RU2517527C1 - Diagnostic technique for malignant growth generalisation - Google Patents

Abstract

FIELD: medicine.

SUBSTANCE: technique involves preparing blood serum; preparing at least 10 serum samples by applying the serum on glass slide; drying simultaneously under a cover glass at room temperature and the relative humidity of 70-75% for 7-10 days; studying by polarised light microscopy and detecting anisotropic spherolites. If observing the anisotropic spherolites having a curved structure, the malignant growth generalisation is diagnosed. If observing the anisotropic spherolites having a granular structure or a combination of the spherolite with the included microspherolite with the granular structure, no generalisation is diagnosed.

EFFECT: technique allows diagnosing the malignant growth generalisation at different stages of the disease and carrying out the targeted correction of the therapy that improves the clinical effectiveness in the patients and prolongs their life expectancy.

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Description

The invention relates to medicine, namely to laboratory diagnostics, and can be used to diagnose the course of malignant growth.

A known method for predicting the generalization of the process in malignant bone tumors, based on the determination of the concentration of vitamin E in the blood at the stages of neoadjuvant autoplasmochemotherapy. With a decrease in the concentration of vitamin E during the treatment process, a generalization of the process is noted, and with an increase in the concentration or the absence of changes in this indicator, the stabilization of the process is noted (RF Patent No. 2427323, IPC G01N 33/48, publ. 2011).

However, this method has significant limitations associated with:

- the type and localization of the tumor - the course of the disease is evaluated only in patients with malignant bone tumors (osteogenic sarcoma and Ewing sarcoma);

- age of patients - only for children and adolescents;

- type of treatment - neoadjuvant autoplasmochemotherapy.

Closest to the claimed method is a method for the diagnosis of malignant neoplasms (RF Patent No. 2235323, IPC G01N 33/48, publ. 2004), including the study of blood serum dried at room temperature, while 0.01-0.02 ml of blood serum is applied on a glass slide, and drying is carried out under a coverslip at a relative humidity of 55-60% for 72-96 hours, and the study is carried out under microscopy in polarized light. If anisotropic spherulites with single small multi-colored inclusions are detected in the central part of the spherulite, the preclinical stage of malignant growth in the body is diagnosed; when anisotropic spherulites with pronounced color inclusions are detected, the early stage of malignant neoplasm is diagnosed.

However, this method has the disadvantage that it is limited to identifying the nature of the tumor process mainly at the initial stage of malignant growth.

The objective of the invention was to eliminate these drawbacks, shortening the time for detecting changes in the tumor tissue during or after the combined treatment (surgery and radiation or chemotherapy) or monotherapy (only radiation or chemotherapy), by identifying a marker indicating the transformation of tumor cells into a more aggressive clone followed by generalization of the malignant tumor.

To this end, in a method for diagnosing the generalization of a malignant neoplasm, including obtaining blood serum, preparing a sample by applying blood serum to a glass slide, drying it under a coverslip at room temperature, followed by microscopic examination in polarized light and revealing anisotropic spherulites, it is proposed to carry out simultaneous drying by at least 10 samples of blood serum at a relative humidity of 70-75% for 7-10 days. Moreover, when detecting anisotropic microspherulites with a wavy structure, the generalization of a malignant neoplasm is diagnosed, and when detecting anisotropic microspherolites with a granular structure or a combination of spherulite with microspherulite with a granular structure included in it, the absence of generalization is diagnosed.

The proposed method proceeds from the notion that the generalization of a malignant neoplasm is a qualitative change in one or more properties of neoplasia, aimed at increasing the already existing differences between normal and neoplastic tissue. Generalization is characterized by increasing instability of the genome, leading to aneuploidy and other chromosomal aberrations (Gantsev Sh.Kh., Khusnutdinov Sh.M. Pathology and morphological characteristics of tumor growth. M .: Medical Information Agency, 2003. - 208 p.). The body accumulates the number of cells with these changes, which leads to accelerated proliferation, increased invasiveness and metastasis. The altered tumor cell genome provides the production of protein molecules having a specific conformation, which determines the activation of many proto-oncogenes and a number of enzymes, which underlies the growth of aggressiveness of the corresponding cell clones. A significant part of these molecules is excreted into the general circulation and concentrated in the blood serum. It is these molecules that, when serum is dehydrated, are the systemic and local organizers of specific tumor markers.

