Anthony L. DePass

Plant Polyphenolics as Anti-Invasive Cancer Agents

Because invasion is, either directly or via metastasis formation, the main cause of death in cancer patients, development of efficient anti-invasive agents is an important research challenge. We have established a screening program for potentially anti-invasive compounds. The assay is based on organotypic confronting cultures between human invasive cancer cells and a fragment of normal tissue in three dimensions. Anti-invasive agents appeared to be heterogeneous with regard to their chemical nature, but plant alkaloids, polyphenolics and some of their synthetic congeners were well represented. Even within this group, active compounds were quite diverse: (+)-catechin, tangeretin, xanthohumol and other prenylated chalcones, 3,7-dimethoxyflavone, a pyrazole derivative, an isoxazolylcoumarin and a prenylated desoxybenzoin. The data gathered in this system are now applied in two projects. Firstly, structure-activity relationships are explored with computer models using an artificial neural network approach, based on quantitative structural descriptors. The aim of this study is the prediction and design of optimally efficient anti-invasive compounds. Secondly, the metabolism of orally ingested plant polyphenolics by colonic bacteria is studied in a simulator of the human intestinal microbial ecosystem (SHIME) and in human intervention trials. This method should provide information on the final bioavailability of the active compounds in the human body, with regard to microbial metabolism, and the feasibility of designing pre- or probiotics that increase the generation of active principles for absorption in the gastro-intestinal tract. The final and global aim of all these studies is to predict, synthesize and apply in vivo molecules with an optimal anti-invasive, and hence an anti-metastatic activity against cancer.

Model cell culture system for defining the molecular and biochemical events mediating terminal differentiation of human melanoma cells.

Cancer cells are commonly less differentiated than their normal progenitors; a phenotype that correlates with loss of specialized functions and an increased capability to self‐renew. Melanoma is an ideal model to analyze cancer progression and differentiation since a well‐characterized process of step‐wise tumor progression has been defined. Our lab previously described a combinatorial in vitro treatment protocol to induce terminal differentiation of human melanoma cells using a low dose of the PKC activator Mezerein (Mez) combined with interferon‐β (IFN‐β), which also activates IFN‐stimulated gene expression in addition to the re‐differentiation program. In principle, using an alternate way to induce terminal differentiation not including IFN‐β would be more compatible with gene expression profiling. A higher concentration of Mez alone induced terminal differentiation of HO‐1 human melanoma cells as measured by morphological, growth and biochemical assays. Pre‐treatment with the PKC inhibitor GF109203x blocked changes associated with differentiation and inhibited the ability of Mez to force irreversible/terminal differentiation. By combining this efficient method of inducing terminal differentiation with microarray analyses we now identify potential regulators of this process and demonstrate utility of this novel in vitro model in which to study the molecular determinants and mechanisms of human melanoma differentiation. J. Cell. Physiol. 218: 304–314, 2009.

Specificity of calreticulin transacetylase to acetoxy derivatives of benzofurans: effect on the activation of platelet nitric oxide synthase.

Calreticulin Transacetylase (CRTAase) catalyzes the transfer of acetyl group(s) from polyphenolic acetates (PAs) to functional proteins, such as Glutathione S-transferase (GST), NADPH Cytochrome c reductase and Nitric Oxide Synthase (NOS) resulting in the modulation of biological activities. A comparison of the specificities of the acetoxy derivatives of coumarins, biscoumarins, chromones, flavones, isoflavones and xanthones has been carried out earlier by us with an aim to study the effect of nature and position of the acetoxy groups on the benzenoid ring and the position of the carbonyl group with respect to oxygen/nitrogen heteroatom for the catalytic activity of CRTAase. In this communication for the first time, we have studied the influence of differently substituted benzofurans on the CRTAase activity to study the effect of the replacement of pyran ring of coumarin with furan ring, presence of carbonyl at C-3, substitution of C-3 carbonyl group with acetoxy group and presence of various substituents (OAc/OH/Cl) on the benzenoid ring. It was observed that acetoxy derivatives of benzofurans lead to inhibition of ADP induced platelet aggregation by the activation of platelet Nitric Oxide Synthase catalyzed by CRTAase. Accordingly, the formation of NO in platelets by 3-oxo-2,3-dihydrobenzofuran-6,7-diyl diacetate (3a) was found to be comparable with that of model polyphenolic acetate (PA), 7,8-diacetoxy-4-methylcoumarin (DAMC).

