Identification Of Specific Mutations Research Articles

Phenotype-Genotype Correlations in Three Different Cases of Adult-Onset Genetic Focal Segmental Glomerulosclerosis.

This study highlights the importance of a combined diagnostic approach in the diagnosis of rare diseases, such as adult-onset genetic FSGS. We present three adult patient cases evaluated with kidney biopsy for proteinuria, chronic kidney disease, and hypertension, which were suggestive of adult-onset genetic FSGS. Renal biopsy samples and formalin-fixed, paraffin-embedded fetal kidneys were evaluated using standard light microscopical stainings, direct immunofluorescence on cryostat sections, and electron microscopy. Clinical exome sequencing was performed for each case, and 45 FSGS-related genes were analyzed. Identifying mutations in the PAX2, ACTN4, and COL4A5 genes have prompted a re-evaluation of the previous histopathological examinations. The PAX2 mutation led to a thinner nephrogenic zone and decreased number of glomeruli, resulting in oligohydramnios during fetal development and oligomeganephronia and adaptive focal-segmental glomerulosclerosis in adulthood. The ACTN4 mutation caused distinct electron-dense aggregates in podocyte cell bodies, while the COL4A5 mutation led to segmental sclerosis of glomeruli with marked interstitial fibrosis and tubular atrophy. The identification of specific mutations and their histopathological consequences can lead to a better understanding of the disease and its progression, as well as potential treatment options.

Analysis of random mutations in Salmonella Gallinarum dihydropteroate synthase conferring sulfonamide resistance.

In bacteria and primitive eukaryotes, sulfonamide antibiotics block the folate pathway by inhibiting dihydropteroate synthase (FolP) that combines para-aminobenzoic acid (pABA) and dihydropterin pyrophosphate (DHPP) to form dihydropteroic acid (DHP), a precursor for tetrahydrofolate synthesis. However, the emergence of resistant strains has severely compromised the use of pABA mimetics as sulfonamide drugs. Salmonella enterica serovar Gallinarum (S. Gallinarum) is a significant source of antibiotic-resistant infections in poultry. Here, a sulfonamide-resistant FolP mutant library of S. Gallinarum was generated through random mutagenesis. Among resistant strains, substitution of amino acid Arginine 171 with Proline (R171P) in the FolP protein conferred the highest resistance against sulfonamide. Substitution of Phe28 with Leu or Ile (F28L/I) led to modest sulfonamide resistance. Structural modeling indicates that R171P and Phenylalanine 28 with leucine or isoleucine (F28L/I) substitution mutations are located far from the substrate-binding site and cause insignificant conformational changes in the FolP protein. Rather, in silico studies suggest that the mutations altered the stability of the protein, potentially resulting in sulfonamide resistance. Identification of specific mutations in FolP that confer resistance to sulfonamide would contribute to our understanding of the molecular mechanisms of antibiotic resistance.

Molecular determinants for cold adaptation in an Antarctic Na+/K+-ATPase

Enzymes from ectotherms living in chronically cold environments have evolved structural innovations to overcome the effects of temperature on catalysis. Cold adaptation of soluble enzymes is driven by changes within their primary structure or the aqueous milieu. For membrane-embedded enzymes, like the Na+/K+-ATPase, the situation is different because changes to the lipid bilayer in which they operate may also be relevant. Although much attention has been focused on thermal adaptation within lipid bilayers, relatively little is known about the contribution of structural changes within membrane-bound enzymes themselves. The identification of specific mutations that confer temperature compensation is complicated by the presence of neutral mutations, which can be more numerous. In the present study, we identified specific amino acids in a Na+/K+-ATPase from an Antarctic octopus that underlie cold resistance. Our approach was to generate chimeras between an Antarctic clone and a temperate ortholog and then study their temperature sensitivities in Xenopus oocytes using an electrophysiological approach. We identified 12 positions in the Antarctic Na+/K+-ATPase that, when transferred to the temperate ortholog, were sufficient to confer cold tolerance. Furthermore, although all 12 Antarctic mutations were required for the full phenotype, a single leucine in the third transmembrane segment (M3) imparted most of it. Mutations that confer cold resistance are mostly in transmembrane segments, at positions that face the lipid bilayer. We propose that the interface between a transmembrane enzyme and the lipid bilayer is a critical determinant of temperature sensitivity and, accordingly, has been a prime evolutionary target for thermal adaptation.

