Close
today i learned (til)

Possible New Therapies for Deadly Neural Tube Defects!

December 3, 2023

Neural Tube Defects (NTDs) are a type of birth defect that can impact the brain, spine, or spinal cord during the early stages of fetal development (1). Common NTDs that fetuses develop include Spina Bifida, Anencephaly, Encephalocele, and Iniencephaly (5,6). NTDs have a strong global prevalence, impacting approximately 322,000 pregnancies worldwide annually (2). In the United States alone, NTDs occur in approximately 3,000 babies annually, with 1,000 of those affected experiencing Anencephaly (1,3,4). NTDs are particularly challenging to treat early, with no effective treatments, as the defects typically begin developing during the first month of pregnancy, which is when some women may not even know they are pregnant (1). Unfortunately, by the time Neural Tube Defects are correctly and definitively diagnosed, it becomes extremely difficult to effectively treat it. Prior to researching more, I had never heard of Neural Tube Defects! However, being a Biomedical Engineer, I only felt that it was my role and responsibility to contribute at least a little bit of the principles that I have learned to come up with new therapies! Who knows? Maybe, it can help someone one day. So, in this article, I decided to highlight what genes I thought of targeting so far. While this may turn out to be a nothing burger, hopefully it’s at least an interesting read for you all!

Previous literature that I read through established the FOX1, ANKDR6, and WDR4 genes as significant contributors to the development of neural tube defects. Based on this, I came up with two methods by which normal functioning of these genes can be restored, and NTD development can be prevented, both carried out before the 6th week of the first trimester of pregnancy, since that is when the neural tube closes. Each of the genes being targeted plays a significant role in normal transcription regulation/signaling mechanisms that, when disrupted, lead to NTD’s. ANKDR6 mutations affect normal reciprocal mechanisms, causing motor neuron pools to be deactivated, hindering muscle innervation (7). WDR34 mutations affect the PCP signaling pathway, impacting muscle regeneration and neuron orientation (9,10,11); 3). FOX1 gene mutations lead to the formation of abnormal FOX1 proteins, which affects downstream transcriptional regulation of NTF mechanisms (8,12). The rationale for having two methods is that a comparison of both the methods will provide insight into which one is more effective in preventing NTDs over the long-term, especially given the known limitations of synthetic oligonucleotides for gene editing and removal. These aims will be tested on pregnant mice, whose fetuses have mutations in these three genes. 

As you can see below, based on the research that I did, I compiled a few more details for each of my aims. These are works in progress, but using my creative mind, this is what I was able to think of!

Aim 1: Utilize synthetic oligonucleotides with Homology Directed Repair to edit genetic mutations in the FOX1, ANKDR6, and WDR4 genes to prevent the formation of Neural Tube Defects.

  1. Mice will be randomly assigned to treatment, placebo, and control groups. 
  2. For the treatment group, complementary DNA Oligonucleotides to the genes will be created, while those unrelated to the three genes will be created for the control group and none will be created for the placebo group (13).
  3. Synthesize Repair DNA template strands, with Fluorescent Reporter Genes (GFP, BFP, MCherry), using promoters. They will then be ligated/joined with the oligonucleotides (14,20,21).
  4. The oligonucleotides with the fluorescent reporter genes will be successfully delivered to the somatic cells of the fetuses, through lipid nanoparticles (18,19). This will facilitate a double strand break at the ends of each of the three genes, which will promote their deletion, due to the HDR Mechanisms present in the cell (15,16,17).
  5. After enough time has passed, these cells will be harvested, and inserted through a flow cytometer. MFI will be measured, with a higher MFI, correlating to a higher success of gene removal (23).

Aim 2: Utilize Crispr-Cas9, with HDR via a Lipid Nanoparticle delivery system to edit genetic mutations in the FOX1, ANKDR6, and WDR4 genes to prevent the formation of Neural Tube Defects.

  1. Mice will be randomly assigned to treatment and control groups.
  2. For the treatment group, guide RNA strands that correspond to the three genes will be created, while nonspecific guide RNA strands will be created for the control group (16).
  3. Synthesize Repair DNA template strands, with GFP, BFP, MCherry, using promoters (14). The Cas9 proteins, the gRNA and the repair template DNA will be delivered, through lipid nanoparticles (18), to the somatic cells of the fetuses. This will facilitate the breakage of the strands containing these genes, and their substitution with strands that lack these genes and express proteins that induce fluorescence (15,16,17). 
  4. After enough time has passed, these cells will be harvested, and inserted through a flow cytometer. Mean fluorescence intensity will be measured, with a higher MFI, correlating to a higher success of gene removal.

