Sanaa Ansari, M.A.
HPV Vaccine Now and in the Future abstract
Advisor: Carol Lin
Swathi Ashok, M.A.
Improving E. coli MsbA Crystallization by Epitome Engineering abstract
Advisor: John F. Hunt
Keenan T. Bashour, M.A.
Developing Dendritic Cells for Cancer Vaccination abstract
Advisor: Robert Tillman
Pin-Jung Chen, M.A.
A Review of Two Currently Approved Monoclonal Antibody Therapeutics for Breast Cancer abstract
Advisor: Ron Prywes
I-Ju Chiang, M.A.
A New Chapter on the Role of Small RNAs in Tumorigenesiss abstract
Advisor: Alexander Tzagoloff
Ethan Christ, M.A.
The Warburg Effect and Its Role in Cancer Detection and Therapy abstract
Advisor: Liang Tong
Kristina DiPietrantonio, M.A.
Tricyclic Indole Derivatives Function as a Privileged Scaffold abstract
Advisor: Brent R. Stockwell
Chelsea Durgan, M.A.
The Role of Transplastomic Plants in Agricultural Biotechnology and the Development of Oral Vaccines abstract
Advisor: Michael A. Lawton
Shai Gilad, M.A.
Prenatal Screening and Diagnosis of Down Syndrome (Trisomy 21) abstract
Advisor: Reba Goodman
Yih Lin Goh, M.A.
Communicating Microchemostat for Studying Bacterial Language abstract
Advisor: John March
Gregory W. Hruby, M.A.
Combination Therapy with Ipilimumab and Prostate Cryoablation for Prostate Cancer abstract
Advisor: Aaron Katz
David M. Jacobs, M.A.
Pharmacogenomics abstract
Advisor: Howard L. McLeod
Erik Koppes, M.A.
RED and SMU-1 Regulate Alternative Splicing of HSPG-2 pre-mRNA abstract
Advisor: James Manley
Justin Kuk, M.A.
Current Progress in Induced Pluripotent Stem Cells abstract
Advisor: Clark Hung
Chia-Yu Kuo, M.A.
Activin: A Critical Signaling Pathway in Tumorigenesis abstract
Advisor: Gloria H. Su
Tatiana Kurchuk, M.A.
Potential of Natural Killer Cells in Cancer Therapy abstract
Advisor: Ron Prywes
Chia-Chin Lee, M.A.
Stem Cell Therapy For Parkinson’s Disease abstract
Advisor: Alexander Tzagoloff
Martin A. Lehr, M.A.
Designing a Universal Vaccine Against Pandemic Flu abstract
Advisor: Chloe Bulinski
Shay G. McLean, M.A.
Challenges and Prospects for Small Molecule Biologics Development for Rheumatoid Arthritis abstract
Advisor: Carol Reiss
Ted C. Mehalic II, M.A.
The Development of IPSCs and a Comparison to Embryonic Stem Cells for Therapy abstract
Advisor: Jeremy Mao
Jennifer Myaeng, M.A.
Aberrant Dendritic Spine Morphologies in Nonspecific X-linked Mental Retardation and Fragile-X Syndrome abstract
Advisor: Tae-Wan Kim
Natasha Qaisar, M.A.
Role of Bone Marrow-Derived Cells in the Microenvironmetal Regulation of Tumor Metastasis abstract
Advisor: Carol Lin
Linda Sanders, M.A.
A Multidimensional Study of the Oncogenome to Achieve Personalized Cancer Treatment abstract
Advisor: James Russo
Zaher Shabani, M.A.
Rapid Purification of Recombinant Plasmid DNA from Escherichia coli via Aqueous Three-Phase Partitioning abstract
Advisor: William W. Ward
Shruti Sharma, M.A.
Retroviruses: Why HIV Continues to Be an Enigma abstract
Advisor: Vincent R. Racaniello
Steven J. Swanson, M.A.
Recent Discoveries of Long Noncoding RNA abstract
Advisor: Daniel Kalderon
Bushra Zaidi, M.A.
