Could the Cure be Worse Than the Disease? An Ebola Antivirus Study

Kaitlyn Olsen
7th Hour - Kerwin

Abstract: The purpose of this project is to use gene and protein databases to determine if drugs that bind with cholesterol transporter NPC1 protein with the intent of stopping new cellular infection by the Ebola virus might have lethal side effects. Will identify the two most closely related proteins (NPC1 homolog 1 and NPC1 homolog 2) to NPC1 that might also bind with the drug and determine what the possible effects might be. The hypothesis is supported that the drugs will not be lethal but might cause female reproductive harm of some kind.

Keywords: virus, genetics, protein, NPC1, Ebola

Purpose: The purpose of this research is to help decide whether or not a drug designed to block the Ebola virus from infecting cells will be damaging to the patient. I chose to do this study because I want to know whether or not Ebola’s antivirus could be more damaging than the disease itself. The “Ebola virus is an aggressive pathogen that causes a highly lethal hemorrhagic fever syndrome in humans and nonhuman primates” (Sullivan, Yang, and Nabel). Today, only the symptoms of an Ebola infection are treated and the body is left to fight the infection on its own, since there is no cure at the moment (Schoenstadt).

"Not very much is known about the pathogenesis of filoviruses. But we do know that Ebola attacks cells important to the function of lymphatic tissues. It can be found in fibroblastic reticular cells (FRC) among the loose connective tissue under the skin and in the FRC conduit (FRCC) in lymph nodes. This allows the virus to rapidly enter the blood and leads to disruption of lymphocyte homing at high endothelial venules (HEV)” (Waterman).

The Ebola virus uses the Niemann-Pick C1(NPC1) protein to gain entry into the endosomes and causes them to burst, releasing the virus into the cell.

"NPC1 has been well known in the biomedical literature. Primarily associated with cholesterol metabolism, this protein, when mutated, causes a rare genetic disorder in children, Niemann-Pick disease. Using cells derived from these patients, the group found that this mutant form of NPC1 also completely blocks infection by the Ebola virus” (Cooke and Shanks).

Once I have researched all of the information that I need about the NPC1 protein by reading research papers and published articles, I will be able to draw a conclusion about whether or not the antivirus is worse than the disease itself.

So are you doing this in a survey form? (Berry)
What kind of problems does this cause and how do they already cure this now? (Angela Jaszarowski)
Explain more on why you are interested in this and why you chose this topic. (Kylie Ashton)
Explain more on how you are going to do the research and you could maybe try to look at interviews of patients with this disease. (Lauren Martel 1/10/13)

Hypothesis:
If a drug binding the NPC1 protein is given to a person with Ebola, then it will prevent new cellular infection by the Ebola virus without causing patient death.

Materials:
  • Computer with an Internet connection
  • Academic and peer-reviewed medical journals and sources
  • Gene/Protein databases

Procedure: I will begin my ISP by researching information about the Ebola Virus and NPC1 gene in research logs and gene databases and any other source that has reliable information about the topic. Since scientists were actually thinking about using the NPC1 gene to be a antivirus to the Ebola Virus, I will also research why they thought this would be a good idea. After I have researched as much information about each of them as I can, I will draw a conclusion about if the side effects of the antivirus, if any, are worse than the disease.


Independent Variable: NPC1-binding drugs
Dependent Variable: Interference with normal cellular and bodily functions (and degree of compatibility with life)

Background Information:
Ebola Virus
  • Ebola is a virus that is usually fatal in most humans and primates (Questions).
  • It was named after where it was first recognized, the Ebola river in the Democratic Republic of the Congo in Africa (Questions).
  • It was first recognized in 1976 (Questions).
  • Cases of the Ebola Hemorrhagic Fever (Ebola HF) have been reported in the Democratic Republic of the Congo, Gabon, Sudan, the Ivory Coast, Uganda, and the Republic of the Congo. No case of the disease in humans has ever been reported in the United States (Questions).
  • Researchers have hypothesized that the initial patients get infected when they come in contact with an animal carrying the virus (Questions).
  • Most deaths occur within 8-16 days of initial infection (Schaffer).
  • The virus only contains 7 genes but kills up to 90% of the people it infects (Questions).
  • Experts fear that terrorists could use the virus as a weapon (Questions).
  • Its patients usually suffer from fevers, fatigue, seizures, delirium, and bleeding from the eyes, ears, and mouth (Questions).
external image vR2iuisrcUgbNzqQLAGSKRUPY2jUB_kasbPm8a2IzvmmO4Hy8EoyWhIQDHi6Jdtk2tJe7ywUpq_MFXIhUhP5VSv8TmIt2WbMscvyb80FQv5HBF6uvLPq4qZo
*About 25-90% of all cases die. __http://www.cdc.gov/ncidod/dvrd/spb/mnpages/dispages/ebola/ebolatable.htm__ __http://www.nytimes.com/2012/01/17/health/npc1-protein-may-give-ebola-its-opening.html?_r=2__&

