Benny Abraham Kaipparettu

Graphene Quantum Dots Derived from Carbon Fibers

Graphene quantum dots (GQDs), which are edge-bound nanometer-size graphene pieces, have fascinating optical and electronic properties. These have been synthesized either by nanolithography or from starting materials such as graphene oxide (GO) by the chemical breakdown of their extended planar structure, both of which are multistep tedious processes. Here, we report that during the acid treatment and chemical exfoliation of traditional pitch-based carbon fibers, that are both cheap and commercially available, the stacked graphitic submicrometer domains of the fibers are easily broken down, leading to the creation of GQDs with different size distribution in scalable amounts. The as-produced GQDs, in the size range of 1-4 nm, show two-dimensional morphology, most of which present zigzag edge structure, and are 1-3 atomic layers thick. The photoluminescence of the GQDs can be tailored through varying the size of the GQDs by changing process parameters. Due to the luminescence stability, nanosecond lifetime, biocompatibility, low toxicity, and high water solubility, these GQDs are demonstrated to be excellent probes for high contrast bioimaging and biosensing applications.

Multilevel Genomics-Based Taxonomy of Renal Cell Carcinoma

On the basis of multidimensional and comprehensive molecular characterization (including DNA methalylation and copy number, RNA, and protein expression), we classified 894 renal cell carcinomas (RCCs) of various histologic types into nine major genomic subtypes. Site of origin within the nephron was one major determinant in the classification, reflecting differences among clear cell, chromophobe, and papillary RCC. Widespread molecular changes associated with TFE3 gene fusion or chromatin modifier genes were present within a specific subtype and spanned multiple subtypes. Differences in patient survival and in alteration of specific pathways (including hypoxia, metabolism, MAP kinase, NRF2-ARE, Hippo, immune checkpoint, and PI3K/AKT/mTOR) could further distinguish the subtypes. Immune checkpoint markers and molecular signatures of T cell infiltrates were both highest in the subtype associated with aggressive clear cell RCC. Differences between the genomic subtypes suggest that therapeutic strategies could be tailored to each RCC disease subset.

Mechanisms of estrogen receptor antagonism toward p53 and its implications in breast cancer therapeutic response and stem cell regulation

Estrogen receptor α (ERα) plays an important role in the onset and progression of breast cancer, whereas p53 functions as a major tumor suppressor. We previously reported that ERα binds to p53, resulting in inhibition of transcriptional regulation by p53. Here, we report on the molecular mechanisms by which ERα suppresses p53’s transactivation function. Sequential ChIP assays demonstrated that ERα represses p53-mediated transcriptional activation in human breast cancer cells by recruiting nuclear receptor corepressors (NCoR and SMRT) and histone deacetylase 1 (HDAC1). RNAi-mediated down-regulation of NCoR resulted in increased endogenous expression of the cyclin-dependent kinase (CDK)-inhibitor p21Waf1/Cip1 (CDKN1A) gene, a prototypic transcriptional target of p53. While 17β-estradiol (E2) enhanced ERα binding to p53 and inhibited p21 transcription, antiestrogens decreased ERα recruitment and induced transcription. The effects of estrogen and antiestrogens on p21 transcription were diametrically opposite to their known effects on the conventional ERE-containing ERα target gene, pS2/TFF1. These results suggest that ERα uses dual strategies to promote abnormal cellular proliferation: enhancing the transcription of ERE-containing proproliferative genes and repressing the transcription of p53-responsive antiproliferative genes. Importantly, ERα binds to p53 and inhibits transcriptional activation by p53 in stem/progenitor cell-containing murine mammospheres, suggesting a potential role for the ER–p53 interaction in mammary tissue homeostasis and cancer formation. Furthermore, retrospective studies analyzing response to tamoxifen therapy in a subset of patients with ER-positive breast cancer expressing either wild-type or mutant p53 suggest that the presence of wild-type p53 is an important determinant of positive therapeutic response.

