Project
Undertaken / Involved in the laboratory:
1. Effect of ‘Quinacrine’ on human lung, colon and breast cancer:
(Funded by DBT, Cancer Mission Pilot Grant, Govt. of India, GOA Cancer Society, India):
Non-small cell lung cancer which accounts for nearly 80% of all lung cancer cases is highly metastatic and invasive in nature even in initial stages of diagnosis. As per statistical data the 5-year survival rate after diagnosis is roughly around 50% followed by death due to metastatic spread. In spite of its clinical importance, the underlying cellular and molecular mechanisms of cancer metastasis are only partially understood. Thus, improved knowledge of how cancer cells acquire metastatic traits is necessary to unravel novel drug targets. Quinacrine (QC), a derivative of 9-aminoacridine is shown to have anti-cancer properties in breast, colon and gastric cancer cells. However, its effect on lung cancer cells has not been reported yet. We have investigated the effect of QC on two NSCLC cell lines namely, A549 (type II alveolar carcinoma) and NCI H520 (squamous cell carcinoma). Our ongoing work reveled significant modulation in the cellular morphology of NSCLC cells with increasing dose due to exposure to QC. Starting from formation of stress fibers and lamellipodial extensions at lower concentration exposures to rounding-up to a spheroid structure in higher concentration exposures; indicating changes in the cytoskeletal elements. There were changes in the expression pattern of small GTPases at mRNA as well as protein level. QC causes generation of reactive oxygen species (ROS) and activation of p38MAPK and SAPK/JNK pathway leading to mitochondrial apoptotic process of cell death. Results of Hoechst staining and FACS analysis showed that QC increased cell death in a dose-dependent manner by decreasing the G1 and of G2 cell populations while increasing the sub- G1 population which accounts for fragmented DNA. Collectively, the results presented here suggest QC has efficient anti-cancer potential against NSCLC cells via not only activation of p53 signaling pathway, but also via induction of G1/S arrest. We have also observed the differences of susceptibility in two different human colon cancer cell lines namely, HCT116 and INT407. Presence of stress granules, rounding-off of the cells and loss of cell-cell contact characterized the cellular effects. Expression pattern of smGTPases in both at the mRNA level as well as at the protein level reveled significant amount of alteration in these genes. Significant difference in the expression pattern of HSP70 and p53 was also observed. HCT116 and INT407 wound healing assay gave comparative results showing decrease in contact inhibition property depends on concentration of QC.
2. Nanoparticles and its effect at cellular and molecular level:
(Funded by DAE, Govt. of India)
Metal oxides (MeOx) are exponentially being used in a wide range of applications and are the largest class of commercially produced nanomaterials. This presents unprecedented human exposure. Thus, understanding nanoparticle induced cellular stress can greatly help design strategies to combat them. Scores of studies have been carried out to understand the effects of MeOx nanoparticle exposure on human alveolar cells, which are highly susceptible to aerosolized matter. There is a huge redundancy of information generated, also, a lack of a comprehensive conglomeration of this information. We have built an understanding of the various cellular responses as a direct consequence of MeOx nanoparticle exposure on human alveolar (A549) cells. Detailed accounts of cellular morphology modulation, generation of reactive oxygen species (ROS) and oxidative stress, inflammation and cytokine release, genotoxic and epi-genotoxic insults, toxicological trend, nanoparticle internalization, modes of cell death, protein synthesis, and membrane damage among others are discussed. Finally, to aid predictability of the highly dynamic and multifactorial nature of this toxicity, we have hypothesized models that describe the ensuing mechanisms based on common patterns.
Martin and Sarkar, 2018, Nanotoxicology
3. Effect of ‘Snake venom’ toxicity on human cancer cells: [Along with Prof. Dibakar Chakrabarty] (Funded by ICMR, , Govt. of India):
‘Daboialectin’, a low molecular weight C-type lectin (18.5 kDa) isolated from Russell's viper venom showed cytotoxic effects on human broncho-alveolar carcinoma derived (A549) cell lines. Daboialectin induced inhibition of A549 cell growth was time and concentration dependent with severe morphological changes by altering the functions of small GTPases such as Rac, Rho and cdc42. ROS induced DNA damage may result in apoptosis by inducing disruption of membrane integrity, blebbing and nuclear disintegration by activating caspases. Our results indicate that Daboialectin, a snake c type lectin (Snaclec) isolated from RVV alters morphology of A549 cells via regulation of cytoskeleton through RHOGTPases. On other hand, the HSP70 and some other anti-apoptotic proteins required for the survival of cancer cells was found to be down-regulated in presence of Daboialectin. Daboialectin was also found to be inhibitory to anti-adhesive and anti-invasive to A549 cells in vitro. Daboialectin is the ?rst Snaclec reported to induce cytoskeletal changes through regulation of RHO-GTPases and blocking anti-apoptotic pathway for a cancer cell line.
@Toxicon, 2018
4. Effect of vitamin B12 on cadmium toxicity on human cells cultured in vitro: (Funded by BITS Pilani as well as student supported by CSIR, Govt. of India):
Cadmium is a heavy metal element known to be toxic to the biological system and is present in significant quantities in cigarette smoke that can cause considerable pulmonary damage. Methylcobalamin (MeB12) is a vitamin B12 analogue with known antioxidant properties, whose effect on metal induced stress recovery is not explored much. This study was initiated to see whether MeB12 can reduce cadmium induced stress/ toxicity in A549 cells. A549 cells were grown in MeB12 containing media followed by exposure to increasing concentrations of CdCl2. Surprisingly, not only did MeB12 not help the cells reduce the metal induced stress, but also enhanced their sensitivity towards cadmium. Microscopic analysis, cytotoxic assays and DNA damage studies revealed that, MeB12 enhanced cadmium induced cell death in human A549 cells. An interaction study using isobolographic analysis revealed a synergistic relation between these two compounds. Studies on the promoter methylation of metallothionenin genes; MT1A and MT2A revealed that MT1A promoter is already heavily methylated in A549 cell line, while MT2A appeared to be methylated only in the presence of CdCl2 and MeB12. This suggests that the epigenetic modification of MT2A is one of the contributing factors towards the enhanced cadmium sensitivity in A549 cells in the presence of MeB12.
