Emerging role of liquid biopsy in rat sarcoma virus mutated metastatic colorectal cancer:A case report

BACKGROUND In patients with metastatic colorectal cancer(mCRC),the treatment options are limited and have been proved to be affected by rat sarcoma virus(RAS)mutational status.In RAS wild-type(wt)patients,the combination of antiepidermal growth factor receptor(EGFR)monoclonal antibodies with chemotherapy(CT)is more effective than CT alone.On the other hand,RAS-mutated patients are not eligible for treatment with anti-EGFR antibodies.CASE SUMMARY Eleven patients with initially RAS-mutated mCRC were followed from diagnosis to May 2022.At the time of cell-free DNA determination,five patients had undergone one CT line,five patients had undergone two CT lines,and one patient had undergone three CT lines(all in combination with bevacizumab).At the second and third treatment lines[second line(2L),third line(3L)],patients with neo-RAS wt received a combination of CT and cetuximab.In neo-RAS wt patients treated with anti-EGFR,our findings indicated an increase in progression-free survival for both 2L and 3L(14.5 mo,P=0.119 and 3.9 mo,P=0.882,respectively).Regarding 2L overall survival,we registered a slight increase in neo-RAS wt patients treated with anti-EGFR(33.6 mo vs 32.4 mo,P=0.385).At data cut-off,two patients were still alive:A RAS-mutated patient undergoing 3L treatment and a neo-RAS wt patient who received 2L treatment with anti-EGFR(ongoing).CONCLUSION Our case series demonstrated that monitoring RAS mutations in mCRC by liquid biopsy may provide an additional treatment line for neo-RAS wt patients.

The Mutated Acetolactate Synthase Gene from Rice as a Non-Antibiotic Selection Marker for Transformation of Bamboo Cells

Previously, we developed a particle bombardment-mediated transformation protocol in Phyllostachys nigra bamboo by expressing hygromycin phosphotransferase gene (HPT) and neomycin phosphotransferase II gene (NPT II). Although these marker genes could introduce to several tissue cultured organs (e.g. leaves, buds, and calli) of Phyllostachs bamboo species, some organs showed a high susceptibility and/or a low selectivity to hygromycin and kanamycin. In this report, therefore, we describe advantages and technical details for generating stable transgenic bamboo cells using the particle bombardment method with the mutated-acetolactate synthase gene (mALS) from rice (W548L/S627IOsALS) as a non-antibiotic selection marker. A facile and efficient transformation was achieved with the mALS gene and enhanced fluorescent protein gene (mCherry). Approximately 490 and 1400 mCherry-expressing cells/dish/shot in average were observed in both P. bambusoides and P. nigra under fluorescent stereo-microscope. Stable transgenic bamboo cell lines were generated in a selection medium supplemented with 0.1 μM of bispyribac-sodium (BS) as ALS inhibitor. The integration of mALS gene was identified by in vivo ALS enzyme assay and a PCR-restriction fragment length polymerphism (RFLP) based detection procedures.

Mutated recombinant human glucagon-like peptide-1 induces differentiation of PC12 cells

Glucagon-like peptide-1 （GLP-1） and its long-acting analogues have neuroprotective and neurotrophic properties and are emerging as potential treatments for neurodegenerative diseases. Its short half-life has limited the application of GLP-1 in the clinic. We generated a mutated form of human GLP-1 （mGLP-1） using site-directed mutagenesis and gene recombination techniques, and found that these modifications significantly prolonged the biological half-life of GLP-1 compared with native GLP-1 （nGLP-1）. This study investigated the role of mGLP-1 on inducing PC12 cell differentiation, mGLP-1 induced PC12 cell differentiation with neurite outgrowth and increased the expression of growth-associated protein-43 and neuronal class III I^-tubulin, and significantly increased cyclic adenosine monophosphate level. No significant difference was found between mGLP-1 and nGLP-I. The results indicate that mGLP-1 activates the GLP-1 receptor, induces PC12 cell differentiation, and has neurotrophic effects.

