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Article Excerpt
Co-Chair: Lisa Haynie, University of Mississippi Medical Center
Co-Chair: Tina Martin, University of Mississippi Medical Center
Co-Vice Chair: Mary Tan, Holmes Community College
Co-Vice Chair: Edwin Swiatlo, University of Mississippi Medical Center
THURSDAY MORNING
Bost Auditorium North
Session I: Oncology Research
8:00 Opening Remarks
8:15 PHENOXAZINES INHIBIT AKT AND INDUCE APOPTOSIS IN CANCER CELLS
K. N. Thimmaiah (1*), H Simmons (1), A Rinaldy (1), L Sylvester (1), P Grisham (1), and Peter J. Houghton (2), (1) Northwest Mississippi Community College, Southaven, Mississippi 38671 and (2) St Jude Children's Research Hospital, Memphis, Tennessee, 38105
Phenoxazines shut down the activation of Akt/mTOR/p70S6/S6 kinase pathway and induce apoptosis to a significant extent in rhabdomyosarcoma cells. Within the series, compounds having 3 carbon alkyl bridge were less potent than compounds with 4 carbon chain length. We proposed that by increasing the alkyl chain length to (-[CH.sub.2])[.sub.5] or (-[CH.sub.2])[.sub.6] at [N.sup.10]-position, the potency will be increased to a significant extent. Towards this goal, thirteen new derivatives have been synthesized and the parent 2-chlorophenoxazine was reacted with chlorobromoalkanes which resulted in [N.sup.10]-(chloroalkyl)-derivatives. Nucleophilic substitution reactions of these compounds with various secondary amines resulted in the desired products which were investigated to determine whether they would inhibit the phosphorylation of Akt. Serum starved cells were exposed to 100 nM-2000 nM phenoxazine derivatives before stimulating with IGF-I (10 ng/mL) for 10 min. Akt or Erk-1/2 phosphorylation was detected. Indeed, an increase in the expression of phosphorylated form of Akt at Ser 473 or Erk-1/2 was observed in response to IGF-I stimulation. These compounds at 100 nM inhibited the phosphorylation of Akt without affecting the phosphorylation of Erk-1/2. The potency of these compounds has been increased by 10-fold, and additional experiments are in progress (Supported by NIH grant CA 115404-01 to KNT).
8:30 COMPARISON OF POTENTIAL CHEMOTHERAPEUTIC AGENTS, 5-FLUORURACIL, GREEN TEA, AND THYMOQUINONE ON COLON CANCER CELLS
Anne A. Norwood*, Mary Tan, Marilyn May, Michelle Tucci, and Hamed Benghuzzi, University of Mississippi Medical Center, Jackson, MS 39216
Antioxidants have been found to be quite successful in deterring certain disease processes for years, especially cancer. Antioxidants protect the body by neutralizing the free radicals and donating one of their own electrons, thus ending the scavenger reaction. Epigallocatechin-3-gallate (EGCG), the most abundant catechin found in green tea, is a valuable scavenger of reactive oxygen species in vitro as well as in vivo. Thymoquinone (TQ), a major active component of black seed (Nigella sativa), is also known for its powerful scavenger abilities as an inhibitor of oxidative stress and has been utilized in the Middle East for centuries for healing properties. These two potent antioxidants when compared to the chemotherapeutic drug of choice, 5-fluorouracil (5-FU), have demonstrated incredible chemotherapeutic responses to the SW-626 cell line. The objective of this study was to evaluate and compare the effects of SW-626 colon cancer cells after a 24, 48, and 72 hour incubation periods with low, medium, and high doses of EGCG, TQ, and 5-FU. Cell viability, cell number, cellular morphology, and cellular metabolism were compared for the control and treatment groups. The results of this study evidenced a similar significant decrease in cell number as early as 24 hours in the groups treated with TQ and EGCG compared to 5-FU. Increases in cellular damage were evident after 24, 48, and 72 hours and in all treated groups compared with the control. Reduced cell numbers in the treated groups suggests the possibility that TQ and EGCG may have similar chemotherapeutic effects on cancer cells as 5-FU.
