Mutant Allele Frequency Research Articles

Genetic and Clinicopathologic Characteristics of Papillary Thyroid Carcinoma in the Chinese Population: High BRAF Mutation Allele Frequency, Multiple Driver Gene Mutations, and RET Fusion May Indicate More Advanced TN Stage.

PurposeTo describe the genetic landscape and clinical characteristics of Chinese patients diagnosed with papillary thyroid cancer (PTC) and to determine which high-risk genetic characteristics suggest a likelihood of lymph node metastasis (LNM) and lateral lymph node metastasis (LLNM).Patients and MethodsData from previously untreated patients with PTC collected between May 2018 and December 2020 from 14 hospitals in China were analyzed retrospectively. High-risk pathologic characteristics were defined as T3/T4, N(+), and N1b(+) stages. All patients were tested for 57 genes by second-generation sequencing. The t-test, chi-square test, and Fisher’s exact test were performed for statistical analysis.ResultsOverall, 395 patients were enrolled in this study. The prevalence of BRAF mutation was 78.53%. BRAF mutant allele frequency (MAF) >16.93% was associated with a significantly higher risk of LNM, LLNM, and T3 + T4 stage compared with a low-risk group, defined by a MAF <2.54% (odd ratios [ORs] for each risk=3.38, 3.46, and 8.54, respectively), and an intermediate-risk group, defined by a MAF of 2.54% to 16.93% (ORs=2.04, 2.07, and 4.07, respectively). The population with RET fusion had higher T, N, and N1b stages (ORs for each stage=10.40, 7.60, and 8.77, respectively) compared with a RET-negative population. Similar conclusions about T, N, and N1b stages were observed in relation to multiple driver gene mutations (ORs for each stage=7.48, 2.80, and 7.04, respectively) compared with population without multiple driver mutations. These genetic characteristics may be suggestive of high clinical risk. However, regardless of genetic profiles, patients younger than age 45 years had greater rates of LNM and LLNM.ConclusionThe main driver gene in this study, BRAF, differs significantly between the United States (79% vs 51%) and other countries. The Chinese population in this study that experienced more aggressive tumor biology had a BRAF MAF greater than 16.93%, exhibited RET fusion events, and had multiple driver gene mutations; thus, these traits may be considered high-risk genetic characteristics in PTC that could warrant aggressive treatment in such population.

Comprehensive landscape of BRAF variant classes, clonalities, and comutations in metastatic colorectal cancer using ctDNA profiling.

152 Background: Although BRAF V600E accounts for the majority of the BRAF mutations in mCRC, non-V600 BRAF variants have been shown in recent years to represent a distinct molecular subtype of mCRC. This study provides a comprehensive profile of BRAF V600 and non-V600 variants, their clonalities and co-mutations in mCRC using a large genomic database of circulating tumor DNA (ctDNA). Methods: A systematic analysis of Guardant360 results was performed among ctDNA samples of mCRC patients from September 2014 to May 2021. A variant was defined as clonal if the mutant allele frequency (MAF) was greater than 50% of the highest somatic MAF in the sample; otherwise it was defined as subclonal. A previously validated anti-EGFR exposure score was applied to predict prior anti-EGFR therapies. Co-mutation analysis was conducted with BRAF, KRAS, NRAS, NF1, ERBB2, PIK3CA and SMAD4. Results: 1,733 out of 14,742 mCRC patients had at least one BRAF variant, including 6.5% of patients with BRAF V600 variants, 1.1% with class II variants, 1.9% with class III variants, and 3.2% with unclassified variants. 431 unique BRAF variants were identified in a total of 1,905 BRAF variants. 70.7% of BRAF V600 variants were clonal while most (56.0%-78.8%) class II, III and unclassified BRAF variants were subclonal (Table). Patients with non- BRAF V600 variants tend to be younger and male. The prevalence of BRAF class II and III variants were higher (2.1% and 3.7%) in patients with predicted prior anti-EGFR exposure compared with patients predicted to have no prior exposure (0.8% and 1.4%). BRAF variants of all classes are more likely to be subclonal in patients predicted to have anti-EGFR exposure than those predicted nonexposed ( p &lt;0.05 in all classes, Fisher’s exact test). Among patients with non- BRAF V600 variants, a greater fraction of co-occurring KRAS and NRAS mutations were detected in those predicted to have prior anti-EGFR exposure. In the patients without predicted EGFR exposure, BRAF class II and III variants showed a higher rate of co-occurring KRAS mutations (25.6% and 21.5%) and co-occurring NRAS mutations (5.8% and 2.7%) compared with BRAF V600 variants (2.4% for KRAS and 0.1% for NRAS); however, co-occurring KRAS G12C was only noted in one patient with a BRAF class II variant. The analysis of outcome data by variant class will be presented at the meeting. Conclusions: We noted significant differences between BRAF V600 and class II/III variants using a large genomic database. Within BRAF class II and III variants, the enrichment in patients with predicted anti-EGFR exposure and the high fraction of co-mutations in KRAS/ NRAS suggest a unique therapeutic need for these patients.[Table: see text]

Plasma-informed minimal residual disease (MRD) assay: A multicenter prospective study in Japanese patients with stage II colorectal cancer.

