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Cori, Ryan, & Shaan - did you see this? :)
Thanks,
Gwen Stovall
Nucleic Acid Aptamer Selection Against Endoglin (CD105) for Marker of Overproduction of Endothelial Cells
Here is the link to my final manuscript: https://docs.google.com/document/d/1DRo2m--Hr9V4lYq8MnboE8rvMq-Y5Hyni1zN0zEZ2GU/edit
Proposal: https://docs.google.com/document/d/1kBQJ-g00D5QQ3sC3XXkGz-pPoDAdWXKPS34uYDIXR3A/edit?hl=en_US
Progress report 10/18/11: http://aptamerstream.blogspot.com/2011/10/r50-selection-against-cd105endoglin.html#links
Elena Mora
Figure 1: The role of endoglin in (TGF)-β adapted from Seon, Ben. (Seon Labs)
Cell-surface glycoprotein endoglin (CD105) has been identified as a promising marker for proliferation of human endothelial cells. These tumor-associated cells are optimal targets in therapeutic approaches for human cancer and identifying over-expressed cells (1). They may lead to a deeper understanding in angiogenesis and the use of anti-CD105 as a therapeutic agent (2). Aptamers also would be much more affordable than the antibody therapeutics currently available.
Specific Aim: Several components of the receptor complex Transforming Growth Factor (TGF)-β superfamily, and CD105, are shown to strongly bind together in these cells (3). Studies have also shown that inhibition of CD105 expression enhanced the TGF- β ability to suppress growth of cultured endothelial cells (3). In this respect, an aptamer could possibly serve a likewise function to suppress growth and spreading of malignant tumors.
An aptamer selected against CD105 would be useful in diagnosis and treatment of cancer tumors. Identification with this marker and possibly inhibiting the growth of these destructive cells could potentially be a powerful step towards more curative measures.
Elena Mora
Raybiotech_Catalog: 228-10388-2, Size: 10µg, Price: $ 189.00
RNA Selection Against Burkholderia pseudomallei (BPM) to Accelerate Burkholderia Diagnostics and Aid in Treating Melioidosis
By: Arsany Gadallah
Melioidosis is a commonly misdiagnosed infection that can be presented as a fatal case of pneumonia or sepsis. The presence of melioidosis has been proven to be related to rainfall and extreme weather cases such as in the tropics (Baker and Tahani D, 2011). It has been spreading across the globe from northern Australia, several African areas and Asian countries to neighboring countries within the Americas at a fast rate. The causative agent of melioidosis is Burkholderia pseudomallei (BPM), which is a motile soil-borne bacterium that infects humans and animals, inducing symptoms such as chest pain and engendering cases of sepsis (Cheng and Currie, 2005). The disease has a mortality rate of 20-68% (White NJ and Dance DA, 1989), ranging from acute cases to severe and blood-borne cases. Both forms of the disease, acute and chronic forms, have called for attention from public health institutions, pointing out melioidosis as an endemic. Regarding the slow response to the current treatment of the disease, global researchers have been looking for faster, more responsive therapeutics to minimize the morbidity of the disease.
A vaccine, however, is being developed through extravagant, costly techniques, which would cause economic hardships to many infected areas (Cheng and Currie, 2005). Therefore, institutions are looking for cheaper and faster methods to treat melioidosis. One way to carry out such a task is to utilize nucleic acid oligonucleotides. Through the usage of aptamers, identification of the bacterium is possible. BimA, a trimetric Autotransporter/ macromolecule secreted by BMP (Brown and Iverson, 2011), which is an integral protein on the outer surface of B. pseudomallei, is functionalized with enough Histidine tags and actin-nucleating factors for a SELEX process using Nickel-NTA beads (Lazar and Stevens, 2011). Through selection, a fast therapeutic use of aptamers can help in treating the infection with accurate resources.
Specific aim 1: An aptamer, a precise nucleic acid sequence, is needed for selection against BMP through Nickel-NTA beads. The beads should help the protein and the RNA to meet and bind. The RNA sequence has to bind Burkholderia pseudomallei at the BPS site in order to activate a signal transduction that would aid in indentifying the bacteria and in accelerating the diagnosis and treatment of the infection (melioidosis). This therapeutic advantage of aptamers would be a congenial treatment, considering the precision and efficiency of SELEX rounds. If the selection is successful and the goals are achieved, inhibition of the Burkholderia bacteria would be easier and faster. Similarly, the same process can be applied onto Burkholderia mallei targets and similar results can be expected in treating glanders, since both B. pseudomallei and B. mallei are closely related.
Figure 1. An aptamer is needed to bind the Burkholderia pseudomallei (BPM) bacterium at the surface (BimA) for identification, which would later help identifying the bacterium and avoiding the enhancement of melioidosis.