Clinical, Pathologic, and Molecular Characterization of Acanthosis Nigricans Maligna: A Comprehensive Analysis of a Dermatologic Entity and its Association with Underlying Neoplastic Conditions

Acanthosis nigricans maligna (ANM) is a distinctive dermatologic entity presenting with cutaneous hyperpigmentation and epidermal thickening, characteristically located in flexural areas and skin folds. Despite its apparent cutaneous nature, ANM is frequently associated with underlying neoplastic conditions, implying profound clinical significance. In this study, we undertook a comprehensive evaluation of ANM from clinical, pathological and molecular perspectives with the aim of elucidating its etiopathogenic mechanisms and its relationship with neoplastic processes. From a clinical standpoint, we retrospectively analyzed a cohort of patients diagnosed with ANM, exploring the clinical and epidemiologic features that could indicate the presence of an underlying malignant condition. In addition, histopathologic studies were performed on samples of affected skin to discern the microscopic alterations associated with ANM and their correlates with neoplastic processes. In a molecular phase, an in-depth analysis of genetic and molecular markers in skin samples and associated tumor tissues was carried out using advanced sequencing techniques. This allowed the identification of specific mutations, chromosomal alterations and gene expression profiles that could contribute to both the occurrence of ANM and the promotion of the microenvironment conducive to tumor development. The results obtained indicate a significant correlation between the presence of ANM and the abnormal activation of cell signaling pathways, as well as the altered expression of regulatory factors of cell growth and differentiation. Our study highlights the importance of considering ANM as a potential cutaneous marker of underlying neoplastic processes and suggests that its early detection could facilitate the early identification of early stage malignant conditions. Taken together, this comprehensive analysis provides an in-depth view of ANM and its clinical, pathological and molecular relevance in the context of neoplastic diseases.

CT and MRI of Gastrointestinal Stromal Tumors: New Trends and Perspectives.

Gastrointestinal stromal tumors (GISTs) are defined as mesenchymal tumors of the gastrointestinal tract that express positivity for CD117, which is a c-KIT proto-oncogene antigen. Expression of the c-KIT protein, a tyrosine kinase growth factor receptor, allows the distinction between GISTs and other mesenchymal tumors such as leiomyoma, leiomyosarcoma, schwannoma and neurofibroma. GISTs can develop anywhere in the gastrointestinal tract, as well as in the mesentery and omentum. Over the years, the management of GISTs has improved due to a better knowledge of their behaviors and risk or recurrence, the identification of specific mutations and the use of targeted therapies. This has resulted in a better prognosis for patients with GISTs. In parallel, imaging of GISTs has been revolutionized by tremendous progress in the field of detection, characterization, survival prediction and monitoring during therapy. Recently, a particular attention has been given to radiomics for the characterization of GISTs using analysis of quantitative imaging features. In addition, radiomics has currently many applications that are developed in conjunction with artificial intelligence with the aim of better characterizing GISTs and providing a more precise assessment of tumor burden. This article sums up recent advances in computed tomography and magnetic resonance imaging of GISTs in the field of image/data acquisition, tumor detection, tumor characterization, treatment response evaluation, and preoperative planning.

Gold nanoparticles from magnetite for the detection of amyloid proteins in neurodegenerative diseases

Introduction. Currently, neurodegenerative diseases (ND) are the fourth leading cause of death worldwide that pose a great challenge in the development of tools for early diagnosis. Thus, advances in science seek sensitive and selective detection systems and this manuscript will highlight the importance of nanotechnology.
 Material and methods. A literature review was conducted on the representative findings of NPs technologies in neurodegenerative diseases. Articles written in both English and Spanish were included. References between 2015-2021 were also taken into account.
 Results. One of the most representative techniques, AuNP was specifically implemented, together with a magnetic center composed of magnetite, which has as a specific ligand with a C-terminal cysteine domain present in the B-amyloid protein, which adhere directly to the surface of the NPs, characterizing the anomalous protein. Subsequently, by means of nanosensors capable of detecting and measuring different concentrations, these pathologies are identified at an early stage.
 Conclusions. Today, along with the advent of biotechnology, it has been possible to design techniques with NPs that allow the identification of specific mutations and provide diagnosis in individuals. In the investigative models of AuNP, it is possible to infer that the capabilities that make them representative focus on their magnetism and biofunctionality, by specifically binding to amyloid peptides and other ligands present in the protein, which are the major components of amyloid plaques used in these studies.