Mutated Leader Sine-Cosine Algorithm for Secure Smart IoT-Blockchain of Industry 4.0

In modern scenarios,Industry 4.0 entails invention with various advanced technology,and blockchain is one among them.Blockchains are incorporated to enhance privacy,data transparency aswell as security for both large and small scale enterprises.Industry 4.0 is considered as a new synthesis fabrication technique that permits the manufacturers to attain their target effectively.However,because numerous devices and machines are involved,data security and privacy are always concerns.To achieve intelligence in Industry 4.0,blockchain technologies can overcome potential cybersecurity constraints.Nowadays,the blockchain and internet of things(IoT)are gaining more attention because of their favorable outcome in several applications.Though they generate massive data that need to be effectively optimized and in this research work,deep learning-based techniques are employed for this.This paper proposes a novel mutated leader sine cosine algorithm-based deep convolutional neural network(MLSC-DCNN)in order to attain a secure and optimized IoT blockchain for Industry 4.0.Here,an MLSC is hybridized using a mutated leader and sine cosine algorithm to enhance the weight function and minimize the loss factor of DCNN.Finally,the experimentation is carried out for various simulation measures.The comparative analysis is made for Best Tip Selection Method(BTSM),Smart Block-Software Defined Networking(SDN),and the proposed approach.The evaluation results show that the proposed approach attains better performances than BTSM and SDN.

New Ebola virus in 2014:do identified mutated amino acids in the viral sequences sense?

To the editor,The problem of new Ebola virus outbreak in 2014 is thepresent big consideration of medical society.The new infectionoccurs in West Africa and causes infection in thousands oflocal people,and the trend of the worldwide expansion leads to serious concerns of the medical society. The reason why

Kinetochore protein MAD1 participates in the DNA damage response through ataxia-telangiectasia mutated kinase-mediated phosphorylation and enhanced interaction with KU80

Objective:Mitotic arrest-deficient protein 1(MAD1)is a kinetochore protein essential for the mitotic spindle checkpoint.Proteomic studies have indicated that MAD1 is a component of the DNA damage response(DDR)pathway.However,whether and how MAD1 might be directly involved in the DDR is largely unknown.Methods:We ectopically expressed the wild type,or a phosphorylation-site--mutated form of MAD1 in MAD1 knockdown cells to look for complementation effects.We used the comet assay,colony formation assay,immunofluorescence staining,and flow cytometry to assess the DDR,radiosensitivity,and the G2/M checkpoint.We employed co-immunoprecipitation followed by mass spectrometry to identify MAD1 interacting proteins.Data were analyzed using the unpaired Student'st-test.Results:We showed that MAD1 was required for an optimal DDR,as knocking down MAD1 resulted in impaired DNA repair and hypersensitivity to ionizing radiation(IR).We found that IR-induced serine 214 phosphorylation was ataxia-telangiectasia mutated(ATM)kinase-dependent.Mutation of serine 214 to alanine failed to rescue the phenotypes of MAD1 knockdown cells in response to IR.Using mass spectrometry,we identified a protein complex mediated by MAD1 serine 214 phosphorylation in response to IR.Among them,we showed that KU80 was a key protein that displayed enhanced interaction with MAD1 after DNA damage.Finally,we showed that MAD1 interaction with KU80 required serine 214 phosphorylation,and it was essential for activation of DNA protein kinases catalytic subunit(DNA-PKcs).Conclusions:MAD1 serine 214 phosphorylation mediated by ATM kinase in response to IR was required for the interaction with KU80 and activation of DNA-PKCs.