Using the NPC1 protein to stop the Ebola virus
  • Researchers developed a mimic of the Ebola virus called rVSV-GP-EboV that’s not deadly but carried one of the same proteins as the Ebola virus. They used it to safely find which cells the virus couldn’t affect (Schaffer).
  • Researchers noticed that when cells without the NPC1 gene (which would not produce the NPC1 protein) were exposed to the ebola virus, they didn’t become infected. They also noticed that these cells were also resistant to the Marburg virus, a virus closely related to Ebola (Schaffer).
  • Researchers determined Ebola enters a cell using the NPC1 protein (Single).
  • They later tested cells in mice with low amounts of NPC1 and recorded that some became sick, but most of them survived (Schaffer).
  • They also did tests to see if an antidepressant, imipramine, used to block the NPC1 would also stop the Ebola virus. It did (Schaffer).

NPC1
  • The NPC1 protein plays a role in the movement of cholesterol and other types of lipids across cell membranes (NPC1).
  • It’s located on the long arm of chromosome 18 (out of 46) (NPC1).
  • Niemann-Pick Disease is a rare degenerative disorder that is caused by a mutation in the gene. The disease causes cells to become clogged with cholesterol and die and lipids to build up in the liver, spleen, and brain (Schaffer).

Research Step 1:
I researched the closest related proteins related to the NPC1 protein (gene sequence included at bottom of page). The two most closest-related proteins are displayed in the “Query Coverage of NPC1 Protein” graph below. If an Ebola antiviral drug were to accidentally attach to one of the Homolog’s endosomes, it could cause unwanted problems within the patient’s body, such as the kidneys shutting down. If this would happen, the cure would be no better than the disease because it would kill the patient.
external image kvTG4JqSLjenGF03EDDJK7zAb1MAM_X90s8YqC13w7WCxL4fiAfdGsmQ5NVDh6zywR1OB5q6whQH1h0JJ8BSCj2EPo0sDgOkleT9CLYieviCkRbcY1q0PVDI
*The query coverage is the percentage of amino acids shared with the NPC1 protein.These are the two most closely-related non target proteins. <http://blast.ncbi.nlm.nih.gov/Blast.cgi>

Research Step 2:
I researched which organs and tissues in the body could be affected if the drug did bind to one of the non-target proteins (instead of NPC1), and concluded that it would most likely affect female reproductive organs and reproductive tissues by using microarry results to find out which tissues and organs would be effected. The graph below is an example of what I looked at on http://amazonia.transcriptome.eu/expression.php?probeId=202679_at#. I looked to see which areas on the graph had the highest bars and areas such as the oocytes (part of the reproductive system) did.

external image -VWtzY7jRnY_rZhl1cTF1nn2EkpphsX5xtsn_SIlFVlmz8rAY-wfipU0XD-l-FFcvJVGP1r0MJH7jMfBSAT0KZAJal6FNe9uyUpOI_OEF4XGi4vlFr0_By86
*The organs and tissues that would be affected if a drug was created to stop the Ebola virus by binding with the NPC1 protein. __http://amazonia.transcriptome.eu/expression.php?geneId=Hs.715623__#


For interpretive benefit, the graph above color-codes each microarray signal by the part of body. The colors are defined below.
external image AVjdQjCZ8LA3RWU-czT4QgJTnqSwame8euwOzjKbZxAi41SKn_tQRXhaPKL3jeU10b5z75x5ClOga5GB9yZEiHnOgusJDQj7UVb71mh5EqLV-QDmC8MnEUmf
*Color wheel for information in the microarray graph above.
<http://amazonia.transcriptome.eu/expression.php?probeId=202679_at>

Analysis:
  • According to the information I have found, the Homolog 1 and Homolog 2 proteins are the most closely related proteins to the NPC1 protein. If the drug were to block either of these proteins, it would most likely harm the reproductive organs and reproductive tissues of female patients. For both male and female patients, there is a small chance that it could affect any part of the body.

Conclusion: The hypothesis that a drug binding the NPC1 protein is given to a person with Ebola will prevent new cellular infection by the Ebola virus without causing patient death, is correct and accepted. The patient will not die because of the drug, however, it is possible that female patients may have reproductive issues and/or sterility.


Limitations:
  • This bioinformatics research is limited to the two proteins most closely related to the NPC1 protein. A more comprehensive search that looks at more proteins should be executed.
  • Bioinformatics research is theoretical. In order to know what side effects a drug designed to block the Ebola virus would have and if it would ultimately be successful in preventing new cellular infection by the Ebola virus, it would need pharmaceutical testing.
  • Research is still being tested by scientists, so some information in this project may be proven incorrect.
  • The hypothesis would not change given these limitations; however, a considerably more thorough and lengthy study is required to provide the necessary scientific confidence in the results.