161 Background: ̃80% of stage II colorectal cancer (CRC) can be cured by surgery alone. However, adjuvant chemotherapy is recommended for patients with high risk features such as bowel obstruction, &lt; 12 lymph nodes examined and T4 tumors. Traditional pathological grading and biomarkers such as carcinoembryonic antigen has limited sensitivity. Several reports indicated circulating tumor DNA (ctDNA) may represent a promising prognostic factor to assess MRD as a factor for prediction of recurrence after surgery. Here, we present a proof-of-concept study for the development of a novel plasma-based highly sensitive Next Generation Sequencing (NGS) panel using SafeSEQ technology in operable CRC Japanese patients. Methods: This multicenter prospective study recruited patients diagnosed as operable clinical stage II CRC (n = 46) with pre- and post- (4̃6 weeks) operative plasma samples collected between Nov, 2019 and Jan, 2021. ctDNA were extracted and a 14-gene NGS panel was used to analyze single nucleotide variants (SNVs) and Indels covered by gene-specific amplicons. MRD variant was defined as same variant detected in both pre- and post- operative plasma samples. Tissue NGS by a 500-gene panel was also performed in a small number of tissue samples (n = 5) to compare the concordance of plasma and tissue variants. Results: Pre- and post-operative ctDNA status of 46 patients were analyzed. ctDNA positive was observed in 69.6% (32/46, 95%CI 55.2, 81.0) pre- and 34.8% (16/46, 95%CI 22.7, 49.3) post- samples. AKT1, CTNNB1, NRAS, POLE and PPP2R1A mutation were not detected in this study. TP53 mutation was most frequently detected in both pre- (22/46) and post- (11/46) samples, whereas APC mutation was ranked 2nd in pre- (15/46) but none in post- samples. A combined 96 variants were detected in all samples, in which 76 of them were &lt; 0.5% mutant allele frequency (MAF). MRD variants were detected in 17.4% (8/46, 95%CI 8.82, 30.99) post- samples. Evaluation of positive percentage agreement between tissue and pre- plasma samples in three patients show that a total 7 variants detected in plasma, and 3 of them were detected in tissue samples. Conclusions: This study assesses the feasibility of a plasma-informed NGS panel by evaluating pre- and post- operative plasma samples. The presence of variants with &lt; 0.5% MAF detected in this study indicate a highly sensitive method is required for accurate MRD detection. Further observation is required to explore the relationship between MRD variant and clinical outcome such as 2-year progression-free survival.

An observational study to evaluate RAS mutations in circulating tumor DNA after standard chemotherapies for metastatic colorectal cancer patients with tumors harboring RAS mutation: RASMEX study (JACCRO CC-17).

TPS230 Background: RAS status in tumor tissue is a predictive factor for the efficacy of anti-epidermal growth factor receptor (EGFR) antibodies therapy in patients (pts) with metastatic colorectal cancer (mCRC). In mCRC pts with tumors harboring RAS mutation, anti-EGFR antibodies therapy has the lack of clinical benefit; thus, the survival of RAS mutant mCRC pts is shorter compared to RAS wild mCRC pts. Recently, several studies have shown that colorectal cancer tissue had heterogeneity of gene profiling, and the analysis of circulating tumor DNA (ctDNA) in blood samples could detect the change of gene alterations in tumors which were caused by chemotherapy. Bouchahda et al., demonstrated that nearly half of the pts with RAS mutant mCRC had no detectable RAS mutation in ctDNA after first-line chemotherapy; moreover, some of the pts had clinical benefits from post-anti-EGFR antibodies therapy. Therefore, anti-EGFR antibodies therapy might become a novel treatment option for RAS mutant mCRC pts without RAS mutations in ctDNA after chemotherapies. Other report also showed the frequency of no RAS mutations in ctDNA was about 1%. There have been few studies to prospectively evaluate the RAS status in ctDNA for mCRC pts with RAS mutant tumors treated with standard chemotherapies. We, therefore, conducted an observational study to evaluate RAS mutations and the mutation allele frequency in ctDNA for mCRC pts with RAS mutant tumors who were treated with first- or second-line chemotherapy. Methods: This study is a multi-center observational/translational study in 67 facilities. The key eligibility criteria are as follows: 1) Eastern Cooperative Oncology Group Performance status 0-1, 2) histologically proven unresectable mCRC, 3) RAS mutation in tumor tissue, 4) refractory or intolerable after response to prior fluoropyrimidine-containing regimen. OncoBEAM RAS CRC kit is used to investigate RAS status in ctDNA just after first- or second-line treatment in enrolled pts. The primary endpoint is the frequency of pts without RAS mutations in ctDNA. Secondary endpoints include mutation allele frequency of RAS in ctDNA and clinical outcomes of pre- and post-treatments (overall response rate, disease control rate, overall survival, and progression-free survival). As the exploratory analysis, gene alterations related to the resistant mechanism of anti-EGFR antibodies (BRAF, PIK3CA, ERBB2, MET) are analyzed in pts without RAS mutations in ctDNA. We assume that no RAS mutations are observed in ctDNA in at least 1 % pts. We expect that one patient with no RAS mutations in ctDNA is detected in 100 pts; thus, the sample size of 300 pts is set for our study. Accrual is starting in April 2021. Clinical trial information: UMIN000043442.