Mutated Clones of Caladium Humboldtii ＇Phraya Savet＇ from in vitro Culture and Occurrence of Variants from Somatic Hybridization between Two Caladium Species

Variations in Caladium humboldtii cv. ＇Phraya Savet＇ from in vitro culture were observed. Callus and small shoots were induced from unexpanded leaf segments cultured on modified MS medium supplemented with 2.69 p.M l-Naphthalene acetic acid （NAA） and 17.76 μM N6-Benzyladenine （BA）. Shoots were transferred onto modified MS medium supplemented with 8.88 μM BA for shoot multiplication. Subsequently, roots were induced on MS without growth regulator. The regenerated plantlets were vigorously grown in glasshouse conditions. From 4 morphological groups （leaf color ratio, leaf color, petiole and leaf pattern）, the regenerated ＇Phraya Savet＇ caladium plants were divided into 6 types. The occurrence of variants was 52%. Most of the mutated plants were observed from only leaf color ratio （green ： white） and leaf shape. The most significance for commercial value from mutated clones was round leaf obtaining 20%. Characteristics of new clones from somatic hybridization between C humboldtii cv. ＇Phraya Savet＇ and C. bicolor cv. ＇Suvarnabhum＇ using thin cell layer technique from in vitro calli were investigated. Each thin cell layer of induced callus, about 0.5 - 1 mm thick, from both caladiums was alternately laid on the top of each other for 8 layers. Subsequently, the combination of thin cell layers was cultured and the regenerated plantlets were grown in glasshouse conditions. From 3 morphological groups （leaf pattern, leaf color and petiole）, the regenerated caladium plants were found dissimilarly to both original caladiums at 85 percent with 8 types of different characters. Somatic hybridization between C. humboldtii cv. ＇Phraya Savet＇ and C. bicolor cv. ＇Suvarnabhum＇ gave rise to a number of most hybrids with all conserving C. bicolor characters.

Response of BRCA1-mutated gallbladder cancer to olaparib: A case report

gallbladder cancer(gbc), although considered as a relatively rare malignancy, is the most common neoplasm of the biliary tract system. the late diagnosis and abysmal prognosis present challenges to treatment. the overall 5-year survival rate for metastatic gbc patients is extremely low. BRC A1 and BRCA2 are the breast cancer susceptibility genes and their mutation carriers are at a high risk for cancer development, both in men and women. Olaparib, an oral poly ADP-ribose polymerase inhibitor, has been approved by the Food and Drug Administration and the European commission for the treatment of ovarian cancer with any BRCA1/2 mutations. the first case of BRCA1-mutated gbc patient who responded to olaparib treatment is reported here.

Structural Variation Analysis of Mutated Nannochloropsis oceanica Caused by Zeocin Through Genome Re-Sequencing

Zeocin can cause double strand breaks of DNA and thus may be employed as a mutagen. In this study, two strains of Nannochloropsis oceanica, the wild and the Zeocin-tolerant strains, were re-sequenced to verify such function of Zeocin, The results showed that Zeocin can mutate the N. oceanica genome and cause the structural variation. Zeocin either swept away or selected the alleles of genes functioning in ubiquitin-mediated proteolysis, alpha-linolenic acid metabolism, ascorbate and aldarate metabolism, ribosome biogenesis, and circadian rhythm, indicating that N. oceanica may have adjusted its metabolic performances for protein, carbohydrate, and lipid, and changed its ribosome biosynthesis and living rhythm to survive in Zeocin containing medium. In addition, Zeocin caused mutation may have influenced the expression of a set of tanscription factors. It was concluded that Zeocin effectively caused the structural variation of the genome of N. oceanica, and forced the microalgae to select out the alleles of a set of genes around these variations in order to adapt to Zeocin containing medium. Further studies on the genetic basis of the phenotypic adaptation of this haploid and asexual microalga and the application of Zeocin to its genetic improvement are very important.