Identification of a Prominin-1 Expressing In Vitro Stem Cell in the Adult Mammalian Spinal Cord abstract
Advisor: Fiona Doetsch
HPV Vaccine Now and in the Future
Sanaa Ansari, M.A. 2009
Cervical cancer is the second leading cause of cancer mortality among women worldwide. The discovery of human papillomavirus (HPV) as it’s etiological agent and, more recently, the development of a prophylactic vaccine for the prevention of infection with some of the cancers associated HPV types represent a dramatic success of molecular medicine and herald a new era of cancer prevention and control. There are currently two prophylactic vaccines commercially available that have been approved by several countries worldwide. These vaccines have shown outstanding efficacy and safety results in the clinical trails, however their success will only be evident in the years to come. Since they have only been recommended for young girls before their sexual debut it precludes the millions of sexually active women who are at the risk of developing cervical cancer. Nonetheless, targeting sexually naive women in the public health domain, specifically in developing countries that have no cervical cancer screening, has the potential to greatly reduce the burden associated with HPV. However it is ironic that due to the prohibitive cost of these vaccines, the populations who need it the most will be the ones who are deprived of it. Today researchers are continually striving to develop new methods for the production and delivery of these vaccines to help reduce costs. In addition to these preventative strategies, therapeutic strategies remain paramount in order to treat the millions of women who are currently infected by HPV. Although the development of therapeutic interventions has not been as successful as expected, there is no reason to believe they will not do so in the future. The only plausible way this disease can be conquered is by concerted reduce costs. In addition to these preventative strategies, therapeutic strategies remain paramount in order to treat the millions of women who are currently infected by HPV. Although the development of therapeutic interventions has not been as successful as expected, there is no reason to believe they will not do so in the future. The only plausible way this disease can be conquered is by concerted efforts of both the prophylactic and therapeutic vaccines along with timely screening tests like the Pap smear.
Improving E. coli MsbA Crystallization by Epitome Engineering
Swathi Ashok, M.A. 2009
The formation of diffraction-quality crystals is a major limiting step in the determination of protein structure by X-ray crystallography. Here a novel study was conducted based on the first large-scale statistical study of crystallization that analyzes the records of 679 proteins purified by the NIH-funded Northeast Structural Genomics Consortium. The approach relies on the concept of surface entropy reduction (SER), i.e., the replacement of clusters of flexible, solvent-exposed residues with residues with lower conformational entropy. This strategy minimizes the loss of conformational entropy upon crystallization and renders crystallization thermodynamically favorable. To test these findings, these residues were introduced into E. coli MsbA which is an ATP-binding-cassette (ABC) transporter responsible for the lipid-A transport and has not previously formed diffraction-quality crystals. A total of 29 mutations were designed and 14 single-mutant and 10 double-mutant plasmids. All mutant proteins were expressed, purified, and screened for crystallization. The screen results provide suggestive evidence of crystallization enhancement by surface-entropy-reducing (SER) substitutions, not only the standard SER substitutions of lysine and glutamic acid with alanine that have previously been performed but also non-standard SER substitutions such as that of methionine with alanine. Our screen results also provide suggestive evidence of crystallization enhancement by substitutions with phenylalanine and glycine. 6 mutant proteins with a combination of non-standard SER substitutions and substitutions with phenylalanine and glycine have produced crystals in numerous conditions, almost twice the number of hits produced by the wildtype MsbA. Moreover, mutant crystals not only form in more conditions but also have novel morphologies and grow faster. However, the most promising mutation obtained involved the mutation MD276AG, the plasmid labeled 10. Even double mutations with 10 resulted in a large number of additional hits in the crystal screens. A direct comparison of the 10 mutant with the wild-type MsbA using both apo and nucleotides conditions on two different screens produced very interesting results. These crystals also have unique morphologies; however their diffraction quality is yet to be pursued.
Developing Dendritic Cells for Cancer Vaccination
Keenan T. Bashour, M.A. 2009
The use of cultured dendritic cells (DCs) to combat cancer has been a clinical aspiration for the past decade. With the federal approval of the first DC-based cancer treatment, Provenge, renewed attention has been given to DCs for use in adjuvant vaccine therapy. Natural regulators of the immune system, DCs have the potential to elicit an effective immune response against established tumors. Taking the success of DCs further now requires a reevaluation of the roles and mechanisms of DCs, both in vivo and in culture. Further study of DC subclasses, maturation protocols and delivery methods are necessary in the development of novel DC therapies. This discussion also evaluates the current state of clinical protocols relevant to cancer, their relevance to DC-based therapies and the expected shift to combination-based strategies when fighting cancer.
A Review of Two Currently Approved Monoclonal Antibody Therapeutics for Breast Cancer
Pin-Jung Chen, M.A. 2009
In the past quarter-century, monoclonal antibodies (mAbs) have become the most attractive therapeutic candidates for treating a variety of human diseases, especially cancer, due to their target specificity and low toxicity profiles. At this time, mAbs are the only clinically used immunotherapeutics in cancer therapy. Now, Herceptin (trastuzumab) and Avastin (bevacizumab) are United States Food and Drug Administration (USFDA) approved humanized mAbs that have shown activity in women with breast cancer. Herceptin and Avastin are designed to bind the most promising targets, the HER2 receptor and VEGF, respectively, which are known to play critical roles in breast tumor growth and survival. The two therapeutics represent important milestones in the development of antibody-based therapy. This review focuses on the development rationales, mechanisms of actions, and current status of these two antibody-based therapeutics in the treatment of patients with breast cancer. As the mAbs are predicted to be an expanding class of therapeutics in cancer treatment, the successful experiences gained from Herceptin and Avastin could be good references for further innovations of mAb therapeutics for cancer treatments in the future.
