Finishing the euchromatic sequence of the human genome

The sequence of the human genome encodes the genetic instructions for human physiology, as well as rich information about human evolution. In 2001, the International Human Genome Sequencing Consortium reported a draft sequence of the euchromatic portion of the human genome. Since then, the international collaboration has worked to convert this draft into a genome sequence with high accuracy and nearly complete coverage. Here, we report the result of this finishing process. The current genome sequence (Build 35) contains 2.85 billion nucleotides interrupted by only 341 gaps. It covers ∼99% of the euchromatic genome and is accurate to an error rate of ∼1 event per 100,000 bases. Many of the remaining euchromatic gaps are associated with segmental duplications and will require focused work with new methods. The near-complete sequence, the first for a vertebrate, greatly improves the precision of biological analyses of the human genome including studies of gene number, birth and death. Notably, the human enome seems to encode only 20,000-25,000 protein-coding genes. The genome sequence reported here should serve as a firm foundation for biomedical research in the decades ahead

31st Annual Meeting and Associated Programs of the Society for Immunotherapy of Cancer (SITC 2016) : part two

Background The immunological escape of tumors represents one of the main ob- stacles to the treatment of malignancies. The blockade of PD-1 or CTLA-4 receptors represented a milestone in the history of immunotherapy. However, immune checkpoint inhibitors seem to be effective in specific cohorts of patients. It has been proposed that their efficacy relies on the presence of an immunological response. Thus, we hypothesized that disruption of the PD-L1/PD-1 axis would synergize with our oncolytic vaccine platform PeptiCRAd. Methods We used murine B16OVA in vivo tumor models and flow cytometry analysis to investigate the immunological background. Results First, we found that high-burden B16OVA tumors were refractory to combination immunotherapy. However, with a more aggressive schedule, tumors with a lower burden were more susceptible to the combination of PeptiCRAd and PD-L1 blockade. The therapy signifi- cantly increased the median survival of mice (Fig. 7). Interestingly, the reduced growth of contralaterally injected B16F10 cells sug- gested the presence of a long lasting immunological memory also against non-targeted antigens. Concerning the functional state of tumor infiltrating lymphocytes (TILs), we found that all the immune therapies would enhance the percentage of activated (PD-1pos TIM- 3neg) T lymphocytes and reduce the amount of exhausted (PD-1pos TIM-3pos) cells compared to placebo. As expected, we found that PeptiCRAd monotherapy could increase the number of antigen spe- cific CD8+ T cells compared to other treatments. However, only the combination with PD-L1 blockade could significantly increase the ra- tio between activated and exhausted pentamer positive cells (p= 0.0058), suggesting that by disrupting the PD-1/PD-L1 axis we could decrease the amount of dysfunctional antigen specific T cells. We ob- served that the anatomical location deeply influenced the state of CD4+ and CD8+ T lymphocytes. In fact, TIM-3 expression was in- creased by 2 fold on TILs compared to splenic and lymphoid T cells. In the CD8+ compartment, the expression of PD-1 on the surface seemed to be restricted to the tumor micro-environment, while CD4 + T cells had a high expression of PD-1 also in lymphoid organs. Interestingly, we found that the levels of PD-1 were significantly higher on CD8+ T cells than on CD4+ T cells into the tumor micro- environment (p < 0.0001). Conclusions In conclusion, we demonstrated that the efficacy of immune check- point inhibitors might be strongly enhanced by their combination with cancer vaccines. PeptiCRAd was able to increase the number of antigen-specific T cells and PD-L1 blockade prevented their exhaus- tion, resulting in long-lasting immunological memory and increased median survival

Why Patients Decline Digital Breast Tomosynthesis? Results From a Patient Survey in an Urban Academic Breast Center.

BACKGROUND: Despite the advantages of reduced callback rates, higher sensitivity, and higher specificity associated with digital breast tomosynthesis (DBT) over traditional full-field digital mammography (FFDM), many patients declined DBT at our urban academic breast center. Most states also do not have mandated insurance coverage for DBT. METHODS: A patient survey was conducted at our breast center from February 2017 to April 2017. All patients were informed regarding the potential benefits of DBT as well as the potential additional charge related to DBT, which depended on the insurance coverage. The survey aimed to examine why the patient declined the DBT option. Reasons included cost, increased radiation risk, anxiety about newer technology, discomfort associated with the exam, lack of education about potential benefits, or patient belief that she will not benefit from DBT. We also inquired if patients would change their opinion about DBT if cost were removed. Patients answered each question by rating their responses on a scale of 1-5, from strongly disagree, disagree, neutral, agree, and strongly agree. RESULTS: Potential additional cost associated with tomosynthesis had the highest influence on patient decision to decline DBT with an average score of 4.68 out of 5. Other factors as described above had less impact on the patient decision with an average score ranging from 1.53-1.72 out of 5. CONCLUSION: Potential out-of-pocket cost for patients remains one of the major obstacles in adoption of DBT as standard of care for breast cancer screening

Tip60 HAT activity mediates APP induced lethality and apoptotic cell death in the CNS of a Drosophila Alzheimer's disease model.