A mutated CRYGD associated with congenital coralliform cataracts in two Chinese pedigrees

AIM:To investigate the causal gene mutation and clinical characteristics for two Chinese families with autosomal dominant congenital coralliform cataract.METHODS:Two Chinese pedigrees with congenital cataract were investigated.Routine ophthalmic examinations were performed on all patients and non-affected family members.Peripheral blood samples were collected,and the genomic DNAs were extracted.The coding regions of proband’s DNAs were analyzed with cataract gene panel.The identified mutation was amplified by polymerase chain reaction,and automated sequencing was performed in other members of two families to verify whether the mutated gene was co-segregated with the disease.RESULTS:Congenital coralliform cataract was inherited in an autosomal dominant mode in both pedigrees.For each family,more than half of the family members were affected.All patients presented with severe visual impairment after birth as a result of bilateral symmetric coralliform lens opacification.An exact the same defect in the same gene,a heterozygous mutation of c.70 C>A(p.P24 T)in exon 2 of γ Dcrystallin gene,was detected in both probands from each family.Sanger sequencing analysis demonstrated that the mutated CRYGD was co-segregated in these two families.CONCLUSION:A c.70 C>A(p.P24 T)variant in CRYGD gene was reconfirmed to be the causal gene in two Chinese pedigrees.It is known that mutated CRYGD caused most of the congenital coralliform cataracts,suggesting that the CRYGD gene is associated with coralliform congenital cataract.

Symptomatic Val122del mutated hereditary transthyretin amyloidosis: Need for early diagnosis and prioritization for heart and liver transplantation

Background: Hereditary transthyretin(ATTRv) amyloidosis is an autosomal dominant disease linked to transthyretin gene mutations which cause instability of the transthyretin tetramer. After dissociation and misfolding they reassemble as insoluble fibrils(i.e. amyloid). Apart from the common Val30 Met mutation there is a very heterogeneous group of non-Val30 Met mutations. In some cases, the clinical picture is dominated by a rapidly evolving restrictive and hypertrophic cardiomyopathy. Methods: A case series of four liver recipients with the highly clinically relevant, rare and particularly aggressive Val122 del mutation is presented. Medical and surgical therapeutic options, waiting list policy for ATTRv-amyloidosis, including the need for heart transplantation, and status of heart-liver transplantation are discussed. Results: Three patients needed a staged(1 patient) or simultaneous(2 patients) heart-liver transplant due to rapidly progressing cardiac failure and/or neurologic disability. Domino liver transplantation was impossible in two due to fibrotic hepatic transformation caused by cardiomyopathy. After a follow-up ranging from 3.5 to 9.5 years, cardiac(allograft) function was maintained in all patients, but neuropathy progressed in three patients, one of whom died after 80 months. Conclusions: This is the first report in(liver) transplant literature about the rare Val122 del ATTRv mutation. Due to its aggressiveness, symptomatic patients should be prioritized on the liver and, in cases with cardiomyopathy, heart waiting lists in order to avoid the irreversible neurological and cardiac damage that leads to a rapid lethal outcome.

Prediction of mutation position, mutated amino acid and timing in hemagglutinins from North America H1 influenza A virus

This study was trying to predict the mutations in H1 hemagglutinins of influenza A virus from North America including the predictions of mu-tation position, the predictions of would-be-mutated amino acids and the predictions of time of occurrence of mutations. The results paved a possible way for accurate, precise and reliable prediction of mutation in proteins from influenza A virus.

Muscle hypertrophy in transgenic mice due to over-expression of porcine myostatin mutated at its cleavage site

Myostatin, a member of the transforming growth factor beta（TGF-β） superfamily, is a dominant inhibitor that acts to limit skeletal muscle growth and development. In this study, we generated transgenic mice that express porcine myostatin containg mutations at its cleavage site（RSRR） to evaluate its effect on muscle mass. Results showed that the weight of four skeletal muscles including gastrocnemius, rectus femoris, tibialis anterior, and pectoralis increased by 17.83 and 28.39%, 21.76 and 28.70%, 34.31 and 41.62%, 53.21 and 27.54% in transgenic male and female mice, respectively, compared to their corresponding non-transgenic control mice. Measurement of muscle fiber size and number indicated that the mean myofiber size increased by 50.73 and 61.30% in transgenic male and female mice respectively compared to the non-transgenic controls. However, there was no difference in the number of myofiber between transgenic and non-transgenic male mice. These results clearly demonstrated that the increase in skeletal muscle mass in transgenic mice is caused by hypertrophy instead of hyperplasia.