Solitary fibrous tumor: features of the clinical course, radiation and laboratory diagnostics, therapy: A review

Solitary fibrous tumor (SFT) is a relatively rare mesenchymal neoplasm, mainly affecting serous membranes and soft tissues.It is distinguished by a long-term course, a variable risk of metastasis, the possibility of occurrence in any anatomical region, as well as a low-specific histological structure.Moreover, the last two features of SFT are the main causes of diagnostic errors in morphological verification of a neoplasm. Despite the fact that in the twentieth century SFT was mentioned in the scientific literature quite rarely and, as a rule, in the form of single observations, in the last two decades, due to the widespread use of new molecular genetic research methods and the identification of specific mutations in mesenchymal tumors, the quality and accuracy have significantly increased. making a diagnosis. In addition, the clinical observations accumulated in recent decades in a considerable number of cases made it possible to assess in more detail the clinical and morphological parameters of the SFT associated with the risk of local recurrence and the appearance of distant metastases. This article analyzes the modern literature on epidemiology, molecular pathology, features of radiation and morphological diagnostics, aspects of the genesis of solitary fibrous tumor, as well as the choice of optimal treatment for patients.

Clinical utility of liquid biopsy in breast cancer: A systematic review.

Advancements in genetic sequencing techniques along with the identification of specific mutations and structural changes in multiple cancer genes, make it possible to identify circulating tumor cells and cell free nucleic acids as blood-based biomarkers, serving as a liquid biopsy (LB) with great utility for the diagnosis, treatment and follow-up of patients with neoplasms. This systematic review focuses on the clinical utility of LB in patients with breast cancer (BC). Articles published between 1990 and 2021 were included. Databases searched: Trip Database, WoS, EMBASE, PubMed, SCOPUS, and Clinical Keys. Variables studied: Publication year, country, number of cases, primary study design, LB detection methods, genes found, overall survival, disease-free survival, stage, response to treatment, clinical utility, BC molecular type, systemic treatment and methodological quality of primary studies. Of 2619 articles, 74 were retained representing 12 658 patients, mainly cohort studies (66.2%), the majority were from China (15%) and Japan (12.2%). All primary studies described clinical stage and type of systemic treatment used. Most used biomarker detection method: DNA (52.7%) and type of analysis: quantification of total cfDNA (35.1%). PIK3CA mutation was most frequent (62.9%). Evidence suggests clinically useful applications of BC. Though heterogeneous, publications suggest that LB will constitute part of the standard diagnostic-therapeutic process of BC.

Cyclic Neutropenia, Congenital and Idiopathic Neutropenia

Background: Cyclic neutropenia (CyN) is an autosomal dominant disease usually attributable to mutations in the gene for neutrophil elastase, ELANE . The diagnosis depends upon serial blood counts (at the minimum every other day or three days a week for 6 weeks) showing periods of severe neutropenia (absolute neutrophil count [ANC] < 0.2 x 109/L) recurring at approximately 21 day intervals. The diagnosis can be confirmed by Lomb periodogram analysis. Genetic testing is helpful but not diagnostic because of the overlap of the mutational patterns in ELANE for CyN and severe congenital neutropenia (SCN).Methods: We reviewed clinical and laboratory data for 87 patients enrolled in the Severe Chronic Neutropenia International Registry (SCNIR) with a diagnosis of CyN in 41 kindreds. Their family histories indicated that there were 117 other neutropenic family members. For this review we focused on 53 CyN patients in 15 kindreds who met the following criteria: two or more patients within the kindred had 10 or more CBCs available for analysis. Using data collected before treatment with G-CSF by the patients' primary physicians, we examined graphically displayed serial ANC's for each affected family member and performed periodogram analysis. We compared patterns of ANC fluctuations within families and for patients of similar ages within these families.Findings and implications: •Within families, if 1 patient has a mutation in ELANE, all other neutropenic members will have the same mutation in ELANE.•Within families if the index case has typical and diagnostic oscillations in blood neutrophils, other affected family members often have neutropenia without clear or diagnostic oscillations in their counts. If the patient has a mutation in ELANE, the diagnosis of SCN is sometimes assigned.•In familial CyN the oscillations tend to dampen with lower peaks and somewhat less severe neutropenia in older adults (> 30 years of age). Unless these patients have genetic testing showing a mutation in ELANE or another gene associated with neutropenia, they may be given the diagnosis of chronic idiopathic neutropenia.•The Lomb periodogram is very useful for diagnosing CyN. It is a web-based tool readily accessible on the SCNIR web site, https://depts.washington.edu/registry/. However, it is necessary to collect sufficient number of counts (usually at least 20 counts spaced at regular intervals) to use this method to make the diagnosis of CyN. Although visually there was cycling in at least one member of the 15 families, only 5 families had a member who had CyN with a 95 or 99 percent confidence level based on the Lomb periodogram.Conclusions:Careful family history and serial measurements of ANCs are the foundations for the diagnosis of CyN. Genetic sequencing and identification of specific mutations in ELANE are helpful for prognosis. The Lomb periodogram is helpful, but requires a robust series of counts to assure that cycling is present with a high (95%) confidence level. DisclosuresDale:Genzyme (now owned by Sanofi-Aventis): Consultancy, Patents & Royalties, Research Funding; Genkyotex: Other: DSMB (work completed 6/2015); Hospira: Consultancy; Prolong: Consultancy; Boheringer-Ingelheim: Consultancy; Coherus: Consultancy; Omeros: Other: DSMB; Shire: Other: Independent Review Board; X4Pharma: Research Funding; Philips: Research Funding; Wolter Kluwer: Other: Editor, Current Opinions in Hematology; WedMD/Medscape: Membership on an entity's Board of Directors or advisory committees; National Institutes of Health: Research Funding; University of Washington: Employment, Research Funding; American College of Physicians: Other: Editor and author; GlaxoSmithKline: Equity Ownership; Johnson&Johnson: Equity Ownership; Cellerant: Membership on an entity's Board of Directors or advisory committees; Sanofi Aventis: Consultancy, Editor, Current Opinions in Hematology, Honoraria; Amgen: Consultancy, Research Funding.

