Drug and Alcohol Rehabilitation: In Vivo Cell Reprogramming Towards Pluripotency by Virus-Free Overexpression of Defined Factors.

In Vivo Cell Reprogramming towards Pluripotency by Virus-Free Overexpression of Defined Factors.

Filed under: Drug and Alcohol Rehabilitation

PLoS One. 2013; 8(1): e54754
Yilmazer A, de Lázaro I, Bussy C, Kostarelos K

The ability to induce the reprogramming of somatic mammalian cells to a pluripotent state by the forced expression of specific transcription factors has helped redefine the rules of cell fate and plasticity, as well as open possibilities for disease modeling, drug screening and regenerative medicine. Here, we hypothesized that the non-viral forced expression of the four originally discovered defined factors (OKSM) in adult mice could result in in vivo reprogramming of cells in the transfected tissue in situ. We show that a single hydrodynamic tail-vein (HTV) injection of two plasmids encoding for Oct3/4, Sox2, Klf4 and c-Myc respectively, are highly expressed in the liver tissue of Balb/C adult mice. Hallmark pluripotency markers were upregulated within 24-48 h after injection, followed by down-regulation of all major hepatocellular markers. Generation of transcriptionally reprogrammed cells in vivo was further confirmed by positive staining of liver tissue sections for all major pluripotency markers in Balb/C mice and the Nanog-GFP reporter transgenic strain (TNG-A) with concomitant upregulation of GFP expression in situ. No signs of physiological or anatomical abnormalities or teratoma formation were observed in the liver examined up to 120 days. These findings indicate that virus-free expression of OKSM factors in vivo can transcriptionally reprogram cells in situ rapidly, efficiently and transiently, absent of host tissue damage or teratoma formation.
HubMed – drug

Global Transcriptional Profiling of Longitudinal Clinical Isolates of Mycobacterium tuberculosis Exhibiting Rapid Accumulation of Drug Resistance.

Filed under: Drug and Alcohol Rehabilitation

PLoS One. 2013; 8(1): e54717
Chatterjee A, Saranath D, Bhatter P, Mistry N

The identification of multidrug resistant (MDR), extensively and totally drug resistant Mycobacterium tuberculosis (Mtb), in vulnerable sites such as Mumbai, is a grave threat to the control of tuberculosis. The current study aimed at explaining the rapid expression of MDR in Directly Observed Treatment Short Course (DOTS) compliant patients, represents the first study comparing global transcriptional profiles of 3 pairs of clinical Mtb isolates, collected longitudinally at initiation and completion of DOTS. While the isolates were drug susceptible (DS) at onset and MDR at completion of DOTS, they exhibited identical DNA fingerprints at both points of collection. The whole genome transcriptional analysis was performed using total RNA from H37Rv and 3 locally predominant spoligotypes viz. MANU1, CAS and Beijing, hybridized on MTBv3 (BuG@S) microarray, and yielded 36, 98 and 45 differentially expressed genes respectively. Genes encoding transcription factors (sig, rpoB), cell wall biosynthesis (emb genes), protein synthesis (rpl) and additional central metabolic pathways (ppdK, pknH, pfkB) were found to be down regulated in the MDR isolates as compared to the DS isolate of the same genotype. Up regulation of drug efflux pumps, ABC transporters, trans-membrane proteins and stress response transcriptional factors (whiB) in the MDR isolates was observed. The data indicated that Mtb, without specific mutations in drug target genes may persist in the host due to additional mechanisms like drug efflux pumps and lowered rate of metabolism. Furthermore this population of Mtb, which also showed reduced DNA repair activity, would result in selection and stabilization of spontaneous mutations in drug target genes, causing selection of a MDR strain in the presence of drug pressures. Efflux pump such as drrA may play a significant role in increasing fitness of low level drug resistant cells and assist in survival of Mtb till acquisition of drug resistant mutations with least fitness cost.
HubMed – drug

Suppression of AKT Anti-Apoptotic Signaling by a Novel Drug Candidate Results in Growth Arrest and Apoptosis of Hepatocellular Carcinoma Cells.

Filed under: Drug and Alcohol Rehabilitation

PLoS One. 2013; 8(1): e54595
Cuconati A, Mills C, Goddard C, Zhang X, Yu W, Guo H, Xu X, Block TM

Hepatocellular carcinoma (HCC) is the third most common cause of cancer fatalities worldwide, with limited treatment options and five year survival rates of between <5 and 15%. To address this medical need, we conducted a screen of a drug-like small molecule library for HCC-selective cytotoxins. We report here the identification of a disubstituted aminothiazole termed HBF-0079, with remarkable selective toxicity for HCC-derived cell lines versus non-HCC liver lines and most other cancer lines. HBF-0079 caused irreversible growth arrest and apoptosis of the HCC lines Huh7, Hep3B, HepaRG as well as the hepatoblastoma line HepG2, with CC(50) values from ?0.7-7.7 µM, while more than 45 µM was needed to achieve CC(50) values for the immortalized normal hepatocyte lines THLE-2 and PH5CH. Of the sixty cancer lines from the National Cancer Institute panel, only five exhibited >50% growth inhibition by HBF-0079. In Huh7 cells, HBF-0079 induced cell cycle arrest in G1 and concomitant apoptosis, and its effects were irreversible after removal of the compound. These observations corroborate a loss of AKT phosphorylation at the mTORC2-targeted residue S473, with concurrent loss of phosphorylation of the mTORC1 targets SK6 and 4EBP1 in Huh7 but not PH5CH cells. Finally, growth of Hep3B-derived tumors in a murine xenograft model was significantly repressed by the compound through either systemic or intratumoral administration of formulated HBF-0079. The potential for development of this drug candidate is discussed.
HubMed – drug

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