The document discusses the challenges of implementing next-generation sequencing (NGS) in newborn screening (NBS), emphasizing the ethical, psychological, and economic implications. Although NGS holds promise for expanding genetic screening, concerns regarding reliability, informed consent, and the potential for non-actionable information raise significant issues. Key challenges include high costs, the need for extensive bioinformatics support, and the ethical responsibility to ensure that screening results positively impact newborns without causing harm.

1. Challenges of using Next
Generation Sequencing for
Newborn Screening
Dr.Amir Abdelazim Ahmed
amiral6666@gmail.com

2. Introduction
For 50 years, newborn screening (NBS) programs in the worldwide have
successfully identified disease in pre-symptomatic babies using blood
samples collected on filter paper (dried blood spots, DBS) shortly after
birth
We celebrate daily as the national program of mandatory newborn
screening (NBS) has been a huge success. It has virtually eliminated the
tragedy of intellectual disability from many disorders . It has eliminated
death or profound neurologic sequelae also.
https://www.hrsa.gov/advisory-committees/heritable-disorders/rusp/index.html

3. Ethical principles for screening programs set out by Wilson
and Jungner in Principles and Practice of Screening for Disease
(1968)
• if the targeted condition is an important health problem, whose natural
history is well-understood, and whose symptoms are amenable to early
intervention and effective treatment. Accordingly, the mere availability of
a reliable test would not justify routine screening for a condition, unless
such screening could be shown to provide direct medical benefit to those
who test positive for the condition.
Introduction
New era in NBS, including several conditions which do not meet traditional screening criteria .

4. • If this can be accomplished for the metabolic and
endocrine disorders, could there be even greater benefit
from NBS for genetic disorders in general, including non-
metabolic genetic disorders?
• True, we do not have preventive therapies for
chromosomal aberrations or most other genetic
abnormalities but there could be many other benefits
from neonatal diagnosis, such as information for the
family to prepare for progressive disability in the child,
for genetic counseling for family planning, for prenatal
or preconceptual diagnosis in future pregnancies, for
prevention of needless and expensive diagnostic
odysseys in the child, and still other potential benefits
(Landau et al. 2014).
Introduction

5. • In recent years, next-generation sequencing (NGS) technology has
greatly reduced the cost and effort needed to obtain accurate DNA
sequence data from exomes, and even the whole human genome, in
one massively parallel process. This technology is capable of quickly
allowing the assembly of a single multi-gene panel relevant to
newborn screening.
Introduction

6. Newborn screening
Genetic Newborn
screening???

7. Should NBS be expanded into genetic screening?
• The sequencing is unlikely to be totally reliable and errors will almost
certainly occur (Zhu and Xiong 2012).
• Depending on the sequencing platform used, some variations will be
missed (Clark et al. 2011)
• Interpretation will vary among screening laboratories, one program assigning a
variation as a pathological mutation and another program considering the
variation inconsequential. (Cooper and Shendure 2011)
A major concern is the possibility that some parents might opt out of NBS entirely from
their general opposition to DNA examination or fear that the detection of genetic
variations in their newborn will jeopardize obtaining health insurance or life insurance, or
even school acceptance and future employment (Landau et al. 2014).
Genetic NBS will almost certainly require informed consent.

8. Challenges of
NGS for NBS
Psychological outcomes
Ethical outcomes
Economic
considerations
Medical and technical
considerations
Non-actionable
information
Time considerations
We need
database
We need
high
throughput

9. Medical and technical
considerations
• Next generation sequencing (NGS) encompasses several technologies
of rapid, high yield, parallel DNA sequencing.
• WES examines only a small part of the genome with high coverage of
the coding regions but is superior to WGS in finding DNA changes of
known and important clinical significance. Because WES requires
capture and enrichment of the exome, it may fail to capture certain
exons and thus has the potential for false negative results in coding
regions
WGS has an important advantage over WES. While it also covers the exome, it
identifies not only variations in the coding regions but also sequence changes in
non-coding regions that may modify gene expression, thus increasing the likelihood
of establishing a genetic diagnosis.
too much
information

10. Non-actionable information
• For example ::: when parents can get infant screening results that
suggest, for example, a 10% increase in the chance of being
diagnosed with a certain disease, such as autism or Alzheimer’s
disease, then the question is what will be the psychological economic
and overarching ethical consequences of this non-actionable
information. Thus, the question remains in the clinical genetic testing
arena,
• how much information is too much information? For the parents? For
the children? For the genetic professional?

11. Medical and technical
considerations

12. Psychological outcomes
• NGS could detect genetic variations in genes encoding high-
penetrant, adult-onset disorders. These include cancer (e.g. breast,
ovarian and colon), neurological disorders (e.g. CADASIL syndrome),
connective tissue (e.g. Marfan syndrome), arrhythmias and many
others.
The potential ethical and practical problems of dealing with this knowledge will need to be very
seriously considered. Since the parents have the ethical and legal authority to make medical
decisions on behalf of their newborn, they are expected to make decisions that are in the best
interests of their children.
Testing newborns for late-onset genetic diseases denies newborns the option of deciding
about testing later in life, thus denying them future adult autonomy and confidentiality.
potential difficulties in their future ability to form relationships.
mental health issues for parents who receive the information of their newborn
being a carrier of a serious, untreatable disease.

