RU2650964C1 - Method of personified prescribing of targeted therapy agents in patients with metastatic kidney cancer in preoperative mode - Google Patents

Abstract

FIELD: medicine.

SUBSTANCE: invention relates to medicine, namely to oncology, and relates to the personified administration of targeted therapy agents in patients with metastatic kidney cancer in a preoperative regimen. To do this the activity of proteasomes, calpaines, the content of NF-kB-nuclear factor kB p50, HIF-1α - a factor activated by hypoxia, content of the vascular endothelial growth factor, its receptor, phospho-m-TOR, the MSKCC coefficient, the degree of gradation of the Furman cell differentiation, criterion T and on the basis of the obtained values calculate the discriminant functions Y1, Y2, Y3, Y4 by the formulas. At Y1> Y2, the lack of effect on targeted therapy is predicted, and for Y1 <Y2 - effective targeted therapy is predicted. In case of Y1<Y2, the functions Y3 and Y4 are calculated: for Y3>Y4, targeted therapy with pazopanib is prescribed, and for Y3<Y4, targeted treatment with everolimus is prescribed.

EFFECT: method provides effective antitumor therapy due to individual approach to the choice of targeted therapy.

1 cl, 2 tbl, 2 ex

Description

The invention relates to medicine, specifically to oncology, and can be used for the personified appointment of targeted therapy of agents in patients with metastatic kidney cancer in preoperative mode.

The development of clear cell renal cancer is associated with mutational changes in the von Hippel-Lindau protein [8], an increase in the content of nuclear factor HIF-1, overexpression of endothelial growth factor (VEGF), and activation of tyrosine kinase pathways [7, 11]. Targeted therapy is by far the most effective treatment for metastatic kidney cancer (mCCR) [1]. In clinical practice, it uses both tyrosine kinase inhibitors and m-TOR inhibitors, in particular everolimus (Afinitor, Novartis).

A search is currently underway for molecular markers to predict the effectiveness of targeted drugs in patients with metastatic renal cell carcinoma, as existing Motzer and Heng prognostic scales based on clinical and laboratory indicators are considered to a greater extent with respect to the possible outcome of the disease [8, 10]. There are attempts to predict the possible effectiveness of targeted therapy, in particular with the use of bevacizumab (Avastin). Such criteria include factors such as a history of nephrectomy, elevated levels of neutrophils and alkaline phosphatase in the blood [2]. However, in this work, molecular markers mediating the development and progression of the disease are not taken into account. Regarding the possibility of choosing drugs (affinitor or pazopanib) based on clinical, laboratory and medical history data, there is no such information. From the presented positions, the search for new predictor markers associated with the effectiveness and choice of targeted therapy for metastatic renal cell carcinoma is extremely relevant.

In previous studies, it was shown that the effectiveness of the use of targeted agents, such as tyrosine kinase inhibitors, m-TOR blockers, is associated with a change in the complex of molecular parameters. An objective response to the treatment of kidney cancer patients with these drugs is observed at high levels of transcription factors NF-κB, HIF-1, growth factor VEGF, as well as increased activity of proteasomes in the tumor before treatment [3, 4]. The presence of similar changes in molecular parameters with the effectiveness of using both tyrosine kinase inhibitors and m-TOR blockers is justified by their influence on the general biological processes of oncogenesis.

Closest to the claimed is the method described by Deprimo S.E. et al., where the determination of the effectiveness of the use of a tyrosine kinase inhibitor is assessed by the level of VEGF - vascular endothelial growth factor in serum [5]. These factors were analyzed separately for each group of drugs, without substantiating the conditions for their choice. Moreover, the data obtained are presented either by markers evaluated in blood serum, or in the form of qualitative immunohistochemical indicators, which does not allow to obtain high accuracy and information content.

A new technical task is the identification of new prognostic criteria associated with the possible effectiveness of treatment with targeted agents: tyrosine kinase inhibitors and m-TOR blockers.

