Early diagnostics and detection of malignancies is an important medical problem. Various methods are used for cancer diagnostics, such as X-ray, endoscopic, ultrasonographic, morphologic, as well as measurement of the DNA concentration in blood or serum of malignant patients. However, these methods are inefficient for the detection of malignant transformation and at early disease stages.
A method of early diagnostics of cancer is known, which is based on the measurement of blood levels of intracellular nucleic acids bound to cellular surface of hemocytes (Russian Federation Patent No. 2251696, published on May 10, 2005). The zero value of this parameter is attributed to malignancy.
A main drawback of the aforementioned method is in that it is intricate.
Another method of diagnostics of malignancies is known which is based on the measurements of blood biochemical values such as levels of vitamins A and E and iron-dependent organ-specific antibody-like proteins possessing false SOD-activities (Russian Federation Patent No. 2021612, published on Oct. 15, 1994).
However, this method is time-consuming, highly expensive, and requires special equipment to be used.
Most closely related to the claimed method is a method for qualitative early express diagnostics of malignancy (see Russian Federation Patent No. 2309405, published on Oct. 27, 2007) that involves the following steps: collection of a blood sample; addition of a reagent containing a polar solvent and a non-polar water-soluble alcohol; filtration of the mixture; separation of the upper colored water-alcoholic solution (a supernatant); and evaluation of the reaction outcome by comparing the color of the filtrate with colored controls prepared in a similar way from the blood of a knowingly healthy subject and malignant patients at different stages of the disease.
However, the above-described method does consider an immune reaction of the body because it is based on a non-specific reaction. Furthermore, it requires a considerable amount of blood (25 mL) to be collected, which can make the patients feel worse.
The goal of the present invention is to develop a simple, efficient and precise method of early express diagnostics of malignancies which would consider the body's immune reaction to the presence of malignant cells and their activities products and also be of low cost.
To achieve the goal, the method of precise early express diagnostics of malignancies, which involves collecting a blood sample, mixing it with a reagent and evaluating the reaction outcome, is modified, in accordance with the present invention, by additionally preparing 0.12-0.5% suspensions of citrated blood and washed erythrocytes, both from the same patient; using, as a reagent, a supernatant liquid obtained as a result of growing a porcine embryo kidney finite cell culture in anaerobic conditions; separately mixing suspensions of citrated blood and washed erythrocytes with the reagent and incubating at 4-8° C. for 12-15 h; and separately evaluating the outcomes of reactions of citrated blood and washed erythrocytes. Based on this method, the patient is concluded malignant if the reaction of erythrocytes is positive while the reaction of citrated blood is either negative or positive, as well as if the reaction of erythrocytes is negative and the reaction of citrated blood is positive.
To the authors' knowledge, no data are available in the related art patents and scientific technical literature on an efficient, simple, and quick method of qualitative preliminary express diagnostics of malignancies using a reagent in the form of a supernatant liquid obtained as a result of growing a porcine embryo kidney finite cell culture (SPEV-line) in anaerobic conditions.
The method is based on—evaluating the outcomes of reactions of both citrated blood and washed erythrocytes obtained from the same patient with the reagent in the form of a supernatant liquid obtained as a result of growing a porcine embryo kidney finite cell culture (SPEV-line), and—the phenomenon of immune response regulation.
Previous studies demonstrated the presence of C and D onco-RNA-viruses in the latent form in the SPEV cell culture that was grown in anaerobic conditions (see Laskavyi, V. N., Dyakonova, L. P., and Melnikova, G. V. Culture and accumulation of oncogenic viruses in anaerobic conditions. Veterinamyi Vrach, 2004, no. 2, pp 32-38). In the studies, the cells were grown in the Eagle 199 medium for 3 to 5 days at a medium-to-air phase ratio of 1:5.
The authors have used a supernatant liquid obtained as a result of growing SPEV cells containing C and D viruses in latent form for malignancies diagnostics. The SPEV cells culture is commonly used as a substrate to isolate and accumulate various viral agents.
The phenomenon of immune response regulation registered as Discovery No. 385 dated Jun. 28, 2006, consists in that a system of self-regulation of an organism's immunity involves, apart from T- and B-lymphocytes, red blood cells. Being a part of the hematopoietic system, the immunocompetent cells are capable of interacting with other components of hematopoiesis (in particular, with erythroids) and experience regulatory effects caused by the latters.
The mechanism of reaction of citrated blood and washed erythrocytes is based on a natural interaction between erythrocytes having immunoglobulins or immunoglobulin complexes on their surfaces with a reagent, being a supernatant liquid obtained as a result of growing SPEV cells in the anaerobic conditions (for 3 to 5 days at a medium-to-air phase ratio of 1:5) which contains C and D onco-RNA-viruses in the latent form.
Thus, the immunoglobulins absorbed on the surface of erythrocytes can interact with activities products of the malignant cells. It is this interaction that appears as reactions of citrated blood and erythrocytes with the reagent.
When malignant cells are in excess, the reaction with citrated blood may not occur. However, washed erythrocytes which are free of activities products of malignant cells can enter into a reaction with the reagent, giving a positive reaction.
There is known a method of diagnostics of infectious diseases in animals that is based on evaluation of the outcomes of reaction of citrated blood and agglutination of erythrocytes, both from the same animal and in the presence of the same antigen (Russian Federation Patent No. 2111492, published on May 20, 1998). However, the mentioned method is designed to diagnose infectious diseases in animals, in particular, viral infections such as swine fever and transmissible gastroenteritis. As a reagent, the method uses viral antigens in the form of supernatant liquid obtained as a result of growing the corresponding virus in the SPEV cell culture as substrate.
No data are available on the use of a supernatant liquid obtained as a result of growing SPEV cells alone (without any virus) in anaerobic conditions for the diagnostics of malignancies.
Based on the aforesaid, it can be concluded that the claimed invention meets the “inventive level” criteria.