13. Increased knowledge is not an absolute good:
• parents may feel guilty for passing on harmful mutations to their
children or stigmatized as having the potential to do so.
as the child or their parents may be required to disclose the information in various
circumstances, it may cause discrimination against the child in several circles of life
including education, employment and medical insurance

14. Ethical outcomes
•Do we support genetic determinism in which all
information is present at the beginning with a clear
path and destiny and no ultimate freedom of choice?
Thus, in choosing NGS newborn screening and
administering this test early on, we will be breaking the
commitment to free choice (Chadwick, 2011; Rosoff, 2012;
Botkin et al., 2014; Gannet, 2014).

15. Time considerations
•Compared to the typical short turnaround time of
metabolic screening (a few days), the time for
receiving results of WES and WGS can be weeks.
A major challenge would be the bioinformatics required for analyzing
the huge amount of data generated, especially when dealing with rare
or novel genetic changes. Even if we focus only on known disease-
related genes, the expected number of variants with incomplete
knowledge regarding their clinical significance that would be detected
carries a major interpretive challenge.

16. Economic considerations
• the cost of genomic sequencing is much too high for application to
newborn screening but is rapidly decreasing and, in the near future,
may approach the price at which it could be used for newborn
screening.
Additional family testing and followup that would likely be required, at least in some cases, could add
substantial additional costs to the already high cost of NGS (Sboner et al., 2011; Beckmann 2015).
The cost of delivery of this sensitive information to the newborn’s parents,
pre- and post-testing, is also a major factor in the decision to implement NGS
screening to newborns.
proper counselling of the parents will be a lengthy, difficult and expensive
process as many more genetic personnel will have to be recruited for this
purpose alone (Ormond et al., 2010; Feero, 2013).

17. Potential role of NGS for NBS
• Primary test for current disorders
• Primary test for disorders where there is no suitable biochemical test
available
• Use as 2nd tier test
• Use as 3rd tier test
• Obtain a clearer picture of disease severity ,
• expected onset (earlly or late) and treatment regimen

18. We need
database
• For disease included
• Development of geneotype-phenotype
database
• Important clinical resource
• Enhance disease understanding

19. We need high throughput
• Compare DNA extracted from Venous blood
and DNA extracted from Dried blood spot
(DBS)
• Comparing same sequence quality from DBS
DNA and from VB DNA
• Many studies found that
• DNA extracted from DBS sufficient quality and quantity also it found that
no significant loss of variant information between VB and DBS
• Method used for DBS extraction cost effective and can be automated

20. NGS is not likely to fit within available public healthcare budgets
WGS and WES also have the potential to provide significant information that does not directly benefit newborns
screen for targets that do not meet traditional screening criteria, prompting calls for more evidence-based decision
making. One example is screening for lysosomal storage disorders, including Krabbe disease,
Currently, laboratories cannot interpret most WGS-generated data, as it remains quite difficult to understand the
significance of many variants in the coding regions, which make up only 2% of the genome. The noncoding
regions are vast and present an even greater challenge.
NGS testing would require a significant overhaul of equipment and personnel. Few laboratories in the U.S. are
currently capable of performing NGS testing at scale
Expanded NGS also would pose another important issue— consent
NGS-NBS likely will be implemented at some point in the future, whether it replaces traditional NBS or serves as a
technological supplement for screening disorders within the existing framework. However, the technology is well
ahead of policy, and NGS likely will exacerbate current NBS challenges. These include costs, standards for which
genes to include and which variants to report, infrastructure, and education of the medical community.

22. lab workflow
60 min to punch 96
well (6mm punch)
30 min for 96 well
using Biomek FX robot
program
Ampliseq Library pre.
On Biomek FX robot
manually to uniform
coverage
Ion Chef (11h
overnight )
Ion S5 : 2.5 h run
time per chipc
Need high throughput
bioinformatics analysis
NBS currently takes place at centralized state laboratories, under standardized conditions
using methods that measure enzyme activity, metabolites, or other molecules. DNA testing is
not a routine part of the process, with only a few babies in some states receiving a DNA test as
a second-tier follow-up on abnormal results, such as for CFTR genotyping after a failed or
inconclusive screen for cystic fibrosis by immunoreactive trypsinogen on dried blood spots

23. Example of current timeline for lab receive 2
plate(96) per day about 1000 per week
• Doubling up on automation equipment would increase sample high throughput capability

24. Example from
New York
For 3rd tier in
cystic fibrosis

25. To summarize
while availability and costs have made NGS a potential attractive mode
of screening, its implementation as a general medical practice for
newborn screening is still premature.
We are not ready for instituting population-based newborn screening
because adequate detection rates are compromised by the
heterogeneity of DNA changes found and the clinical course of the
disease remains difficult to predict in many cases. We have to make
sure, beyond all doubt, that knowledge of the screening results will
benefit the newborns and is free of potential harmful consequences
later in life or of detrimental impacts on child–parent–society
relationships.