Hybrid 2D nanomaterials as dual-mode contrast agents in cellular imaging

The design of multifunctional nanofluids is highly desirable for biomedical therapy/cellular imaging applications.[1–4] The emergence of hybrid nanomaterials with specific properties, such as magnetism and fluorescence, can lead to an understanding of biological processes at the biomolecular level.[1] Various hybrid systems have been analyzed in the recent past for several possible biomedical applications.[5–9] Carbon-based hybrid systems such as carbon nanotubes with various nanoparticles are being widely tested for their biological applications because of their ability to cross cell membranes and their interesting thermal and electrical properties.[10,11] Graphene oxide (GO) is a fairly new graphene-based system with a 2D carbon honeycomb lattice decorated with numerous functional groups attached to the backbone: these functional groups make it an excellent platform for further attachment of nanoparticles and synthesis of hybrid materials. Cell viability studies on GO have been recently attempted, showing biocompatibility. [12,13] Moreover, the intrinsic photoluminescence (PL) properties of GO can be utilized for cellular imaging.[13] The large surface area and non-covalent interactions with aromatic molecules make GO an excellent system for biomolecular applications and drug attachment.

Highly luminescent-paramagnetic nanophosphor probes for in vitro high-contrast imaging of human breast cancer cells.

Highly luminescent-paramagnetic nanophosphors have a seminal role in biotechnology and biomedical research due to their potential applications in biolabeling, bioimaging, and drug delivery. Herein, the synthesis of high-quality, ultrafine, europium-doped yttrium oxide nanophosphors (Y(1.9)O(3):Eu(0.1)(3+)) using a modified sol-gel technique is reported and in vitro fluorescence imaging studies are demonstrated in human breast cancer cells. These highly luminescent nanophosphors with an average particle size of ≈6 nm provide high-contrast optical imaging and decreased light scattering. In vitro cellular uptake is shown by fluorescence microscopy, which visualizes the characteristic intense hypersensitive red emission of Eu(3+) peaking at 610 nm ((5)D(0)-(7)F(2)) upon 246 nm UV light excitation. No apparent cytotoxicity is observed. Subsequently, time-resolved emission spectroscopy and SQUID magnetometry measurements demonstrate a photoluminescence decay time in milliseconds and paramagnetic behavior, which assure applications of the nanophosphors in biomedical studies.

Fatty Acid Oxidation-Driven Src Links Mitochondrial Energy Reprogramming and Oncogenic Properties in Triple-Negative Breast Cancer

Transmitochondrial cybrids and multiple OMICs approaches were used to understand mitochondrial reprogramming and mitochondria-regulated cancer pathways in triple-negative breast cancer (TNBC). Analysis of cybrids and established breast cancer (BC) cell lines showed that metastatic TNBC maintains high levels of ATP through fatty acid β oxidation (FAO) and activates Src oncoprotein through autophosphorylation at Y419. Manipulation of FAO including the knocking down of carnitine palmitoyltransferase-1A (CPT1) and 2 (CPT2), the rate-limiting proteins of FAO, and analysis of patient-derived xenograft models confirmed the role of mitochondrial FAO in Src activation and metastasis. Analysis of TCGA and other independent BC clinical data further reaffirmed the role of mitochondrial FAO and CPT genes in Src regulation and their significance in BC metastasis.

Crosstalk from Non-Cancerous Mitochondria Can Inhibit Tumor Properties of Metastatic Cells by Suppressing Oncogenic Pathways

Mitochondrial-nucleus cross talks and mitochondrial retrograde regulation can play a significant role in cellular properties. Transmitochondrial cybrid systems (cybrids) are an excellent tool to study specific effects of altered mitochondria under a defined nuclear background. The majority of the studies using the cybrid model focused on the significance of specific mitochondrial DNA variations in mitochondrial function or tumor properties. However, most of these variants are benign polymorphisms without known functional significance. From an objective of rectifying mitochondrial defects in cancer cells and to establish mitochondria as a potential anticancer drug target, understanding the role of functional mitochondria in reversing oncogenic properties under a cancer nuclear background is very important. Here we analyzed the potential reversal of oncogenic properties of a highly metastatic cell line with the introduction of non-cancerous mitochondria. Cybrids were established by fusing the mitochondria DNA depleted 143B TK- ρ0 cells from an aggressive osteosarcoma cell line with mitochondria from benign breast epithelial cell line MCF10A, moderately metastatic breast cancer cell line MDA-MB-468 and 143B cells. In spite of the uniform cancerous nuclear background, as observed with the mitochondria donor cells, cybrids with benign mitochondria showed high mitochondrial functional properties including increased ATP synthesis, oxygen consumption and respiratory chain activities compared to cybrids with cancerous mitochondria. Interestingly, benign mitochondria could reverse different oncogenic characteristics of 143B TK- cell including cell proliferation, viability under hypoxic condition, anti-apoptotic properties, resistance to anti-cancer drug, invasion, and colony formation in soft agar, and in vivo tumor growth in nude mice. Microarray analysis suggested that several oncogenic pathways observed in cybrids with cancer mitochondria are inhibited in cybrids with non-cancerous mitochondria. These results suggest the critical oncogenic regulation by mitochondrial-nuclear cross talk and highlights rectifying mitochondrial functional properties as a promising target in cancer therapy.