A phase 1b/2 trial of the PLK1 inhibitor onvansertib in combination with FOLFIRI-bev in 2L treatment of KRAS-mutated (mKRAS) metastatic colorectal carcinoma (mCRC).

100 Background: CRC is a common cancer world-wide, accounting for ̃10% of cancer cases and mortality. Treatment options are limited, and survival is poor for pts with advanced disease, particularly those with mKRAS. After failure of 1L treatment for mCRC, regardless of KRAS mutation status, the ORR for FOLFIRI-bev is 5-13%, with PFS 4-6 mos, and OS 10-12 mos. Onvansertib is a highly selective, ATP-competitive, orally bioavailable PLK1 inhibitor that is synergistic with irinotecan and with 5FU in xenograft models of mKRAS CRC. We present preliminary safety, efficacy, and biomarker data from an ongoing Ph1b/2 trial of onvansertib + FOLFIRI-bev in pts with mKRAS mCRC progressing after 1L treatment with fluoropyrimidine + oxaliplatin, +/- bev. Methods: Pts with mCRC with a KRAS mutation detected by a CLIA-certified lab were eligible. In the Ph1b portion of the study, onvansertib was given on a 3+3 dose escalation at 12, 15 or 18 mg/m2 on days 1-5 and 15-19 of each 28-day cycle in combination with FOLFIRI-bev. The MTD was 15 mg/m2 and was chosen as the RP2D. The primary endpoint for the Ph2 was ORR, and radiographic response was assessed every 8 wks per RECIST v1.1. Safety was evaluated continuously, and AEs were recorded using CTCAE v5.0. Baseline and post-treatment blood samples were collected for biomarker analyses, including mutant allele frequency (MAF) of the pt’s known KRAS mutation. Results: As of 16Sep2021, a total of 50 pts had been treated: 18 on the Ph1b and 32 on the Ph2, including 35 pts at the RP2D, and median follow up was 4.7 mos (range 0.4-18). Of the 50 pts, 26 remain on treatment, as do 24 of 35 RP2D pts. The combination was well-tolerated: fatigue, neutropenia, and nausea were the most common treatment-emergent adverse events (TEAE) and were generally low-grade. Neutropenia was managed by removing the 5FU bolus from subsequent cycles of FOLFIRI and adding growth factor. Of the 50 pts, 44 were evaluable for efficacy, including 31 of 35 RP2D pts. ORR was 36% for the total group (1CR and 15 PR in 44 pts) and 35% for the RP2D group (1 CR and 10 PR in 31 pts). First responses were seen between 2 and 6 months after the start of therapy. Responses were observed across different KRAS variants. Pts achieving a CR or PR showed the greatest decreases in plasma MAF after the first cycle of therapy. Of the 50 pts, 24 pts have discontinued for the following reasons: progressive disease (13), toxicity (4), patient decision (4), proceeding to potentially curative surgery or other localized therapy (3). Conclusions: The combination of onvansertib with FOLFIRI-bev was well tolerated: observed TEAEs have been generally low-grade and manageable. The combination has demonstrated a promising ORR in 2L treatment of mCRC pts harboring various KRAS mutations, and efficacy was correlated with early changes in plasma mKRAS. Updated safety, efficacy, and biomarker analyses will be presented. Clinical trial information: NCT03829410.

Evaluation of molecular methods for plasma detection of EGFR mutations in non-small cell lung cancer.

