Research Article: Promoters with Cancer Cell-Specific Activity for Melanoma Gene Therapy

Date Published: , 2011

Publisher: A.I. Gordeyev

Author(s): V.V. Pleshkan, I.V. Alekseenko, M.V. Zinovyeva, T.V. Vinogradova, E.D. Sverdlov.

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Abstract

Melanoma is one of the most aggressive tumors. It develops from pigment-forming cells (melanocytes) and results in a high number of lethal outcomes. The use of genetic constructs with the ability to specifically kill melanoma cells, but not normal cells, might increase the lifespan of patients, as well as improve their quality of life. One of the methods to achieve a selective impact for therapeutic genes on cancer cells is to utilize a transcriptional control mechanism using promoters that are specifically activated only in cancerous cells. In this review, promoters of the genes that are preferentially expressed in melanoma cells are described. These promoters, and other highly melanoma-specific regulatory elements, reduce the unspecific expression of therapeutic genes in normal tissues. Moreover, cancer-specific promoters and their elements are advantageous for the development of universal anticancer drugs. Examples of the use of double promoters that have a high potential as instruments in cancer gene therapy are also given in this review.

Partial Text

A steady increase in deaths caused by malignant skin melanoma has recently been in evidence around the world, including in Russia. Malignant melanoma belongs to the most aggressive variety of tumors; the five-year survival rate of patients is less than 50%. Melanoma especially stands out by its early metastazing; therefore, chemotherapy and radiation therapy are not very effective against the disease [1–3].

The most well-studied promoter modules that control the specific expression of the therapeutic gene in melanoma cells, with recent widespread application, are promoters of the tyrosinase gene ( TYR ) or the gene encoding the melanoma inhibitory activity ( MIA ), sometimes combined with distal elements of other promoters and/or the enhancers [21, 22].

Another approach that makes it possible to specifically control transgene expression in melanoma cells is the use of promoters that are specific not only in melanoma, but in other tumor cells as well. The examples of such tumor-specific promoters (TSP) include promoters of the genes TERT , Cox-2, CXCR4 , and  BIRC5 , for which overexpression of the genes controlled by them in numerous tumor types and the absence or a minimal expression in the normal tissues is typical.

The controlling elements of the known promoters that have been characterized are used to construct de novo specific promoter modules. For instance, artificial promoters based on the elements of the promoters of the human tyrosinase and α-fetoprotein (AFP1) genes possessing strong and specific expression in melanoma cell lines were constructed [88]. As previously mentioned, the tyrosinase promoter contains the M-box, the conservative element that is common to melanocyte-specific promoters [89]. This element was used in combination with the elements from the 5’-region of the AFP1 promoter – the GRE element, which is specific for the cell cycle and the AP1-binding element. Several efficient melanocyte-specific promoters were obtained upon different combinations of one or several copies of fragments of the tyrosinase and α-fetoprotein promoters – the M-box, AP1 and GRE elements. These promoters were selectively active in the B16 melanoma line, but not in the HeLa cell line [88]. The length of the artificial constructs was not more than 300 bp; the promoter consisting of three GRE, three AP1 elements, and two M-boxes was the most efficient one. It was ascertained that if the number of regulatory elements of the promoter in the chimeric construct is higher than eight units, a loss in promoter specificity is observed [88]. It seems that the activity of synthetic promoters is dependent both upon the number of regulatory elements and upon the vector. The optimal number of regulatory elements has to be selected in each case. For example, it was shown in the above-mentioned study by Rothfels et al . [22] that it is sufficient to bind four copies of the enhancers of the mouse tyrosinase gene in order to increase the activity of both the MIA and TYR promoters.

Melanoma treatment is associated with a number of difficulties, including the high resistance of melanoma cells and early metastazing, which determine the unfavourable prognosis. The necessity to affect the metastatic loci disseminated over the entire organism requires the systemic administration of antimelanoma agents, which involves a certain risk that other cells of the organism can be affected. Gene therapy, which has known rapid development, offers new methods that allow to increase the specificity of the effect on melanoma cells whilst simultaneously decreasing the probability of damaging healthy cells. The use of melanoma-specific promoters makes it possible to specifically affect melanoma cells. These methods have recently gone into the stage of development and are permanently being improved to find the most efficient solutions, starting with the selection of optimal regulatory elements, the designing of constructs based on these elements, and ending with the search for new vectors, with both natural viruses or artificially constructed systems for packing the genetic material being used as such vectors [94, 95]. One can hope that a simple and efficient system for the elimination of melanomas and its metastases will be designed.

 

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