My hope is that my approach of editing mutated versions of these genes in fetus somatic cells and replacing them, can inspire some scientists to pursue this further, and help to prevent NTD development and save lives. The problem is that, right now at least, if the genes were not edited or replaced, children would inevitably not survive the pregnancy, due to the lack of any effective treatments. These types of diseases simply do not have effective cures, and they require out of the box thinking to make any progress. Conventional wisdom does not always work in science! There are limitations, and challenges down the road, however. Edits will not automatically be passed on to future generations, since they do not affect the germline cells, which means that the offspring of someone who has this gene in their genome will unfortunately still suffer the consequences (19). This is a problem that I unfortunately could not find a way to get around in my methods, but hopefully I have provided a good starting point for other researchers to look into! If you made it this far, kudos to you and hopefully this inspires you to look into these horrible diseases as well!

Sources:

  1. Cleveland Clinic Medical Professional. Neural tube defects (NTDs): What They Are, Causes & Prevention. Cleveland Clinic. March 30, 2022. Accessed October 28, 2023. https://my.clevelandclinic.org/health/diseases/22656-neural-tube-defects-ntd. 
  2. Kancherla V. Neural tube defects: a review of global prevalence, causes, and primary prevention. Child’s Nervous System. 2023/07/01 2023;39(7):1703-1710. doi:10.1007/s00381-023-05910-7
  3. Neural Tube Defects. March of Dimes. February 2022. Accessed October 28, 2023. https://www.marchofdimes.org/find-support/topics/planning-baby/neural-tube-defects#:~:text=Screening%20tests%20for%20NTDs%20include,to%2022%20weeks%20of%20pregnancy. 
  4. MedlinePlus [Internet]. Bethesda (MD): National Library of Medicine (US); [updated 2020 Jun 24]. Anencephaly; [updated 2019 Oct 01; reviewed 2018 Jun 01; cited 2023 Oct 29]; [about 5 p.]. Available from: https://medlineplus.gov/genetics/condition/anencephaly/. 
  5. King, Jesse, “Anencephaly”. Embryo Project Encyclopedia ( 2013-02-13 ). ISSN: 1940-5030 https://hdl.handle.net/10776/4215
  6. Amorosi S, D’Armiento M, Calcagno G, et al. FOXN1 homozygous mutation associated with anencephaly and severe neural tube defect in human athymic Nude/SCID fetus. Clinical Genetics. 2008;73(4):380-384. doi:https://doi.org/10.1111/j.1399-0004.2008.00977.x
  7. Rai S, Leydier L, Sharma S, Katwala J, Sahu A. A quest for genetic causes underlying signaling pathways associated with neural tube defects. Review. Frontiers in Pediatrics. 2023-May-22 2023;11 doi:10.3389/fped.2023.1126209 
  8. Greene NDE, Stanier P, Copp AJ. Genetics of human neural tube defects. Human Molecular Genetics. 2009;18(R2). doi:10.1093/hmg/ddp347 
  9. Wu G, Huang X, Hua Y, Mu D. Roles of planar cell polarity pathways in the development of neutral tube defects. Journal of Biomedical Science. 2011;18(1). doi:10.1186/1423-0127-18-66 
  10. Butler MT, Wallingford JB. Planar cell polarity in development and disease. Nature Reviews Molecular Cell Biology. 2017;18(6):375-388. doi:10.1038/nrm.2017.11 
  11. Hou P-S, hAilín D, Vogel T, Hanashima C. Transcription and beyond: Delineating Foxg1 function in cortical development and disorders. Frontiers in Cellular Neuroscience. 2020;14. doi:10.3389/fncel.2020.00035 
  12. Wong L-C, Singh S, Wang H-P, Hsu C-J, Hu S-C, Lee W-T. Foxg1-related syndrome: From clinical to molecular genetics and pathogenic mechanisms. International Journal of Molecular Sciences. 2019;20(17):4176. doi:10.3390/ijms20174176 
  13. Sauer NJ, Mozoruk J, Miller RB, et al. Oligonucleotide‐directed mutagenesis for precision gene editing. Plant Biotechnology Journal. 2015;14(2):496-502. doi:10.1111/pbi.12496 
  14. Li S, Chen L, Peng X, et al. Overview of the reporter genes and reporter Mouse Models. Animal Models and Experimental Medicine. 2018;1(1):29-35. doi:10.1002/ame2.12008 
  15. Smirnikhina SA, Zaynitdinova MI, Sergeeva VA, Lavrov AV. Improving homology-directed repair in genome editing experiments by influencing the cell cycle. International Journal of Molecular Sciences. 2022;23(11):5992. doi:10.3390/ijms23115992 
  16. Nambiar TS, Billon P, Diedenhofen G, et al. Stimulation of CRISPR-mediated homology-directed repair by an engineered rad18 variant. Nature Communications. 2019;10(1). doi:10.1038/s41467-019-11105-z 
  17. Chen J, Li S, He Y, Li J, Xia L. An update on precision genome editing by homology-directed repair in plants. Plant Physiology. 2022;188(4):1780-1794. doi:10.1093/plphys/kiac037 
  18. Givens BE, Naguib YW, Geary SM, Devor EJ, Salem AK. Nanoparticle-based delivery of CRISPR/Cas9 genome-editing therapeutics. The AAPS Journal. 2018;20(6). doi:10.1208/s12248-018-0267-9 
  19. Han JP, Kim M, Choi BS, et al. In vivo delivery of CRISPR-Cas9 using lipid nanoparticles enables antithrombin gene editing for sustainable hemophilia A and B therapy. Science Advances. 2022;8(3). doi:10.1126/sciadv.abj6901 
  20. Falzarano MS, Passarelli C, Ferlini A. Nanoparticle delivery of antisense oligonucleotides and their application in the exon skipping strategy for Duchenne muscular dystrophy. Nucleic Acid Therapeutics. 2014;24(1):87-100. doi:10.1089/nat.2013.0450 
  21. Moss KH, Popova P, Hadrup SR, Astakhova K, Taskova M. Lipid nanoparticles for delivery of therapeutic RNA oligonucleotides. Molecular Pharmaceutics. 2019;16(6):2265-2277. doi:10.1021/acs.molpharmaceut.8b01290 
  22. Blackburn L, Brigden T, Hall A, Johnson E. Somatic genome editing: An overview. PHG Foundation. 2019. Accessed November 18, 2023. https://www.phgfoundation.org/briefing/somatic-genome-editing-overview.
  23. Bahreyni-Toossi MT, Dayyani M, Mahmoudi M, Azimian H. Flow cytometric assessment of DNA double-strand break and repair kinetics in prediction of intrinsic radiosensitivity. Iran J Basic Med Sci. 2022;25(9):1104-1109. doi:10.22038/IJBMS.2022.65178.14350
0Likes
About Author

