Hirendra N. Banerjee

Epigenetic mechanisms in cancer

After the completion of the human genome, a need was identified by scientists to look for a functional map of the human genome. Epigenomics provided functional characteristics of genes identified in the genome. Epigenetics is the alteration in gene expression (function) without changing the nucleotide sequence. Both activation and inactivation of cancer-associated genes can occur by epigenetic mechanisms. The major players in epigenetic mechanisms of gene regulation are DNA methylation, histone deacetylation, chromatin remodeling, small noncoding RNA expression and gene imprinting. In the last few years, epigenetic mechanisms have been studied in a number of tumor types and epigenetic markers have been identified that are suitable for cancer detection, diagnosis, follow-up of treatment and screening high-risk populations. One interesting aspect of epigenetics is the reactivation of genes by successful reversion of some epigenetic changes using chemicals. The reversibility of epigenetic aberrations has made them attractive targets for cancer treatment with modulators that demethylate DNA and inhibit histone deacetylases, leading to the reactivation of silenced genes. In this article, we have described the current status of this powerful science and discussed the challenges in the clinical fields where epigenetic approaches in cancer are applied.

Use of nanotechnology for the development of novel cancer biomarkers

Novel nanotechnologies can complement and augment existing genomic and proteomic techniques employed to analyze variations across different tumor types, thus offering the potential to distinguish between normal and malignant cells. Sensitive biosensors constructed out of nanoscale components (e.g., nanocantilevers, nanowires and nanochannels) can recognize genetic and molecular events and have reporting capabilities, thereby offering the potential to detect rare molecular signals associated with malignancy. Such signals may then be collected for analysis by nanoscale harvesters that selectively isolate cancer-related molecules from tissues. Another area with near-term potential for the early detection of cancer is the identification of mutations and genomic instability.

Deciphering the finger prints of Brain Cancer Astrocytoma in comparison to Astrocytes by using near infrared Raman Spectroscopy

To explore the biochemical differences between brain cancer cells Astrocytoma and normal cells Astrocyte, we investigated the Raman spectra of single cell from these two cell types and analyzed the difference in spectra and intensity. Raman spectrum shows the banding pattern of different compounds as detected by the laser. Raman intensity measures the intensity of these individual bands.The Raman spectra of brain cancer cells was similar to those of normal cells, but the Raman intensity of cancer cells was much higher than that of normal cells.The Raman spectra of brain cancer Astrocytoma shows that the structural changes of cancer cells happen so that many biological functions of these cells are lost. The results indicate that Raman spectra can offer the experimental basis for the cancer diagnosis and treatment.

Application of nanotechnology in cancer.

Nanotechnology refers broadly to a field of applied science and technology whose unifying theme is the control of matter on the molecular level in scales smaller than 1 micrometer, normally 1 to 100 nanometers, and the fabrication of devices within that size range. In the last five years this technology has been improved tremendously in disease diagnosis and prognosis and maximum research and clinical work has been completed in cancer. The use of various pharmaceutical nanocarriers has become one of the most important areas of nanomedicine. Novel nanotechnologies can complement and augment existing genomic and proteomic techniques to analyze variations across different tumor types, thus offering the potential to distinguish between normal and malignant cells. Sensitive biosensors constructed of nanoscale components (e.g., nanocantilevers, nanowires, and nanochannels) can recognize genetic and molecular events and have reporting capabilities, thereby offering the potential to detect rare molecular signals associated with malignancy. Such signals may then be collected for analysis by nanoscale harvesters that selectively isolate cancer-related molecules from tissues. The implication of nanotechnology in cancer is discussed in this article with an emphasis on biomarker detection, imaging studies for diagnosis, and its role in therapeutic intervention.

Search for a diagnostic/prognostic biomarker for the brain cancer glioblastoma multiforme by 2D-DIGE-MS technique

The prognosis of patients with glioblastoma multiforme, the most malignant adult glial brain tumor, remains poor in spite of advances in treatment procedures, including surgical resection, irradiation, and chemotherapy. Genetic heterogeneity of glioblastoma warrants extensive studies to gain a thorough understanding of the biology of this tumor. While there have been several studies of global transcript profiling of glioma with the identification of gene signatures for diagnosis and disease management, translation into clinics is yet to happen. In the present study, we report a novel proteomic approach by using two-dimensional difference gel electrophoresis followed by spot picking and analysis of proteins/peptides by Mass spectrometry. We report at least ten different novel proteins/peptides as identified by this technique which are differentially expressed in this cancer and could be of further importance for diagnostic, therapeutic, and prognostic approaches.

Effects of 2-amino-4,6-dinitrotoluene on p53 tumor suppressor gene expression.

