness. Among the 68 pregnant women who underwent amniocentesis, 51 women developed chromosomal abnormalities, with trisomy 21 syndrome being the majority (23/51,45.10%). The diagnostic efficacy of noninvasive prenatal DNA screening combined with nuchal translucency thickness measurement in the diagnosis of fetal chromosomal aneuploidy reached the ideal level.Conclusion:Noninvasive prenatal DNA screening combined with nuchal translucency thickness measurement has a high clinical application value. The combined method can be used as the main prenatal DNA screening method for pregnant women and it can effectively avoid the birth of children with chromosomal abnormalities.
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Objective:To investigate pathogenic genes and inheritance patterns in 3 consecutive collodion babies in a family.Methods:The proband was diagnosed as a collodion baby due to extensive dry and chapped skin all over the body at birth. Phenotypes of the proband's parents were normal, but their first and second children presented with dry and chapped skin at birth and died a few days after birth. DNA was extracted from peripheral blood samples of the patient and her parents for whole-exome capture sequencing, and candidate mutations were verified by Sanger sequencing.Results:Compound heterozygous mutations in the ALOX12B gene were identified in the infant, including a missense mutation c.1405 C>T (p.R469w) inherited from her father and a frameshift mutation c.68_69insC (p.L24fs) inherited from her mother.Conclusions:The infant was diagnosed with hereditary ichthyosis, which was inherited in an autosomal recessive manner. The missense mutation c.1405 C>T and frameshift mutation c.68_69insC in the ALOX12B gene may contribute to the clinical phenotype of this infant, and the frameshift mutation had not been reported in China or other countries.
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Objective:To analyze pathogenic mutations in a child with ankyloblepharon-ectodermal defects-cleft lip/palate syndrome.Methods:Clinical data were collected from a patient with ankyloblepharon-ectodermal defects-cleft lip/palate syndrome, and DNA was extracted from peripheral blood samples from the patient and his parents. High-throughput sequencing was performed in the patient by using a gene panel targeting hereditary skin diseases, aiming to determine sites of disease-causing gene mutations. Then, Sanger sequencing was conducted to bidirectionally verify the mutations in the patient and his parents.Results:The male patient aged 3 years and 9 months, and presented with extensive erythema, scales, erosions as well as repeated infections and erosions of the scalp after birth. Reticulated hyper- and hypopigmented patches and scars left on the trunk and limbs after healing of erosions. Physical examination also showed sparse scalp hair, absence of most eyebrows and eyelashes, cleft palate, dysplastic teeth, dystrophic finger and toe nails, and deformed ears without ankyloblepharon. Genetic testing of the patient showed a novel heterozygous missense mutation c.1790T>A (p.Ile597Asn) in the TP63 gene, which had not been reported previously and was rated as pathogenic according to the American College of Medical Genetics and Genomics guidelines. This mutation was not identified in either of his parents.Conclusion:The novel heterozygous missense mutation c.1790T>A in the TP63 gene probably contributes to ankyloblepharon-ectodermal defects-cleft lip/palate syndrome in the patient, which expands genotypic and phenotypic spectrum of this disease.