8:45 POTENTIAL EFFECT OF A-ESTRADIOL IN HUMAN JURKAT T-CELLS
Michael Johnson (1*), Danielle Wells (1), Erika Brown (2), Clement Yedjou (2), and Joseph A. Cameron (2), (1) Hinds Community College, Raymond, MS 39154 and (2) Jackson State Universtiy, Jackson, MS 39216
a-estradiol is the most potent estrogen of a group of endogenous estrogen steroids, which includes estrone and estriol. This steroid hormone is the most potent natural estrogen, produced mainly by the ovary, placenta, and in smaller amounts by the adrenal cortex, and the male testes. Although a-estradiol protects the renal and cardiovascular systems, the mechanisms involved remain unclear. In this research, we performed both MTT assay and trypan blue exclusion test to evaluate the effect of a-estradiol in HL-60 and Jurkat T-cells; and to compare the sensitivity of these two cells types. The results from both MTT assay and trypan blue exclusion test demonstrated that low, physiological levels of a-estradiol induce cellular proliferation in Jurkat T-cells. At higher dose of exposure (16 uM), a-estradiol decreases the viability of Jurkat T-cells compared to the control cells. Similar trend was obtained with the trypan blue exclusion test using the hemacytometer to count the cells manually. In summary, the results of the present study demonstrate that physiological levels of a-estradiol induce cell growth, and cellular proliferation of HL-60 and Jurkat T-cells.
9:00 COMPARISON OF GENE EXPRESSION IN MALIGNANT GASTRIC AND ENDOMETRIAL TUMORS
Sireesha Chinthaparthi Reddy (1*), Margot Kaelbling (2), Warren May (2), John Cleary (2), (1) Mississippi College, Clinton, MS 39058 and (2) University of Mississippi Medical Center, Jackson, MS 39216
We aimed to (1) identify genes that are differentially expressed in gastric adenocarcinoma, the second most common cause of cancer-related death worldwide, versus uninvolved tissue of the same patients and (2) assess if genes that are differentially expressed in gastric tumors were similarly expressed in endometrial tumors that were analyzed previously (MK). Total RNA was isolated from six tissue panels. RNAs from tumor and uninvolved tissue were separately reverse-transcribed and simultaneously hybridized onto a glass microarray spotted with 19,008 human ESTs. Detection was with Cy3[TM] (tumor) and Cy5[TM] (uninvolved tissue). Scanned arrays were analyzed with QuantArray[R] software. Statistical analysis involved correction of unequal dye intensity, bas[e.sub.2] transformation, LOESS normalization, SAM-type analysis, and volcano plots. Spots lacking gene information were eliminated. Seventeen genes were upregulated and 63 were downregulated in at least three of the six analyses. Two upregulated genes (CASP4 and ARL6IP) are at loci reported by other investigators to have gained chromosomal material in gastric tumors while six downregulated genes (SYNPO2, COX7C, SNAPC3, CDH5, DHX33, and CNN1) had chromosomal loss. Genes that were differentially regulated in both our gastric and endometrial tumor studies were CASP4 and ARL6IP (upregulated) and SYNPO2 (downregulated). Supported by NIH grant RR106476 from the MFGN INBRE Program of the National Center for Research Resources and the Cooperative Human Tissue Network.
9:15 PHARMACOPHORE MODEL FOR ANTIMALARIAL FARNESYLTRANSFERASE INHIBITION
Prasanna Sivaprakasam*, Aihua Xie, and Robert J. Doerksen, University of Mississippi, University, MS 38677
A very promising anticancer and antimalarial target is the heterodimeric zinc-containing protein farnesyltransferase (FT), which is one of the key enzymes in post-translational modification of proteins by prenylation, an important mechanism of cellular regulation. In order to help understand the balance of forces responsible for protein-ligand interaction, a useful approach is to prepare a pharmacophore model which summarizes the key interaction elements from amongst a group of diverse ligands. We selected representative antimalarial FT inhibitors including a potent benzophenone derivative and tetrahydroquinolines plus the known antimalarials chloroquine and artemisinin and prepared pharmacophore models using Catalyst software. One hydrogen bond acceptor and two hydrophobic features were found to be essential for in vitro growth inhibition of Dd2 strain of Plasmodium falciparum (Pf). We also developed a 3D pharmacophore model exclusively for benzophenones that are reported to be FT as well as Pfinhibitors. Several pharmacophore models either for FT inhibition or antimalarial activity exist in literature but to our knowledge this is the first report on a pharmacophore model for antimalarial FT inhibition. Our results showed four important pharmacophoric elements for antimalarial FT inhibitory activity of these benzophenones: one ring aromatic, one hydrophobic and two hydrogen bond accepting features. We will use these models for identifying novel lead antimalarial FT inhibitors using virtual screening.