***The target is on the “Recommended and Available” list of targets provided by Dr. Katy Brown, found in the -80⁰C freezer. The protein is sold by the company ATCC (ATCC.org) for $215 per 5 ug genomic DNA. Catalog # BAA-244D-5***
References:
1. “Groundwater Seeps Facilitate Exposure to Burkholderia pseudomallei.” Appl Environ Microbiol. 2011 Aug 26, Baker A, Tahani D, Gardiner C, Bristow KL, Greenhill AR, Warner J.
2. PLoS Negl Trop Dis. 2010 Nov 30;4(11):e900: “The epidemiology and clinical spectrum of melioidosis: 540 cases from the 20 year Darwin prospective study. Currie BJ, Ward L, Cheng AC.
3. J Infect Dis. 1989 May;159(5):890-9. "Melioidosis: a major cause of community-acquired septicemia in northeastern Thailand." Chaowagul W, White NJ, Dance DA, Wattanagoon Y, Naigowit P, Davis TM, Looareesuwan S, Pitakwatchara N.
4. Faraday Discuss. 2011;149:23-36; discussion 63-77. "Development of reagents and assays for the detection of pathogenic Burkholderia species."Qazi O, Rani M, Gnanam AJ, Cullen TW, Stead CM, Kensing H, McCaul K, Ngugi S, Prior JL, Lipka A, Nagy JM, Gregory CW,Judy BM, Harding SV, Titball RW, Sidhu SS, Trent MS, Kitto GB, Torres A, Estes DM, Iverson B, Georgiou G, Brown KA.
5. Front Microbiol. 2011;2:151. Epub 2011 Jul 15. "Autotransporters and Their Role in the Virulence of Burkholderia pseudomallei and Burkholderia mallei. Lazar Adler NR, Stevens JM, Stevens MP, Galyov EE.
LINK TO PROPOSAL: http://dl.dropbox.com/u/42009520/Proposal%202.docx
LINK TO PROGRESS REPORT 1: http://aptamerstream.blogspot.com/2011/10/progress-discussion-problems-and.html
LINK TO FINAL MANUSCRIPT: http://dl.dropbox.com/u/42009520/Final%20Manuscript.docx
Touria Rguig
08/30/2011
Tr7356
N59, Hemoglobin S
Nucleic Acid Selection against Hemoglobin S to Prevent the Aggregation of Red Blood Cells in Sickle Cell Anemic Patients
Hemoglobin S differs from normal adult hemoglobin called hemoglobin A only by a single amino acid substitution; a valine replacing a glutamine in the 6th position of the beta chain of globin [1]. This single amino acid substitution causes a genetically inherited disease called sickle cell anemia. This disease results in sickle-shaped red blood cells that deliver less oxygen to the body's tissues. These abnormal cells can also get stuck more easily in small blood vessels, and break into pieces that interrupt healthy blood flow.
Valine is a less polar amino acid than glutamate and thus favors hydrophobic interactions between each strand and its neighboring amino acids such as leucine and Histidine [1]. These types of interactions cause the red blood cells to aggregate and become sickled shaped preventing regular blood flow through capillaries and dramatically decreasing the hemoglobin’s high oxygen-binding affinity [1].
Selecting an aptamer against hemoglobin S to inhibit the polymerization of red blood cells caused by valine’s reactivity can prevent the sickling of blood cells. Also, it can potentially restore the high oxygen binding affinity of hemoglobin. Although there are drugs such as “Hydroxyurea” that enhance the quality of life of sickle cell anemic patient, this aptamer could be very specific and causing no detrimental side effects[2].
Specific aim: Synthesis of a specific aptamer that inhibits the aggregation of red blood cells in sickle cell anemic patients.
Hemoglobin can be purchased through Sigma-Aldrich for $204 per 25 mg. The catalog number is H0392-25MG .
References:
1- Lanzkron S, Strouse JJ, Wilson R, et al (June 2008). "Systematic review: Hydroxyurea for the treatment of adults with sickle cell disease". Annals of Internal Medicine 148 (12): 939–55.
2- (2008) Hydroxyurea for Sickle Cell Anemia. New England Journal of Medicine 359:1, 98-99
Here is the link to my proposal: https://docs.google.com/document/d/1so6mZ7vjYMlsYqiFvoKlozjp6_6ktJcX73dZ-vkq9xM/edit?hl=en_US
Here is the link to my progress report: http://aptamerstream.blogspot.com/2011/10/v-behaviorurldefaultvml-o_18.html
Here is a link to my new manuscript: https://docs.google.com/open?id=0By1AdiE2epJ_ZDQzN2E2MTEtZjc1Yy00OTZhLWI1NTgtMjYxY2EzMWU0Mzdl
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