Genetics is nothing but the study of genes, their capacities and impacts. Atomic genetics, formative genetics, populace genetics and quantitative genetics, and human genetics are different sorts of hereditary investigations. Human genetics is the investigation of legacy in people, i.e., it is the investigation of legacy of attributes in kids from guardians. It incorporates the investigation of quality articulation, quality structure, transformation examination, cytogenetic and genomic imaging, genome structure and association, hereditary and physical mapping, malady affiliation contemplates, atomic diagnostics, genetics of complex illnesses, epistatic collaborations, and different ideas. It additionally decides different hereditary components in charge of ailments. It offers different points of interest, for example, distinguishing proof and determination and treatment of different illnesses, for example, malignant growth, cardiovascular sicknesses, and other hereditary maladies.

High risk human papillomaviruses (HPV) plays important roles in the development of cervical cancer, a number of other anogenital cancer and they are increasingly found in oropharyngeal squamous cell carcinoma (OPSCC), however there has not been comprehensive analysis about the role how these viruses play in the development of OPSCC. To characterize the physical status of HPV within OPSCC and to determine the effect this has throughout the host genome, we have performed 30-40X whole genome sequencing (WGS) on the BGI sequencing platform on 34 OPSCCs: 28 of which were HPV positive. We then examined the sequencing data to characterize the HPV copy number and HPV physical status to determine what effect they have on both HPV and human genome structural changes. WGS determined the HPV copy number across the viral genome. HPV copy number ranged from 1 copy to as high as 150 copies in each individual OPSCC. Independent of HPV copy number, most tumors had either a small or a very large deletion in the viral genome. We discovered that these deletions were the result of either HPV integration into the human genome or HPV-HPV sequence junctions. WGS revealed that ~ 70% of these tumors had HPV integrations within the human genome and HPV integration occurred independent of HPV copy number. Individual HPV integrations were found to be highly disruptive resulting in structural variations and copy number changes at or around the integration sites. WGS reveals that there is a great complexity in both HPV sequences present and the HPV integrations events in HPV positive OPSCCs tumors. Thus HPV may be playing different roles in the development of different OPSCCs and this further challenge the HPV-driven carcinogenesis model first proposed for cervical cancer.

Oropharyngeal squamous cell carcinoma (OPSCC) is a cancer of the base of the tongue and tonsils. This cancer was previously found in individuals with a smoking and drinking history but is increasingly being promoted by the presence of high risk types of human papillomavirus (HPV). In an attempt to determine the exact role that HPV plays in the development of this cancer we have been using mate-pair next generation sequencing (MP-Seq). Using MP-Seq we have found that HPV is only integrated into the human genome in 30% of HPV-positive OPSCCs, which is dramatically different than what is found in cervical cancer. MP-Seq not only can determine the physical status of HPV in a given sample, but can also ascertain genome-wide changes in any cancer. We describe how MP-Seq could be used in the clinical management of HPV-positive OPSCC patients. We have also explored another whole genome sequencing strategy that has been developed by the Beijing Genomics Institute. They can currently do whole genome sequencing for a total cost (and this includes analysis) of $600 and are developing methodologies to reduce this to $100. I will describe how these types of whole genome sequencing strategies could be used to direct therapies for these patients including facilitating the use of the liquid biopsy to monitor patients during their clinical treatment.

Introduction. Cervical cancer is the fourth most common female cancer worldwide. The prognosis for women with advanced-stage or recurrent cervical cancer remains poor and response to treatment is variable. Standardized management protocols leave little room for individualization. We report on a novel blood-based liquid biopsy for specific PlIC3CA mutations as a clinically useful biomarker in patients with invasive cervical cancer.Methods. One hundred seventeen Hong Kong Chinese women with primary invasive cervical cancer and their pre-treatment plasma samples were investigated. Two PIK3CA mutations, p.E542K and p.E545K were measured in cell free DNA (cfDNA) extracted from plasma using droplet digital PCR. This liquid biopsy of PIK3CA in cervical cancer was correlated to clinico-pathological features to verify the potential of PIK3CA as a clinically useful molecular biomarker for predicting disease prognosis and monitoring for progression.Results. PIK3CA mutations, either p.E542K or p.E545K, were detected in plasma cfDNA from 22.2% of the patients. PIK3CA mutation status was significantly correlated to median tumor size (p < 0.01). PIK3CA mutations detected in the plasma were significantly associated with decreased disease-free survival and overall survival (p < 0.05).Conclusions. As a liquid molecular biopsy, analysis of circulating PIIC3CA mutations shows promise as a way to refine risk stratification of individual patients with cervical cancer, and provides a platform for further research to offer individualized therapy with the purpose of improving outcomes. (C) 2017 Elsevier Inc. All rights reserved.

