RU2314123C2 - Method for individually selecting and adjusting interferon dose in renal carcinoma patients - Google Patents

Abstract

FIELD: medicine.

SUBSTANCE: method involves determining sensitivity index SI to interferon . SI is defined to be ratio of neutrophilic granulocyte activation index when treating with dose under study and neutrophilic granulocyte activation index without interferon. The IS value is calculated with respect to each of three therapeutic interferon doses: 3 mln IU, 6 mln IU and 15 mln IU. SI>1, minimum dose is to be prescribed on which cellular sensitivity has been observed. SI>1 being observed only on one of the doses, this very interferon dose is to be prescribed. SI≤1 being observed in all cases, interferon therapy is not to be advised.

EFFECT: enhanced effectiveness of treatment.

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Description

The invention relates to medicine, namely to urological oncology. It can be used in the treatment of kidney cancer patients.

Currently, doses and regimens for the administration of interferon to patients with kidney cancer are determined according to the protocols of the Russian Cancer Research Center named after NN Blokhina RAMS [1], based on a combination of clinical and laboratory data characterizing the general condition of the patient. There is no way to individually select the optimal dose of interferon, as well as its correction during therapy, taking into account cellular sensitivity to this drug,

There is a method of chemiluminescent assessment of sensitivity to interferon in patients with kidney cancer [2], in which, in a blood test in vitro, sensitivity to interferon is judged by the deviation of the sensitivity index (IC) from 1.0. IC is the ratio of the neutrophilic granulocyte activation index when exposed to interferon (IA _IFN ) to the neutrophilic granulocyte activation index without interferon (IA). An HDI greater than 1.0 indicates sensitivity to a specific dose of interferon, and an HDI less than or equal to 1.0 indicates a lack of sensitivity to interferon. The disadvantage of this method is the impossibility of individual selection and correction of the therapeutic dose of interferon.

The objective of the invention is to increase the effectiveness of interferon therapy in patients with kidney cancer.

The problem is solved in that according to the invention, the selection of a dose of interferon for each patient is carried out twice - before treatment and before the start of the next course of interferon therapy. The selection is carried out from three therapeutic doses of interferon - 3 million ME; 6 million ME and 15 million IU - by in vitro chemiluminescent blood test. For each studied dose, a sensitivity index (IC) is determined, which is the ratio of the neutrophilic granulocyte activation index when exposed to the studied doses of interferon ( _IFN IA) to the neutrophil granulocyte activation index without interferon (IA). Those. ICH = IA _IFN / IA. The sensitivity to different doses of interferon is judged by the deviation of the IC from 1.0. The course of treatment is chosen depending on the indicators of ICh. If the ICh value is higher than 1.0 for all three doses of interferon, the patient is recommended the minimum of three doses; if the ICh value is higher than 1.0 for two out of three doses of interferon, the patient is recommended the minimum of these two doses; if the ICh value is higher than 1.0, only one dose is recommended for the patient; and if the ICh value is less than or equal to 1.0 for all three doses, interferon therapy is not recommended.

The index of activation of neutrophilic granulocytes upon exposure to interferon (IA _IFN ) is the ratio of the area under the chemiluminescence curve of zymosan-induced neutrophilic granulocytes upon exposure to interferon (S _INFind ) and the area under the spontaneous chemiluminescence curve of neutrophilic granulocytes upon exposure to interferon (S _IFNspon ). The activation index of neutrophilic granulocytes without interferon (IA) is the ratio of the area under the chemiluminescence curve of zymosan-induced neutrophilic granulocytes (S _ind ) and the area under the spontaneous chemiluminescence curve of neutrophilic granulocytes (S _spon ).

Those. IA _IFN = S _IFNind / S _{IFN sp} ; IA = S _ind / S _spon .

The high reactivity of neutrophilic granulocytes, their ability to quickly restructure in response to the effects of agents of various nature, allows us to consider these cells as an indicator of impaired immune homeostasis. In addition, after activation, neutrophilic granulocytes themselves become powerful effectors of cascade reactions, determining the development of inflammation and the manifestation of the cytotoxic activity of this cell population, in particular, to tumor cells. In addition, neutrophilic granulocytes carry specific receptors for interferon on their surface, which makes it possible to evaluate the effect of interferon on these cells in in vitro experiments.

