pls explain this easily and detail well thanks. The challenges of selective accu

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pls explain this easily and detail well

thanks.

The challenges of selective accumulation at the tissue of interest, cellular internalization and endosomal escape are common to the delivery of all nucleic acid therapeu- tics. However, DNA delivery must also provide transport into the nucleus to allow access to the transcriptional machinery (FIG. 1). It was reported nearly 30 years ago that direct microinjection of plasmid DNA that encoded thymidine kinase into the nuclei of thymidine kinase- deficient cells resulted in expression of the kinase in 50-100% of the nuclei, as detected by the incorporation of H-thymidine into DNA following autoradiographic analyses39. However, in >1,000 cells that received cyto- plasmic injections of the same plasmid DNA, no thymi dine kinase activity was detected. The importance of the

Explanation / Answer

Selective accumulation is a process where drugs are uniformly distributed in a specific body tissue of interest. There are two ways by which the challenge of selective accumulation can be overcome. They are, 1) Cellular Internalization and 2) Endosomal Escape.

To deliver a DNA as a therapeutic, the DNA must be selectively accumulated into the nucleus because the transcriptional machinery occurred in the nucleus. As an example, it is stated that around 30 years ago, to express Thymidine Kinase in Thymidine Kinase deficient cells, the gene of Thymidine Kinase is incorporated in a plasmid DNA and the construct directly transferred into the nucleus of the gene-deficient cell by the uses of Microinjection. Microinjection is gene transfer tool where a DNA can directly transfer into a host without any genetic engineer or any gene transfer vector. The success rate of transferring the thymidine kinase gene is checked by H3 tagged thymidine, where H3 tagged where the Thymidine is present and detected with the help of autoradiography. As a result, it is detected that 50-100% of the nuclei, which are deficient in thymidine kinase, are now incorporated with thymidine kinase by selective accumulation process.

In this process around >1000 cells failed to incorporate the plasmid DNA contains the Thymidine Kinase gene, because of the nuclear envelope of these cells. They are too rigid that the microinjection failed to transfect those cells. Thus Nuclear uptake of the synthetic Gene of Interest(GOI) is the rate-limiting step or the slowest step of 'Selective Accumulation' process.

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