4th year Biomedical Engineering Student at Georgia Tech, looking to go into the Premed track.

You May Also Like

today i learned (til)
Battle of the Hands: Mittens vs. Gloves
today i learned (til)
The Pot That Keeps on Giving: Perpetual Stew
Articles, Story Time, today i learned (til), vintage science
Vinyl is Better than Digital Music
Articles, today i learned (til)
Alex or Alexa: Gender Norms in Technology

Your journey starts here! Let’s talk!

ThemeRex © 2026. All rights reserved.
  Timothy Durham was convicted of rape when a test showed his DNA genotype matched the DNA recovered from the crime scene. Eye witnesses testified that Durham was in a different state at the time of the incident, but he wasn’t exonerated from his 3000 year sentence until follow up DNA analysis showed that the preliminary forensic analysis used in the trial was misinterpreted (Thomas et al.). DNA testing serves an invaluable part in our criminal justice system, but it comes with complications and bias that can lead to wrongful convictions and misinformed juries.  

Background:

Forensic genetics aims to identify the origin of a biological sample. It can be traced back to Karl Landsteiner who discovered human ABO blood groups. With blood groups, suspects could be eliminated based on their blood type. For example, if a suspect had type A blood and there was type B blood left at the scene by the perpetrator, the investigators could rule out the suspect. Unfortunately, the reverse didn’t work. If the suspect and perpetrator both had the same blood type, it can’t be concluded that the suspect was guilty. Blood identifiers evolved into 8 distinct categories which improved the ability to eliminate suspects, but could only conclude that (if the system matched) it might be the perpetrator or this person might be related to the perpetrator (Jobling and Gill). The field evolved in the 1980s with the discovery of Minisatellites, lines of repetitive DNA that don’t encode proteins. Minisatellite patterns in DNA create what we know today as the DNA fingerprint, and it has a much higher accuracy rate than previous methods (Augustyn et al.). Around the same time, a breakthrough for victims of sexual assault was discovered. In the case of assault, bodily fluids of both the victim and the assaulter can get mixed. A process called differential lysis was created which could selectively increase the concentration of sperm in a fluid sample thereby allowing for accurate DNA testing (Jobling and Gill). Forensics evolved to use single-locus probe testing after several years because of its ease of interpretation. It looked at specific minisatellites that had high variability between subjects. This was the first DNA-based method to be used in a criminal investigation. Two murder homicides from several years apart were connected using DNA evidence. The evidence was then used to exonerate an innocent man, and the guilty party was found when he tried to evade mandatory DNA testing for all men in the small town. His guilt was then reinforced when the DNA profiles matched, and he gave a confession (Jobling and Gill). Finding DNA at a crime scene doesn’t guarantee that you’ll find a murderer. There are issues with non-pure samples and limited samples that make DNA work difficult, and accreditation of facilities and new techniques and take time (Jobling and Gill).  