High-risk human papillomaviruses (HPVs) are known to be associated with different anogenital cancers including cervical, anal, penile, and vaginal cancers. They are also found to be responsible for the dramatic increases in oropharyngeal squamous cell carcinoma (OPSCC) observed in the United States and Europe. The model for how high-risk HPVs induce cancer formation comes from studies of cervical cancer which usually involves integration of the HPV into the human genome and subsequent changes due to induced chromosomal instability. Recent work, discussed here, however suggests that this model may not be completely correct. In addition, we summarize studies now done in OPSCC which demonstrate that the role of HPV in these cancers may be different from that in cervical cancer. Finally, we propose new models for how HPV may be involved in the formation of these 2 cancers.

KEYWORDS: Next generation sequencing (NGS)mate-pair sequencing (MP-Seq)cancer genomesgenomic alterationsliquid biopsy

Human papillomaviruses (HPV) are responsible for the development of almost all cervical cancers. HPV is also found in 85% of anal cancer and in 50% of penile, vulvar, and vaginal cancers, and they are increasingly found in a subset of head and neck cancers, i.e., oropharyngeal squamous cell carcinomas (OPSCC). The model for how HPV causes cancer is derived from several decades of study on cervical cancer, and it is just presumed that this model is not only completely valid for cervical cancer but for all other HPV-driven cancers as well. Next-generation sequencing (NGS) has now provided the necessary tools to characterize genomic alterations in cancer cells and can precisely determine the physical status of HPV in those cells as well. We discuss recent discoveries from different applications of NGS in both cervical cancer and OPSCCs, including whole-genome sequencing and mate-pair NGS. We also discuss what NGS studies have revealed about the different ways that HPV can be involved in cancer formation, specifically comparing cervical cancer and OPSCC. (C) 2017 S. Karger AG, Basel

Common fragile sites (CFS) are chromosome regions that are prone to form gaps or breaks in response to DNA replication stress. They are often found as hotspots for sister chromatid exchanges, deletions, and amplifications in different cancers. Many of the CFS regions are found to span genes whose genomic sequence is greater than 1 Mb, some of which have been demonstrated to function as important tumor suppressors. CFS regions are also hotspots for human papillomavirus (HPV) integrations in cervical cancer. We used mate‐pair sequencing to examine HPV integration events and chromosomal structural variations in 34 oropharyngeal squamous cell carcinoma (OPSCC). We used endpoint PCR and Sanger sequencing to validate each HPV integration event and found HPV integrations preferentially occurred within CFS regions similar to what is observed in cervical cancer. We also found that many of the chromosomal alterations detected also occurred at or near the cytogenetic location of CFSs. Several large genes were also found to be recurrent targets of rearrangements, independent of HPV integrations, including CSMD1 (2.1Mb), LRP1B (1.9Mb), and LARGE1 (0.7Mb). Sanger sequencing revealed that the nucleotide sequences near to identified junction sites contained repetitive and AT‐rich sequences that were shown to have the potential to form stem‐loop DNA secondary structures that might stall DNA replication fork progression during replication stress. This could then cause increased instability in these regions which could lead to cancer development in human cells. Our findings suggest that CFSs and some specific large genes appear to play important roles in OPSCC. © 2016 Wiley Periodicals, Inc.

Background. Although alcohol and tobacco use are known risk factors for development of squamous cell carcinoma in the head and neck, human papillomavirus (HPV) has been increasingly associated with this group of cancers. We describe the case of a married couple who presented with HPV-positive oropharynx squamous cell carcinoma within two months of each other. Methods. Tumor biopsies were positive for p16 and high-risk HPV in both patients. Sanger sequencing showed a nearly identical HPV16 strain in both patients. Both patients received chemoradiation, and one patient also underwent transoral robotic tongue base resection with bilateral neck dissection. Results. Both patients showed no evidence of recurrent disease on follow-up PET imaging. Conclusions. New head and neck symptoms should be promptly evaluated in the partner of a patient with known HPV-positive oropharynx cancer. This case expands the limited current literature on concurrent presentation of HPV-positive oropharynx squamous cell carcinoma in couples.