A new technical result is an increase in the accuracy and information content of the method. In order to achieve a new technical result in a method for the personified administration of targeted therapy agents in patients with metastatic kidney cancer in the preoperative mode by examining biologically active molecules in the tumor tissue, the total proteasome activity, calpain activity, content of NF-kB - nuclear factor kB p50, are determined according to the invention. HIF-1α - hypoxia-activated factor, vascular endothelial growth factor, vascular endothelial growth factor receptor, phospho-m-TOR, coefficients MSKCC event, Furman grade, criterion T, and Y1 and Y2 are calculated: according to the equations

Y1 = -67.01 + 0.33⋅X1 + 0.63⋅X2 + 0.04⋅X3 + 0.17⋅X4 + 1.51⋅X5-0.26⋅X6 + 2.33⋅X7-0.95 ⋅X8 + 34.86⋅X9 + 15.79⋅X10;

Y2 = -62.28 + 0.40⋅X1 + 0.36⋅X2 + 0.07⋅X3-0.06⋅X4 + 1.53⋅X5 + 0.33⋅X6 + 2.47⋅X7-0.91 ⋅X8 + 25.56⋅X9 + 18.64⋅X10,

Where

X1 - total proteasome activity, UE / mg protein;

X2 - calpain activity, UE / mg protein;

X3 — VEGF content, pg / mg protein;

X4 is the content of HIF-1α, PU / mg protein in the well;

X5 — NF-κB p50 content, UE / mg protein per well;

X6 — NF-κB p65 content, UE / mg protein per well;

X7 — phospho-m-TOR content, pg / mg protein;

X8 — VEGFR2 content, pg / mg protein;

X9 is the content of the MSKCC coefficient, where 1 is a favorable prognosis of the disease, 2 is intermediate, 3 is unfavorable;

X10 - the degree of gradation according to Furman;

and if Y1 <Y2, effective targeted therapy is predicted, after which the functions Y3 and Y4 are calculated according to the equations

Y3 = -53.02 + 0.19⋅X1 + 0.52⋅X2 + 0.05⋅VX3 + 0.22⋅X4 + 0.81⋅X5-0.32⋅X6 + 2.44⋅X7-0 56⋅X8 + 35.92⋅X9-0.32⋅X11;

Y4 = -44.03 + 0.28⋅X1 + 0.10⋅X2 + 0.11⋅X3-0.10⋅X4 + 0.56⋅X5 + 0.35 X6 + 2.87⋅X7-0, 36⋅X8 + 27.07⋅X9-3.29⋅X11,

where X11 is the size of the kidney tumor according to TNM, where 1 is the size of the tumor up to 7 cm, corresponding to the criterion T1; 2 - tumor size of more than 7 cm, corresponding to the criterion T2; 3 - meets the criterion T3; 4 - meets the criterion T4;

and with Y3> Y4, targeted therapy with pazopanib is prescribed, and with Y3 <Y4, targeted therapy with everolimus is prescribed.

The method is as follows.