If tumor markers are detected by the method of regional dehydration under standard conditions (relative humidity 55-60% for 48-72 hours, prototype), then with delayed structuring of blood serum (7-10 days), which occurs in conditions of increased relative humidity - 70-75 %, optimal conditions are created for the full transition of highly sensitive liquid crystal structures of the lipid-water, lipid-protein-water systems to the crystalline state, in which local specific tumor markers of malignant generalization are formed Tumor-governmental - mikrosferolity undulating structure.

Figure 1, 4, 7, 9, 11, 13 presents microspherulites with the presence of granular structures; figure 2, 5, 8, 12 presents a combination of spherulite and microspherulite with the presence of granular structures; figure 3 presents the combination of spherulite and microspherulite with the presence of wavy structures; figure 6, 10 presents isolated microspherulites with the presence of wavy structures.

The method is as follows.

A patient with any localization of a malignant tumor before treatment is started, during the course of radiation or chemotherapy take blood from a vein or pulp of a finger in the morning, strictly on an empty stomach and receive blood serum by settling or centrifuging it. You can also take blood from a wound during surgery to remove a tumor and get blood serum from it for examination.

Next, blood serum is placed between the coverslip and the slide. Drying is carried out at a relative humidity of 70-75% for 7-10 days. The number of samples for microscopy in polarized light and the detection of a marker for the generalization of malignant neoplasms should be at least 10.

Preliminary studies on a representative number of patients (274 people) showed that 10 samples or more are guaranteed to identify a marker for the generalization of malignant neoplasms.

The following microscopic studies of blood serum samples in polarized light with the detection of anisotropic spherulites and among them microspherulites with a wavy or granular structure make it possible to identify the presence or absence of the process of generalization of malignant neoplasms. Simultaneously with the proposed studies, clinical observations were made in the control group of patients and confirmation of the data of the proposed method was obtained.

Example 1

Patient P., 57 years old. Diagnosis: malignant neoplasm of the left breast, early stage (T2N0M0). Upon admission to the hospital using the above-described technology, 10 blood serum samples were prepared; in a study of 10 of them, separately located granular microspherulites were found (Fig. 1), as well as combinations of spherulite with granular microspherulite (Fig. 2).

Conclusion: signs of generalization of malignant neoplasms were not detected.

During the operation, blood was taken from the tumor site. Re-conducted studies on the proposed method. Were prepared 10 samples of blood serum, in 3 of them were detected microspherulites with a wavy structure in combination with spherulite (figure 3).

Conclusion: signs of generalization of a malignant neoplasm were found in the tumor focus.

After removal of the tumor and a monthly study in a patient in the blood serum from the ulnar vein for 2.5 years, microspherulites, both isolated and in combination with spherulites, were not detected, which indicated a favorable outcome of the disease.

Example 2

Patient S., 69 years old. Diagnosis: laryngeal cancer, late stage (T3N2M0). Upon admission to the hospital, analogously to example 1, 10 blood serum samples were prepared, in 10 of them separately located granular microspherolites were found (Fig. 4), as well as combinations of spherulite with granular microspherulite (Fig. 5).

Radiotherapy was prescribed by the attending physician. After the end of the first course of radiation therapy (SOD 37 Gy - total total dose), 10 blood serum samples were prepared, in 4 of them single microspherulites with the presence of wave structures were detected (Fig.6).

Conclusion: the patient after receiving SOD 37 Gy showed signs of generalization of the malignant neoplasm.