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Objective:To detect mutations in a pedigree containing two brothers with oculocutaneous albinism (OCA) by whole-exome sequencing and Sanger sequencing.Methods:Clinical data were collected from a pedigree with OCA, and DNA was extracted from peripheral blood samples obtained from the proband and other family members. The whole-exome coding region of the proband was directly sequenced by whole-exome sequencing technology to identify potential pathogenic mutations, and Sanger sequencing was conducted to verify the gene mutations.Results:Both the proband and his younger brother presented with generalized white skin, golden-yellow hair, bilateral nystagmus, photophobia, translucent iris, conjunctival congestion, and refractive errors of both eyes. The proband′s parents, grandparents, maternal grandparents, and children were all phenotypically normal, and his parents′ marriage was non-consanguineous. Three heterozygous mutations were identified in the OCA2 gene of both the proband and his younger brother, including a nonsense mutation c.1290T>A, and 2 missense mutations c.1363A>G and c.1204T>C. The mutation c.1204T>C has not been previously reported, and was a novel gene mutation in the OCA2 gene. In addition, 1 heterozygous mutation c.1204T>C was identified in the OCA2 gene in the proband′s father and daughter, 2 heterozygous mutations c.1290T>A and c.1363A>G were found in the OCA2 gene in the proband′s mother, and 1 heterozygous mutation c.1290T>A was identified in the OCA2 gene in the proband′s son and the daughter of the proband′s younger brother.Conclusions:Three gene mutations were identified in the OCA2 gene in the 2 patients with OCA, and the nonsense mutation c.1290T>A may be the pathogenic mutation causing the clinical phenotype of this family. These findings expand the pathogenic mutational spectrum of the OCA gene.
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Objective:To analyze gene mutations in and clinical phenotypes of 4 children with recessive dystrophic epidermolysis bullosa (RDEB) .Methods:Clinical data were collected from 4 children with RDEB, and DNA was extracted from peripheral blood samples of the children and their parents. A multi-gene panel targeting congenital epidermolysis bullosa was used for high-throughput sequencing. After identification of causative gene mutations, Sanger sequencing was performed to bidirectionally verify the mutations in the patients and their parents.Results:Genetic testing showed 8 compound heterozygous mutations in the COL7A1 gene in the 4 cases. Case 1 was diagnosed with moderate generalized RDEB, and inherited a paternal mutation c.7828C>T and a maternal mutation c.448G> A; case 2 was also diagnosed with moderate generalized RDEB, and inherited a paternal mutation c.3625_3635del and a maternal mutation c.2726_2728del; case 3 was diagnosed with localized RDEB, carrying a paternal mutation c.682+1G>A and an allelic mutation c.5532+1G>A, but the mutation c.5532+1G>A was not identified in the DNA extracted from the parental peripheral blood samples; case 4 was diagnosed with severe generalized RDEB, and inherited a paternal mutation c.5196delC and a maternal mutation c.500_501insAGGG. Among these mutations, c.2726_2728del and c.5196delC had not been reported previously.Conclusions:All the 4 children with RDEB carried compound heterozygous mutations in the COL7A1 gene, which may be the cause of RDEB. The phenotypes of biallelic frameshift mutation carriers appearred more severe than those of carriers of compound heterozygous mutations composed of biallelic splice site, missense and nonsense, frameshift and amino acid deletion or insertion mutations.
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Objective:To report 3 cases of rare subtypes of hereditary epidermolysis bullosa.Methods:Clinical data were collected from the probands and their relatives, whole-exome sequencing was performed to screen disease-causing mutations in the probands, and Sanger sequencing or qPCR was conducted to verify the mutations in patients and their relatives.Results:Case 1 mainly presented with linear red scars on the back, and the proband, her mother with similar clinical manifestations and her asymptomatic daughter all carried a mutation c.4573G>A (p.Gly1525Arg) in the COL7A1 gene. Case 2 presented with generalized reticular pigmentation all over the body and occasional blisters restricted to the hand and foot, and carried a de novo mutation c.74C>T (p.Pro25Leu) in the KRT5 gene. Case 3 presented with pigmentation abnormalities mainly located at the sun-exposed sites and incomplete syndactyly of the left hand, and carried homozygous deletion mutations in exons 2-6 of the FERMT1 gene, which were inherited from her asymptomatic parents. Case 1 was diagnosed with dominant dystrophic epidermolysis bullosa pruriginosa, case 2 was diagnosed with epidermolysis bullosa simplex with mottled pigmentation, and case 3 was diagnosed with Kindler epidermolysis bullosa. Conclusion:The clinical manifestations of epidermolysis bullosa vary greatly, and gene detection is very important for confirmation of diagnosis of its rare types.