A New Chapter on the Role of Small RNAs in Tumorigenesiss
I-Ju Chiang, M.A. 2009
MicroRNAs are short non-protein-coding RNAs as negative regulators in various gene expressions to control cell division, cell differentiation, or cell death. As they are processed by sequential enzymatic activities, mature miRNAs will bind to the 3’ untranslated region (UTR) of target mRNAs to destabilize mRNAs or to repress gene translation. Here, it only mentions about different models in gene translational repression. For example, they may block translational factor binding to the 5’ m7Gppp cap structure or inhibit the joining of two ribosomal units. However, the exact mechanism of how miRNAs regulate target mRNAs is still unknown. After more and more miRNAs have been characterized, it has indicated that the same miRNAs can regulate various targets; also, the target can be regulated by various miRNAs. Since miRNAs play a key role in various biological processes, recent evidences have shown dysregulated miRNAs contribute to many diseases, including cancer. Several factors explained the dysregulation in miRNAs, such as changes in copy number of miRNAs, epigenetic modifications, abnormal functions of miRNA-accessory genes and proteins, and mutations in miRNAs; dysregulated miRNAs act as oncogenes or tumor suppressors or both. The same miRNA playing different roles is determined by cancer or tissue specific and surrounding target mRNA to be regulated. With advanced techniques in quantifying miRNAs, certain dysregulated miRNAs are always correlated with specific cancer type; therefore, miRNA expression profile can be useful in prognosis and diagnosis in cancers. What’s more, later in the future, it can be developed as an effective therapeutic molecule to treat cancer.
The Warburg Effect and Its Role in Cancer Detection and Therapy
Ethan Christ, M.A. 2009
The Warburg Effect is a cellular phenomenon in cancer cells discovered by Otto Warburg in 1924. His findings showed that in normoxic conditions tumor cells primarily use glycolysis for energy production instead of mitochondrial oxidative phosphorylation like normal cells. This breakthrough has been the basis for much research. It has resulted in a successful and widely-used cancer detection method, the positron emission tomography (PET) scan. The PET scan uses radioactive isotopes and the fact that cancer cells exhibit higher rates of glycolysis to pinpoint tumors with advanced imaging tools. Furthermore, Warburg’s work helped to show the potential for beneficial pharmaceuticals that could be developed by inhibiting certain chemical mechanisms of glycolysis to specifically target and kill cancer cells. This review covers research that has used the Warburg effect as a premise and the heretofore indications and applications of the Warburg effect.
Tricyclic Indole Derivatives Function as a Privileged Scaffold
Kristina DiPietrantonio, M.A. 2009
Current methods for designing and assembling small molecule libraries for high-throughput applications aim to maximize chemical diversity. In contrast, we found that focusing libraries around a specific scaffold can be more effective. We observed that tricyclic indole derivatives (TIDs) yielded higher hit rates in four different assays compared to compounds selected on traditional diversity considerations. To investigate this enrichment further, we assembled a small compound library composed exclusively of TIDs. Screening this library in the same assays yielded active compounds at an increased rate compared to traditionally selected compounds. Analysis of the TID library revealed favorable positions for substitutions, providing guidance in the selection of TIDs for inclusion in screening collections. Thus, we suggest that TID-based chemical libraries are particularly fruitful for use in high-throughput screening.
The Role of Transplastomic Plants in Agricultural Biotechnology and the Development of Oral Vaccines
Chelsea Durgan, M.A. 2009
Agricultural biotechnology has been greatly influenced in the past few decades by advances in recombinant DNA technology. Genetically modified plants have attracted particular attention due to their potential to boost nutrition, scale-up food production, and improve crop resistance to hostile conditions. Moreover, by producing foreign proteins in a more timely and cost efficient manner, genetically modified plants have the capacity to revolutionize the production of pharmaceuticals and industrial materials, and to help promote the use of biofuels. A relatively new area of plant biotechnology focuses on developing crops suitable for oral vaccination. However, many early attempts to design a plant vaccine concentrated on engineering the nuclear genome, leading to complications such as low protein expression, gene silencing, pleiotropic effects, position effects, and inadvertent gene transfer to other organisms. More recent studies have attempted to circumvent these problems by engineering the plastid genome (plastome) in lieu of the nuclear genome. Though scientists have exploited several techniques to successfully obtain antigens and other foreign proteins from the plastome, those techniques conforming to an approach known as Chloroplast genome-Clean Gene Transformation Technology (C-CGTT) are most likely to be employed in the future development of oral plant vaccines. C-CGTT techniques abnegate the use of antibiotic resistance genes in plastome engineering, greatly reducing the risk of negative health and environmental consequences.