Sequential chemotherapy and icotinib as first-line treatment for advanced epidermal growth factor receptor-mutated non-small cell lung cancer

BACKGROUND Icotinib could have potential effect and tolerability when used sequentially with chemotherapy for advanced epidermal growth factor receptor(EGFR)-mutated non-small cell lung cancer(NSCLC).AIM To evaluate the efficacy and safety of chemotherapy followed by icotinib maintenance therapy as first-line treatment for advanced EGFR-mutated NSCLC.METHODS This multicenter,open-label,pilot randomized controlled trial enrolled 68 EGFRmutated stage IIIB/IV NSCLC patients randomized 2:3 to the icotinib alone and chemotherapy+icotinib groups.RESULTS The median progression-free survival in the icotinib alone and chemotherapy+icotinib groups was 8.0 mo(95%CI:3.84-11.63)and 13.4 mo(95%CI:10.18-16.33),respectively(P=0.0249).No significant differences were found in the curative effect when considering different cycles of chemotherapy or chemotherapy regimen(all P>0.05).CONCLUSION A sequential combination of chemotherapy and EGFR-tyrosine kinase inhibitor is feasible for stage IV EGFR-mutated NSCLC patients.

Mutated elements of a complex promoter (Amh) can help to demonstrate the role of certain elements in controlling differential gene expression

Amh is a single copy gene which is expressed in different ways during mammalian development. Several potential promoter elements have been identified using physiological experimentation and on the basis of interspecific sequence comparison. The role of putative promoter elements in controlling gene expression has been investigated by many workers over the last two decades and here by individually mutating each element. Expression was measured in vitro in cells of Sertoli descent by flowcytometry using EGFP as a reporter gene. Three lines of murine cells were used;pre- and post-pubertal Sertoli and granulosa cells. Differences between the three lines of cells, support the view that differentiation in this in vitro model system is likely to be at the level of available transcription factors at given points in development.

Descriptively probabilistic relationship between mutated primary structure of von Hippel-Lindau protein and its clinical outcome

In this study, we use the cross-impact analysis to build a descriptively probabilistic relationship between mutant von Hippel-Lindau protein and its clinical outcome after quantifying mutant von Hippel-Lindau proteins with the amino-acid distribution probability, then we use the Bayes-ian equation to determine the probability that the von Hippel-Lindau disease occurs under a mutation, and finally we attempt to distinguish the classifications of clinical outcomes as well as the endocrine and nonendocrine neoplasia induced by mutations of von Hippel-Lindau protein. The results show that a patient has 9/10 chance of being von Hippel-Lindau disease when a new mutation occurs in von Hippel- Lindau protein, the possible distinguishing of classifications of clinical outcomes using mod-eling, and the explanation of the endocrine and nonendocrine neoplasia in modeling view.

Unexpected discovery of 2 cases of hepatocyte nuclear factor 1α-mutated infracentimetic adenomatosis

We present 2 cases of hepatocyte nuclear factor 1α (HNF1α)-mutated adenomatosis, discovered for reasons unrelated to this disease, and identified using immunohistochemical methods. These new tools may further our understanding of the link between adenomas/adenomatosis subtypes and their complications, and their association with other abnormalities.

The human δ2 glutamate receptor gene is not mutated in patients with spinocerebellar ataxia

The human glutamate receptor delta 2 gene （GRID2） shares 90%homology with the orthologous mouse gene. The mouse Grid2 gene is involved with functions of the cerebellum and sponta-neous mutation of Grid2 leads to a spinocerebellar ataxia-like phenotype. To investigate whether such mutations occur in humans, we screened for mutations in the coding sequence of GRID2 in 24 patients with familial or sporadic spinocerebellar ataxia and in 52 normal controls. We de-tected no point mutations or insertion/deletion mutations in the 16 exons of GRID2. However, a polymorphic 4 nucleotide deletion （IVS5-121_-118 GAGT） and two single nucleotide polymor-phisms （c.1251G〉T and IVS14-63C〉G） were identiifed. The frequency of these polymorphisms was similar between spinocerebellar ataxia patients and normal controls. These data indicate that spontaneous mutations do not occur in GRID2 and that the incidence of spinocerebellar ataxia in humans is not associated with GRID2 mutation or polymorphisms.