Abstract2-Amino-4,6-dinitrotoluene (2-Am-DNT) and its isomers are the most common metabolites of 2,4,6-trinitrotoluene (TNT). It is desirable to know the toxicity of this metabolite particularly because of its role in carcinogenicity and because it could contaminate drinking water. We used MCF-7 human breast cancer cells which have p53 tumor suppressor gene in wild type form in both the loci. Immunoblotting with p53 antibody showed enhanced p53 level in treated cells compared to untreated control cells. Similarly, p53 DNA-protein binding assays (gel-shift) showed accumulation of p53 protein in treated cells. This is the first report which shows p53 accumulation in 2-Am-DNT treated cells providing evidence of potential carcinogenic effects of 2-Am-DNT.

Molecular Diagnosis of Helicobacter Pylori Strain by 16S rDNA PCR Amplification and Direct Sequencing

Aim Rapid detection of H.pylori strains by PCR-Sequencing. Methods 16S rDNA amplification by PCR from template genomic DNA, confirmation of amplicon size by agarose gel electrophoresis, sequencing of amplicons by automated sequencer, analysis of sequences by NCBI –BLAST software. Results The PCR –Sequencing and analysis of the sequence data by BLAST resulted in detection of the strain to be of H.pylori strain#26695. Conclusion The pathogenicity of H.pylori depends on the strain of the bacteria, PCR-Sequencing and analysis of the sequence data by BLAST can be a very quick and useful diagnostic method of the pathogen.

The effect of novel rhenium compounds on lymphosarcoma, PC-3 prostate and myeloid leukemia cancer cell lines and an investigation on the DNA binding properties of one of these compounds through electronic spectroscopy.

Despite the tremendous success of cisplatin and other platinum-based anticancer drugs, severe toxicity and resistance to tumors limit their applications. It is believed that the coordination (formation of covalent bond) of the metal (platinum) to the nitrogen bases of DNA cause the ruptures of the cancer as well as normal cells. A search for anticancer drugs with different modes of action resulted in the synthesis of variety of novel compounds. Many of them are in clinical trials now. Recently we synthesized a series of novel rhenium pentylcarbonato compounds (PC1-PC6). The rhenium atom in each compound is coordinated (bonded) to a planar polypyridyl aromatic ligand, thereby forcing each compound to intercalate between the DNA bases. We have investigated the DNA binding properties of one of the PC-series of compounds (PC6) using electronic spectroscopy. The UV absorption titration of PC6 with DNA shows hypochromic effect with concomitant bathochromic shift of the charge transfer band at 290 nm. These results suggest that the compound PC6 binds to DNA through intercalation. It is therefore likely that the other PC-series of compounds will behave in a similar manner. Thus it is expected that these compounds will exhibit negligible or no side effect. We have observed that the PC-series of compounds are strong cytotoxic agents against lymphosarcoma (average GI50 ≈ 2±2.6 µM), PC-3 prostate (average GI50 ≈ 3±2.8 µM) and myeloid leukemia (average GI50 ≈ 3±2.8 µM) cancer cell lines. The average GI50 values of the PC-series of compounds are 2-3 less than the corresponding GI50 values of cisplatin. Also each of the PC-series of compounds exhibits less toxicity than cisplatin in the glomerular mesangial cells.

Synthesizing a Cellulase like Chimeric Protein by Recombinant Molecular Biology Techniques

In order to meet the Renewable Fuels Standard demands for 30 billion gallons of biofuels by the end of 2020, new technologies for generation of cellulosic ethanol must be exploited. Breaking down cellulose by cellulase enzyme is very important for this purpose but this is not thermostable and degrades at higher temperatures in bioreactors. Towards creation of a more ecologically friendly method of rendering bioethanol from cellulosic waste, we attempted to produce recombinant higher temperature resistant cellulases for use in bioreactors. The project involved molecular cloning of genes for cellulose-degrading enzymes based on bacterial source, expressing the recombinant proteins in E. coli and optimizing enzymatic activity. We were able to generate in vitro bacterial expression systems to produce recombinant His-tag purified protein which showed cellulase like activity.

Metabolomic Approaches in Cancer Epidemiology

Metabolomics is the study of low molecular weight molecules or metabolites produced within cells and biological systems. It involves technologies such as mass spectrometry (MS) and nuclear magnetic resonance spectroscopy (NMR) that can measure hundreds of thousands of unique chemical entities (UCEs). The metabolome provides one of the most accurate reflections of cellular activity at the functional level and can be leveraged to discern mechanistic information during normal and disease states. The advantages of metabolomics over other “omics” include its high sensitivity and ability to enable the analysis of relatively few metabolites compared with the number of genes and messenger RNAs (mRNAs). In clinical samples, metabolites are more stable than proteins or RNA. In fact, metabolomic profiling in basic, epidemiologic, clinical, and translational studies has revealed potential new biomarkers of disease and therapeutic outcome and has led to a novel mechanistic understanding of pathogenesis. These potential biomarkers include novel metabolites associated with cancer initiation, regression, and recurrence. Unlike genomics or even proteomics, however, the degree of metabolite complexity and heterogeneity within biological systems presents unique challenges that require specialized skills and resources to overcome. This article discusses epidemiologic studies of altered metabolite profiles in several cancers as well as challenges in the field and potential approaches to overcoming them.