Epidermal growth factor receptor (EGFR) mutations are detected in non-small cell lung cancer (NSCLC) and associated with responses to therapy with tyrosine kinase inhibitors (TKIs). We compared the analytical performances of two real-time PCRs and droplet digital PCR (ddPCR) to detect EGFR mutations using plasma. Plasma EGFR tests were performed using 86 plasma samples from 75 prospectively enrolled NSCLC patients with early and advanced stages. Analytical performances of plasma-using two real-time PCR, Cobas EGFR mutation v2 and PANAMutyper, EGFR kit, and ddPCR were evaluated based on the tissue EGFR test results. The frequencies of EGFR mutations and acquired T790M mutation after TKI therapy were also assessed. The incidence of all EGFR mutations was 52.3% (23/44) in tissue and was up to 43.2% (19/44) in plasma. The Cobas detection rates of three EGFR mutations (exon 19 deletions, L858R, and T790M) in plasma were similar to those in tissue. The Cobas showed a higher detection rate (76.7%) than that by the PANAMutyper (60.5%). Sensitivity for T790M mutation was lower than the sensitivity for the exon 19 deletions or L858R in both tests. Mutant allele frequency measured by ddPCR was significantly correlated with the semi-quantitative values of the Cobas. Plasma EGFR tests showed similar detection rates for common EGFR mutations compared to the tissue EGFR tests. Cobas showed higher sensitivity in detection of EGFR mutations in body fluids than the PANAMutyper. Real-time PCR using plasma or body fluids could be a suitable first test for the detection of EGFR mutations.

Acoustic Array Biochip Combined with Allele-Specific PCR for Multiple Cancer Mutation Analysis in Tissue and Liquid Biopsy.

Regular screening of point mutations is of importance to cancer management and treatment selection. Although techniques like next-generation sequencing and digital polymerase chain reaction (PCR) are available, these are lacking in speed, simplicity, and cost-effectiveness. The development of alternative methods that can detect the extremely low concentrations of the target mutation in a fast and cost-effective way presents an analytical and technological challenge. Here, an approach is presented where for the first time an allele-specific PCR (AS-PCR) is combined with a newly developed high fundamental frequency quartz crystal microbalance array as biosensor for the amplification and detection, respectively, of cancer point mutations. Increased sensitivity, compared to fluorescence detection of the AS-PCR amplicons, is achieved through energy dissipation measurement of acoustically "lossy" liposomes binding to surface-anchored dsDNA targets. The method, applied to the screening of BRAF V600E and KRAS G12D mutations in spiked-in samples, was shown to be able to detect 1 mutant copy of genomic DNA in an excess of 104 wild-type molecules, that is, with a mutant allele frequency (MAF) of 0.01%. Moreover, validation of tissue and plasma samples obtained from melanoma, colorectal, and lung cancer patients showed excellent agreement with Sanger sequencing and ddPCR; remarkably, the efficiency of this AS-PCR/acoustic methodology to detect mutations in real samples was demonstrated to be below 1% MAF. The combined high sensitivity and technology-readiness level of the methodology, together with the ability for multiple sample analysis (24 array biochip), cost-effectiveness, and compatibility with routine workflow, make this approach a promising tool for implementation in clinical oncology labs for tissue and liquid biopsy.

Real-Life Data From the Largest Pediatric Familial Mediterranean Fever Cohort.

Familial Mediterranean fever (FMF) is the most common monogenic autoinflammatory disease manifesting phenotypic heterogeneity. It is a clinically diagnosed disease supported by MEditerranean FeVer (MEFV) gene mutation analysis. However, the phenotype-genotype correlation is not yet established clearly. We aimed to determine the clinical findings, phenotype-genotype correlation, and treatment outcomes within a large pediatric FMF cohort. The medical charts of children with FMF who were diagnosed and followed up at the eight pediatric rheumatology units were reviewed retrospectively. All patients in the cohort were analyzed for sequence variants in exon 2,3,5 and 10 of the MEFV gene. Patients without any mutations or with polymorphisms including R202Q were excluded. A total of 3,454 children were involved in the study. The mean ± standard deviation of current age, age at symptom onset, and age at diagnosis were 12.1 ± 5.2, 5.1 ± 3.8, and 7.3 ± 4.0 years, respectively. Of 3,454 patients, 88.2% had abdominal pain, 86.7% had fever, 27.7% had arthritis, 20.2% had chest pain, 23% had myalgia, and 13.1% had erysipelas-like erythema. The most common MEFV mutation patterns were homozygous (32.5%) and heterozygous (29.9%) mutations of exon 10. Homozygous M694V was present in 969 patients (28.1%). Allele frequencies of common mutations were M694V (55.3%), M680I (11.3%), V726A (7.6%), and E148Q (7.2%). Children carrying homozygous or compound heterozygous exon 10 mutations had an earlier age of disease onset (4.6 vs. 5.6 years, p = 0.000) and a higher number of attacks per year (11.1 vs. 9.6, p = 0.001). Although 8% of the patients had a family history of amyloidosis, 0.3% (n = 11) had the presence of amyloidosis. M694V homozygosity was detected in nine patients who developed amyloidosis. Colchicine resistance was present in 4.2% of our patients. In this largest pediatric cohort reviewed and presented to date, patients with exon 10 mutations, particularly the M694V homozygous mutation, have been demonstrated earlier disease onset, annual attack count, and more frequent colchicine-resistant cases. Although E148Q is considered as a polymorphism in some populations, it was identified as a disease-causing mutation in our cohort. Secondary amyloidosis is still happening in adults however, it is extremely rare among children, presumably due to increased awareness, tight control, and the availability of anti-IL1 agents in colchicine-resistant cases.