Integrating Biomarkers and Targeted Therapy Into Colorectal Cancer Management.

There have been substantial advances in the treatment of metastatic colorectal cancer (mCRC) over the past 15 years. Molecular characteristics of mCRC and identification of specific mutations can serve as predictive and prognostic indicators of disease and response to targeted therapies. When incorporated into clinical decision-making, these biomarkers can serve as critical tools in personalizing therapy to ensure the best outcomes. Additional improvements in the survival of patients with mCRC will be made possible with the identification of new predictive molecular biomarkers and their evaluation using rational and innovative clinical trials.

Molekulargenetische Hochdurchsatzanalysen beim Urothelkarzinom – Möglichkeiten der klinischen Anwendung

The application of next-generation high-throughput techniques for the identification of molecular alterations in tumour tissue has greatly improved our knowledge of the biology of urothelial carcinoma. Gene expression analysis performed as part of the Cancer Genome Atlas project (TCGA) enabled the identification of molecular subtypes with distinct biologic features. The knowledge gained through these analyses could lead to a significant change in the clinical management of patients with urothelial carcinoma through targeted applications. Initial studies indicate that the determination of molecular subtypes could inform the decision on whether or not to use neoadjuvant chemotherapy. Moreover, drugs with effects limited to patients with specific genetic alterations are currently under investigation in clinical trials. The identification of specific mutations in circulating tumour DNA (ctDNA) using liquid biopsies enables the characterisation of metastatic tumours without a tissue biopsy. Moreover, circulating tumour DNA could be used in the aftercare of patients with urothelial carcinoma.

Whole Exome Sequencing of Residual Disease in Multiple Myeloma: Searching for Novel Therapeutic Targets