To predict the outcome of the disease in the tumor tissue obtained by performing a diagnostic biopsy before surgical treatment, the total proteasome activity, calpain activity, NF-kB content (nuclear factor kappa-light-chain-enhancer of activated B cells - nuclear transcription factor kB) are determined p50, HIF-1α (hypoxia inducible transcription factor), vascular endothelial growth factor (VEGF) content, VEGFR2, phospho-m-TOR. Tissue samples before determining the activity of enzymes are stored at a temperature of -800C and thawed no more than 1 time. Before determining the activity of the studied enzymes, the frozen tumor tissue is homogenized to a powder state in liquid nitrogen. The homogenate is centrifuged at 2400g and 40 ° C to obtain a pellet that is resuspended in 50 μl of 50 mM Tris-HCl buffer (pH 7.5) and then centrifuged at 14000g and 40 ° C. The obtained supernatant is used to determine the expression of the transcription factor HIF-1α (hypoxia inducible transcription factor), the content of vascular endothelial growth factor (VEGF), its receptor VEGFR2 and serine / threonine protein kinase phospho-m-TOR. Molecular indices are determined by enzyme immunoassay. The MSKCC coefficient is determined by 5 independent parameters: status on the Karnowski scale, lactate dehydrogenase level, adjusted blood calcium, hemoglobin level and lack of a history of nephrectomy. The MSKCC coefficient of 1 is estimated as a favorable prognosis of the disease, 2 - intermediate, 3 - unfavorable. Also, with a standard histological examination of the tumor, the Furman gradation degree, criterion T are determined and the discriminant functions Y1, Y2, Y3, Y4 are calculated according to the above formulas included in the two models. At the first stage, the value of the equations Y1, Y2 is determined to predict the effectiveness of treatment with targeted drugs. When Y1 <Y2, high targeted therapy is predicted, after which they proceed to the next stage - the choice of drug. To do this, evaluates the value of the functions Y3, Y4. For Y3> Y4, effective targeted therapy with tyrosine kinase inhibitors is predicted, and for Y3 <Y4, effective targeted therapy with m-TOR blockers is predicted.

The informational content of the selected criteria was confirmed by the presence of a relationship between the activity of intracellular proteinases - proteasomes and calpains, the content of transcription, growth factors and phospho-m-TOR with the effectiveness of therapy with tyrosine kinase inhibitors - Votrient and m-TOR blocker - everolimus in patients with metastatic kidney cancer. The study included 40 patients with clear cell renal cancer treated at the Department of General Oncology at the Oncology Research Institute of Tomsk Scientific Research Center in the period from 2011 to 2016. Comprehensive treatment of patients involved a 2-month course of preoperative targeted therapy with a tyrosine kinase inhibitor, a dose of pazopanib 800 mg 1 time (17 patients) and everolimus at a dose of 10 mg (23 patients) daily per day, followed by the surgical stage in the amount of nephrectomy / resection of the kidney. The average time between the end of pazopanib and surgical treatment was 6 days (4 to 9 days). The direct effectiveness of targeted therapy was evaluated based on the results of a clinical examination, CT / MRI data on the RECIST scale.

An objective response to Votrient treatment was 88.4%: in 7 (26.9%) patients, a partial regression of the primary tumor and metastases was recorded, in 16 (61.5%) the stabilization of the process was noted, and in 3 (11.5%) - progression . In the group of patients who received an affinitor, the objective response was slightly lower and reached a value of 69%: in 3 (21.4%) patients, a partial regression of the primary tumor and metastases was recorded, in 8 (42.8%), the process was stabilized, and 3 (21.4%) - progression.

An objective response to treatment with a tyrosine kinase inhibitor in patients with kidney cancer was observed at high levels of transcription factors NF-κB, HIF-1, growth factor VEGF, as well as increased proteasome activity in the tumor before treatment. The effectiveness of this drug is associated with a significant decrease in the studied parameters after targeted therapy. In turn, an objective response to treatment with an m-TOR inhibitor in patients with kidney cancer is noted at high levels of transcription factors NF-κB, HIF-1, growth factor VEGF, its receptor VEGFR2, as well as increased proteasome activity in the tumor before treatment.

The quality of the obtained functions was assessed in 12 patients not included in the main sample. The sensitivity and specificity of the obtained discriminant model for assessing the estimated effectiveness of targeted therapy was 97.5% and 91.7%, respectively, and diagnostic accuracy was 92.8%. The use of a model for choosing the type of targeted therapy has a sensitivity of 77.4%, specificity of 96.4% and diagnostic accuracy of 90.1%.

Table 1. Indicators included in the discriminant model for predicting the effectiveness of targeted therapy in patients with disseminated kidney cancer in the preoperative mode.

Table 2. Indicators included in the discriminant model for predicting the effectiveness of targeted therapy depending on the choice of targeted drug in patients with disseminated kidney cancer.

