Non-invasive prenatal testing (NIPT): limitations on the way to become diagnosis

Abstract

With the discovery of existing circulating cellfree fetal DNA (ccffDNA) in maternal plasma and the advent of next-generation sequencing (NGS) technology, there is substantial hope that prenatal diagnosis will become a predominately non-invasive process in the future. At the moment, non-invasive prenatal testing (NIPT) is available for high-risk pregnancies with significant better sensitivity and specificity than the other existing non-invasive methods (biochemical and ultrasonographical). Mainly it is performed by NGS methods in a few commercial labs worldwide. However, it is expected that many other labs will offer analogous services in the future in this fastgrowing field with a multiplicity of in-house methods (e.g., epigenetic, etc.). Due to various limitations of the available methods and technologies that are explained in detail in this manuscript, NIPT has not become diagnostic yet and women may still need to undergo risky invasive procedures to verify a positive finding or to secure (or even expand) a negative one. Efforts have already started to make the NIPT technologies more accurate (even at the level of a complete fetal genome) and cheaper and thus more affordable, in order to become diagnostic screening tests for all pregnancies in the near future.

Quality assurance and standardization in view of non-invasive prenatal testing (NIPT)

“…Although NIPT is not regarded as a genetic "test", but as "screening" [2,11], the whole process of isolating and analysing the cffDNA should be carried out in a human molecular genetics laboratory. Therefore, the requirements outlined above apply to NIPT [1,2] (Figure 1).…”

“…This is done by a straight forward determination of fetal DNA in the sample [4]. During analysis, the maternal DNA is still present in the same tube as the cffDNA [2]. Thus, it is essential for further analysis to implement strategies to measure the quantity of the fetal DNA [4,22].…”

“…The most common method to analyze cffDNA is next-generation sequencing (NGS) [2]. For the implementation of NGS in diagnostics, the need for quality standards was noted years ago and guidelines have already been published [25,26].…”

“…Subsequently, an estimation of the fetal set of chromosomes with regards to the current tested trisomies (13,18,21) [1,2] could be given. It is superfluous to emphasize the importance of validation for this purpose.…”

“…With the appearance of cell-free fetal DNA analysis, the interpretation and implementation of quality assurance parameters such as quality controls and validation has to be made by a molecular genetics laboratory. To harmonize and improve the quality of new methods, best practice guidelines have to be developed over time [1][2][3][4].…”