Although the rates of cervical squamous cell carcinoma have been declining, the rates of cervical adenocarcinoma are increasing in some countries. Outcomes for advanced cervical adenocarcinoma remain poor. Precision mapping of genetic alterations in cervical adenocarcinoma may enable better selection of therapies and deliver improved outcomes when combined with new sequencing diagnostics. We present whole-exome sequencing results from 15 cervical adenocarcinomas and paired normal samples from Hong Kong Chinese women. These data revealed a heterogeneous mutation spectrum and identified several frequently altered genes including FAT1, ARID1A, ERBB2 and PIK3CA. Exome sequencing identified human papillomavirus (HPV) sequences in 13 tumors in which the HPV genome might have integrated into and hence disrupted the functions of certain exons, raising the possibility that HPV integration can alter pathways other than p53 and pRb. Together, these provisionary data suggest the potential for individualized therapies for cervical adenocarcinoma based on genomic information.

DNA viruses are known to be associated with a variety of different cancers. Human papillomaviruses (HPV) are a family of viruses and several of its sub-types are classified as high-risk HPVs as they are found to be associated with the development of a number of different cancers. Almost all cervical cancers appear to be driven by HPV infection and HPV is also found in most cancers of the anus and at least half the cancers of the vulva, penis and vagina, and increasingly found in one sub-type of head and neck cancers namely oropharyngeal squamous cell carcinoma. Our understanding of HPVs role in cancer development comes from extensive studies done on cervical cancer and it has just been assumed that HPV plays an identical role in the development of all other cancers arising in the presence of HPV sequences, although this has not been proven. Most invasive cervical cancers have the HPV genome integrated into one or more sites within the human genome. One powerful tool to examine all the sites of HPV integration in a cancer but that also provides a comprehensive view of genomic alterations in that cancer is the use of next generation sequencing of mate-pair libraries produced from the DNA isolated. We will describe how this powerful technology can provide important information about the genomic organization within an individual cancer genome, and how this has demonstrated that HPVs role in oropharyngeal squamous cell carcinoma is distinct from that in cervical cancer. We will also describe why the sequencing of mate-pair libraries could be a powerful clinical tool for the management of patients with a DNA viral etiology and how this could quickly transform the care of these patients.

Objective Assess human papilloma virus (HPV) transcriptional activity in inverted Schneiderian papillomas (IPs).Study Design Case series with chart review.Setting Academic tertiary care center.Subjects and Methods Retrospective clinicopathologic review of 19 cases of IP in patients undergoing surgical excision from 1995 to 2013 at Mayo Clinic in Rochester, Minnesota. Surgical pathology archival material was histopathologically reviewed using hematoxylin and eosin-stained slides. Formalin-fixed, paraffin-embedded material from each case was evaluated for p16 expression using immunohistochemistry as well as HPV DNA and E6/E7 messenger RNA (mRNA) transcription using polymerase chain reaction (PCR) and in situ hybridization (via RNAscope technology), respectively.Results Eight patients were female (42%), with an average age of 53 years (range, 23-82 years). Three demonstrated malignancy, and 5 subsequently recurred. Average follow-up was 49 months (range, 0-200 months), and 1 patient died from squamous cell carcinoma arising from the IP. RNAscope detected HPV mRNA transcripts exclusively within IP in 100% of cases; however, in 11 patients (58%), less than 1% of cells exhibited transcriptional activity. Only 2 of 19 cases (11%) demonstrated mRNA activity in 50% or more cells. HPV DNA was detected in only 2 specimens by PCR.Conclusions This study reveals wide prevalence but limited transcriptional activity of HPV in IP. No correlation between HPV transcriptional activity and progression, recurrence, or malignant transformation was identified. These data suggest that transcription of HPV may contribute to the pathogenesis of IP, but prospective data are needed to definitively demonstrate this connection. These results also suggest that RNAscope may be more sensitive than PCR in detecting HPV activity in IP.

WWOX is a gene that spans an extremely large chromosomal region. It is derived from within chromosomal band 16q23.2 which is a region with frequent deletions and other alterations in a variety of different cancers. This chromosomal band also contains the FRA16D common fragile site (CFS). CFSs are chromosomal regions found in all individuals which are highly unstable. WWOX has also been demonstrated to function as a tumor suppressor that is involved in the development of many cancers. Two other highly unstable CFSs, FRA3B (3p14.2) and FRA6E (6q26), also span extremely large genes, FHIT and PARK2, respectively, and these two genes are also found to be important tumor suppressors. There are a number of interesting similarities between these three large CFS genes. In spite of the fact that they are derived from some of the most unstable chromosomal regions in the genome, they are found to be highly evolutionarily conserved and the chromosomal region spanning the mouse homologs of both WWOX and FHIT are also CFSs in mice. Many of the other CFSs also span extremely large genes and many of these are very attractive tumor suppressor candidates. WWOX is therefore a member of a very interesting family of very large CFS genes.

