Illumination for the tumor
Krasnoyarsk is developing a test system for cancer markers based on luminous hybrid proteins
Early and accurate diagnosis of oncological diseases remains one of the urgent problems of healthcare. Krasnoyarsk scientists have created genetically engineered hybrid proteins, including a fluorescent label, which can be used to diagnose cancer using enzyme immunoassay.
Today, different approaches are used to diagnose oncological diseases. One of them is the determination of the so–called tumor markers – tumor proteins in the patient's blood samples. Such, for example, as survivin or melanoma inhibitory activity protein (MIA).
Survivin, which belongs to the inhibitors of apoptosis ("cell suicide"), plays an important role in the processes of cell division and migration and participates in the regulation of tumor processes. In healthy tissues of an adult, this protein is not present, but it is present in the cells of almost all malignant tumors. The MIA protein is primarily associated with melanoma, a very aggressive neoplasm originating from pigmented skin cells. It is produced mainly by the cells of this tumor, as well as some others.
Currently, enzyme immunoassay (ELISA) based on the antigen-antibody reaction is used to determine these and other cancer markers. In this case, we are talking about a very specific binding of a certain part of the tumor protein from a blood sample with a test antibody included in the diagnostic kit. These antibodies in a certain concentration are applied to the wells on the surface of the immunological tablet, where they bind to the target molecule.
Such diagnostic kits have previously been distinguished by a considerable cost, and for MIA protein and survivin in our country they are not produced at all. In addition, the kits available on the market are not intended for use in clinical practice. They belong to the so-called "sandwich type", i.e. they use two antibodies aimed at different parts of the protein, which affects the cost of analysis, since obtaining antibodies is a time–consuming and expensive process.
Scientists from the Institute of Biophysics of the Krasnoyarsk Scientific Center of the Siberian Branch of the Russian Academy of Sciences and Siberian Federal University are now engaged in creating original test systems for fast and accurate detection of tumor markers in the blood. Using genetic engineering methods, they created bifunctional hybrid proteins consisting of the photoprotein obelin and one of two cancer markers – survivin or MIA. Representing an artificial "analogue" of tumor proteins, they can also bind to specific antibodies.
The analysis is based on the competition between hybrid proteins and cancer proteins from blood serum for binding to antibodies on the surface of the immunological tablet. To detect the reaction, the phenomenon of bioluminescence is used: the luminescent mark is an obelin, which glows with a bright blue light when interacting with calcium. Test hybrid proteins are always added in the same amount, and the amount of the target tumor protein, for obvious reasons, can vary greatly. If the latter is not enough, it will lose in this competition and the bioluminescent signal will be higher, and vice versa.
The scientists tested the effectiveness of hybrid proteins on a model test system with commercial antibody preparations. The test was successful: as a result, it was possible to determine the concentration of MIA and survivin cancer markers in the range close to the diagnostically significant. And the almost instantaneous and bright reaction of the bioluminescence of the obelin provides fast and accurate detection of the target, which is important in the diagnosis and monitoring of diseases.
Hybrid proteins were not only active, but also stable. They can be stored frozen in solution for up to six months: the loss of bioluminescent activity during this time was only 10-15%.
According to one of the authors of the work, Candidate of Biological Sciences E.E. Bashmakova (IBF SB RAS), "using the example of two cancer markers, we have shown good prospects for using these hybrids as luminous labels. The test developed by us is carried out in one stage and uses only one antibody, which significantly reduces the cost of analysis."
Article by Bashmakova et al. N-extended photoprotein obelin to competitively detect small protein tumor markers is published in the journal Biochemical and Biophysical Research Communications.
It should be noted here that this development itself was originally aimed at creating a simpler and cheaper test for conducting scientific research. Currently, work is underway to replace imported antibodies in this system with other specific molecules that could bind the cancer protein. Time will tell how successful these searches will be. After all, modern science is inherently cosmopolitan, and any attempt at self-isolation negatively affects, first of all, its technical and material base and, accordingly, the effectiveness of scientific research.
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