The activation index is an indicator of the ratio of the integral characteristics of the synthesis of reactive oxygen species by neutrophilic granulocytes in the measured time range obtained from stimulated cells and cells in a state of relative rest.

A sensitivity index above 1.0 characterizes a positive functional reaction of neutrophilic granulocytes to the corresponding dose of interferon. The sensitivity index is 1.0 in cases where the response of cells to a dose of interferon is absent. A sensitivity index below 1.0 is determined by the inhibitory effect of a dose of interferon on the cell and is accompanied by the development of a negative functional reaction.

The method is as follows. Prior to treatment of the patient and before the start of the next course of interferon therapy, an IC is calculated for each of the three studied doses of interferon - 3 million ME; 6 million ME; 15 million ME. For this, neutrophilic granulocytes are isolated from the venous blood of the examined patient, their number is calculated in the Goryaev chamber, a chemiluminescent suspension analysis is performed [3], the IC is calculated for each dose of interferon, and the course of treatment with interferon is selected depending on the indicators of the IC.

To isolate neutrophilic granulocytes from the venous blood of the examined patient, 1 ml of polyglucin is added to 5 ml of blood with heparin. The mixture is incubated for 30 min at 37 ° C to accelerate the deposition of red blood cells. The resulting cell supernatant is washed twice in Hanks solution without phenol red for 10 min at 400 g. The supernatant is drained, the remaining neutrophilic granulocytes are diluted in 1 ml of Hanks solution. Count the number of neutrophilic granulocytes in the Goryaev chamber. The concentration of granulocytes is adjusted to a level of 2 million / ml. A chemiluminescent suspension analysis is performed. For this, control and experimental samples are prepared. The control sample for recording spontaneous chemiluminescence has the following composition: 200 μl of a suspension of neutrophilic granulocytes, 20 μl of donor serum (blood group AB, Rh factor negative), 240 μl of Hanks solution (without phenol red) and 50 μl of luminol at a concentration of 100 μg / ml . Experimental sample No. 1 contains 200 μl of a suspension of neutrophilic granulocytes, 20 μl of donor serum, 220 μl of Hanks solution, 20 μl of reaferon at a concentration of 1 million IU / ml (corresponding to a therapeutic dose of 3 million ME) and 50 μl of luminol. Experimental sample No. 2 contains 200 μl of a suspension of neutrophilic granulocytes, 20 μl of donor serum, 200 μl of Hanks solution, 40 μl of reaferon (corresponding to a treatment dose of 6 million ME) and 50 μl of luminol. Experimental sample No. 3 contains 200 μl of a suspension of neutrophilic granulocytes, 20 μl of donor serum, 140 μl of Hanks solution, 100 μl of reaferon (corresponding to a therapeutic dose of 15 million ME) and 50 μl of luminol.

Spontaneous chemiluminescence is recorded for 45 minutes. Then, 40 μl of zymosan (a non-specific stimulator of the production of reactive oxygen species) at a concentration of 2 mg / ml was added to the samples and zymosan-induced chemiluminescence of neutrophilic granulocytes was recorded. Chemiluminescence is recorded on a chemiluminescent analyzer, for example, "CL3606M". The control of the chemiluminescent analyzer and the registration of the results are carried out through a computer. The chemiluminescence curves of the experimental (with reaferon) and control (without reaferon) samples are obtained, each of which has two peaks. The first peak on each curve corresponds to the maximum level of spontaneous chemiluminescence, the second - induced.

The values of the area under the first (S _spon ) and second (S _ind ) peak of the chemiluminescent curve in the control sample are determined. The neutrophilic granulocyte activation index in the control sample is calculated:

IA = S _ind / S _spon

The area under the first (S _{IFN spon} ) and second (S _IFNind ) peak of the chemiluminescent curve in the experimental samples is _determined . The neutrophilic granulocyte activation index in the experimental sample is calculated:

IA _IFN = S _IFNind / S _{IFN spon}

Calculate the sensitivity index for each experimental sample:

ICH = IA _IFN / IA

An IF exceeding 1.0 indicates the sensitivity of neutrophilic granulocytes to a specific dose of interferon. In this case, the patient is recommended the minimum dose of the studied, for which cellular sensitivity was detected. So, with an ICh value above 1.0 for all three doses of interferon, the patient is prescribed the minimum of these three doses; if the ICh value is higher than 1.0 for two of the three doses of interferon, the patient is recommended the minimum of these two doses, and if the ICh value is higher than 1.0, only this dose is recommended for the patient only for one of the three doses of interferon.