Thus, the results obtained indicate that high efficiency in the treatment with tyrosine kinase inhibitor - pazopanib and m-TOR blocker - everolimus in patients with kidney cancer is associated with initially high levels of transcription factors NF-κB, HIF-1, growth factor VEGF. The data obtained allow use them as markers associated with the effectiveness of targeted therapy and the ability to select one or another targeted agent, which allows them to be used when choosing them.

Clinical examples

Example 1. Patient A, 71 years old, turned to the Tomsk Research Institute of Oncology, where, according to a comprehensive examination, he was diagnosed with clear cell renal cell carcinoma, stage G ₂ T ₂ N ₁ M ₁ . The prognosis of the disease according to the criteria of MSKCC was assessed as intermediate. The study of tumor tissue obtained by performing a diagnostic biopsy showed that the level of HIF-1α was 12 UE / mg protein per well, the content of VEGF and VEGFR2 was 85.14 and 15 pg / mg protein. The expression of phospho-m-TOR was 30.82 pg / mg protein. The activity of the proteasome is 88.1 UE / mg protein per well, calpain - 65.1. The concentration of NFkb50 is 23 UE / mg protein per well, and NFkb65 is 33.7.

Y1 was 135.1; and the coefficient Y2 = 199.7. Function Y1 <Y2, which indicated the expected effectiveness of preoperative targeted therapy in this patient. Y3 was 150.32 and Y4 was 138.73. Y3> Y4, which allowed us to predict the effectiveness of tyrosine kinase inhibitors. Treatment of the patient included a 2-month course of targeted therapy with a VEGFR inhibitor - pazopanib at a dose of 800 mg once a day, after which its effectiveness was evaluated according to the RECIST criteria. During therapy, a partial regression of both the main focus and metastases was noted. The patient underwent a resection of the kidney followed by the appointment of pazopanib. The observation period for the patient at the moment is 26 months, there are no signs of disease progression.

Example 2. Patient D., 66 years old, turned to Tomsk Research Institute of Oncology, where, according to the results of a comprehensive examination, clear cell renal cell carcinoma, stage G ₂ T ₁ N ₁ M _1, was diagnosed. The prognosis of the disease according to the criteria of MSKCC was assessed as intermediate. A study of tumor tissue obtained by performing a diagnostic biopsy showed that the level of HIF-1α was 3.27 UE / mg protein per well, the content of VEGF and VEGFR2 was 6.79 and 28.27 pg / mg protein. The expression of phosphor-m-TOR was 12.58 pg / mg protein. The activity of proteasomes is 2.9 U / mg protein per well, calpains - 0.8. The concentration of NFkb50 is 7.24 UE / mg protein per well, and NFkb65 is 6.03.

The coefficient Y1 was - 47.57; and the coefficient Y2 = 43.36. Moreover, Y1> Y2. The obtained levels indicated the alleged low effectiveness of preoperative targeted therapy in this patient. However, further treatment of the patient was carried out without taking into account the data calculated using the discriminant model. The patient underwent a 2-month course of preoperative targeted therapy with an m-TOR inhibitor, everolimus, at a dose of 10 mg once a day, after which its effectiveness was evaluated according to the RECIST criteria. During therapy, the disease progressed with the development of multiple organ failure and the death of the patient within 4 months.

Example 3. Patient K., 51 years old, turned to the Tomsk Research Institute of Oncology, where, according to the results of a comprehensive examination, she was diagnosed with clear cell renal cell carcinoma, stage G ₂ T ₁ N0M ₁ . Moreover, the prognosis of the disease according to the criteria of MSKCC was assessed as intermediate. The study of tumor tissue obtained by performing a diagnostic biopsy showed that the level of HIF-1α was 7.12 UE / mg protein per well, the content of VEGF and VEGFR2 was 52.59 and 35.24 pg / mg protein. The expression of phosphor-m-TOR was 7.43 pg / mg protein. The activity of proteasomes is 33.3 U / mg protein per well, calpains - 4.1. The concentration of NFkbp50 is 20.63 UE / mg protein per well, and NFkbp65 is 15.37.