Prenatal Screening and Diagnosis of Down Syndrome (Trisomy 21)
Shai Gilad, M.A. 2009
Over the last three decades, prenatal screening and diagnosis of Down Syndrome (Trisomy 21) have slowly evolved. New biochemical markers have been introduced to improve the sensitivity of screens, ultrasound images have been recognized to provide prognostic value and improved protocols have been developed. Despite these advances, confirmatory invasive diagnostic tests such as Amniocentesis and Chorionic Villus Sampling are still the gold standard for high risk pregnancies and these carry a real risk of miscarriage. Academia and industry have been hard at work to develop a non-invasive definitive prenatal diagnostic. The findings that nucleated fetal cells as well as cell-free fetal DNA and RNA circulate in maternal serum have provided the basis for a range of efforts that are now finally coming to fruition. While efforts focused on the extraction of fetal cells from maternal serum dominated the 1990s, it now appears that techniques relying on the analysis of cell-free DNA and RNA such “Chromosome Dosage Analysis” and “SNP Allelic Ratio Analysis” have prevailed. Indeed, one company, Sequenom, appears to have taken the lead and is set to introduce the first approved non-invasive diagnostic for Trisomy 21 in June 2009. This diagnostic and ones to follow are widely anticipated to revolutionize prenatal diagnosis of Trisomy 21 and prenatal genetic diagnostics in general.
Communicating Microchemostat for Studying Bacterial Language
Yih Lin Goh, M.A. 2009
Environmental contamination is the main issue facing the industrialized world today. Bioremediation has slowly emerged as one of the key strategies to treat the environmental pollutants over past 10 years. Nonetheless, the microbial ecology within an environment affects the effectiveness of bioremediation. Microbial interactions can hamper or enhance remediation efforts. A more effective bioremediation process can be designed by understanding the interactions within the microbial ecology and harnessing synergistic relationships.
We propose to adapt a three channel hydrogel-based microfluidic device, into a communicating microchemostat that can be used for monitoring bacterial behavior over a longer period of time. We plan to use the principle of quorum sensing from Vibrio harveyi to test the communication set up between microchemostats. This cheap and effective tool can also be manipulated into different settings, such as (1) For studying cellular growth kinetics, (2) For studying motility of Vibrio, (3) For studying biofilm formation and (4) For studying bacterial interspecies communication. For the long term goals, we plan to develop a better bioremediation process over longer period of time with minimal resources.
Combination Therapy with Ipilimumab and Prostate Cryoablation for Prostate Cancer
Gregory W. Hruby, M.A. 2009
As cancer treatments become more diverse, new combinations of both medical and surgical therapies are being utilized to improve oncological outcomes. Recurrent prostate cancer offers an ideal population for novel combinatorial strategies. Salvage cryoablation of the prostate after primary radiotherapy treatment is an acceptable second line therapy. However, after 7 years of follow up 41% of patients are expected to relapse. These patients are ideal to introduce combination therapies. Cryoablation has been shown to impart further oncologic control by inducing antitumor immunomodulation. Induction and maintenance of the cytotoxic T-cell response relies heavily on CD4+ cells, which through either direct antigen recognition or recognition of antigen presenting cells (APC) can also produce antitumor activity. Ipilimumab is an ideal medical therapy to apply in combination with salvage prostate cryoablation. Ipilimumab is an anti-CTLA-4 antibody which binds CTLA-4 preventing T-Cell down regulation. It is possible that when Ipilimumab is placed in combination with salvage prostate cryoablation, the influx of APCs with ablated prostatic samples which up regulates T-cell activation, will be further up regulated by impairing the CTLA-4 down regulation of T-cells. We propose a single arm, single center study for prostate cancer patients who have failed primary radiotherapy treatment and who are considering an androgen ablative therapy to study the efficacy and safety of both Ipilimumab and salvage prostate cryoablation.
David M. Jacobs, M.A. 2009
Pharmacogenomics studies the interaction of individual genetic makeup with drug therapy to predict response. Genetic variation between people helps account for the fact that the same drug does not affect everyone uniformly. The early years of pharmacogenomics focused on understanding and characterizing how genes impact drug response. Pharmacogenomics is still in its infancy, though, because the harder task of applying it to drug development and integrating it into a tangled healthcare system based upon many traditional approaches to medicine is still ongoing. Using K-RAS as a case in point, pharmacogenomics challenges current regulatory frameworks, as rapid technological advances outpace the evolution of legal standards. Without medical community and regulatory authority harmonization, oncologists are unsure how to appropriately treat colorectal cancer patients who harbor the mutant allele of K-RAS with cetuximab and panitumumab, two monoclonal antibodies targeting the EGFR. Despite current obstacles, pharmacogenomic analysis is increasingly being built into clinical trial design, providing further evidence that it is a distinct scientific field with practical application across numerous segments of the current healthcare value chain. Even skeptics of pharmacogenomics could agree that if every challenge represents an opportunity, pharmacogenomics has the potential to transform drug development and the practice of medicine.