Introduction:Multiple myeloma (MM) is a plasma cell dyscrasia causing damage of multiple organs with fatal consequences for patients. Despite the success of modern therapies eliminating a vast bulk of the aberrant cells, surviving residual clones eventually lead to the relapse of the disease. Accumulation of genomic alterations during the stage of minimal residual disease (MRD) likely contributes to a selective grow advantage and survival under the drug pressure. Identification of specific mutations in MM patients with MRD can provide unique opportunities to target the residual plasma cell clones. Here we present the first whole exome sequencing (WES) analysis of 22 MM samples of patients with MRD that identified 814 mutated genes with 4% of genes previously implicated in the pathogenesis of MM.Methods:Aberrant plasma cells (A-PCs) and peripheral blood (PB) were collected from patients after signing informed consent form. Presence of MRD was assessed with EuroFlow protocol and A-PCs were sorted out from bone marrow according to their pathological immunophenotype based on the expression of antigens CD38, CD45, CD19 and CD56. DNA from A-PCs was isolated and amplified by Repli-g Single cell kit (QIAGEN). Sequencing libraries were prepared using SureSelect Human All Exon V6 Kit (Agilent Technologies) and sequenced by Macrogen Inc. on Illumina HiSeq 4000 platform with average coverage 50x and 2x 100bp read length. Sequencing data were processed using the Bcbio framework following the standard workflow for tumor-matched-normal studies. Specifically, reads were mapped to the human reference genome GRCh37, successively marking duplicates using Picard. Germline mutations were identified using GATK HaplotypeCaller, whereas somatic mutations were identified using MuTect2 reporting as significant variants observed in at least 5 reads and minimum allele frequency of 10%. Variants in homopolymer regions longer than 5 nucleotides were filtered out. The final set of calls were further characterised by assessing their functional impact using snpEff and by annotating each variant using data from 1000 Genomes Phase 3, ExAC, and ClinVar. We then used OncodriveCLUST to identify putative oncogenic genes, and later compared these results with a literature curated list of MM driver genes (Weaver & Tariman, 2017).Results:Our dataset comprises 22 samples from 21 patients (one patient was sampled in two time points) with MM MRD, who received bortezomib-based regimen (age 41-71, average 59 years, 11/22 males, 10/22 females). 8 patients reached complete response, 9 patients had very good partial response and 4 patients had partial response. In our analysis, we detected 1,014 tumour somatic variants (8-287 per sample, median 36), most of them being missense mutations (676/1014), splice site mutations (145/1014) and frameshift insertions (134/1014). The variants affected a total of 814 genes, 97 genes were shared in at least two samples. The most frequently mutated genes were KIAA1211 (11/22), the immunoglobulin gene IGLV3-1 (8/22), apoptotic chromatin condensation inducer ACIN1 (7/22) and CCR4-associated factor 3 CNOT3 (7/22). We also identified 32 genes known to be mutated in MM in 64% of our samples (14/22). We found mutations shared by at least 2 samples in KRAS (4/22), DIS3 (3/22), TRAF3 (3/22), NRAS (2/22), ANK2 (2/22), BRAF (2/22) and RBM15 (2/22). Further analysis with OncodriveCLUST identified 18 putative oncogenic genes (FDR < 0.1), including KRAS, DIS3, ACIN1.Conclusion:We presented the first whole exome study of MM MRD, providing a characterisation of the mutations observed in A-PCs. We overcame problem with low amount of A-PCs in this disease stage by using whole genome amplification and a highly customised bioinformatic analysis pipeline. Our study suggests that A-PCs are characterised by new MM MRD specific set of mutated genes, along with the presence of mutations in well-known multiple myeloma cancer driver genes. This offers a great potential for design of novel precise treatments targeting MRD after standard MM therapies.Supported by Ministry of Health of the Czech Republic (17-30089A, CZ-DRO-FNOs/2016) and Ministry of Education of the Czech Republic (SGS18/PřF/2017-2018) DisclosuresKryukov:JSC BIOCAD: Employment. Maisnar:Celgene: Honoraria, Membership on an entity's Board of Directors or advisory committees; Novartis: Honoraria; Bristol Myers Squibb: Membership on an entity's Board of Directors or advisory committees; Amgen: Honoraria, Membership on an entity's Board of Directors or advisory committees; Janssen: Honoraria, Membership on an entity's Board of Directors or advisory committees; Takeda: Membership on an entity's Board of Directors or advisory committees. Hajek:Celgene: Consultancy, Honoraria, Research Funding; Janssen: Consultancy, Honoraria, Research Funding; Takeda: Consultancy, Honoraria, Research Funding; Bristol Myers Squibb: Consultancy, Honoraria; Amgen: Consultancy, Honoraria, Research Funding; Novartis: Research Funding.

Abstract 4283: A structure network approach to predict the dynamics and structural stability effects of germline PTEN mutations associated with cancer versus autism phenotypes

Abstract Individuals with germline mutations in the tumor suppressor gene phosphatase and tensin homolog (PTEN), irrespective of clinical presentation, are referred to as PTEN hamartoma tumor syndrome (PHTS). PHTS confers a high risk of breast, thyroid and other cancers. Yet, PTEN mutations are found in up to 20% of individuals with autism spectrum disorder (ASD) and macrocephaly. What remains unclear is why mutations in one gene can lead to seemingly disparate phenotypes. Thus, we sought to identify differences in cancer- vs. ASD-associated PTEN mutations by investigating putative structural effects. We utilized a computational theoretical approach combining protein structure network (PSN), normal mode analysis (NMA), and elastic network models (ENMs) to interrogate 6 PTEN mutations from patients with PHTS-associated cancer, 6 mutations from those with ASD only, 4 mutations shared across both phenotypes, and 1 mutation with both ASD and cancer. Combined with in silico prediction methods we calculated structural stability changes induced by each mutation. All 6 cancer-associated mutations showed decreases in structure stability and increased dynamics across domain interface. In contrast, 5 of 6 ASD-associated mutations showed only localized destabilization. We coupled ENM-NMA with PSN and found PTEN R130G (cancer only) and R173C (shared in patients with both ASD and cancer), play a role in inter-residue signal propagation, indicating both residue positions are crucial to structural stability. Our data suggest that ASD-associated mutations affect local structural dynamics, whereas cancer-associated mutations mediate long-range perturbations significantly altering structural stability and salient communication pathways. These approaches lend insight into altered structural effects of germline PTEN mutations associated with PTEN-ASD and PTEN-cancer, potentially aiding in identification of specific mutations that contribute to each phenotype. Moreover, this work may illuminate the shared and separate molecular features that contribute to autism or cancer – providing a deeper understanding of genotype-phenotype relationships for germline PTEN mutations. The ability to accurately predict PTEN-ASD may spare patients the need to undergo high-risk cancer surveillance that is now performed in all with PHTS. Citation Format: Iris N. Smith, Stetson Thacker, Charis Eng. A structure network approach to predict the dynamics and structural stability effects of germline PTEN mutations associated with cancer versus autism phenotypes [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 4283.

