Utilization of Benchtop Next Generation Sequencing Platforms Ion Torrent PGM and MiSeq in Noninvasive Prenatal Testing for Chromosome 21 Trisomy and Testing of Impact of In Silico and Physical Size Selection on Its Analytical Performance

OBJECTIVES: The aims of this study were to test the utility of benchtop NGS platforms for NIPT for trisomy 21 using previously published z score calculation methods and to optimize the sample preparation and data analysis with use of in silico and physical size selection methods. METHODS: Samples from 130 pregnant women were analyzed by whole genome sequencing on benchtop NGS systems Ion Torrent PGM and MiSeq. The targeted yield of 3 million raw reads on each platform was used for z score calculation. The impact of in silico and physical size selection on analytical performance of the test was studied. RESULTS: Using a z score value of 3 as the cut-off, 98.11% - 100% (104-106/106) specificity and 100% (24/24) sensitivity and 99.06% - 100% (105-106/106) specificity and 100% (24/24) sensitivity were observed for Ion Torrent PGM and MiSeq, respectively. After in silico based size selection both platforms reached 100% specificity and sensitivity. Following the physical size selection z scores of tested trisomic samples increased significantly-p = 0.0141 and p = 0.025 for Ion Torrent PGM and MiSeq, respectively. CONCLUSIONS: Noninvasive prenatal testing for chromosome 21 trisomy with the utilization of benchtop NGS systems led to results equivalent to previously published studies performed on high-to-ultrahigh throughput NGS systems. The in silico size selection led to higher specificity of the test. Physical size selection performed on isolated DNA led to significant increase in z scores. The observed results could represent a basis for increasing of cost effectiveness of the test and thus help with its penetration worldwide

The application of circulating tumor cell and cell-free DNA liquid biopsies in ovarian cancer

Ovarian cancer is the deadliest gynecological cancer. 70% of the cases are diagnosed at late stages with already developed metastases due to the absence of easily noticeable symptoms. Early-stage ovarian cancer has a good prognosis with a 5-year survival rate reaching 95%, hence the identification of effective biomarkers for early diagnosis is important. Advances in liquid biopsy-based methods can have a significant impact not just on the development of an efficient screening strategy, but also in clinical decision-making with additional molecular profiling and genetic alterations linked to therapy resistance. Despite the well-known advantages of liquid bi-opsy, there are still challenges that need to be addressed before its routine use in clinical practice. Various liquid biopsy-based biomarkers have been investigated in ovarian cancer; however, in this review, we are concentrating on the current use of cell-free DNA (cfDNA) and circulating tumor cells (CTCs) in disease management, focusing on their emerging importance in clinical practice. We also discuss the technical aspects of these workflows. The analysis of cfDNA is often chosen for the detection of mutations, copy number aberrations, and DNA methylation changes, whereas CTC analysis provides a unique opportunity to study whole cells, thus allowing DNA, RNA, and protein-based molecular profiling as well as in vivo studies. Combined solutions which merge the strengths of cfDNA and CTC approaches should be developed to maximize the potential of liquid biopsy technology

Rare and unique adaptations to cancer in domesticated species : an untapped resource?

Strong and ongoing artificial selection in domestic animals has resulted in amazing phenotypic responses that benefit humans, but often at a cost to an animal's health, and problems related to inbreeding depression, including a higher incidence of cancer. Despite high rates of cancer in domesticated species, little attention has been devoted to exploring the hypothesis that persistent artificial selection may also favour the evolution of compensatory anticancer defences. Indeed, there is evidence for effective anti-cancer defences found in several domesticated species associated with different cancer types. We also suggest that artificial selection can favour the “domestication” of inherited oncogenic mutations in rare instances, retaining those associated to late and/or less aggressive cancers, and that by studying these seemingly rare anticancer adaptations, novel cancer treatments may be found

Rare and unique adaptations to cancer in domesticated species: An untapped resource?

Strong and ongoing artificial selection in domestic animals has resulted in amazing phenotypic responses that benefit humans, but often at a cost to an animal's health, and problems related to inbreeding depression, including a higher incidence of cancer. Despite high rates of cancer in domesticated species, little attention has been devoted to exploring the hypothesis that persistent artificial selection may also favour the evolution of compensatory anticancer defences. Indeed, there is evidence for effective anti-cancer defences found in several domesticated species associated with different cancer types. We also suggest that artificial selection can favour the "domestication" of inherited oncogenic mutations in rare instances, retaining those associated to late and/or less aggressive cancers, and that by studying these seemingly rare anticancer adaptations, novel cancer treatments may be found

Cancer Susceptibility as a Cost of Reproduction and Contributor to Life History Evolution

International audienceReproduction is one of the most energetically demanding life-history stages. As a result, breeding individuals often experience trade-offs, where energy is diverted away from maintenance (cell repair, immune function) toward reproduction. While it is increasingly acknowledged that oncogenic processes are omnipresent, evolving and opportunistic entities in the bodies of metazoans, the associations among reproductive activities, energy expenditure, and the dynamics of malignant cells have rarely been studied. Here, we review the diverse ways in which age-specific reproductive performance (e.g., reproductive aging patterns) and cancer risks throughout the life course may be linked via trade-offs or other mechanisms, as well as discuss situations where tradeoffs may not exist. We argue that the interactions between host-oncogenic processes should play a significant role in life-history theory, and suggest some avenues for future research

Z score values of trisomic and euploid samples before and after <i>in silico</i> size selection of sequencing reads.

<p><b>□</b> –trisomic samples, <b>○</b> –euploid samples, horizontal line—mean z score value). Dotted lines represent the standard limit for identification of a trisomic sample (z score = 3). A—Ion Torrent PGM analyzed samples, B—MiSeq analyzed samples.</p

Z score values of samples calculated by three different previously published methods.

<p><b>□</b> –trisomic samples, <b>○</b> –euploid samples, horizontal line—mean z score value of trisomic samples. Dotted lines represent the standard limit for identification of a trisomic sample (z score = 3). A—Ion Torrent PGM analyzed samples, B—MiSeq analyzed samples.</p

Determination of optimal <i>in silico</i> size selection limit for sequencing reads to be used in z score calculation.

<p>Dotted lines represent the standard limit for identification of a trisomic sample (z score = 3). A—Ion Torrent PGM analyzed samples, B—MiSeq analyzed samples.</p

Z score values calculated from reads without size selection (all), after <i>in silico</i> size selection (IS) and after physical size selection (P).

<p>Horizontal lines represent mean z score value calculated from 3, 2 and 1 million raw reads (3m, 2m, 1m). Dotted lines represent the standard limit for identification of a trisomic sample (z score = 3). A—Ion Torrent PGM analyzed samples, B—MiSeq analyzed samples.</p