Hot Nano-particles in Polar or Paramagnetic Liquids Interact as Monopoles.

When neutral nano-particles are heated or cooled in a polar liquid, they will interact with each other as if they carry an electrostatic charge that is proportional to the temperature difference between the particle and the surrounding fluid. The same should hold for suspensions liquids of asymmetric ferromagnetic particles, in which case the heated nano-particles should behave as magnetic monopoles. However, the analogy with electrostatics/magnetostatics is not complete: heated/cooled nano-particles do not move under the influence of an applied homogeneous field. They should, however, interact as monopoles with each other and should move in inhomogeneous fields.This is the author accepted manuscript. The final version is available from the American Chemical Society via https://doi.org/10.1021/acs.jpcb.6b0184

Microarray-based survey of CpG islands identifies concurrent hyper- and hypomethylation patterns in tissues derived from patients with breast cancer

Maintenance of CpG island methylation in the genome is crucial for cellular homeostasis and this balance is disrupted in cancer. Our rationale was to compare the methylation of CpG islands in tissues (tumor, healthy breast and blood) from patients with breast cancer. We studied 72 genes in 103 samples using microarray hybridization and bisulfite sequencing. We observed tumor specific hyper- or hypomethylation of five genes; COL9A1, MT1A, MT1J, HOXA5 and FLJ45983. A general drop of methylation in COL9A1 was apparent in tumors, when compared with blood and healthy breast tissue. Furthermore, one tumor displayed a complete loss of methylation of all five genes, suggesting overall impairment of methylation. The downstream, evolutionary conserved island of HOXA5 showed hypomethylation in 18 tumors and complete methylation in others. This CpG island also displayed a semimethylated state in the majority of normal breast samples, when compared to complete methylation in blood. Distinct methylation patterns were further seen in MT1J and MT1A, belonging to the metallothionein gene family. The CpG islands of these genes are spaced by 2 kb, which shows selective methylation of two structurally and functionally related genes. The promoters of FLJ45983 and MT1A were methylated above 25% in 18 primary and metastatic tumors. Concurrently, there was also >10% methylation of healthy breast tissue in 11 and 5 samples, respectively. This suggests that the methylation process for the latter two genes takes place already in normal breast cells. Our results also point to a considerable heterogeneity of epigenetic disturbance in breast cancer. This article contains Supplementary Material available at http://www.interscience.wiley.com/jpages/1045-2257/suppmat

High prevalence of somatic PIK3CA and TP53 pathogenic variants in the normal mammary gland tissue of sporadic breast cancer patients revealed by duplex sequencing

The mammary gland undergoes hormonally stimulated cycles of proliferation, lactation, and involution. We hypothesized that these factors increase the mutational burden in glandular tissue and may explain high cancer incidence rate in the general population, and recurrent disease. Hence, we investigated the DNA sequence variants in the normal mammary gland, tumor, and peripheral blood from 52 reportedly sporadic breast cancer patients. Targeted resequencing of 542 cancer-associated genes revealed subclonal somatic pathogenic variants of: PIK3CA, TP53, AKT1, MAP3K1, CDH1, R81, NCOR1, MED12, CBFB, T8X3, and TSHR in the normal mammary gland at considerable allelic frequencies (9 x 10(-2) - 5.2 x 10(-1)), indicating clonal expansion. Further evaluation of the frequently damaged PIK3CA and TP53 genes by ultra-sensitive duplex sequencing demonstrated a diversified picture of multiple low-level subclonal (in 10(-2)-10(-4) alleles) hotspot pathogenic variants. Our results raise a question about the oncogenic potential in non-tumorous mammary gland tissue of breast-conserving surgery patients

Plasma protein changes reflect colorectal cancer development and associated inflammation

Introduction: Colorectal cancer (CRC) is the third most common malignancy and the second leading cause of death worldwide. Efficient non-invasive blood-based biomarkers for CRC early detection and prognosis are urgently needed. Methods: To identify novel potential plasma biomarkers, we applied a proximity extension assay (PEA), an antibody-based proteomics strategy to quantify the abundance of plasma proteins in CRC development and cancer-associated inflammation from few mu L of plasma sample. Results: Among the 690 quantified proteins, levels of 202 plasma proteins were significantly changed in CRC patients compared to age-and-sex-matched healthy subjects. We identified novel protein changes involved in Th17 activity, oncogenic pathways, and cancer-related inflammation with potential implications in the CRC diagnosis. Moreover, the interferon gamma (IFNG), interleukin (IL) 32, and IL17C were identified as associated with the early stages of CRC, whereas lysophosphatidic acid phosphatase type 6 (ACP6), Fms-related tyrosine kinase 4 (FLT4), and MANSC domain-containing protein 1 (MANSC1) were correlated with the late-stages of CRC. Discussion: Further study to characterize the newly identified plasma protein changes from larger cohorts will facilitate the identification of potential novel diagnostic, prognostic biomarkers for CRC