Arylalkyl ketones, benzophenones, desoxybenzoins and chalcones inhibit TNF-α induced expression of ICAM-1: structure-activity analysis

The interaction between leukocytes and the vascular endothelial cells (EC) via cellular adhesion molecules plays an important role in the pathogenesis of various inflammatory and autoimmune diseases. Small molecules that block these interactions have been targeted as potential therapeutic agents against acute and chronic inflammatory diseases. In an effort to identify potent intercellular cell adhesion molecule‐1 (ICAM‐1) inhibitors, a large number of arylalkyl ketones, benzophenones, desoxybenzoins and chalcones and their analogs (54 in total) have been synthesized and screened for their ICAM‐1 inhibitory activity. The structure‐activity relationship studies of these compounds identified three potent chalcone derivatives and also demonstrated the possible mechanism for their ICAM‐1 inhibitory activities. The most active compound was found to be 79.

Inositol 1,4,5 trisphosphate is inactivated by a 5-phosphatase in stamen hair cells of Tradescantia

Abstract. Inositol 1,4,5 trisphosphate [Ins(1,4,5)P3] is produced from the hydrolysis of phosphatidylinositol 4,5 bisphosphate, and as part of a second-messenger signal transduction mechanism, induces release of Ca2+ from internal stores in both plant and animal systems. It is less well established how the active Ins(1,4,5)P3 is inactivated. Studies in animal cells have demonstrated two separate metabolic pathways. Ins(1,4,5)P3 can be hydrolyzed by a 5-phosphatase or phosphorylated by a 3-kinase, resulting in the formation of Ins(1,4)P2 and Ins(1,3,4,5)P4, respectively, neither of which is able to mobilize intracellular Ca2+. Plant cell extracts have been reported to have hydrolytic and kinase activities that produce Ins(1,4)P2, and Ins(4,5)P2 and Ins(1,4,5,6)P4 from Ins(1,4,5)P3. These results offer little insight into the enzyme activities in the intact plant cell since the observed activities might be confined to intracellular compartments that have little if any impact on the signaling events within the cytosol that require Ins(1,4,5)P3. To resolve the mechanism of Ins(1,4,5)P3 inactivation, we microinjected stamen hair cells of Tradescantia virginiana L. with nonhydrolysable analogs of Ins(1,4,5)P3 that have been previously shown to cause Ca2+ release from intracellular stores. Our results indicate a sustained cytosolic [Ca2+] increase when cells were injected with the 5-phosphatase-insensitive 5-monophosphorothioate derivative of Ins(1,4,5)P3, in contrast to a brief transient when injected with the 3-kinase-insensitive 3-fluoro-3-deoxy Ins(1,4,5)P3 analog. We conclude that the 5-phosphatase pathway is the preferred pathway for Ins(1,4,5)P3 inactivation in the stamen hair cells of Tradescantia.

Chick heart invasion assay

Tumors are microecosystems in which a continuous cross talk between cancer cells and host cells decides on the invasive behavior of the tumor cell population as a whole (Mareel et al., Encyclopedia of cancer, San Diego, CA, Academic Press, 1997). Both compartments secrete activating and inhibitory factors that modulate activities such as cell-extracellular matrix (ECM) interaction, cell-cell adhesion, remodeling of the ECM, and cell motility. For this reason, confrontations of cancer cells with a living normal host tissue in organ culture have been introduced by several groups: Wolff and Schneider in France (Wolff and Schneider, C R S Soc Biol (Paris) 151:1291-1292, 1957), Easty and Easty in the United Kingdom (Easty and Easty, Nature 199:1104-1105, 1963), and Schleich in Germany (Schleich et al., J Natl Cancer Inst 56:221-237, 1976). Embryonic chick heart fragments in organ culture maintain many histological features of their tissue of origin: They are composed of myocytes, fibroblasts, and endothelial cells, and their ECM contains fibronectin, laminin, and several collagen types. Moreover, the fragments remain contractile, and this activity allows the monitoring of their functional integrity during organ culture.