Biased expression of mutant alleles in cancer-related genes in esophageal squamous cell carcinoma.

Recent progress of large-scale international studies has provided comprehensive catalogs of somatic mutations in cancers. Additionally, it has become evident that allelic imbalance in the abundance of somatic mutations between DNA and RNA were pervasive in various types of cancer. However, the allelic imbalance of the abundance of somatic mutations in esophageal squamous cell carcinoma (ESCC) has not been fully analyzed. We performed exome sequencing for 25 Japanese patients with ESCC to detect a comprehensive catalog of somatic mutations in ESCC. Additionally, we performed mRNA sequencing to evaluate the allelic imbalance of the identified somatic mutations at the transcriptional level by comparing the mutant allele frequencies between RNA and DNA. The exome sequencing showed that TP53 and ZNF750 were significantly mutated genes. The expression levels of TP53 and ZNF750 were different depending on the mutation status. In almost all the tumors with missense mutations in TP53 and ZNF750, the mutant allele frequencies were higher in the RNA sequencing than those in the exome sequencing, indicating that the mutant alleles were preferentially expressed. By examining the allelic imbalances for all the identified missense mutations, we demonstrated that genes showing preferential expressions of the mutant alleles were involved in the pathways including cell cycle, cell death, and chromatin modification. The results of this study suggest that the allelic imbalance of the abundance of somatic mutations plays important roles in the initiation and progression of ESCC by modulating cancer-related biological pathways.

Predictive values of mutational variant allele frequency in overall survival and leukemic progression of myelodysplastic syndromes.

The implication of mutational variant allelic frequency (VAF) has been increasingly considered in the prognostic interpretation of molecular data in myeloid malignancies. However, the impact of VAF on outcomes of myelodysplastic syndromes (MDS) has not been extensively explored. Targeted next-generation sequencing was performed in 350 newly diagnosed MDS cases. The associations of mutational VAF of each gene with overall survival (OS) and leukemia-free survival (LFS) were examined by multivariate Cox regression after univariate analysis. Shorter OS was independently associated with DNMT3A VAF (HR 1.020 per 1% VAF increase; 95% CI 1.005-1.035; p = 0.011) and TP53 VAF (HR 1.014 per 1% VAF increase; 95% CI 1.006-1.022; p = 0.001). LFS analyses revealed that TET2 VAF (HR 1.013 per 1% VAF increase; 95% CI 1.005-1.022; p = 0.003) and TP53 VAF (HR 1.012 per 1% VAF increase; 95% CI 1.004-1.021; p = 0.005) were independently associated with faster leukemic transformation. Furthermore, we established nomograms to predict OS and LFS, respectively,by integrating independent mutational predictors into the revised International Prognostic Scoring System. Our study highlights that VAF of certain genes should be incorporated into routine clinical prognostication of survival and leukemic transformationof MDS.

Polymorphism of coupled indels in porcine TNNC2 alters its transcript splicing and is associated with meat quality traits.

The TNNC2 gene encodes the fast-skeletal C subunit of the troponin complex that plays a vital role in the regulation of striated muscle contraction and could be a candidate gene for pork quality. Here, we identified coupled insertion/deletion (indel) variants, a 17-bp insertion and an 11-bp deletion, in porcine TNNC2. The coupled indel variants provide an alternative splicing donor site and cause a 42-bp truncation in the first exon of TNNC2-201, leading to increased expression of TNNC2-201. Polymorphism of the two indel variants is associated with the average backfat thickness (p=3.16×10-3 ), pH value 24h post-slaughter (p=4.31×10-4 ), intramuscular fat (IMF) content (p=1.54×10-2 ), and myofiber cross-sectional area (p=2.86×10-2 ) of longissimus dorsi in a population of 425 Duroc (♂)×Luchuan (♀) pigs. In an independent population of 1,304 commercial hybrid pigs, we further confirmed that it is associated with the IMF content (p=1.75×10-4 ), pH value 45min post-slaughter (p=6.34×10-3 ), and drip loss (p=2.88×10-2 ) of the longissimus dorsi muscle. An increased frequency of the mutant allele is linked to increased IMF content, smaller myofibers, and a relatively moderate pH value. Furthermore, we detected a mutant allele frequency of 96.67% in Luchuan pigs and 86.67% in Tongcheng pigs, whereas the frequency was 0.91% in Duroc pigs, 2.04% in Landrace pigs, and 0% in Yorkshire and Pietrain pigs, indicating its opposing distributions in lean-type and Chinese local pig breeds. The present results establish coupled indel variants of TNNC2 as a novel molecular marker for meat quality improvement.