Make a point on left ventricular noncompaction

Hypertrabeculation is a feature of the left ventricle that, by itself, does not coincide with left ventricular non compaction (LVNC), which represents a specific cardiomyopathy. Nowadays, in the absence of gold standard diagnostic criteria, the clinician must integrate imaging aspects together with medical history. The family inheritance for LVNC, presence of neuromuscular disorders, symptoms or signs of heart failure, thromboembolic events, unexplained syncope, pathological findings at rest ECG, Holter ECG, stress test, systolic/diastolic dysfunction at rest echocardiogram, late gadolinium enhancement at cardiac magnetic resonance, and identification of specific mutations are all considered features useful for the diagnosis. Many aspects are not fully understood: multicenter studies, registers and observational studies are needed for a better comprehension of the pathology, adequate risk stratification and targeted follow-up.

Liquid biopsy of cancer: a multimodal diagnostic tool in clinical oncology.

Over the last decades, the concept of precision medicine has dramatically renewed the field of medical oncology; the introduction of patient-tailored therapies has significantly improved all measurable outcomes. Liquid biopsy is a revolutionary technique that is opening previously unexpected perspectives. It consists of the detection and isolation of circulating tumor cells, circulating tumor DNA and exosomes, as a source of genomic and proteomic information in patients with cancer. Many technical hurdles have been resolved thanks to newly developed techniques and next-generation sequencing analyses, allowing a broad application of liquid biopsy in a wide range of settings. Initially correlated to prognosis, liquid biopsy data are now being studied for cancer diagnosis, hopefully including screenings, and most importantly for the prediction of response or resistance to given treatments. In particular, the identification of specific mutations in target genes can aid in therapeutic decisions, both in the appropriateness of treatment and in the advanced identification of secondary resistance, aiming to early diagnose disease progression. Still application is far from reality but ongoing research is leading the way to a new era in oncology. This review summarizes the main techniques and applications of liquid biopsy in cancer.

Molecular characterization of hepatitis C virus in end-stage renal disease patients under hemodialysis.

New direct-acting antiviral (DAA) agents are in development or already approved for the treatment of chronic hepatitis C virus (HCV) infection. The effectiveness of these drugs is related to the previous existence of resistant variants. Certain clinical conditions can allow changes in immunological characteristics of the host and even modify genetic features of viral populations. The aim of this study was to perform HCV molecular characterization from samples of end-stage renal disease patients on hemodialysis (ESRD-HD). Nested PCR and Sanger sequencing were used to obtain genetic information from the NS5B partial region of a cohort composed by 86 treatment-naïve patients. Genomic sequences from the Los Alamos databank were employed for comparative analysis. Bioinformatics methodologies such as phylogenetic reconstructions, informational entropy, and mutation analysis were used to analyze datasets separated by geographical location, HCV genotype, and renal function status. ESRD-HD patients presented HCV genotypes 1a (n = 18), 1b (n = 16), 2a (n = 2), 2b (n = 2), and 3a (n = 4). Control subjects were infected with genotypes 1a (n = 11), 1b (n = 21), 2b (n = 4), and 3a (n = 8). Dataset phylogenetic reconstruction separated HCV subtype 1a into two distinct clades. The entropy analysis from the ESRD-HD group revealed two amino acid positions related to an epitope for cytotoxic T lymphocytes and T helper cells. Genotype 1a was found to be more diverse than subtype 1b. Also, genotype 1a ERSD-HD patients had a higher mean of amino acids changes in comparison to control group patients. The identification of specific mutations on epitopes and high genetic diversity within the NS5B HCV partial protein in hemodialysis patients can relate to host immunological features and geographical distribution patterns. This genetic diversity can affect directly the new DAA's resistance mechanisms.

The Current Landscape of Anaplastic Lymphoma Kinase (ALK) in Non-Small Cell Lung Cancer: Emerging Treatment Paradigms and Future Directions.