Radiation therapy was adjusted and continued to an SOD of 70 Gy. After the end of radiation therapy and the subsequent monthly follow-up of the patient in this way for 1.5 years, microspherolites in all samples of blood serum from the cubital vein were absent. Thus, the second course of radiation therapy up to an SOD of 70 Gy was effective in suppressing a particularly malignant clone of tumor cells.

Example 3

Patient M., 45 years old. Diagnosis: cerebral glioblastoma, late stage (T3N0M0). Upon admission to the hospital, 10 blood serum samples were studied, in 10 of them separately located granular microspherulites were found (Fig. 7), as well as combinations of spherulite with granular microspherulite (Fig. 8).

After removal of the tumor, radiation therapy was prescribed. After the end of the first course of radiation therapy (SOD 37 Gy), studies were repeated on the proposed method of 10 blood serum samples, in 6 of them single microspherolites with granularity were detected (Fig. 9). Conclusion: signs of generalization of malignant neoplasms were not detected.

Radiation therapy was continued until an SOD of 69 Gy. After the end of radiation therapy, 10 blood serum samples were also prepared, in 2 of them microspherolites with the presence of wave structures were detected (Fig. 10). Conclusion: signs of generalization of a malignant neoplasm were revealed.

Further monitoring of the patient showed the presence of metastases in his lungs and 6 months after the end of the course of radiation therapy a fatal outcome occurred.

Example 4

Patient B., 55 years old. Diagnosis: laryngeal cancer, late stage (T3N0M0). Upon admission to the hospital using the above-described technology, 10 blood serum samples were prepared, in 10 of them separately located granular microspherulites were found (Fig. 11), as well as combinations of spherulite with granular microspherulite (Fig. 12).

Surgical treatment was prescribed by the attending physician, followed by chemotherapy .. After the completion of the first chemotherapy course, 10 blood serum samples were re-prepared, 5 of which revealed microspherolites with granular structures (Fig. 13). Conclusion: signs of generalization of malignant neoplasms were not found.

Chemotherapy was continued. After completing the courses of chemotherapy and following the monthly follow-up of the patient in this way, microspherolites were absent for 1.5 years in all blood serum samples from the cubital vein, which indicated a favorable outcome of the disease.

The identification of morphological signs of the generalization of malignant neoplasms is very important for assessing the effectiveness of the therapy and timely treatment correction in order to destroy the altered genome of tumor cells, leading to an increase in the aggressiveness of certain cell clones; elucidation of the reasons for the selection of an aggressive clone of tumor cells and the development of new methods of treating cancer patients that do not allow the process to be transferred to the generalization of malignant neoplasms.

At the stage of preliminary studies during the dynamic observation of 32 patients with laryngeal cancer and 12 patients with different localization, it was found that microspherolite with a wavy structure was revealed in 9 patients in the blood serum, which indicated the generalization of malignant neoplasms. Of the 7 patients with laryngeal cancer in five people, continued growth of a malignant neoplasm was noted by clinicians 1-3 months later than was established morphologically, i.e. the proposed method. Of the 44 patients observed, a fatal outcome was predicted only in the group with a determination of the generalization of the process by the proposed method. In the future, five out of 9 people died during the first year of observation, two - after a year and a half, two more are under observation.

The proposed method allows at different stages of the disease to timely diagnose the generalization of the tumor process and carry out targeted therapy correction, which increases the efficiency of treatment of patients and their life expectancy.

Claims (1)

A method for diagnosing the generalization of a malignant neoplasm, including obtaining blood serum, preparing a sample by applying blood serum to a slide, drying it under a coverslip at room temperature, followed by microscopic examination in polarized light and identifying anisotropic spherulites, characterized in that they are simultaneously dried by at least 10 blood serum samples at a relative humidity of 70-75% for 7-10 days, and with the detection of anisotropic microspheres LTL undulating structure generalization diagnosed malignancy and the detection anisotropic mikrosferolitov with a grainy texture, or a combination with spherulite mikrosferolitom incorporated therein with a granular structure diagnosed lack of generalization.

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