Prevalence and molecular characterization of alpha and beta-Thalassemia mutations among Hakka people in southern China.

Our aim was to investigate molecular features of thalassemia for proper clinical consultation and prevention in Heyuan. In our research, a total of 25,437 positive screening subjects were further subjected to a genetic analysis of α-thalassemia (α-thal) and β-thalassemia (β-thal). The deletion of α-thal mutation was tested by Gap-PCR, while the non-deletion of α-thal and β-thal mutation were identified by the PCR-reverse dot blot (PCR-RDB) technique. Nested PCR detected Hkαα/-- SEA and Hkαα/αα. Among the 25,437 positive screening subjects, 44.09% (11216/25437) subjects were bearers of thalassemia variations, and 30.85% (7847/25437) subjects showed α-thal changes alone. Among the 23 genotypes with α-thal mutation alone, the three common genotypes were --SEA/αα(68.34%), -α3.7/αα(16.44%), and -α4.2/αα(6.38%). Of the 11.50% (2924/25437) subjects and 29 genotypes with β-thal mutation alone, the three common genotypes were βCD41-42/βN(36.22%), βIVS-II-654/βN(30.88%), and β-28/βN(13.47%). Additionally, of the 1.75% (445/25437) subjects and 55 genotypes showed both α- and β-thal mutations. We also identified 269 cases of Hb H and six patients of Hkαα. Furthermore, the common genotypes of α-thal and β-thal mutations were consistent with allele frequencies of mutations. Our study establishes molecular features of thalassemia among Hakka people in Heyuan. It will be useful for developing strategies to prevent thalassemia.

Effect of MTTP -493G/T, I128T, Q95H and Q244E polymorphisms on hepatic steatosis in patients with chronic hepatitis

BackgroundChronic hepatitis C is characterized by a progressive deterioration of liver function and is involved in metabolic complications, such as hepatic steatosis. ObjectiveThe aim of this study was to investigate the role of host and viral characteristics associated with -493G/T (rs1800591), I128T (rs3816873), Q95H (rs61733139), and Q244E (rs17599091) Single Nucleotide Polymorphisms (SNPs) in the Microsomal Triglyceride Transfer Protein (MTTP) gene on hepatic steatosis in chronic hepatitis C. MethodsSNPs were genotyped by PCR-RFLP and analyzed in combination with host and viral characteristics by multiple logistic regression in different genetic models of inheritance. ResultsThe authors analyzed 236 patients with chronic hepatitis C, and 53% had hepatic steatosis. The mutated allele frequencies were > 5%, and the genotypes were in Hardy-Weinberg equilibrium (p ≥ 0.05). It was observed that patients with HCV genotype 3 infection (OR = 2.74, 95% CI 1.24‒6.06, p = 0.013), female sex (OR = 2.28, 95% CI 1.21‒4.28, p = 0.011) and moderate- and high-intensity liver inflammatory activity (A2-A3) (OR = 3.61, 95% CI 1.86‒7.01, p < 0.001) alone exhibited a higher risk of steatosis. The results of multiple logistic regression analysis for interaction showed that for the -493G/T SNP, when the GT/TT genotype (dominant model) and the GT genotype (codominant model) were each combined with HCV genotype 3 infection, an 11.51-fold (95% CI 2.08‒63.59, p = 0.005) and a 15.69-fold (95% CI 2.46‒99.85, p = 0.004) increased risk of steatosis, respectively, was observed. For the I128T SNP, when both the IT/TT genotype (dominant model) and the IT genotype (codominant model) were combined with HCV genotype 3 infection, an 8.51-fold (95% CI 1.59‒45.54, p = 0.012) and an 8.40 fold (95% CI 1.51‒46.91, p = 0.015) increased risk of steatosis, respectively, was observed. ConclusionThe present study showed that the viral genotype combined with the -493G/T and I128T SNPs in the MTTP gene influences hepatic steatosis.

Mature plasmacytoid dendritic cells associated with acute myeloid leukemia show similar genetic mutations and expression profiles to leukemia cells.