RED and SMU-1 Regulate Alternative Splicing of HSPG-2 pre-mRNA
Erik Koppes, M.A. 2009
The alternative splicing factors RED and SMU-1 are herein shown to enhance the inclusion of exon 70 into the hspg-2 messenger RNA. Previous RNAi work in the Manley lab recognized RED and SMU-1 as alternative splicing factors regulating hspg-2 mRNA. I have carried out in vitro splicing reactions of 32P labeled hspg-2 minigene transcripts containing exon 69 to 71 with and without the addition of recombinant RED and SMU-1. Fluorescently tagged RED and SMU-1 localize in the nucleus of HeLa and 293 cells. The evidence presented in this thesis indicates that SMU-1 and RED interact and regulate activity of the spliceosomal machinery associated with hspg-2 pre-mRNA.
Current Progress in Induced Pluripotent Stem Cells
Justin Kuk, M.A. 2009
Somatic cells can be reprogrammed to a pluripotent state through overexpression of a set of transcription factors to produce induced pluripotent stem cells (iPSC). The purpose of this thesis is threefold: First, to outline various methods to produce and verify iPSC. In particular the recent developments to circumvent genomic integration, omit C-myc and identify of small molecules that augment efficiency will be in the spotlight as they are the key to developing a safe, sufficient supply of iPSC. Secondly, recent proof of concept studies demonstrating iPSC utility for studying establishing models for disease specific investigations, pharmaceutical screening and enabling future applications in regenerative medicine will be reviewed. Finally, having reviewing the state of the art, a critical assessment of future directions that remain to be pursued will be detailed.
Activin: A Critical Signaling Pathway in Tumorigenesis
Chia-Yu Kuo, M.A. 2009
Members of TGF-β superfamily have been proved to be important regulators of many essential cellular processes, including proliferation, differentiation, cell cycle control, migration and apoptosis. Activin, a member of TGF-β superfamily, is involved in many physiological processes, such as embryonic development, follicle development, skin morphogenesis, wound repair, ectodermal organogenesis, and angiogenesis as well as in many human diseases. Cancers which are highly correlated to activin signaling pathway are observed to have many mutations in TGF-β/activin signaling. Activin was first purified from porcine follicular fluid and was found to have FSH secretion stimulating function. Subsequent studies showed that activin was widely expressed in many tissues. With the parallel signaling pathway with TGF-β, activin is always considered as an important pathway in tumorigenesis. This article reviews many aspects of acitivn, including the signal cascade of activin, activin-related proteins, its role in development and tumorigenesis. Inactivation of activin signaling is observed to abrogate the anti-proliferation and apoptosis effects of activin in many cancer types. However, activin also has been reported to have tumor stimulating effects in esophagus carcinoma. The complex networks that activin is involved in tumorigenesis remain unclear. Future experiments to further our understanding of the networks which may identify additional therapeutic targets for cancers and other human diseases.
Potential of Natural Killer Cells in Cancer Therapy
Tatiana Kurchuk, M.A. 2009
Natural Killer (NK) cells are lymphocytes that exhibit cytotoxicity towards tumor and virus-infected cells without prior sensitization. Since their discovery in the 1970’s, NK cells have demonstrated a great potential in targeting and killing cancer cells. Much research and trials have been performed to harness this potential for effective cancer therapies. Following decades of research and animal trials, there are NK cell therapies that are emerging to be very promising. One combination therapy currently in at Phase I clinical trial at the University of Minnesota utilizes allogeneic NK cells to attack leukemic and lymphomic cells in combination with NK-proliferative IL-2 and target-specific Rituxan that utilizes the ADCC mechanism of action. Another combination strategy that has not been tested yet is the use of engineered NK cells in combination with high-intensity focused ultrasound (HIFU) and all-trans-retinoic acid (ATRA), as well as IL-2 cytokine, to infiltrate and destroy solid tumors. While much progress has been made for the cure of some cancers, there are many more that currently do not have adequate therapies available. It is the hope that through research and better understanding of the interaction between NK cells and cancer, more effective therapies will be designed for curing cancer or extending the life of the patient with minimum side-effects.