Common fragile sites (CFSs) are large chromosomal regions that are hot-spots for alterations especially within cancer cells. The three most frequently expressed CFS regions (FRA3B, FRA16D and FRA6E) contain genes that span extremely large genomic regions (FHIT, WWOX and PARK2, respectively), and these genes were found to function as important tumor suppressors. Many other CFS regions contain extremely large genes that are also targets of alterations in multiple cancers, but none have yet been demonstrated to function as tumor suppressors. The loss of expression of just FHIT or WWOX has been found to be associated with a worse overall clinical outcome. Studies in different cancers have revealed that some cancers have decreased expression of multiple large CFS genes. This loss of expression could have a profound phenotypic effect on these cells. In this review, we will summarize the known large common fragile site genes and discuss their potential relationship to cancer development.

Common fragile sites (CFSs) are large regions with profound genomic instability that often span extremely large genes a number of which have been found to be important tumor suppressors. RNA sequencing previously revealed that there was a group of six large CFS genes which frequently had decreased expression in oropharyngeal squamous cell carcinomas (OPSCCs) and real-time reverse transcriptase polymerase chain reaction experiments validated that these six large CFS genes (PARK2, DLG2, NBEA, CTNNA3, DMD, and FHIT) had decreased expression in most of the tumor samples. In this study, we investigated whether the decreased expression of these genes has any clinical significance in OPSCCs. We analyzed the six CFS large genes in 45 OPSCC patients and found that 27 (60%) of the OPSCC tumors had decreased expression of these six genes. When we correlated the expression of these six genes to each patient's clinical records, for 11 patients who had tumor recurrence, 10 of them had decreased expression of almost all 6 genes. When we divided the patients into two groups, one group with decreased expression of the six genes and the other group with either slight changes or increased expression of the six genes, we found that there is significant difference in the incidence of tumor recurrence between these two groups by Kaplan-Meier plot analysis (P < .05). Our results demonstrated that those OPSCC tumors with decreased expression of this select group of six large CFS genes were much more likely to be associated with tumor recurrence and these genes are potential prognostic markers for predicting tumor recurrence in OPSCC.

BACKGROUND:Inherited polyneuropathies often go undiagnosed. We investigated whole exome sequencing (WES) in utility to identify the genetic causes of diverse forms of inherited polyneuropathies without genetic diagnosis. METHODS:WES was applied to 24 cases from 15 kindreds. These kindreds had earlier unsuccessful candidate gene testing and five probands were initially thought to have acquired neuropathy. We assessed the efficacy of WES in screening 74 known neuropathy genes and 5195 reported pathogenic mutations for hereditary motor and sensory neuropathy, distal hereditary motor neuropathy, hereditary sensory and autonomic neuropathy, complicated hereditary spastic paraplegia, and select hereditary metabolic neuropathies. RESULTS:Pathogenic mutations were identified in five kindreds: (1) ATL1-p.Val253Ile; (2) LITAF-p.Gly112Ser; (3) MFN2-p.Arg94Gln; (4) GARS-p.Ile334Phe; and (5) BSCL2-p.Ser 90Leu. Complexities in establishing inheritance, difficulties in selecting candidate genes and high cost of gene testing all hindered earlier gene discoveries. WES expanded the phenotypic spectrum of the identified known mutations. Possible causal mutations in known genes (SPTLC1, DCTN1, REEP1) were identified in three kindreds. In the remaining seven kindreds, multiple rare or novel variants were identified in novel genes not previously linked with neuropathy. Our results demonstrate an average sequencing depth of 140×, >98% coverage and >10× sequencing depth for 93% (range 89%-96%) of the diverse neuropathy genes and their known mutations. CONCLUSIONS:Diverse inherited neuropathy patients without genetic discovery by candidate gene testing have a favourable probability of receiving a genetic diagnosis by WES. Frequently, atypical phenotypes account for earlier failed candidate approaches, and WES is demonstrated to expand the clinical spectrum of known pathogenic mutations.