Indications of post-zygotic copy number variation in a region between <i>IL10Rβ</i> and <i>IFNAR1</i>.

<p>Results from eight Illumina genotyping experiments are shown using blood DNA from two pairs of monozygotic twins (panel A for twin 012_01 versus co-twin 012_02, and in panel B for twin 159201 versus co-twin 159202) and two unrelated individuals, where two different tissues were analyzed from each subject (Panels C and D; subjects ML36 and SK58, respectively). Abbreviations BL, PT and UM indicate peripheral blood DNA, primary breast tumor and healthy morphologically normal breast tissue from a patient affected with breast cancer, respectively. Illumina 610 or 660W SNP arrays were used, which also contain so called “intensity only probes” (often with cvni-prefix), only useful for copy number analyses. Therefore, only Log R Ratio (LRR) windows of Illumina experiments are shown here, since the B Allele Frequency (BAF) values are not informative for this type of probes. LRR values below and above zero suggest a deletion or a gain, respectively. The four array probes showing variation between the studied samples are labeled as red dots in yellow fields.</p

Variable length of alleles within hypervariable region showing post-zygotic variation.

<p>Panel <b>A</b> shows post-zygotic mosaicism in healthy and phenotypically concordant monozygotic twin pair 148341/148342, with five alleles observed in twin 148341, and three alleles present in co-twin 148342. Similarly, panel <b>B</b> displays post-zygotic variation in another monozygotic twin pair 004_01/004_02. In total 5 different alleles are shown on this gel and only one of them is overlapping between both twins. Panel <b>C</b> illustrates post-zygotic mosaicism in breast cancer patient SK58. There are three different alleles in DNA from morphologically normal breast tissue (UM), two alleles in blood cells (BL) and three alleles in primary tumor (PT). In panels <b>A</b>, <b>B</b> and <b>C</b>, Taq DNA polymerase was used for initial PCR amplification from genomic DNA, as indicated by suffix “T” in the ID of each plasmid clone. In panel <b>D</b>, Phusion DNA polymerase confirmed post-zygotic mosaicism in monozygotic twin pair 148341/148342, as indicated by suffix “Ph” in the ID of each plasmid clone. The length of inserts in all plasmid clones was estimated after EcoRI digestion releasing the insert, and using 1% agarose gel. BL, PT and UM indicate peripheral blood DNA, primary breast tumor and healthy morphologically normal breast tissue from a patient affected with breast cancer, respectively.</p

Size and distribution of 14 HVR-alleles identified by sequencing and gel electrophoresis.

<p>The •/× indicate that the size of the allele was determined by both agarose gel images and sequencing, whereas filled circles (•) denote that the allele size was estimated from agarose gel images.</p><p>The two most common alleles (HVR1098 and HVR1700) are highlighted in bold and underlined text.</p><p>The sample indicated by a single asterisk (*) are from breast cancer patients. BL, PT and UM indicate peripheral blood DNA, primary breast tumor and healthy morphologically normal breast tissue from a patient affected with breast cancer, respectively.</p><p>The samples indicated by two asterisks (**) are monozygotic twin pairs.</p

Summary of validation of somatic variation in the <i>IFNAR1</i> locus.

<p>Summary of Illumina SNP genotyping, which suggested structural variation within the hypevariable region and results from subsequent confirmation using Sanger sequencing and agarose gel electrophoresis. One (*) and two (**) asterisks after the subject ID indicate patients with breast cancer and pairs of monozygotic twins, respectively. BL, UM and PT stand for DNA from peripheral blood cells, healthy morphologically normal breast tissue from a patient affected with breast cancer and primary breast tumor, respectively. “Seq” indicate that the somatic mosaicism was verified by Sanger sequencing while “Gel” shows that it was confirmed by estimation of allele sizes from agarose gel.</p