Calreticulin transacetylase mediated upregulation of thioredoxin by 7,8-diacetoxy-4-methylcoumarin enhances the antioxidant potential and the expression of vascular endothelial growth factor in peripheral blood mononuclear cells

Extensive research carried out in our group on polyphenolic acetates (PAs) substantiated the potential role of PAs in causing diverse biological and pharmacological actions. Our earlier investigations firmly established the calreticulin transacetylase (CRTAase) catalyzed activation of nitric oxide synthase (NOS) by PAs. In this report, we have studied the effect of 7,8-diacetoxy-4-methylcoumarin (DAMC, a model PA) and other acetoxy coumarins on the thioredoxin and VEGF expression in human peripheral blood mononuclear cells (PBMCs), with a view to substantiate our earlier observation that DAMC was a superb inducer of angiogenesis. Real time RT-PCR analysis revealed the enhanced expression of thioredoxin reductase (TRXR) and diminished expression of thioredoxin interacting protein (TRXIP) leading to the increased expression and activity of thioredoxin (TRX) in PBMCs due to the the action of DAMC. The fact that TRX activity of PBMCs was enhanced by various acetoxy coumarins in tune with their affinity to CRTAase as substrate, suggested the possible activation of TRX due to acetylation. The overexpression of thioredoxin was found to correlate with that of VEGF as proved by real time RT-PCR and VEGF -ELISA results, apart from the DAMC-caused enhanced production of NO acting as an inducer of VEGF. Moreover, the intracellular ROS levels were also found to be reduced drastically, by DAMC thus reducing the oxidative stress in cells. These observations strongly evidenced the crucial role of TRX in DAMC-induced tissue angiogenesis with the involvement of VEGF.

Institutional Interventions That Remove Barriers to Recruit and Retain Diverse Biomedical PhD Students.

The faculty and student populations in academia are not representative of the diversity in the U.S. population. Thus, research institutions and funding agencies invest significant funds and effort into recruitment and retention programs that focus on increasing the flow of historically underrepresented minorities (URMs) into the science, technology, engineering, and mathematics (STEM) pipeline. Here, we outline challenges, interventions, and assessments by the University of Texas MD Anderson Cancer Center UTHealth Graduate School of Biomedical Sciences (GSBS) that increased the diversity of the student body independently of grade point averages and Graduate Record Examination scores. Additionally, we show these efforts progressively decreased the attrition rates of URM students over time while eliminating attrition in the latest cohort. Further, the majority of URM students who graduate from the GSBS are likely to remain in the STEM pipeline beyond the postdoctoral training period. We also provide specific recommendations based on the data presented to identify and remove barriers that prevent entry, participation, and inclusion of the underrepresented and underserved in the STEM pipeline.

The Competence of 7,8-Diacetoxy-4-Methylcoumarin and Other Polyphenolic Acetates in Mitigating the Oxidative Stress and their Role in Angiogenesis