Stem Cell Therapy For Parkinson’s Disease
Chia-Chin Lee, M.A. 2009
Parkinson’s disease (PD) is one of the most common neurodegenerative disorders, affecting more than 2% of the population over age 65. The disease is manifested by the degeneration of dopamine neurons in the substantia nigra region of the midbrain. Patients are usually diagnosed at a late age, after most dopamine neurons have been destroyed and the body has begun to display abnormal voluntary motor functions. The most prominent features of PD are bradykinesia, rigidity, postural instability, and tremor. Currently, there are no cures for PD, and no adequate long-term treatment. Stem cell transplantation is under investigation as one possible treatment, based on the idea that replacing the lost dopaminergic neurons would relieve and reverse the symptoms of the disease. Several labs have successfully developed methods to induce embryonic stem cells into dopamine neurons, and have demonstrated functional recovery in animal models. Genetic factors and other components related to the pathogenesis of PD have been investigated, and manipulated to generate dopamine neurons from stem cells. When these neurons are transplanted into animals with induced PD, they are able to release dopamine and improve motor functions. Strategies to generate dopamine neurons from human embryonic stem cells are still in their infancy, but initial experimentation gives hope that an unlimited supply of dopamine neurons will eventually be widely available in neurotransplantation for PD patients.
Designing a Universal Vaccine Against Pandemic Flu
Martin A. Lehr, M.A. 2009
Dr. Edward Jenner designed the first vaccine in 1796 after learning that local female milkmaids who had contracted cowpox virus were resistant to being infected by smallpox, a more severe version of the virus. Dr. Jenner’s vaccine discovery laid the groundwork for virologists to better understand how viruses function in order to design novel vaccines. While the advances made in vaccine technology are laudable, researchers have not solved the curing or prevention of many common and deadly viruses. Of particular interest to vaccine developers is seasonal influenza, which has the capability to mutate each season making it challenging for drug developers to rapidly develop new vaccines to address the needs of the current season. Additionally, the influenza vaccines currently in usage are not very effective and almost always contain adjuvants, carriers that can lead to serious allergic reactions. Therefore, there is an unmet need for an influenza vaccine that is potent, rapidly manufactured, and does not have to be seasonally produced. A flagellin-linked M2e universal vaccine might be a novel alternative to current vaccine therapies and could become the future standard of care. The immunogenicity of flagellin, combined with the stable expression of an M2e epitope on surface of the influenza virus, makes a flagellin-linked M2e recombinant fusion protein an ideal candidate for becoming the first universal flu vaccine.
Challenges and Prospects for Small Molecule Biologics Development for Rheumatoid Arthritis
Shay G. McLean, M.A. 2009
Rheumatoid arthritis (RA) is a highly debilitating, incurable autoimmune disease of the joint lining which produces pain and swelling and reduced mobility of the patient’s joints. Autoimmune attack on the synovium leads to destruction of the ligaments, tendons, and muscles supporting the joint and can cause bone erosion and sometimes deformation. With increased understanding of the benefits of early aggressive treatment for RA and the resulting need for long term chronic therapy, opportunity exists to provide new treatments that offer greater convenience, less safety risk and lower cost to patients. One category of treatments with potential to meet that need is the small molecule DMARDs currently in development for RA. This review will discuss the current understanding of potential treatment targets in RA and the progress to date for small molecule DMARDs aiming at those targets. Nearly 200 oral agents that have been investigated in RA have failed or been discontinued in the RA patient population, which provides evidence of the difficulties of these research efforts. Despite these many discontinuations and failures, two leading oral DMARD compounds are progressing to Phase III trials within the next year. Pfizer's oral JAK3 inhibitor, CP-690,550, has demonstrated consistent efficacy similar to the anti-TNFs both as a monotherapy and with background methotrexate and in both biologic-naïve and biologic-experienced patients. Rigel's Syk kinase inhibitor, R788, showed impressive efficacy in early Phase II studies but failed to show significant improvement in its latest trial looking at harder-to-treat patients who had previously failed a biologic DMARD. If a novel small molecule DMARD can show consistent efficacy and a safety profile not worse than the anti-TNFs, it will likely carve out a market and compete for use in RA patients.
The Development of IPSCs and a Comparison to Embryonic Stem Cells for Therapy
Ted C. Mehalic II, M.A. 2009
Recent breakthroughs in stem cell research have brought about a new type of cell called induced pluripotent stem cells (IPSCs). For the first time researchers are able to coerce a terminally differentiated somatic cell back into a pluripotent state by introducing a few select transgenes to the cell. These IPSCs are capable of differentiating into cell types from each of the three different embryonic germ layers. By using a variety of other factors, it’s possible to differentiate IPSCs into a tissue of choice. When this procedure is applied to a diseased individual, novel drugs could be developed by testing drug combinations and different factors in vitro on a tissue specific disease, a procedure that previously has been extremely limited.
The recent successes of generating IPSCs from several different types of cells and the wide age range of the donors indicate the versatility of IPSCs for an assortment of therapies. The ability of converting a donor’s somatic cell into any one of the adult cell types means that IPSCs also have a huge potential for autologous cell therapy. Despite the rapid fame that IPSC have achieved, the mechanisms by which IPSC are generated are not completely understood and it is not ready for clinical therapy yet.