Signatures of post-zygotic structural genetic aberrations in the cells of histologically normal breast tissue that can predispose to sporadic breast cancer

Sporadic breast cancer (SBC) is a common disease without robust means of early risk prediction in the population. We studied 282 females with SBC, focusing on copy number aberrations in cancer-free breast tissue (uninvolved margin, UM) outside the primary tumor (PT). In total, 1162 UMs (1-14 per breast) were studied. Comparative analysis between UM(s), PT(s), and blood/skin from the same patient as a control is the core of the study design. We identified 108 patients with at least one aberrant UM, representing 38.3% of cases. Gains in gene copy number were the principal type of mutations in microscopically normal breast cells, suggesting that oncogenic activation of genes via increased gene copy number is a predominant mechanism for initiation of SBC pathogenesis. The gain of ERBB2, with overexpression of HER2 protein, was the most common aberration in normal cells. Five additional growth factor receptor genes (EGFR, FGFR1, IGF1R, LIFR, and NGFR) also showed recurrent gains, and these were occasionally present in combination with the gain of ERBB2. All the aberrations found in the normal breast cells were previously described in cancer literature, suggesting their causative, driving role in pathogenesis of SBC. We demonstrate that analysis of normal cells from cancer patients leads to identification of signatures that may increase risk of SBC and our results could influence the choice of surgical intervention to remove all predisposing cells. Early detection of copy number gains suggesting a predisposition toward cancer development, long before detectable tumors are formed, is a key to the anticipated shift into a preventive paradigm of personalized medicine for breast cancer.De 2 första författarna delar förstaförfattarskapet.</p

Graphical summary of variation in a presumptive regulatory VNTR containing region.

<p>Panel <b>A</b> shows an overview of approximately 2 Mb locus on 21q, around four genes encoding functionally related receptors; <i>IFNAR2</i>, <i>IL10Rβ</i>, <i>IFNAR1</i> and <i>IFNGR2</i>. Panel <b>B</b> is zooming on the position of the hypervariable region (HVR, red box), which is located approximately 4 kb upstream from the transcription start site of the <i>IFNAR1</i> gene and is flanked by CpG-islands (green boxes). The last three and the first three exons of <i>IL10Rβ</i>, and <i>IFNAR1</i>, respectively, are shown as grey boxes. Panel <b>C</b> is showing the size and position of HVR according to the most common allele (HVR1098, see below panel D) in relation to the CpG island. Positions of PCR and sequencing primers used in the analysis of the locus are also displayed. Yellow boxes indicate the position of the non-repetitive anchor 1 (A1) and anchor 2 (A2) sequences, that are immediately flanking the repeated segments and were used for alignments of sequence reads. Panel <b>D</b> shows a summary of eight HVR-alleles from the studied samples, which were identified based on Sanger sequencing results of PCR fragments sub-cloned in plasmids. The displayed alleles are ordered from longest to shortest according to size from anchor 1 (A1) to anchor 2 (A2) sequences. Summary of sizes for all 14 different HVR-alleles is shown in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0067752#pone-0067752-t001" target="_blank">Table 1</a>. Sizes of fragments (in base pairs) are given between non-repetitive A1 and A2 sequences and between primers p1 and p8, which were used for PCR amplification from genomic DNA. Asterisk (*) indicates the most frequent allele (HVR1098), which is in agreement with the reference sequence according to NCBI sequence build 36.3. The allele frequency shown here is taking into account only the nine alleles, where the entire sequence could be unequivocally determined using Sanger sequencing. The most common variation encompasses the variable number of 32 base pair segments; i.e. indel 2, indel 3, indel 4, and indel 5. The latter indel 5 is composed of 6 repeated 32 base pair segments (HVR1066). However, there are also indels containing shorther segments; e.g. indel 1, indel 6 and indel 7. Panel <b>E</b> illustrates the positions of two of the four probes from Illumina beadchips, which are aligned onto the NCBI reference sequence for this locus (top sequence with an asterisk, representing HVR1098). The two probes shown here are from Illumina 610 SNP array; cnvi0010761 (green) and cnvi0010759 (blue). All four Illumina probes from <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0067752#pone-0067752-g001" target="_blank">Figure 1</a>, which were used for initial identification of variation in this region are located within hypervariable region. As shown here for two of these four probes, the probeA sequences (as called by Illumina and used for capturing of genomic DNA on beadchips) are shifted only by two bases. The core 32 bp repeat motif is shown in brackets.</p