The common fragile sites (CFSs) are large regions of profound genomic instability found in all individuals. The frequent deletions and other alterations in these regions in multiple cancers has led to the discovery of a number of extremely large genes contained within these regions and several of the large CFS genes have already been demonstrated to function as tumor suppressors involved in the formation of many different cancers. To study the large CFS genes in oropharyngeal squamous cell carcinoma (OPSCC), we did RNA seq analysis from 11 head and neck cancer patients. This revealed that there are six large CFS genes which consistently had decreased expression in the tumor samples compared to their matched normal tissues. These six genes are PARK2, DLG2, NBEA, CTNNA3, DMD , and FHIT . PARK2 and FHIT are located within the two most frequently expressed CFSs and both have been demonstrated to function as tumor suppressors, while the other large genes are found to have frequent alterations in multiple cancers. Validation experiments using real time PCR indicated that over 60% of OPSCC tumors showed decreased expression for all six genes. Both HPV‐positive and HPV‐negative OPSCCs had similar proportions with loss of expression of these genes. Our results suggest that this selected group of large genes might serve as potential tumor suppressors involved in the development of OPSCCs. Further studies are needed to investigate whether the decreased expression observed is due to genomic instability within the CFS regions or the selection for alterations of specific large CFS genes. © 2014 Wiley Periodicals, Inc.

Background: Human papillomavirus (HPV) is now recognized to be very important in the pathogenesis of oropharyngeal squamous cell carcinoma (OPSCC). It is not clear yet whether the physical status of HPV in OPSCC is similar to what is found in cervical cancer.Study design: We performed genome-wide mate pair next generation sequencing from 20 OPSCCs patients, thirteen of which were positive for HPV16 to determine the HPV physical status and its relationship to HPV oncogene E6 and E7 expression.Results: This high throughput approach detected HPV integration events and also determined the bridged HPV coverage in each sample. Two of the HPV16-positive OPSCCs had HPV integration and one of the HPV16-negative OPSCCs had an HPV26 integration. We mapped the site of integration in the HPV genome in all integration events and the integrations were located at E1, E5, E6 and L2 region respectively. One HPV positive OPSCC had two integration events but also had a very high bridged HPV coverage, while the other two just had HPV integrated into the human genome.Conclusion: Our results are thus different from what is routinely observed in cervical cancer where HPV is almost always integrated into the human genome with loss of episomal HPV sequences. Thus more investigation should be carried out to study how episomal HPV alone can contribute to the development of most OPSCCs. (C) 2013 Elsevier B.V. All rights reserved.

Abstract Human papillomavirus (HPV) appears to be responsible for dramatic increases in the incidence of oropharyngeal squamous cell carcinoma (OPSCC) in the United States in the last two decades, in spite of decreases in the incidence of smoking. It is generally assumed that the role of HPV in the development of OPSCC is identical to the one that it plays in cervical cancer where HPV infection followed by integration into the host genome and the subsequent increased expression of the E6 and E7 oncoproteins leads to increased genomic instability. The common fragile sites (CFSs) are highly unstable chromosomal regions that will be particularly sensitive to increased genomic instability and they are also hot-spots for viral integrations in cervical cancer. The CFS regions span many megabases and many of them also span extremely large genes, a number of which have been demonstrated to function as important tumor suppressors. We therefore examined the expression of very large genes in OPSCC. An analysis of RNAseq data generated on 11 OPSCCs and matched normal oropharyngeal tissue from the same patients revealed that some of the tumors had greatly decreased expression for many of the largest genes. The two genes with the greatest decreases in expression in the tumors as compared to matched normal were the known tumor suppressor PARK2 and the putative tumor suppressor DLG2, but three other large CFS genes: CTNNA3, LRP1B and DMD, and one large gene not yet precisely localized within a CFS region: PDE4D, also had much more decreased expression than any of the other large genes. Real-time RT-PCR was used to measure the expression of these six genes in a much larger number of OPSCC tumor-normal pairs. The expression of these six genes was concordant in most OPSCCs. An individual tumor generally had either decreased expression of these six genes, no changes in the expression of these genes or increased expression of all six genes. When the tumors were grouped with respect to which ones were HPV-positive or HPV-negative we observed that 40% of the HPV-positive OPSCCs had greatly decreased expression (more than 100-fold) for all six genes. Half of the HPV-negative tumors had decreased expression of these six genes, but the amount of decreased expression was much more modest (4-16 fold). Most exciting, however was that 7 of the 12 HPV-positive tumors, and three of the four of the HPV-negative patients that had decreased expression of these genes were from patients who had tumor recurrence. In contrast only one of 18 patients whose tumors did not have decreased expression of the large genes had tumor recurrence. The loss of expression of multiple large genes, many of which are derived from CFS regions, could thus be a powerful predictor for which patients will have tumor recurrence. These results also suggest a linkage between HPV-induced chromosomal instability and dramatic decreases in the expression of multiple CFS genes in some OPSCCs. Citation Format: David I. Smith, Ge Gao, Nicole Tombers, Jan Kasperbauer, Vivian Wang. Human papillomavirus and the inactivation of expression of multiple large common fragile site genes in oropharyngeal squamous cell carcinoma. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 582. doi:10.1158/1538-7445.AM2014-582