Aberrant Dendritic Spine Morphologies in Nonspecific X-linked Mental Retardation and Fragile-X Syndrome
Jennifer Myaeng, M.A. 2009
Dendritic spines are sites of excitatory postsynaptic synapse and their morphology affects synaptic plasticity. In the cortex of non-specific X-linked Mental Retardation (N-XMR) and fragile-X syndrome patients, the majority of the spines show abnormal morhphological characteristics as compared to normal patients. Understanding the molecular cause behind this phenotype will help us understand the underlying molecular and cellular cause(s) of these disorders, which will eventually allow the development of effective therapeutics. F-actin polymerization/depolymerization of the cytoskeleton are essential to spine morphology, and actin-binding proteins (ABPs) are shown to regulate F-actin. In N-XMR , the ABP regulatory pathways are disturbed because three N-XMR mutations OPHN1, PAK3, and ARHGEF6 change the activation/deactivation of Rho GTPases, the pathway’s upstream players. Cytoskeleton dynamics area also disrupted in Fragile-X, as a mutation in FMR1 disrupts Rac1 regulation of actin-binding ADF/cofilin. These observations collectively suggest the aberrant regulation of actin polymerization/depolymerization and subsequent alterations in spine morphology contribute significantly to the pathophysiology of N-XMR and FXS and may provide the molecular targets for future treatments.
Role of Bone marrow-derived Cells in the Microenvironmetal Regulation of Tumor Metastasis
Natasha Qaisar, M.A. 2009
Cancer has been long viewed as the manifestation of aberrant genetic and epigenetic alterations leading to pathological conditions such as metastasis, a multistep process, which allows the primary cancer cells to escape from the primary tumor to distinct sites, where it grows to form new tumor. Available evidence is indicating that the specific microenvironment of both primary tumor as well as the metastasizing one is crucial for the initiation, development and progression of tumor and its subsequent transition to malignant forms. The interactive microenvironment provides specific regulatory cues to tumor cells which help them to survive both at local and distinct sites in the host. It is now understood that most of the mechanisms underlying tumor development and progression into a malignant form involve the mobilization of bone-marrow derived cells from the bone marrow to the tumor site, the formation of new tumor vasculature through the process of angiogenesis, the degradation of extracellular stromal matrix, the induction of autocrine and paracrine signaling pathways, cells, and the secretion of growth factors and chemoattractants by the tumor cells to recruit other cells to the tumor site.
An important lineage of the bone marrow-derived cells is the myeloid lineage which is capable of differentiating into a number of cell types such as macrophages, dendritic cells, neutrophils, eosinophils, mast cells, and basophils. These cells are the essential components of both innate and adaptive immune systems. As sentinels of innate immune system, these lineages perform phagocytosis of foreign antigens and mediate inflammation in event of tissue injury. In addition, they are critical for mediating antitumor responses by releasing a number of proinflammatory cytokines and chemokines and activating cells of adaptive immunity against the growing tumor. However, knowledge gathered from studies using transgenic mouse models as well as the analyses of human tumor biopsies have highlighted a more ‘protumor’ than ‘antitumor’ role played by these cells in the tumor microenvironment. It is thought that these cells are recruited by the tumor cells to promote their growth and development though angiogenesis, invasion and metastasis. Further improvement in our knowledge about the role of these cells can help in providing answers to many questions related to the mechanism of tumor development and its progression to metastasis as well as the therapeutic potential of these cells in the development of anticancer therapies, notably the antiangiogenic therapies.
A Multidimensional Study of the Oncogenome to Achieve Personalized Cancer Treatment
Linda Sanders, M.A. 2009
The intention of this thesis is to demonstrate that cancer is amenable to multidimensional genome-wide study and that these genomic approaches will be necessary in taking the next step toward personalizing the treatment of cancer and its many subtypes. It will discuss the current approach to cancer treatment and suggest the reasons for its underachievement.
While personalized medicine is desirable for all diseases, this thesis will review the genetic basis of cancer to demonstrate the need for a personalized approach for this distinct class of disease. Also, because breast cancer has best typified the achievements of an individualized approach, this subtype will frequently be used as a specific example of how personalized medicine can be successfully applied.
Lastly, this thesis will explain the roles that new technologies are playing in achieving personalized medicine in the various “omic” fields of genome-wide analysis. It will then propose an integrative approach to cancer by combining these genomic approaches with the molecular understanding scientists have worked hard to achieve, so as to provide a context for new findings. Finally, future technologies and endeavors that are on the horizon and are likely to play an integral role in enabling personalized medicine for cancer will be discussed.