Additional Information:
Gene sequence for NPC1 Niemann-Pick disease, type C1 [ Homo sapiens ]
Gene ID: 4864, updated on 3-Mar-2013

1 mtarglalgl lllllcpaqv fsqscvwyge cgiaygdkry nceysgppkp lpkdgydlvq
61 elcpgfffgn vslccdvrql qtlkdnlqlp lqflsrcpsc fynllnlfce ltcsprqsqf
121 lnvtatedyv dpvtnqtktn vkelqyyvgq sfanamynac rdveapssnd kalgllcgkd
181 adacnatnwi eymfnkdngq apftitpvfs dfpvhgmepm nnatkgcdes vdevtapcsc
241 qdcsivcgpk pqpppppapw tilgldamyv imwitymafl lvffgaffav wcyrkryfvs
301 eytpidsnia fsvnasdkge asccdpvsaa fegclrrlft rwgsfcvrnp gcviffslvf
361 itacssglvf vrvttnpvdl wsapssqarl ekeyfdqhfg pffrteqlii rapltdkhiy
421 qpypsgadvp fgppldiqil hqvldlqiai enitasydne tvtlqdicla plspyntnct
481 ilsvlnyfqn shsvldhkkg ddffvyadyh thflycvrap aslndtsllh dpclgtfggp
541 vfpwlvlggy ddqnynnata lvitfpvnny yndteklqra qawekefinf vknyknpnlt
601 isftaersie delnresdsd vftvvisyai mflyislalg hmkscrrllv dskvslgiag
661 ilivlssvac slgvfsyigl pltlivievi pflvlavgvd nifilvqayq rderlqgetl
721 dqqlgrvlge vapsmflssf setvafflga lsvmpavhtf slfaglavfi dfllqitcfv
781 sllgldikrq eknrldifcc vrgaedgtsv qasesclfrf fknsysplll kdwmrpivia
841 ifvgvlsfsi avlnkvdigl dqslsmpdds ymvdyfksis qylhagppvy fvleeghdyt
901 sskgqnmvcg gmgcnndslv qqifnaaqld nytrigfaps swiddyfdwv kpqssccrvd
961 nitdqfcnas vvdpacvrcr pltpegkqrp qggdfmrflp mflsdnpnpk cgkgghaays
1021 savnillghg trvgatyfmt yhtvlqtsad fidalkkarl iasnvtetmg ingsayrvfp
1081 ysvfyvfyeq yltiiddtif nlgvslgaif lvtmvllgce lwsavimcat iamvlvnmfg
1141 vmwlwgisln avslvnlvms cgisvefcsh itraftvsmk gsrveraeea lahmgssvfs
1201 gitltkfggi vvlafaksqi fqifyfrmyl amvllgathg liflpvllsy igpsvnkaks
1261 cateerykgt ererllnf

Works Cited:
  1. "AmaZonia! - Probe Page - 202679_at." AmaZonia! - Probe Page - 202679_at. N.p., 11 Mar. 2013. Web. <http://amazonia.transcriptome.eu/expression.php?probeId=202679_at>
  2. "Basic Local Alignment Search Tool." BLAST:. N.p., n.d. Web. 09 Mar. 2013.
  3. Cooke, Robert, and Lori Shanks. "Protein Essential for Ebola Virus Infection Is a Promising Target." Protein Essential for Ebola Virus Infection Is a Promising Target. Harvard Medical School, 26 Aug. 2011. Web. 29 Nov. 2012. <http://hms.harvard.edu/content/protein-essential-ebola-virus-infection-promising-target>
  4. "Genes and Mapped Phenotypes." National Center for Biotechnology Information. U.S. National Library of Medicine, 3 Mar. 2013. Web. 09 Mar. 2013.
  5. "NPC1." NPC1. Genetics Home Reference, 4 Mar. 2013. Web. <http://www.ncbi.nlm.nih.gov/gene/4864>
  6. "Questions and Answers about Ebola Hemorrhagic Fever." Centers for Disease Control and Prevention. Centers for Disease Control and Prevention, 31 July 2012. Web. <http://www.cdc.gov/ncidod/dvrd/spb/mnpages/dispages/ebola/qa.htm>
  7. Schaffer, Amanda. "Key Protein May Give Ebola Virus Its Opening." The New York Times. N.p., 16 Jan. 2012. Web.
  8. Schoenstadt, Arthur. "Ebola Information." Ebola Information. N.p., 12 Oct. 2012. Web. 29 Nov. 2012. <http://ebola.emedtv.com/ebola/ebola-information.html>.
  9. Sullivan, Nancy, Zhi-Yong Yang, and Garry J. Nabel. "Ebola Virus Pathogenesis: Implications for Vaccines and Therapies." N.p., Sept. 2003. Web. 28 Nov. 2012. <http://jvi.asm.org/content/77/18/9733.full.pdf>.
  10. Waterman, Tara. "Ebola Tissue Tropism and Pathogenesis." Ebola Tissue Tropism and Pathogenesis. N.p., 1999. Web. 29 Nov. 2012. <http://virus.stanford.edu/filo/trop.html>.