Histone acetylation of chromatin promotes dynamic transcriptional responses in neurons that influence neuroplasticity critical for cognitive ability. It has been demonstrated that Tip60 histone acetyltransferase (HAT) activity is involved in the transcriptional regulation of genes enriched for neuronal function as well as the control of synaptic plasticity. Accordingly, Tip60 has been implicated in the neurodegenerative disorder Alzheimer's disease (AD) via transcriptional regulatory complex formation with the AD linked amyloid precursor protein (APP) intracellular domain (AICD). As such, inappropriate complex formation may contribute to AD-linked neurodegeneration by misregulation of target genes involved in neurogenesis; however, a direct and causative epigenetic based role for Tip60 HAT activity in this process during neuronal development in vivo remains unclear. Here, we demonstrate that nervous system specific loss of Tip60 HAT activity enhances APP mediated lethality and neuronal apoptotic cell death in the central nervous system (CNS) of a transgenic AD fly model while remarkably, overexpression of Tip60 diminishes these defects. Notably, all of these effects are dependent upon the C-terminus of APP that is required for transcriptional regulatory complex formation with Tip60. Importantly, we show that the expression of certain AD linked Tip60 gene targets critical for regulating apoptotic pathways are modified in the presence of APP. Our results are the first to demonstrate a functional interaction between Tip60 and APP in mediating nervous system development and apoptotic neuronal cell death in the CNS of an AD fly model in vivo, and support a novel neuroprotective role for Tip60 HAT activity in AD neurodegenerative pathology

Genetic Testing and Screening Recommendations for Patients with Hereditary Breast Cancer.

Professionals who specialize in breast imaging may be the first to initiate the conversation about genetic counseling with patients who have a diagnosis of premenopausal breast cancer or a strong family history of breast and ovarian cancer. Commercial genetic testing panels have gained popularity and have become more affordable in recent years. Therefore, it is imperative for radiologists to be able to provide counseling and to identify those patients who should be referred for genetic testing. The authors review the process of genetic counseling and the associated screening recommendations for patients at high and moderate risk. Ultimately, genetic test results enable appropriate patient-specific screening, which allows improvement of overall survival by early detection and timely treatment. The authors discuss pretest counseling, which involves the use of various breast cancer risk assessment tools such as the Gail and Tyrer-Cuzick models. The most common high- and moderate-risk gene mutations associated with breast cancer are also reviewed. In addition t

Viability analysis indicates genetic interaction between Tip60 and APP in <i>Drosophila</i>.

<p>The indicated transgene was expressed ubiquitously in the fly using 337-Gal4 driver or pan-neuronally using 179 y-Gal4 driver. The number of F1 progeny that eclosed were counted daily. The percentage of eclosed flies was calculated relative to the wild type control (<i>w¹¹¹⁸</i>). All crosses were carried out in triplicate at 25°C. Overexpression of APP drastically reduced viability to <10% while no effect was observed due to expression of truncated version of APP lacking its C-terminal domain. Overexpression of varying levels of wild type dTip60 (dTip60^WT) also reduced viability in a dose independent manner. However, co-expression of dTip60^WT with APP partially rescued the lethal effects induced by APP expression in a dose dependent manner with the maximum effect observed with high levels of dTip60^WT. In the presence of APP lacking the C-terminus, overexpression of dTip60^WT had similar effects seen in flies that overexpressed dTip60^WT alone.</p

Gene expression changes of dTip60^E431Q misregulated target genes in the different transgenic lines.

a<p>Quantitative RT-PCR analysis was performed for the indicated target genes.</p>b<p>Staged second instar larvae ubiquitously expressing the indicated transgene(s) were used for cDNA preparation. Quantitative RT-PCR reactions were carried out in triplicate and the relative fold change was calculated using the 2−ΔΔCT method using RP49 as control.</p>§<p>Genes that were differentially regulated between flies expressing the Tip60 HAT mutant dTip60^E431Q alone and in conjunction with APP.</p> <p>Gene that were differentially regulated between flies overexpressing dTip60^WT alone or together with APP.</p

Transgenic fly lines used for this study.

a<p>The Tip60 P-element insertion is located on chromosome 3 and the APP P-element insertion is located on chromosome 2.</p>b<p>Indicates where the transgenic fly lines were generated.</p

Generation and characterization of dTip60^E431Q containing APP or APP-dCT double transgenic flies.

<p>The dominant negative HAT defective lines dTip60^E431Q A or dTip60^E431Q B (Lorbeck <i>et al.</i>, 2011) were introduced into an APP or APP dCT background using standard genetic techniques. (<b>A</b>) Histogram depicting qPCR analysis of exogenous levels of dTip60^E431Q in staged F1 second instar larval progeny resulting from a cross between the ubiquitous driver 337 and either dTip60^E431Q (lines A and B), APP; dTip60^E431Q (lines A and B) or APP dCT; dTip60^E431Q (lines A and B). 337-Gal4 crossed to <i>w¹¹¹⁸</i> served as a control. Quantification of the exogenously expressed dTip60^E431Q mRNA levels relative to endogenously expressed dTip60 mRNA was done using the comparative CT method with RP49 as internal control as described in (Lorbeck <i>et al</i>, 2011). Asterisks (*) indicate significant fold change between the lines A and B for each genotype with values of p<0.05; n = 3. Error bars represent standard error of the mean. (<b>B</b>) Semiquantitative RT-PCR analysis of APP or APP dCT expression in the different transgenic lines to confirm APP transgene presence. cDNA was prepared as before from staged second instar larvae ubiquitously expressing dTip60^E431Q with APP or APP dCT (lines A or B in each case) and PCR amplified using primers that flank a 100 bp region in the N-terminal portion of APP. PCR products were visualized using 2% agarose gel containing ethidium bromide. Staged second instar larvae ubiquitously expressing APP or APP dCT were used as controls.</p