At the first stage, the functions Y1 and Y2 were calculated, which amounted to 60.81 and 66.56, respectively. Y1 <Y2, which testified to the expected effectiveness of targeted therapy and allowed moving on to the next step: determining the functions Y3 and Y4. Y3 was 15.56; and the coefficient Y4 is 14.94. Y3> Y4. The data obtained indicated the presumed high effectiveness of preoperative targeted therapy of TKI with Votrient in this patient. Treatment of the patient included a 2-month course of preoperative targeted therapy with a VEGFR inhibitor, pazopanib, at a dose of 800 mg once a day, after which its effectiveness was evaluated according to the RECIST criteria. During therapy, a partial regression of both the main focus and metastases was noted. After preoperative targeted therapy, the patient underwent a resection of the kidney followed by the appointment of pazopanib. The observation period for the patient at the moment is 27 months, there are no signs of disease progression.

Claims (19)

The method of personified prescription of targeted therapy agents in patients with metastatic kidney cancer in the preoperative mode, by examining biologically active molecules in the tumor tissue, characterized in that the total proteasome activity, calpain activity, content of NF-kB - nuclear factor kB p50, HIF-1α - hypoxia-activated factor, vascular endothelial growth factor, vascular endothelial growth factor receptor, phospho-m-TOR, MSKCC coefficient, Furman gradation degree, criterion T and Y1 and Y2 are calculated by the equations. Y1 = -67.01 + 0.33⋅X1 + 0.63⋅X2 + 0.04⋅X3 + 0.17⋅X4 + 1.51⋅X5-0.26⋅X6 + 2.33⋅X7- 0.95 ⋅X8 + 34.86⋅X9 + 15.79⋅X10 Y2 = -62.28 + 0.40⋅X1 + 0.36⋅X2 + 0.07⋅X3-0.06⋅X4 + 1.53⋅X5 + 0.33⋅X6 + 2.47⋅X7- 0.91 ⋅X8 + 25.56⋅X9 + 18.64⋅X10 Where X1 - total proteasome activity, UE / mg protein; X2 - calpain activity, UE / mg protein; X3 — VEGF content, pg / mg protein; X4 is the content of HIF-1α, PU / mg protein in the well; X5 — NF-κB p50 content, UE / mg protein per well; X6 — NF-κB p65 content, UE / mg protein per well; X7 — phospho-m-TOR content, pg / mg protein; X8 — VEGFR2 content, pg / mg protein; X9 - content coefficient MSKCC, where 1 - a favorable prognosis of the disease, 2 - intermediate, 3 - unfavorable; X10 - the degree of gradation according to Furman; and for Y1 <Y2, effective targeted therapy is predicted, after which the functions Y3 and Y4 are calculated to select the targeted agent according to the equations: Y3 = -53.02 + 0.19⋅X1 + 0.52⋅X2 + 0.05⋅VX3 + 0.22⋅X4 + 0.81⋅X5-0.32⋅X6 + 2.44⋅X7- 0 56⋅X8 + 35.92⋅X9 - 0.32⋅X11; Y4 = -44.03 + 0.28⋅X1 + 0.10⋅X2 + 0.11⋅X3-0.10⋅X4 + 0.56⋅X5 + 0.35⋅X6 + 2.87⋅X7- 0 , 36⋅X8 + 27.07⋅X9 - 3.29⋅X11; where X11 is the size of the kidney tumor according to TNM, where 1 is the size of the tumor up to 7 cm, corresponding to the criterion T1; 2 - tumor size of more than 7 cm, corresponding to the criterion T2; 3 - meets the criterion T3; 4 - meets the criterion T4; and with Y3> Y4, targeted therapy with pazopanib is prescribed, and with Y3 <Y4, targeted therapy with everolimus is prescribed.