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Objective:To investigate two Chinese pedigrees with dyschromatosis symmetrica hereditaria (DSH) , and to analyze gene mutations in the pedigrees.Methods:Clinical data were collected from two probands with DSH and other family members in their pedigrees. Peripheral blood samples were obtained from the two probands, their parents and 100 unrelated healthy controls. Gene mutations were detected by using a skin-targeted sequencing panel, and then verified by Sanger sequencing.Results:Case 1, an 18-year-old male patient, presented with millet-sized hyperpigmented and hypopigmented macules scattered on the dorsum of both hands and feet at the age of 5 years, and his mother had similar manifestations. A novel heterozygous frameshift mutation c.1970dupT (p.F657fs) was identified in exon 5 of the ADAR gene in case 1 and his mother, but not found in his father. Case 2, an 8-year-old male patient, presented with mottled rice- to soybean-sized brown hyperpigmented macules and hypopigmented macules on the face and neck, lower back, buttocks, lower limbs, as well as hands and feet, and his father presented with similar manifestations. A known heterozygous frameshift mutation c.2433_2434delAG (p.T811fs) was identified in exon 7 of the ADAR gene in case 2 and his father, but not found in his mother. Neither of the two mutations was identified in the 100 unrelated healthy controls.Conclusion:In this study, a novel mutation c.1970dupT (p.F657fs) in the ADAR gene was identified in a patient with DSH.
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Objective:To identify gene mutations in a family with incontinentia pigmenti, in order to confirm pathogenic mutations.Methods:Clinical data were collected from all patients in a family with incontinentia pigmenti. DNA was extracted from peripheral blood samples obtained from the patients, healthy members in the family, and 100 unrelated healthy controls, and Sanger sequencing was performed for all exons and their flanking sequences of the NEMO gene.Results:Totally, there were 4 patients in the 4-generation family, who all presented with typical skin lesions and different symptoms. Genetic testing indicated that the proband and the other 3 patients all carried a heterozygous nonsense mutation c.1153C>T (p.Gln385X) at position 1153 in exon 8 of the NEMO gene, which led to the substitution of the glutamine codon (CAG) by the termination codon (TAG) at amino acid position 385. The mutation was not identified in the 14 healthy relatives or 100 unrelated healthy controls. The mutation cosegregated with incontinentia pigmenti in the family. Database searching confirmed the mutation to be a novel nonsense mutation, and it was considered as a very strong pathogenic locus according to the American College of Medical Genetic and Genomics guidelines.Conclusion:The mutation c.1153C>T in the NEMO gene is associated with the occurrence of incontinentia pigmenti in this family.
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Objective:To analyze clinical phenotypes and pathogenic mutations of a patient with classic tuberous sclerosis complex.Methods:Clinical data was collected from a patient with classic tuberous sclerosis complex. Next-generation sequencing was performed to screen pathogenic gene variants, and Sanger sequencing to verify the mutations. Minigene plasmids were constructed and transfected into the human renal epithelial cell line 293T, and RNA was extracted for transcriptional analysis.Results:The patient clinically presented with recurrent epileptic seizures, facial angiofibroma, periungual fibroma, pulmonary lymphangioleiomyomatosis, renal angiomyolipoma and multiple osteosclerosis. Next-generation sequencing revealed a suspected pathogenic variant in the TSC2 gene in the patient. Sanger sequencing identified a heterozygous mutation c.336_336+15delGGTAAGGCCCAGGGCG in exon 4 of the TSC2 gene in the patient, but not in his parents or 100 unrelated healthy controls. Moreover, this mutation had not been previously reported. The minigene experiment showed changed mRNA sequence of the TSC2 gene in this patient with loss of the authentic splice site in exon 4 and insertion of a 74-bp intron, which shifted the splice site 90 bp downstream (r.336delins336+16_336+90) .Conclusion:The novel heterozygous mutation c.336_336+15delGGTAAGGCCCAGGGCG in exon 4 of the TSC2 gene can lead to aberrant splicing, and may contribute to tuberous sclerosis complex in this patient.