Mitochondria of highly metastatic breast cancer cell line MDA-MB-231 exhibits increased autophagic properties.

Autophagy is a cellular housekeeping process that removes damaged or unwanted cellular components and recycles them to build new constituents. It is essential for tumor growth under adverse environment. Mitochondria play an important role in the formation of autophagosome and its subsequent docking and fusion with lysosome. To understand the contribution of mitochondria to the regulation of homeostatic autophagy in cancer cells, we used the transmitochondrial cytoplasmic hybrid (cybrid) model. Cybrid system allowed us to compare mitochondria from different cell types including highly metastatic breast cancer cell line MDA-MB-231 (c231), less metastatic breast cancer cell lines: MDA-MB-436 (c436) and MDA-MB-468 (c468), as well as non-cancerous mammary epithelial cell MCF-10A (c10A) in a defined nuclear background. The c231 exhibited lower LC3-II levels but higher ratio of LC3-II/LC3-I than c436, c468 and c10A. In addition, c231 displayed more punctate LC3-positive cells and had lower levels of sequestosome 1 (p62/SQSTM1) than other cybrids. These suggested that mitochondria could contribute to the increased autophagy and autophagic flux in metastatic cancer. This increased autophagy was found to be non-selective autophagy instead of selective mitophagy since LC3 puncta in c231 did not co-localize with mitochondria labeled by Mitotracker red or Tomm 20. The promotion of mitochondrial permeability transition (MPT) in c231 also contributed to increased autophagy. Block of MPT by the inhibition of low-conductance stage of MPT pores resulted in a decrease of LC3 puncta in c231. These results suggested that mitochondria from highly metastatic breast cancer cell line MDA-MB-231 can promote homeostatic autophagy of cancer through opening low-conductance MPT pores.

Functional effects of cancer mitochondria on energy metabolism and tumorigenesis: utility of transmitochondrial cybrids

Reprogramming of energy metabolism is one of the hallmarks of cancer. In normal conditions, cells rely on mitochondrial oxidative phosphorylation to provide energy for cellular activities. Cancer cells are characterized by increased glycolysis and reduced mitochondrial respiratory function. In the past decade, somatic mitochondrial DNA alterations are found to be common in all types of cancers. However, the functional significance of the altered cancer mitochondria is largely unknown. This is because the bulk of cancer properties are regulated by nuclear encoded genes. To overcome this problem, the transmitochondrial cybrid system, which allows the study of the effect of cancer mitochondria in a common nuclear background, has been used. Here we review the accumulating evidence that altered cancer mitochondria affect the respiratory chain function and oncogenic properties in vitro and in vivo using cybrid technologies.

Probing Highly Luminescent Europium-Doped Lanthanum Orthophosphate Nanorods for Strategic Applications

Herein we have established a strategy for the synthesis of highly luminescent and biocompatible europium-doped lanthanum orthophosphate (La0.85PO4Eu0.15(3+)) nanorods. The structure and morphogenesis of these nanorods have been probed by XRD, SEM, and TEM/HRTEM techniques. The XRD result confirms that the as-synthesized nanorods form in a monazite phase with a monoclinic crystal structure. Furthermore, the surface morphology shows that the synthesized nanorods have an average diameter of ∼90 nm and length of ∼2 μm. The HRTEM images show clear lattice fringes that support the presence of better crystal quality and enhanced photoluminescence hypersensitive red emission at 610 nm ((5)D0-(7)F2) upon 394 nm wavelength excitation. Furthermore, time-resolved spectroscopy and an MTT assay of these luminescent nanorods demonstrate a photoluminescent decay time of milliseconds with nontoxic behavior. Hence, these obtained results suggest that the as-synthesized luminescent nanorods could be potentially used in invisible security ink and high-contrast bioimaging applications.