Tumorigenic rearrangements in anaplastic lymphoma kinase (ALK) account for 3-7% of all non-small cell lung cancers (NSCLC). Treatment with targeted tyrosine kinase inhibitors (TKIs) has shown impressive clinical responses. Crizotinib was the first agent approved for front-line therapy of ALK-rearranged NSCLC after it demonstrated superiority to chemotherapy in response rate, duration of response, and progression-free survival. However, eventually all patients progress on crizotinib therapy, with the central nervous system (CNS) being the most common site, which served as the impetus for the development of more potent next-generation ALK inhibitors. Currently, ceritinib, alectinib, and brigatinib are all approved for second-line therapy after progression on or intolerance to crizotinib. Investigations into whether the initiation of a second-generation ALK inhibitor as first-line therapy is the superior treatment paradigm has resulted in the approval of ceritinib as initial therapy. Alectinib has also shown impressive results as front-line therapy, as recently reported in two large randomized studies that compared it to crizotinib. There is a significant need to better understand the drivers of and mechanisms underlying resistance to ALK inhibitors. While specific mutations have been identified, there is currently only limited evidence that the identification of specific mutations should impact selection of the next ALK inhibitor. The best treatment option for patients who become TKI refractory is also unclear, though there is some evidence to suggests that these patients are not responsive to checkpoint inhibitors and may respond better to chemotherapy. Combination therapy with other classes of agents may help to overcome resistance mechanisms and should be investigated further.

PL05.07: A Wise Man’s Conclusion

The past decade has seen more advances in diagnosis and management of lung cancer than were available in the previous 30 years. Fifty years ago, the association of cigarette smoking in lung cancer was firmly established by the Surgeon General's report in the United States. During the past decade, major efforts by IASLC and other organizations have greatly reduced the use of tobacco and, thus, we will be seeing a decrement in morbidity and mortality from lung cancer. However, in the United States last year, there were still 228,000 new cases and 160,000 deaths from lung cancer. It remains the number one cause of cancer death in both American men and women, and the same is true in most developed and developing countries. Over 28% of all cases of cancer death in the United States are due to lung cancer. IASLC has been a leader in updating the TNM classification for non-small cell lung cancer. This allows for uniformity of data results in surgical and adjuvant studies. Cisplatin-based adjuvant chemotherapy has been demonstrated to improve the surgical cure rate by 5-10%. In the future, we hope to be able to identify by molecular, rather than just clinical characteristics, those patients with resected lung cancer who are cured with surgery and do not need to be subjected to adjuvant chemotherapy, as has been similarly accomplished in breast cancer. Also, we hope to have better definition of tumors that are inherently platinum resistant and, therefore, would need alternative strategies to try to improve the surgical cure rate. For the last two decades of the 20th century, chemotherapy has been the backbone for treatment of stage IVB lung cancer. Most studies have been built around platinum combination chemotherapy. Earlier studies pre-platinum utilized inactive drugs such as cyclophosphamide and doxorubicin. In the 1980s, cisplatin and etoposide was a common platinum doublet, and in the 1990's, carboplatin + paclitaxel. A review of phase III trials in North America from 1973-1994 demonstrated very sobering results. Thirty-three trials in 8,434 patients were performed and 23 of these 33 included a platinum compound. Only 5 of the 33 trials demonstrated a statistically significant difference in survival with a median increase of 2 months (range 0.7 to 2.7 months). It thus became clear that we need to personalize therapy rather than giving all patients the same chemotherapy. Modest success was seen by adding Bevacizumab to carboplatin + paclitaxel. Major advances have been made during the past decade with the identification of specific mutations that can be therapeutically exploited. EGFR and ALK were the first to be identified and subsequently ROS-1. Molecular targeted agents demonstrated spectacular responses in the great majority of patients, compared to the usual 25% brief responses that were achieved previously with platinum-based combination chemotherapy. These driver mutations were predominantly in adenocarcinomas and non-smokers or never smokers. More recent mutations have included smokers and non-smokers such as BRAF V600E and MET Exon-14 skipping mutation which can be seen in smokers as well as non-smokers. During the past five years, immunotherapy has been an exciting new addition to the armamentarium for treatment of patients with metastatic lung cancer. Immune checkpoint inhibitors are still in the nascent phase and the optimal duration of therapy for stage IVB disease, combination with other immunotherapeutic agents, chemotherapy, or radiotherapy, as well as use of adjuvant therapy, will be awaited with eager anticipation. Exciting new technology such as CRISPR-cas9 to gene edit PD-1 holds great potential future promise to make these immune checkpoint inhibitors more effective in a larger percentage of patients with lung cancer, as well as those responses being more durable. lung cancer, chemotherapy, immunotherapy, new agents