9:30 Break
9:45 Session II: Posters
THE EFFECTS OF H2O2 ON CELL VIABILITY, CELLULAR MORPHOLOGY AND CELLULAR MEMBRANE DAMAGE USING TYPE II PNEUMOCYTES
Alicia Martin (1*), Michelle Tucci (2), Hamed Benghuzzi (2), and Joseph A. Cameron (3), (1) Hinds Community College, Raymond, MS 39154, (2) University of Mississippi Medical Center, Jackson, MS 39216 and (3) Jackson State University, Jackson, MS 39217
Oxygen supplementation at supraphysiologic levels is necessary in patients with respiratory failure, especially in those with acute lung injury. Oxygen in these settings is a life-preserving supportive measure until the initial pathologic process that elicited the respiratory disease subsides. It has been suggested that the toxic effect of oxygen is mediated by increased reactive oxygen intermediates, such as superoxide anion ([O.sub.2.sup.-]), hydrogen peroxide (H2[O.sub.2]), and hydroxyl radical (O[H.sup.-]). In this study, administration of low, medium and high doses of hydrogen peroxide to mimic oxygen overload or hyperoxia was investigated. In this study, increasing concentrations of [H.sub.2][O.sub.2] predisposed cells to lipid peroxidation, cellular damage, and increased membrane phosphatidylserine a sign of cellular apoptosis. Injury to the respiratory cells resulted in distortion of the alveolar architecture such as morphologic changes characterized by cell flattening/stretching, hyperchromasia, and cellular death. Increasing concentrations resulted in reduced cell numbers with time as well as increased levels of MDA and annexin V staining. Low-level administration of [H.sub.2][O.sub.2] did not induce alteration in cell numbers or cellular damage. The data indicates that type II pneumocytes have cellular machinery capable of detoxifying [H.sub.2][O.sub.2] below 80 FM, and as the dose reaches 220 FM cellular toxicity results and cells undergo cell death by apoptosis as evidenced by an increase in Annexin V staining.
THE EFFECTS OF SUSTAINED DELIVERY OF IGF-1 ON KIDNEY AND TESTICULAR TISSUES USING ADULT MALE RATS AS A MODEL
Phatia Wells (1*), Michelle Tucci (2), and Hamed Benghuzzi (2), and Joseph A. Cameron (3), (1) Hinds Community College, Raymond, MS 39154, (2) University of Mississippi Medical Center, and (3) Jackson State University, Jackson, MS 39217
It's common knowledge that growth hormone (GH) and insulin-like growth factor-1 (IGF-1) are some of the agents used by bodybuilders, athletes. These agents are banned from being used by athletes in most international sport federations, including the International Olympic Committee (IOC). Since 1990, the use of these agents has increased tremendously because of their potential use to produce gains in lean body mass and strength with concurrent loses in fat mass. Supplement companies, sensing the public demand for GH and IGF-1, have released all kinds of supplements that allegedly elevate these anabolic hormones. IGF-1 has been shown in vitro to produce changes in kidney epithelial cells and it is known that GH and IGF-1 play major roles in the development of tissues such as pancreas, kidney and testes. This study evaluated the effects of sustained administration of 5ng/day IGF-1 for 4 weeks on body weights, organ weights and morphology of vital and reproductive organs. Body weights over the 4-week period did not differ from control or sham operated rats. Decreased testicular and kidney weights were observed in the IGF-1 treated rats when compared with control and sham operated rats. Epididymal and seminal vesicle weights were increased at 4 weeks in animals treated with IGF-1. Morphological evaluations revealed structural changes in the proximal tubules of the kidney with increased area and length when compared to control and sham operated animals. In addition to changes within the kidney, there was an increase in the area of the seminiferous tubules as well as marked reduction in sperm by four weeks. The data suggests that doses of 5ng/day IGF-1 does play a significant role in kidney function and can alter reproductive tissues when given over long periods of time.
