Molecular interaction analysis of cigarette smoke carcinogens NNK and NNAL with enzymes involved in DNA repair pathways: An in silico approach

DNA damage occurs almost all the times in cells, but is repaired also continuously. Occurrence of all these mutations and their accumulation in one cell which finally becomes tumorigenic/carcinogenic appears possible if the DNA repair mechanism is hampered. We hypothesize that alterations in DNA repair pathways, either all or at least at one i.e. genetic, translational or posttranslational level, becomes quite imperative for the initiation and progression of Cancer. Therefore, we investigated the interaction capability of some carcinogens with the enzymes involved in the DNA repair mechanisms. Cigarette smoke''s derivatives like NNK and NNAL are well established carcinogens. Hence, we analyzed 72 enzymes involved in the DNA repair Mechanisms for their interactions with ligands (NNK and NNAL). The binding efficiencies with enzymes ranging from +36.96 to -7.47 Kcal/Mol. Crystal Structure of Human Carbonmonoxy-Haemoglobin at 1.25 Å Resolution, PDB ID-1IRD as a +Ve control, showed binding energy -6.31 to -6.68 Kcal/Mol. and Human heat shock factor-binding protein 1, PDB ID- 3CI9 as a -Ve control, showed - 3.91 to +2.09 Kcal/Mol. Binding was characterized for the enzymes sharing equivalent or better interaction as compared to +Ve control. Study indicated the loss of functions of these enzymes, which probably could be a reason for fettering of DNA repair pathways resulting in damage accumulation and finally cancer formation.     

Selective killing of cancer cells by Ashwagandha leaf extract and its component Withanone involves ROS signaling

Background and Purpose

Ashwagandha is a popular Ayurvedic herb used in Indian traditional home medicine. It has been assigned a variety of health-promoting effects of which the mechanisms remain unknown. We previously reported the selective killing of cancer cells by leaf extract of Ashwagandha (i-Extract) and its purified component Withanone. In the present study, we investigated its mechanism by loss-of-function screening (abrogation of i-Extract induced cancer cell killing) of the cellular targets and gene pathways.

Methodology/Principal Findings

Randomized ribozyme library was introduced into cancer cells prior to the treatment with i-Extract. Ribozymes were recovered from cells that survived the i-Extract treatment. Gene targets of the selected ribozymes (as predicted by database search) were analyzed by bioinformatics and pathway analyses. The targets were validated for their role in i-Extract induced selective killing of cancer cells by biochemical and molecular assays. Fifteen gene-targets were identified and were investigated for their role in specific cancer cell killing activity of i-Extract and its two major components (Withaferin A and Withanone) by undertaking the shRNA-mediated gene silencing approach. Bioinformatics on the selected gene-targets revealed the involvement of p53, apoptosis and insulin/IGF signaling pathways linked to the ROS signaling. We examined the involvement of ROS-signaling components (ROS levels, DNA damage, mitochondrial structure and membrane potential) and demonstrate that the selective killing of cancer cells is mediated by induction of oxidative stress.

Conclusion

Ashwagandha leaf extract and Withanone cause selective killing of cancer cells by induction of ROS-signaling and hence are potential reagents that could be recruited for ROS-mediated cancer chemotherapy.     

Second international conference on accelerating biopharmaceutical development

The Second International Conference on Accelerating Biopharmaceutical Development was held in Coronado, California. The meeting was organized by the Society for Biological Engineering (SBE) and the American Institute of Chemical Engineers (AIChE); SBE is a technological community of the AIChE. Bob Adamson (Wyeth) and Chuck Goochee (Centocor) were co‐chairs of the event, which had the theme “Delivering cost‐effective, robust processes and methods quickly and efficiently.” The first day focused on emerging disruptive technologies and cutting‐edge analytical techniques. Day two featured presentations on accelerated cell culture process development, critical quality attributes, specifications and comparability, and high throughput protein formulation development. The final day was dedicated to discussion of technology options and new analysis methods provided by emerging disruptive technologies; functional interaction, integration and synergy in platform development; and rapid and economic purification process development. © This meeting report was written for and published by Landes Bioscience in the journal, mAbs. Reichert JM, Jacob N, Amanullah A. Second International Conference on Accelerating Biopharmaceutical Development: March 9–12, 2009, Coronado, CA USA. mAbs 2009 1(3); http://www. landesbioscience.com/journals/mabs/article/8491 Biotechnol. Prog., 2009     