Rapid Purification of Recombinant Plasmid DNA from Escherichia coli via Aqueous Three-Phase Partitioning
Zaher Shabani, M.A. 2009
The utilization of an aqueous three-phase partitioning (TPP) system in a plasmid DNA purification scheme generates a surprisingly clean, low-viscosity extract enriched with plasmid DNA and free from contaminants such as mRNA, tRNA, gDNA, and almost all protein. Here, a tertiary-butanol/ammonium sulfate aqueous three-phase partitioning method was directly applied to an unlysed whole cell paste of an Escherichia coli strain expressing enhanced green-fluorescent protein, EGFP. TPP was then compared to the classical alkaline-lysis-based plasmid extraction method (Qiagen™ Plasmid Miniprep) and the boiling lysis method – both well-known techniques used for providing small scale, high quality, purified plasmid DNA from Escherichia coli cells for various applications. Two main features of the three-phase partitioning technique are: (1) The apparent isolation of purely supercoiled plasmid DNA from the intermediate phase; and (2) The segregation of open circular and supercoiled plasmid DNA from RNA in the lower aqueous phase.
Retroviruses: Why HIV Continues to Be an Enigma
Shruti Sharma, M.A. 2009
The human immunodeficiency virus, like many other retroviruses, undergoes a rapid rate of evolutionary change in order to survive successfully against the host immune system. The virus displays extensive genetic variation within the host, such as that in the hypervariable region of the env gene, which makes it one of the fastest evolving organisms. These variations and a number of other strategies employed by the virus to evade the human immune system have proved to be one of the major factors impeding effective vaccine development against HIV/AIDS. Therefore, it is of utmost importance to understand the pathogenesis and the interplay between host and viral factors and apply the knowledge of these processes towards the development of an effective antiviral therapy. This thesis is an attempt to summarize some of the findings about HIV/AIDS pathogenesis and recent novel therapeutic approaches to combat the disease.
Recent Discoveries of Long Noncoding RNA
Steven J. Swanson, M.A. 2009
Expression of the human genome is a remarkably complex system of checks and balances. Current estimates call for 22,000 protein-coding genes encoded by roughly 2% of the genome. However, large scale cDNA sequencing and genomic tiling array projects estimate the number of transcripts to be at least one order of magnitude larger than this number of genes. Collectively, these data imply that the pervasive transcription of the remainder of the genome represents either useless transcriptional noise or clear evidence that the currently annotated genome map needs further resolving. Studies aimed at improving our understanding of genomic expression have revealed a new group of transcripts increasingly implicated in myriad pathways. Long noncoding RNA (long ncRNA) has become a catch-all term for these transcripts because of their size and shared biochemical signatures with mRNA, such as 5’ caps and poly-A tails. While the functions of most long ncRNAs are still being deciphered, the evidence suggests a primary function of long ncRNA transcripts regulating gene transcription through a variety of cis and trans, sense and antisense mechanisms. These functions have been observed in various cell types across multiple species and have been linked to common cancers and neurodegenerative diseases. Here, I provide an evaluation of the field and recommend future experimentation needed to achieve a more global understanding of long ncRNA.
Identification of a Prominin-1 Expressing In Vitro Stem Cell in the Adult Mammalian Spinal Cord
Bushra Zaidi, M.A. 2009
The adult mammalian brain has two regions that harbor neural stem cells (NSCs), the subventricular zone (SVZ) of the lateral ventricles and subgranular zone (SGZ) of the hippocampus. However, to date, there has been limited success with prospectively isolating NSCs from non-neurogenic regions, such as the adult mammalian spinal cord. This is because of the low frequency of NSCs in the adult CNS and lack of unique NSC markers, which poses significant challenges to the development of techniques to isolate NSCs. Furthermore, no combination of markers has been identified for prospectively identifying adult NSCs. In this study, we show that a NSC in the adult mammalian spinal cord can be prospectively isolated using a combination of three markers: CD133, CD24, and PSA-NCAM and the combination of markers enriches for in vitro stem cells. Furthermore, we aimed to identify the prominin-1 expressing in vitro NSC in the central canal of the spinal cord. FACS isolation of CD133+/CD24+/PSA-NCAM+ cells resulted in an 8 fold enrichment for neurosphere formation (an assay of in vitro stem cells) as compared to CD133+/CD24+/PSA-NCAM- cells. To further identify these in vitro stem cells we immunostained for glial fibrillary acidic protein (GFAP), an astrocyte marker. To test if astrocytes contributed to neurosphere formation we ablated dividing GFAP-expressing cells in the spinal cord. The ablation of dividing GFAP-expressing cells in the CD133+/CD24+/PSA-NCAM+ spinal cord did not abolish neurosphere formation, suggesting that the in vitro NSCs generating neurospheres in our assay are not GFAP-expressing cells, but a different type of cell. We show that the central canal NSC are CD133+, CD24+, and PSA-NCAM+. The finding that astrocytes in the CD133+/CD24+/PSA-NCAM+ cell fraction are unlikely to be the in vitro NSCs distinguishes these putative spinal cord stem cells from those found in the SVZ. This lays the ground work for future studies to identify the adult NSC in the spinal cord.