An IF equal to or less than 1.0 indicates a lack of sensitivity to a specific dose. If the ICh value is less than or equal to 1.0 for all three doses, interferon therapy is not recommended.

The method of individual selection and correction of the therapeutic dose of interferon in patients with kidney cancer has passed clinical trials based on the Krasnoyarsk Regional Oncology Center.

Example 1. Patient T., 44 years old. The medical history No. 1496/125. He was hospitalized in the urology department of the Krasnoyarsk Regional Oncology Center since 02.03.04. until 03/26/04. with a diagnosis of cancer of the right kidney, T ₃ N ₀ M ₀ . Operation 03.16.04., Performed laparotomy, radical nephrectomy on the right, drainage of the retroperitoneal space. Histological examination No. 11760-77 in the preparation picture of a clear cell variant of renal cell carcinoma. The postoperative period is smooth. A chemiluminescent analysis of the functional activity of neutrophilic granulocytes and cellular sensitivity (IC) to various doses of reaferon in vitro was performed before treatment and before the next course of interferon therapy.

Before treatment Doses of Reaferon 3 million IU 6 million ME 15 million IU ICH 11.63 / 6.73 = 1.73 11.55 / 6.73 = 1.72 12.03 / 6.73 = 1.79

HD for all three doses> 1. In accordance with the proposed method for treatment, a lower dose is selected. A course of reaferonotherapy with a dose of 3 million ME was carried out.

Before starting the next course of interferon therapy Doses of Reaferon 3 million IU 6 million ME 15 million IU ICH 5.04 / 5.71 = 0.88 6.17 / 5.71 = 1.19 5.92 / 5.71 = 0.93

IF of two doses <1. ICH> 1 for only one dose - 6 million IU. A dose increase of up to 6 million ME is recommended.

Example 2. Patient X., 47 years old. The medical history No. 1624/436. I was hospitalized in the urology department of the Krasnoyarsk Regional Oncology Center since 03.03.04. on March 30, 2004. with a diagnosis of cancer of the right kidney, T ₃ N ₀ M ₀ . Operation 23.03.04., Performed a laparotomy, radical nephrectomy on the right, drainage of the retroperitoneal space. Histological examination No. 12459-77 in the preparation, a picture of renal cell carcinoma, clear cell variant, invasion into the capsule. The postoperative period is smooth. A chemiluminescent analysis of the functional activity of neutrophilic granulocytes and cellular sensitivity to reaferon in vitro was performed before treatment and before the next course of interferon therapy.

Before treatment Doses of Reaferon 3 million ME 6 million ME 15 million ME ICH 0.70 / 0.78 = 0.90 1.00 / 0.78 = 1.28 0.54 / 0.78 = 0.69

ICH> 1 for only one dose. A course of treatment with a dose of 6 million.

Before starting the next course of interferon therapy Doses of Reaferon 3 million ME 6 million ME 15 million ME ICH 8.14 / 2.74 = 2.97 5.28 / 2.74 = 1.93 7.82 / 2.74 = 2.85

HD for all three doses> 1. In accordance with the proposed method for treatment, a lower dose is selected. Recommended dose reduction for the next course to 3 million ME.

Example 3. Patient W., 67 years old. The history of the disease №1451 / 119. I was hospitalized in the urology department of the Krasnoyarsk Regional Oncology Center since 03/01/04. March 25, 2004. with a diagnosis of cancer of the left kidney, T ₃ N ₀ M ₀ . Operation 10.03.04., Performed a laparotomy, radical left nephrectomy, splenectomy, drainage of the retroperitoneal space. Histological examination No. 10868-92 in the preparation of a picture of renal cell carcinoma, clear cell variant. The sluggish course of the wound process, wound healing is partially secondary. A chemiluminescent analysis of the functional activity of neutrophilic granulocytes and cellular sensitivity to reaferon in vitro was performed before treatment and before the next course.