PASSIVE IMMUNIZATION WITH PNEUMOLYSIN ANTISERUM AS TREATMENT FOR STREPTOCOCCUS PNEUMONIAE KERATITIS
Sherrina Robinson*, Sherrina N. Dixon, Melissa Sanders, Kathryn S. Monds, Armando R. Caballero, Larry S. McDaniel, and Mary E. Marquart, University of Mississippi Medical Center, Jackson, MS 39216
Streptococcus pneumoniae (pneumococcus) is one of the top three causes of bacterial keratitis. Though pneumococcus can be treated with antibiotics, alternatives are necessary due to the increase in antibiotic resistance. Pneumolysin is a pneumococcal cytotoxin that stimulates the host inflammatory response and forms pores in host cell membranes. Antibiotics are prescribed to treat pneumococcal infections, however, pneumolysin remains and continues to damage the eye. Immunization methods to prevent or treat pneumococcal ocular infections could be beneficial in providing protection against the bacteria. The aim of this study was to determine if passive immunization with antiserum to pneumolysin could protect rabbit corneas from the damage associated with pneumococcal keratitis. Two forms of pneumolysin were used for immunization, heat-inactivated pneumolysin and non-heat-inactivated pneumolysin with a single amino acid change that renders it non-cytolytic. High titer antiserum to each immunogen was produced in rabbits by three monthly subcutaneous injections of each immunogen. Control serum was produced by three monthly subcutaneous injections of PBS. The corneas of new rabbits were injected with [10.sup.5] colony-forming units of S. pneumoniae strain WU2, and then these rabbits were immediately injected intravenously with control serum, antiserum to each of the immunogens. Clinical examination of the corneas was performed at 24, 36, and 48 hours post-infection, and the rabbits were sacrificed. The clinical scores of the corneas from rabbits that received passive immunization with either antiserum to the heat-inactivated toxin or antiserum to the cytolytic-negative toxoid were significantly lower than the scores from rabbits that received control serum. The bacterial colony-forming units recovered from all of the corneas harvested after sacrifice were not significantly different between experimental groups and control groups. These data show that passive immunization with antiserum to pneumolysin is an effective means to treat S. pneumoniae keratitis. (Supported in part by the Howard Hughes Medical Institute)
INFLUENCE OF TERMINALIA CHEBULA ON PHARMACODYNAMIC AND PHARMOCOKINETICS OF GLICLAZIDE IN NORMAL AND DIABETIC RATS
Clareice Stewart (1*), S. Satyanarayana (2), J Rajasekhar (2), T Sailaja (2), Bettaiya Rajanna (1), and Sharada Rajanna (1), (1) Alcorn State, Lorman, MS 39096 and (2) Visakhapatnam, India
It is a fact that countries like India and China with large populations use drugs from oriental medicine along with allopathic drugs. We are also aware of the fact that optimal blood sugar control is needed in diabetes. Diabetes mellitus is a chronic metabolic disorder characterized by persistent hyperglycemia caused by inadequate secretion of the hormone insulin, an inadequate response of target cells to insulin, or a combination of these factors. Albino rats (Wistar strain) of either sex were divided into 3 groups of 6 each and fasted for 18h prior to the experiment. Water was provided to the animals ad libitum. During the experiment they were withdrawn from food and water. Animals in Group-I, II and III were treated with gliclazide with doses equal to pre-determined Therapeutic Dose (TD), 1/2TD & sub TD respectively. Following these treatments, animals in group I, II and III were treated with Terminalia Chebula (TC) at 30 mg/kg, 100mg/kg and 300 mg/kg respectively with a washout period of one week between the treatments. Then, animals in group II were selected and treated with the combination of TC 100 mg/kg + gliclazide 1/2 TD after a washout period of one week. Diabetes was induced by alloxan monohydrate 100 -150 mg / kg body weight i.p. A group of 6 diabetic rats were treated with TC 100mg/kg, gliclazide 1/2 TD and the combination with a washout period of one week between the treatments. Blood samples were collected from retro-orbital plexus at 0, 1, 2, 3, 4, 6, 8, 10 and 12 h. and were analyzed for blood glucose by GOD-POD method using Semi Auto analyzer (Screen Master 3000). Gliclazide levels were estimated by HPLC. The results suggested that Gliclazide & TC produced hypoglycemia dose dependently in rats. Bi-phasic Peak effects were observed in gliclazide and terminalia chebula. TC produced hypoglycemia when given alone and enhanced the effect of gliclazide in normal and diabetic rats. The pharmacokinetics of gliclazide was not altered in the presence of TC. The results of this study indicate that TC enhances the effects of gliclazide, which is of pharmacodynamic in nature and varies with species. Hence, we plan to conduct further studies in humans to establish the clinical significance of the interaction.
SYNTHESIS AND CHARACTERIZATION OF DIORGANOTIN(IV) COMPLEX OF 2-ACETYLPYRAZINE N(4) -3-THIOSEMICARBAZONE
Tramarea Adams (1*), Lugile Sitole (2), Ramaiyer Venkatraman (2), and Joseph A. Cameron (2), (1) Hinds Community College, Raymond, MS 39154 and (2) Jackson State University, Jackson, MS 39216
Thiosemicarbazones exhibit a broad spectrum of pharmacological properties, including antibacterial, antiviral, antifungal, antimalarial, and antineoplastic activities. In solution, thiosemicarbazone molecules can exist in thione--thiol tautomeric form. The unique property of thiosemicarbazone is not only the presence of many electron donors centers in the structure but also the bonding characteristics. As a ligand, thiosemicarbazones are...
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