Whole genome sequencing reveals a 7 base-pair deletion in DMD exon 42 in a dog with muscular dystrophy

Dystrophin is a key cytoskeletal protein coded by the Duchenne muscular dystrophy (DMD) gene located on the X-chromosome. Truncating mutations in the DMD gene cause loss of dystrophin and the classical DMD clinical syndrome. Spontaneous DMD gene mutations and associated phenotypes occur in several other species. The mdx mouse model and the golden retriever muscular dystrophy (GRMD) canine model have been used extensively to study DMD disease pathogenesis and show efficacy and side effects of putative treatments. Certain DMD gene mutations in high-risk, the so-called hot spot areas can be particularly helpful in modeling molecular therapies. Identification of specific mutations has been greatly enhanced by new genomic methods. Whole genome, next generation sequencing (WGS) has been recently used to define DMD patient mutations, but has not been used in dystrophic dogs. A dystrophin-deficient Cavalier King Charles Spaniel (CKCS) dog was evaluated at the functional, histopathological, biochemical, and molecular level. The affected dog’s phenotype was compared to the previously reported canine dystrophinopathies. WGS was then used to detect a 7 base pair deletion in DMD exon 42 (c.6051-6057delTCTCAAT mRNA), predicting a frameshift in gene transcription and truncation of dystrophin protein translation. The deletion was confirmed with conventional PCR and Sanger sequencing. This mutation is in a secondary DMD gene hotspot area distinct from the one identified earlier at the 5′ donor splice site of intron 50 in the CKCS breed.

Abstract 2553: Prevalence of BRCA1 germline mutations in Algerian population: implications for genetic testing and counseling

Abstract Background: Breast cancer is currently the leading cause of cancer morbidity and mortality among Algerian women.In this study, we estimated the prevalence of BRCA1 germline mutations in 192 patients with hereditary breast and/or ovarian cancer and sporadic triple-negative breast cancer, respectively. Materials and Methods: 192 Patients and their families were referred through several public hospitals and private medical clinics which provide oncology services throughout Algeria. Patients were selected for family history of breast and/or ovarian cancer. In addition, in the present study, all women (age under 50 years) with triple-negative breast cancer (TNBC) were considered eligible. BRCA1 was screened by PCR-direct sequencing in all patients including all exons where a mutation was previously found in Algerian population (exons 2, 3, 5 and 11). Results: The analysis of DNA samples of 192 individuals revealed that 11 patients carried pathologic germline mutations in BRCA1 gene (5.7%).Five distinct pathogenic mutations: c.19_47del, c.83_84delTG, c.181T&amp;gt;G, c.798_799delTT and c.2125_2126insA were indentified. Interestingly, the BRCA1 mutation c.83_84delTG detected in previous studies in two unrelated Algerian breast cancer families has been identified in this study in 4 patients with a frequency of 2% (4/192): 3 TNBC patients and one patient with a bilateral ovarian cancer, respectively. The four patients had a strong hereditary breast and/ or ovarian cancer history. The c.181T&amp;gt;G/p.Cys61Gly mutation has been detected here in two young breast cancer patients, a bilateral breast cancer patient with a family history of breast cancer and a young TNBC patient (diagnosed at age 36 years), respectively. To date, the Cys61Gly pathogenic variant is one of the most frequent founder mutation identified in Central European populations, has been previously reported for the first time in two Algerian and Moroccan families. The BRCA1 mutation c.798_799delTT was identified here in a young TNBC patient with a family history of breast cancer. Interestingly, this mutation has been previously detected in several unrelated families from Algeria, Tunisia, Morocco and Italy, respectively. As Mediterranean countries share a common history and migration flow history, BRCA1 mutation c.798_799delTT could be a Mediterranean founder mutation. The BRCA1 mutation c.2125_2126insA has been detected here for the first time in three young TNBC patients with a family history of breast cancer and prostate cancer. Conclusions: The accumulating knowledge about the prevalence and nature of BRCA1 mutations in Algerian population and the identification of specific mutations will contribute in the future to the implementation of genetic testing and counseling for families at risk. Based on our currents results, we recommend using the TNBC immunophenotype as criterion to screen for BRCA1 germline mutations in patients with early onset breast cancer. Citation Format: Farid Cherbal, Hadjer Gaceb, Chiraz Mehemmai, Rabah Bakour, Kamelia Yatta, Ikram Boukoufa, Hassen Mahfouf, Kada Boualga. Prevalence of BRCA1 germline mutations in Algerian population: implications for genetic testing and counseling. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 2553.