Subjectivity of DNA Interpretation:

In a 2009 publication, the National Academy of Science discusses the possibility for subjective influence in lab based forensics. The interpretation bias happens in complex cases where, for example, there could be a mixture of DNA. A study published in 2011 looked specifically at this issue. A gang rape happened in Georgia, and the mixture in DNA from the case was used to prosecute one suspect after another suspect took a plea bargain to testify against him. With DNA mixtures, there are three possible conclusions: suspect excluded, suspect cannot be excluded, and inconclusive. If the suspect is excluded, it means that there’s little to no possibility that his DNA is part of the sample. If the suspect cannot be excluded, it means that there’s a possibility that his DNA is part of the sample. In the trial, the DNA examiners who knew the context of the case concluded that the suspect being prosecuted could not be excluded. The 2011 study gave the DNA evidence from this case and the DNA profiles of the suspects to 17 forensic examiners. These examiners had no context of the case, and they all worked independently. Of the 17, only 1 came to the same conclusion (suspect cannot be excluded) as the examiners who knew the context of the case. Twelve of the 17 concluded that the suspect who was prosecuted and convicted had no relation to the DNA found in the sample. Not only does this show that subjectivity occurs due to the context of the case, but the 17 examiners came up with different findings based on their past experiences meaning that not all DNA analysis is objective. Some organizations like the Scientific Working Group on DNA Analysis Methods say that quantitative statistical evidence should be presented with every categorical conclusion, but the US doesn’t require this (Dror and Hampikian).

False Positives:

There are two categories that cause wrongful identification in a DNA report: coincidental matches and false positives. Coincidental matches are DNA matches between two unrelated people. The probability of these matches is about 1 to 500,000 for white individuals and about 1 to 350,000 for black individuals. This small probability of coincidental matches makes people feel confident in DNA evidence. False positives, on the other hand, are matches between DNA and test DNA that result from issues in the forensics lab. There have been multiple cases of forensics labs accidentally switching victim and suspected rapist DNA which lead to convictions that were later overturned when the lab realized the error. There are also cases of misreading or misunderstanding a sample. In a case from 1993, a man was convicted of rape because someone interpreted a mixed sample of victim and perpetrator DNA as an individual sample. The conviction was again overturned. To prevent false convictions, retesting DNA is allowed, but it’s not an efficient solution. Many people are pushing for statistical information on the probability of false positives, so that the jurors can make accurate judgements based on DNA evidence (Thomas et al.).  

Conclusion:

DNA analysis is a useful tool for the criminal justice system, but subjectivity can be an issue in forensic testing. To combat wrongful convictions, there are organizations like The Innocence Project who use DNA testing to free the innocent, but reformed forensic testing standards should be put in place to prevent subjectivity and to ensure juries are correctly informed during trials. https://www.innocenceproject.org/    

Citation:

Augustyn , Adam, et al. “DNA Finerprinting.” Encyclopedia Britannica , 26 Apr. 2018, www.britannica.com/science/DNA-fingerprinting#ref153186. Dror, Itiel E., and Greg Hampikian. “Subjectivity and Bias in Forensic DNA Mixture Interpretation.” Science & Justice, vol. 51, no. 4, 2011, pp. 204–208., doi:10.1016/j.scijus.2011.08.004. Jobling, Mark A., and Peter Gill. “Encoded Evidence: DNA in Forensic Analysis.” Nature Reviews: Genetics, vol. 5, no. 10, Oct. 2004, pp. 739–751., doi:10.1038/nrg1455. Thompson, William C, et al. “How the Probability of a False Positive Affects the Value of DNA Evidence.” Journal of Forensic Sciences, vol. 48, no. 1, Jan. 2003, pp. 1–8., doi:10.1520/jfs2001171.