Introduction:Mature plasmacytoid dendritic cells (pDCs) proliferation associated with myeloid neoplasms (MPDMN) are recognized as a neoplasm related to fully differentiated pDCs. Although it has been reported for many years, the genomic landscape of MPDMN is poorly understood.Methods:We reported two patients who developed acute myeloid leukemia (French-American-British M5 subtype) coexisted with immunophenotypically mature pDCs proliferation, which fit the diagnosis of MPDMN. We sorted pDCs from myeloid blasts by flow cytometry and performed whole-exome sequencing and RNA sequencing of the two cell populations, respectively.Results:The immunophenotypes of pDCs in both patients were positive for CD123bri, HLA-DR, CD4, CD303, CD304, and negative for CD56, CD34, CD117, and TdT. The variant allele frequency of gene mutations in myeloid blasts and pDCs were similar. The expression data showed myeloid blasts clustered tightly with hematopoietic stem cells, and pDCs from patients clustered tightly with granulocyte-monocyte progenitors/common myeloid progenitor, rather than with pDCs from the GEO platform.Conclusion:Our study suggested that pDCs derived from the leukemic clone, evidenced by a shared mutation profile and similar transcriptional signatures between pDCs and concurrent myeloid blasts.

Mitochondrial sirtuins genetic variations and gastric cancer risk: Evidence from retrospective observational study

The purpose of the present study was to analyze the role of selected polymorphisms of SIRT3 and SIRT5 in gastric carcinogenesis. For this study, 500 blood samples of GC patients and 500 blood samples of healthy individuals were collected. Six selected polymorphisms of mitochondrial sirtuins were analyzed for analysis using Tetra-Arms PCR followed by DNA sequencing. Mutant allele frequencies of selected polymorphisms [rs3782116 (p<0.0001), rs6598072 (p<0.0001) and rs11246020 (p<0.0001), rs938222 (p=0.0136), rs3757261 (p=0.0005) and rs2841511 (p=0.0015)] were observed significant higher in GC patients vs controls. Haplotype analysis was performed, and 51 haplotypes were generated using haploview software. Among these haplotypes, eleven haplotypes were found associated with a significantly increased risk of GC. Furthermore, SNP-SNP interaction showed a significant correlation between studied SNPs and GC risk. Kaplan Meier analysis showed that mutant allele frequencies of selected polymorphisms are linked with a significant decrease in survival of GC patients CONCLUSIONS: It can be concluded that selected SNPs may be associated with enhanced risk of GC and hence can be potential prognostic markers for prognosis and predisposition of GC.

An ancient positively selected BMPRIB missense variant increases litter size of Mongolian sheep populations following latitudinal gradient.

New gene mutation origination is a driving force for the evolution of organisms. The effect of FecB mutation in BMPRIB gene on the litter size of sheep has been well known for a long time, each copy of the mutant allele increases litter size by 0.4-0.5. However, the origin and adaptive evolution mechanism of FecB mutation are still unclear. Here we carried on the thorough analysis on evolutionary features of BMPRIB gene and found that 150 species as a whole is under purifying selection while sheep lineage shows evidence of positive selection. The results of allele age estimation revealed that the FecB mutation in Mongolian sheep of China originated in Mongolian Plateau at about 5000years ago. Due the two shape drops in temperature subsequently, Mongolian sheep migrated from north to south following the northern nomadic people. Accordingly, the FecB mutant allele frequency increased, with the lowest in sheep locating at Mongolian plateau (0.01) and the highest in sheep locating at Yangtze River valley (0.96). In conclusion, the FecB mutation in Mongolian sheep of China originated in Mongolian Plateau at about 5000years ago, and the differentiated litter size of Mongolian sheep might be the result of adaptation to various environments during the migration following latitudinal gradient. This study may well exemplify selection on an ancient variation triggered by drastic ecological shifts, and is also helpful to analyze the adaptive evolution mechanism of economic traits of domestic animals and identify major genes and molecular markers.

A long-term responding epidermal growth factor receptor mutated non-small cell lung cancer patient with extremely high mutation allele frequency.

ENWEndNote BIBJabRef, Mendeley RISPapers, Reference Manager, RefWorks, Zotero AMA Miyazaki K, Sato Y, Satoh H, Hizawa N. A long-term responding epidermal growth factor receptor mutated non-small cell lung cancer patient with extremely high mutation allele frequency. Contemporary Oncology/Współczesna Onkologia. 2022;26(1):88-89. doi:10.5114/wo.2022.115447. APA Miyazaki, K., Sato, Y., Satoh, H., & Hizawa, N. (2022). A long-term responding epidermal growth factor receptor mutated non-small cell lung cancer patient with extremely high mutation allele frequency. Contemporary Oncology/Współczesna Onkologia, 26(1), 88-89. https://doi.org/10.5114/wo.2022.115447 Chicago Miyazaki, Kunihiko, Yoshiharu Sato, Hiroaki Satoh, and Nobuyuki Hizawa. 2022. "A long-term responding epidermal growth factor receptor mutated non-small cell lung cancer patient with extremely high mutation allele frequency". Contemporary Oncology/Współczesna Onkologia 26 (1): 88-89. doi:10.5114/wo.2022.115447. Harvard Miyazaki, K., Sato, Y., Satoh, H., and Hizawa, N. (2022). A long-term responding epidermal growth factor receptor mutated non-small cell lung cancer patient with extremely high mutation allele frequency. Contemporary Oncology/Współczesna Onkologia, 26(1), pp.88-89. https://doi.org/10.5114/wo.2022.115447 MLA Miyazaki, Kunihiko et al. "A long-term responding epidermal growth factor receptor mutated non-small cell lung cancer patient with extremely high mutation allele frequency." Contemporary Oncology/Współczesna Onkologia, vol. 26, no. 1, 2022, pp. 88-89. doi:10.5114/wo.2022.115447. Vancouver Miyazaki K, Sato Y, Satoh H, Hizawa N. A long-term responding epidermal growth factor receptor mutated non-small cell lung cancer patient with extremely high mutation allele frequency. Contemporary Oncology/Współczesna Onkologia. 2022;26(1):88-89. doi:10.5114/wo.2022.115447.