Before starting therapy Doses of Reaferon 3 million ME 6 million ME 15 million ME ICH 3.03 / 2.37 = 1.28 2.17 / 2.37 = 0.92 3.84 / 2.37 = 1.62

IF of two doses> 1. In accordance with the proposed method for treatment, a lower dose is selected. The first course of reaferonotherapy was carried out with a dose of 3 million ME.

Before the next course of interferon therapy Doses of Reaferon 3 million MB 6 million ME 15 million ME ICH 4.79 / 0.70 = 6.73 5.55 / 0.70 = 7.93 2.94 / 0.70 = 4.20

HD for all three doses> 1. In accordance with the proposed method for treatment, a lower dose is selected. The recommended dose of reaferon for the 2nd course was 3 million ME.

Example 4. Patient J., 55 years old. The medical history No. 2425/204. I was hospitalized in the urology department of the Krasnoyarsk Regional Oncology Center since 04/09/04. 05/05/04 with a diagnosis of cancer of the left kidney, T ₃ N ₀ M ₀ . Operation 04/22/04 .: performed laparotomy, radical nephrectomy on the left, drainage of the retroperitoneal space. Histological examination No. 18755-75: in the preparation, a picture of renal cell carcinoma, a mixed version. In the postoperative period, a sluggish course of the wound process was observed. A chemiluminescent analysis of the functional activity of neutrophilic granulocytes and cellular sensitivity to reaferon in vitro was performed before treatment and before the next course.

Before treatment Doses of Reaferon 3 million MB 6 million ME 15 million ME ICH 1.59 / 1.39 = 1.14 1.97 / 1.39 = 1.43 1.97 / 1.39 = 1.42

HD for all three doses> 1. In accordance with the proposed method for treatment, a lower dose is selected. A course of reaferonotherapy with a dose of 3 million IU was carried out.

Before the next course of interferon therapy Doses of Reaferon 3 million ME 6 million ME 15 million ME ICH 1.28 / 1.99 = 0.64 0.94 / 1.99-0.47 1.15 / 1.99 = 0.58

Cellular sensitivity is reduced to all doses studied. Reaferonotherapy with the indicated doses is not recommended.

The proposed method examined 75 people.

The method allows to increase the effectiveness of interferon therapy in patients with kidney cancer due to an individual assessment of cellular sensitivity to various doses of interferon and dose adjustment during therapy.

Information sources

1. Protocols of the Russian Cancer Research Center. N.N. Blokhina RAMS, 2002.

2. Kurtasova L.M., Shkapova E.A., Savchenko A.A. et al. A method for assessing sensitivity to interferon in patients with kidney cancer. RU 2005100610 A, publ. 09/10/2005.

3. Shkapova EA, Kurtasova L. M., Savchenko A. A., Zukov R. A., Krylov G. N. A comparative analysis of the functional activity of neutrophilic granulocytes in blood of patients with renal cell carcinoma before and after surgical treatment // Clinical Laboratory Diagnostics. - 2004. - No. 1. - S. 39-41.

Claims (1)

The method of individual selection and correction of the therapeutic dose of interferon in patients with kidney cancer by chemiluminescent analysis of the functional activity of neutrophilic granulocytes and cellular sensitivity to interferon in a patient’s blood test in vitro with a sensitivity index (IC), which is the ratio of the neutrophilic granulocyte activation index when exposed to interferon (IA _IFN ) of the studied dose to the activation index of neutrophilic granulocytes without interferon (IA), characterized in that before of treatment and before the start of the next course of interferon therapy, the IC is determined for each of the three doses of interferon - 3 million ME, 6 million ME, 15 million ME, and if the IC is higher than 1.0 for all three doses of interferon, the patient is recommended the minimum dose, if the IC is higher 1.0 for two of the three doses of interferon, the patient is recommended the minimum of these two doses, with an ICh value higher than 1.0, for only one of three doses of interferon, the patient is recommended this particular dose, and with an ICh value less than or equal to 1.0 for all three doses of interferon therapy are not recommended.