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Objective:To compare the in vitro susceptibility of fluconazole-resistant Candida albicans strains from superficial and deep infections to 8 antifungal drugs, and to compare drug resistance mutations in these strains. Methods:According to the Clinical and Laboratory Standards Institute (CLSI) protocol M27-A4, 26 deep infection-derived and 33 superficial infection-derived drug-resistant Candida albicans strains were tested for in vitro susceptibility to 8 antifungal drugs (fluconazole, voriconazole, itraconazole, posaconazole, amphotericin B, fluorocytosine, terbinafine, and micafungin) alone or in combination. DNA was extracted from all drug-resistant strains, and mutations in 3 drug resistance genes, including ERG3, ERG11 and FUR1, were detected by PCR. Normally distributed measurement data with homogeneous variance were compared between two groups by using two-independent-sample t test, non-normally distributed measurement data with non-homogeneous variance were compared using Mann-Whitney U test, and enumeration data were compared using chi-square test. Results:The minimum inhibitory concentrations (MICs) of fluconazole, itraconazole, voriconazole, posaconazole and fluorocytosine all significantly differed between the superficial infection group and deep infection group (all P < 0.05) , while there was no significant difference in the MIC of amphotericin B or micafungin between the two groups (both P > 0.05) . The MIC of terbinafine was >64 μg/ml in 96.6% of the above strains, so could not be compared between groups. As combination drug susceptibility testing revealed, the combination of terbinafine with azoles (fluconazole, voriconazole, itraconazole or posaconazole) showed synergistic inhibitory effects against 15 Candida albicans strains (7 strains from deep infections, 8 strains from superficial infections) , with fractional inhibitory concentration (FIC) indices being 0.033 to 0.187; no marked synergistic effect was observed in the combinations between fluorocytosine and azoles, between fluorocytosine and amphotericin B, or between amphotericin B and fluconazole, with the FIC indices being 0.56 to 1.125. The missense mutation V351A in the ERG3 gene was identified in all the 33 (100%) superficial infection-derived strains, as well as in 13 (50%) deep infection-derived strains, and the mutation A353T in the ERG3 gene was identified in 4 (15%) deep infection-derived strains; as for the ERG11 gene, missense mutations identified in the superficial infection-derived strains included I437V (32 strains, 97%) , Y132H (23 strains, 70%) , T123I (16 strains, 48%) , K128T (6 strains, 18%) , D116E (5 strains, 15%) , A114S (4 strains, 12%) , E266D (2 strains, 6%) , G448E (2 strains, 6%) , and G465S (2 strains, 6%) , while missense mutations identified in the deep infection-derived strains included I437V (23 strains, 88%) , E266D (13 strains, 50%) , E260G (5 strains, 19%) , and V488I (4 strains, 15%) ; the missense mutation R101C in the FUR1 gene was identified in 11 (33%) superficial infection-derived strains, but not identified in deep infection-derived strains. Conclusion:The drug susceptibility and drug resistance mutations differed to some extent between superficial infection- and deep infection-derived fluconazole-resistant Candida albicans strains.
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Objective:To investigate the relationship between RAS, BRAF gene mutations and HER2 gene amplification and clinicopathology and prognosis of colorectal cancer.Methods:The clinical data of 268 patients with colorectal cancer were retrospectively analyzed. KRAS, NRAS and BRAF gene mutations and the HER2 gene amplication were detected.Results:The mutation rates of KRAS, NRAS and BRAF were 53.4% , 2.6% and 3.0%, respectively. The amplification rate of HER2 was 6.7%. KRAS gene mutation tended to occur in the right side colon and rectal cancers( χ2=10.824, P=0.004). BRAF gene mutation mainly occurred in the right side colon cancer ( P=0.044). HER2 gene amplification tended to occur in colorectal cancer patients with RAS/BRAF wild-type ( OR=0.322,95% CI:0.117-0.887, P=0.027). Univariate analysis showed that the progress-free survival of colorectal cancer patients with RAS mutation was significantly shorter than that of the patients with wild( χ2=6.153, P=0.013), and there was no significant difference in overall survival time( χ2=1.938, P=0.164).The progress-free survival and overall survival time were shorter in BRAF mutation than in the wild type( χ2=8.090, P=0.004; χ2=11.125, P=0.001). Multivariate analysis showed that BRAF gene mutation was independent risk factor for survival of colorectal cancer patients ( HR=3.536,95% CI:1.305-9.583, P=0.013). Conclusion:BRAF gene mutations was independent risk factor for poor prognosis of colorectal cancer patients.