In vitro Recombination and Identification of Mutated Fragment Corresponding to Regulation Region of mtrR Gene of Neisseria Gonorrhoeae

A site-directed mutant DNA fragment Neisseria Gonorrhoeae （NG） stains to construct the was synthesized and transfected into clinical transformants that contained the corresponding mutagenesis of regulation region of mtrR gene. According to the technique of gene splicing by overlap extension （SOEing）, a DNA segment with specific mutagenesis was constructed by two-step polymerase chain reaction （PCR）. The mutation fragments EF could be used for the next experiment in which the mutation NG strains were induced. By comparing the recombinant EF fragments to the corresponding DNA fragments of clinical NG strains, 2 of these were not compatible completely. The results of sequencing revealed that there was a 9 bp deletion between the 45 to 54 inverted repeat sequence localized within the mtrR promoter. It can be confirmed that the fragments EF are the specifically designed mutant fragments.

A three-year clinical investigation of a Chinese child with craniometaphyseal dysplasia caused by a mutated ANKH gene

BACKGROUND Craniometaphyseal dysplasia(CMD)is a rare genetic disorder.Autosomal dominant CMD(AD-CMD)is caused by mutations in the ANKH gene.Affected individuals typically have distinctive facial features including progressive thickening of the craniofacial bones.Treatment for AD-CMD primarily consists of surgical intervention to release compression of the cranial nerves and the brain stem/spinal cord.To alleviate progression of the clinical course and improve the quality of life in children waiting to undergo the necessary surgery,we investigated clinical changes in a diagnosed patient with AD-CMD over three years.CASE SUMMARY A 17-mo-old boy presented with progressive nasal obstruction,snoring and hearing loss symptoms.Physical examination showed enlargement of the head circumference and clinical features such as wide nasal bridge,paranasal bossing,widely spaced eyes with an increased bizygomatic width,and a prominent mandible.The patient underwent otolaryngological examination,endoscopy,hearing test,laboratory examination of phosphorus and bone metabolism,cranial and femoral computed tomography,X-ray and next-generation sequencing.The patient was diagnosed with AD-CMD due to p.Phe377 deletion(c.1129_1131del)on exon 9 of the ANKH gene.After adherence to a prescribed low-calcium diet,the boy’s alkaline phosphatase(ALP)levels continuously decreased to within the normal range.However,after 14 mo of dietary intervention,his parents altered his diet to an intermittent low-calcium diet to include milk and eggs.The patient’s ALP was slightly higher than normal after the dietary change but remained close to the normal range.His serum osteocalcin changed to within normal levels after dietary regulation for 33 mo.His serum combined beta C-terminal telopeptide of type I collagen also continuously decreased after the nutritional intervention,although still slightly higher than normal levels.Despite fluctuating blood test results,the boy’s nasal symptoms were markedly relieved and steadily improved after dietary intervention.No significant changes were found in the craniofacial bones by cranial radiography.Close monitoring of clinical features is still ongoing.Calcitriol treatment is currently under consideration and a surgical procedure is planned as necessary in the future.CONCLUSION We herein report the first Chinese case of AD-CMD with heterozygous mutation of p.Phe377 deletion(c.1129_1131del)on the ANKH gene.Biochemical alterations were significantly improved after dietary intervention indicating that a lowcalcium diet may be applied in pediatric AD-CMD patients with ANKH mutations to help alleviate phenotypic manifestations and improve the quality of life before surgical intervention.Further large scale studies are needed to replicate these findings and to establish the appropriate timing for nutritional and surgical interventions。