Using All Our Genomes: Blood-based Liquid Biopsies for the Early Detection of Cancer.

The pursuit of highly sensitive and specific cancer diagnostics based on cell-free (cf) nucleic acids isolated from minimally invasive liquid biopsies has been an area of intense research and commercial effort for at least two decades. Most of these tests detect cancer-specific mutations or epigenetic modifications on circulating DNA derived from tumor cells (ctDNA). Although recent FDA approvals of both single and multi-analyte liquid biopsy companion diagnostic assays are proof of the tremendous progress made in this domain, using ctDNA for the diagnosis of early-stage (stage I/II) cancers remains challenging due to several factors, such as low mutational allele frequency in circulation, overlapping profiles in genomic alterations among diverse cancers, and clonal hematopoiesis. This review discusses these analytical challenges, interim solutions, and the opportunity to complement ctDNA diagnostics with microbiome-aware analyses that may mitigate several existing ctDNA assay limitations.

CYP3A4*2 Gene Polymorphism and its association with Clinical Phenotyping (temperament) Concept of Unani Medicine Philosophy in Indian Population

Phenotypic heterogeneity is one of the major concerns in personalized medicine. Variations in drug metabolism rates vary in every individual and have been associated with drug metabolizing gene polymorphism. CYP3A4*2 gene polymorphism in drug metabolism has been reported to be associated with a range of diseases. In this regard, the present investigation is aimed to determine the possible association of Unani philosophy of temperaments (clinical phenotypes) with CYP3A4*2 gene polymorphism in patients and controls. In the present study, 800 subjects were analyzed for CYP3A4*2 gene polymorphism by PCR-RFLP technique in four different clinical phenotypes (Sanguine, Phlegmatic, Choleric and Melancholic) as per unani philosophy. Mutant allelic frequencies of patients and controls of four unani temperaments are sanguine (Damavi) (0.19 vs 0.12), phlegmatic (Balghami) (0.21 vs 0.11), bilious (Safravi) (0.23 vs 0.11) and melancholic (Saudavi) (0.16 vs 0.11) subjects. Safravi (p&lt;0.04) and Balghami (p&lt;0.01) results are statistically significant. Bilious (Safravi) and phlegmatic (Balghami) patients showed significant association with CYP3A4*2 gene polymorphism with 1.76 and 2.04 odds risk. Further such studies are essential in unani regime for providing better treatment for many diseases which require personalized medicine based on phenotype-genotype correlation.

Screening and Functional Analysis of TEK Mutations in Chinese Children With Primary Congenital Glaucoma.

Purposes: Recent studies have suggested that loss-of-function mutations of the tunica intima endothelial receptor tyrosine kinase (TEK) are responsible for approximately 5% of primary congenital glaucoma (PCG) cases in diverse populations. However, the causative role of TEK mutations has not been studied in Chinese PCG patients. Here, we report the mutation spectrum of TEK after screening a large cohort of PCG patients of Chinese Han origin and analyze the identified variants in functional assays. Methods: TEK-targeted next-generation sequencing (NGS) was performed in 200 PCG patients. Candidate variants were prioritized by mutation type and allele frequency in public datasets. Plasmids containing wild type and identified variants of TEK were constructed and used to assess protein expression, solubility, receptor auto-phosphorylation, and response to ligand stimulation in cell-based assays. Results: Ten missense and one nonsense heterozygous variants were detected by NGS in 11 families. The clinical features of TEK variants carriers were comparable to that of TEK-mutated patients identified in other populations and CYP1B1-mutated individuals from in-house database. Functional analysis confirmed four variants involving evolutionarily conserved residues to be loss-of-function, while one variant (p.R1003H) located in tyrosine kinase domain seemed to be an activating mutation. However, our results did not support the pathogenicity of the other five variants (p.H52R, p.M131I, p.M228V, p.H494Y, and p.L888P). Conclusion: We provide evidence for TEK variants to be causative in Chinese PCG patients for the first time. Attention needs to be paid to TEK mutations in future genetic testing.