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Objective:To explore the application of microfluidic chip in detection of hereditary deafness-associated hotspot mutations.Methods:The dedicatedly designed and fabricated microfluidic chip was integrated with kompetitive allele-specific polymerase chain reaction amplification system, scanned via laser-excited confocal fluorescence scanner, and finally analyzed programmatically to acquire the typing results of the 23 mutation sites of the four common genes associated with hereditary hearing loss. Dried blood spots were collected from 276 neonates containing the 131 cases with hearing loss who were born in 2019 in Chengdu (deafness group) and the 145 cases with normal hearing who were born in 2020 in Chengdu (control group), and analyzed by the microfluidic chip to evaluate its clinical performance.Results:By cluster analysis, the microfluidic chip correctly analyzed the 23 positive reference samples and acquired the same typing results as their actual results, with a limit of detection of 1 mg/L. For the 276 newborn blood spots, the detection results of the microfluidic chips were confirmed to be correct by the contrasting methods. Among Deafness Group, 66 (50.4%) tested positive for the selected 23 mutation hotspots; among Control Group, 40(27.6%) were positive. Among these mutations, c.109G>A of the GJB2 gene was the most prevalent one, whose carrier rate in deafness group and control group were 46.6%(61/131) and 23.4% (34/145), respectively.Conclusions:The micro-fluidic chip system was succeeded in fulfilling the hereditary deafness-related mutation detection, and offered many advantages including high specificity, avoiding the amplicon carryover contamination, simplifying the entire experimental operation process and short detection time, so as to better meet the detection requirement of genetic testing for deafness in newborn screening and other fields.
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Objective:To investigate the clinicopathological features, immunohistochemical phenotypes and molecular characteristics of adenosquamous carcinoma of the lung(ASC)in elderly patients.Methods:Clinical data of 72 ASC patients in the Department of Pathology, The Second Affiliated Hospital of Soochow University from January 2009 to December 2020 were retrospectively analyzed, and 48 patients aged ≥60 years were selected.Clinical manifestations, imaging findings, histopathological and immunohistochemical characteristics were collected, and gene mutations were detected by the amplification refractory mutation system(ARMS-PCR).Results:There were 48 patients including 32 males and 16 females with a mean age of 70 years(range: 60-84 years). The maximum diameters of the tumors ranged from 0.3 to 9.0 cm(mean: 2.8 cm). Microscopically, the tumors contained two components, squamous cell carcinoma and adenocarcinoma, with the squamous cell carcinoma tissue showing intercellular bridges and the adenocarcinoma tissue showing papillary, acinar or tubular structures.Immunohistochemistry assays detected varying expression levels of CK7(30/31), CK5/6(20/28), TTF1(12/31), P40(15/17), and P63(12/13). Molecular testing showed that the EGFR mutation rate was 58.8%(10/17)and the ALK fusion mutation rate was 5.9%(1/17), while ROS1 and MET mutations were not detected.All 48 patients underwent surgical resection.Conclusions:ASC cases are relatively rare and prone to misdiagnosis.The diagnosis requires the combination of HE morphology, immunohistochemistry and imaging examination, and surgery is the main treatment option.The mutation rate of the EGFR gene is relatively high in ASC patients.