Overexpressed mortalin (mot-2)/mthsp70/GRP75 and hTERT cooperate to extend the in vitro lifespan of human fibroblasts

The lifespan of human foreskin fibroblasts (HFF5), cultured under standard in vitro conditions (including ambient atmospheric oxygen tension), was extended slightly by expression of exogenous mortalin (mot-2)/mthsp70/Grp75, but not by the catalytic subunit of telomerase, hTERT. Together, mot-2 and hTERT permitted bypass of senescence, a substantial extension of lifespan, and possibly immortalization. This is the first demonstration that mot-2 and telomerase can cooperate in the immortalization process.     

Mortalin imaging in normal and cancer cells with quantum dot immuno-conjugates

Quantum dots are the nanoparticles that are recently emerging as an alternative to organic fluorescence probes in cell biology and biomedicine, and have several predictive advantages. These include their ⑴broad absorption spectra allowing visualization with single light source, ⑵exceptional photo-stability allowing long term studies and ⑶narrow and symmetrical emission spectrum that is controlled by their size and material composition. These unique properties allow simultaneous excitation of different size of quantum dots with a single excitation light source, their simultaneous resolution and visualization as different colors. At present there are only a few studies that have tested quantum dots in cellular imaging. We describe here the use of quantum dots in mortalin imaging of normal and cancer cells. Mortalin staining pattern with quantum dots in both normal and cancer cells mimicked those obtained with organic florescence probes and were considerably stable.     

CARF is a novel protein that cooperates with mouse p19ARF (human p14ARF) in activating p53

The INK4a locus on chromosome 9p21 encodes two structurally distinct tumor suppressor proteins, p16(INK4a) and the alternative reading frame protein, ARF (p19(ARF) in mouse and p14(ARF) in human). Each of these proteins has a role in senescence of primary cells and activates pathways for cell cycle control and tumor suppression. The current prevailing model proposes that p19(ARF) activates p53 function by antagonizing its degradation by MDM2. It was, however, recently shown that stabilization of p53 by p14(ARF) occurs independent of the relocalization of MDM2 to the nucleolus. We have identified a novel collaborator of ARF, CARF. It co-localizes and interacts with ARF in the nucleolus. We demonstrate that CARF is co-regulated with ARF, cooperates with it in activating p53, and thus acts as a novel component of the ARF-p53-p21 pathway.     

Behavioral perinatology: biobehavioral processes in human fetal development

Behavioral perinatology is as an interdisciplinary area of research that involves conceptualization of theoretical models and conduct of empirical studies of the dynamic time-, place-, and context-dependent interplay between biological and behavioral processes in fetal, neonatal, and infant life using an epigenetic framework of development. The biobehavioral processes of particular interest to our research group relate to the effects of maternal pre- and perinatal stress and maternal-placental-fetal stress physiology. We propose that behavioral perinatology research may have important implications for a better understanding of the processes that underlie or contribute to the risk of three sets of outcomes: prematurity, adverse neurodevelopment, and chronic degenerative diseases in adulthood. Based on our understanding of the ontogeny of human fetal development and the physiology of pregnancy and fetal development, we have articulated a neurobiological model of pre- and perinatal stress. Our model proposes that chronic maternal stress may exert a significant influence on fetal developmental outcomes. Maternal stress may act via one or more of three major physiological pathways: neuroendocrine, immune/inflammatory, and vascular. We further suggest that placental corticotropin-releasing hormone (CRH) may play a central role in coordinating the effects of endocrine, immune/inflammatory, and vascular processes on fetal developmental outcomes. Finally, we hypothesize that the effects of maternal stress are modulated by the nature, duration, and timing of occurrence of stress during gestation. In this paper, we elaborate on the conceptual and empirical basis for this model, highlight some relevant issues and questions, and make recommendations for future research in this area.