The potential role of polyphenolic acetate (PA) in causing diverse biological and pharmacological actions has been well studied in our laboratory. Our investigations, for the first time, established the role of calreticulin transacetylase (CRTAase) in catalyzing the acetylation of nitric oxide synthase (NOS) by Pas leading to robust activation of NOS. 7, 8- Diacetoxy-4-methylcoumarin (DAMC) and other acetoxycoumarins augmented the expression of thioredoxin (TRX) and vascular endothelial growth factor (VEGF) in human peripheral blood mononuclear cells (PBMCs). These findings substantiated our earlier observations that DAMC was a superb inducer of angiogenesis. The enhanced expression of thioredoxin reductase (TRXR) and diminished expression of thioredoxin interacting protein (TRXIP) leading to increased expression and activity of TRX in PBMCs due to the action of DAMC was revealed by real time RT-PCR analysis. The possible activation of TRX due to acetylation was confirmed by the fact that TRX activity of PBMCs was enhanced by various acetoxycoumarins in tune with their affinities to CRTAase as substrates. DAMC caused enhanced production of NO by way of acetylation of NOS as mentioned above and thereby acted as an inducer of VEGF. Real time RT-PCR and VEGF ELISA results also revealed the overexpression of TRX. DAMC and other PAs were found to reduce the oxidative stress in cells as proved by significant reduction of intracellular ROS levels. Thus, the crucial role of TRX in DAMC-induced angiogenesis with the involvement of VEGF was established.

Prospective Student Database: Technological Support for Assessing the Effectiveness of Graduate Recruitment Activities

17:le1, 1 To the Editor: As many of the readers of CBE—Life Sciences Education know, academic institutions compete to attract a finite number of graduate students. Prospective students make highstakes decisions regarding graduate education, and educational institutions commit significant and sustained resources to recruit, retain, and train students. Approaches to manage recruitment and admissions would allow data-driven decisions concerning resource allocation. Applicant tracking systems are used in human resource management to provide a central applicant database and a set of processes and tools to help employers manage recruitment and hiring efforts (Gardner et al., 2003; Ngai and Wat, 2006; Alshibly, 2011; Laumer et al., 2015); however, academic institutions have been slow to adopt these tracking methods and instead depend on less sophisticated approaches for reasons of scale, expertise, and cost. We designed and created a Prospective Student Database that is purpose-built for tracking applicant movement through the recruitment and admissions processes in graduate education at The University of Texas MD Anderson Cancer Center UTHealth Graduate School of Biomedical Sciences. The system matches prospective applicants with students who have been admitted and, subsequently, with students who have enrolled at the institution. Admitted and enrolled students from one year can be queried against the list of prospective applicants from the current year and previous years using unique identifiers generated by the system (Codd, 2002). Thus, determination of prospective applicants who are admitted and which of the admitted applicants matriculate provides a clear understanding of recruitment outcomes. The reporting feature of this database provides standard reports that inform recruiters and academic institutions about the success of recruitment activities by event, date, and year of matriculation (Figure 1; Wilson et al., 2018). Tracking the effectiveness of different recruitment strategies and the utility of individual events enables institutions to direct resources toward successful approaches. As proof of principle, our recent work (Wilson et al., 2018) that assesses our graduate schools initiatives on recruiting and retaining underrepresented minority students used this Prospective Student Database and a network of databases (admissions, student, and alumni databases) to track recruitment activities (Prospective Student Database), student metrics (admissions database), candidacy examination outcomes (academic affairs database), attrition (academic affairs database), and career paths (alumni database). Thus, this system can be used by recruiters, faculty, staff, programs, and administrative leaders in graduate education for a complete picture of applicant outcomes. While the Prospective Student Database can be used as a stand-alone application, the real value of this platform is its scalability and adaptability as multi-user Marenda A. Wilson,* Wayne Turner, Anthony L. DePass, and Andrew J. Bean†§¶ Deans’ Office, The University of Texas MD Anderson Cancer Center, UTHealth Graduate School of Biomedical Sciences, Houston, TX 77030; Department of Biology, Long Island University, Brooklyn, NY 11201; §Department of Neurobiology and Anatomy, Cell Biology and Biochemistry, McGovern Medical School at The University of Texas Health Science Center at Houston, Houston, TX 77030; Department of Pediatrics, The University of Texas MD Anderson Cancer Center, Houston, TX 77030 Prospective Student Database: Technological Support for Assessing the Effectiveness of Graduate Recruitment Activities