الفهرس 
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Abstract
The Regulatory Effects of Interleukin-12 on Interleukin-18 and Interferon-Gamma Production in Patients with Breast Cancer
Breast cancer is an important public health problem. It is the commonest form of cancer in the world and ranks first in malignancies in Egypt with an incidence of 36.5%.
Breast cancer like other malignancies results from stepwise genetic alteration of host cells and non genetic changes in the behavior of both malignant and host cells that interact with tumor such as immune alterations.
Cytokines play important role in cellular immunity that is defective in neoplasia. Interleukin-12 (IL-12) is an important cytokine that activates natural killer cells and T cells with anti-angiogenic activities.
Interleukin-18 (IL-18) is a multi functional cytokine, it has a number of immunoregulatory effects. Interferon-gamma (IFN-) production by T-cell and NK cells is important for anti-tumor cascade reaction.
We aimed at this work to investigate the regulatory effects of r IL-12 on IL-18 and IFN- production in non-metastatic and metastatic breast cancer, and the relationships between them are studied before and after in vitro treatment with IL-12.
The patients in this study was conducted from clinical Oncology Unit of Medical Research Institute and they are proven to have breast cancer and don’t suffer from any immunological diseases.
- The patients are divided into two groups non-metastatic and metastatic and ten healthy subjects are taken as control growth.
- The mean age of normal controls was 45.20 ± 8.92 and 60% were in pre- menopausal phase, the non-metastatic patients had mean age of 50.4 ± 10.81 years and 60% were in pre-menopausal phase. All of them had infiltrative ductal carcinoma and the majority of them had positive lymph nodes, estrogen receptors and progesterone receptors (ER, PR).
The mean age for the metastatic group was 47.5 ± 8.25 years. The majority of this group of patients had infiltrative ductal carcinoma and all of them had positive lymph nodes.
Peripheral blood mononuclear cells (PBMC’s) were isolated from patients and normal control by ficoll-hypaque density gradient centrifugation, the separated cells were cultured without and with IL-12 supplementation, the culture supernatants were isolated and stored at -70 oC until cytokines assay, the levels of IL-18 and IFN- we assayed by ELISA technique.
The present study showed that IL-18 levels were significantly decreased in non-metastatic and metastatic patients with breast cancer than normal control which can be explained by impairment of the adaptive immune response by blocking the maturation and function of antigen presenting cells, which are the main cells to produce IL-18 also there is dramatic decrease in natural killer (NK) cells and cytotoxic T lymphocytes (CTL) which constitutively and highly expressed IL-18 receptor (IL-18R) and also attributed to null production of IL-12 which up-regulates IL-18.
Our study indicated that, there is a significant decrease in the mean values of IFN- in non-metastatic and metastatic patients with breast cancer compared to normal control.
This suppression of IFN- production is attributed to presence of immunoregulatory T-cells which have a role in suppression of anti-tumor immunity in cancer patients. These cells and NK cells have molecular mechanisms of immuno-suppression.
The impairment of immune response associated with cancer can be due to presence of classical tumor suppressor proteins which regulate tumor cell immune function genes, as well as tumor cell mutations that eliminate MHC class I expression affect the anti-tumor immune response.
Dendritic cell which are producers of IFN- are dysfunctional and their functions are related to prostaglandin-E2.
When cultures of subjects murder this study were supplemented with IL-12, which is important regulatory cytokine and a key component of immune functions, there was a significant increase in the levels of IL-18 and IFN- production.
After IL-12 supplementation, the mean values of iL-18 were significantly increased form 1.44 to 2.72 ng/ml in normal control and stimulation % was 88.99%. the same results obtained for both non-metastatic and metastatic groups with stimulation percentage 86 and 70.27 respectively.
This can be explained by different mechanisms, resting T and B cells don’t express IL-18 receptor and fail to produce IFN- in response to IL-18. Pre-treatment of T and B cells with IL-12 rendered them more responsive to IL-18 by activation of many transcription factors including STAT4 (signal transduction and activator of transduction).
We can say that by this study there is up-regulation of IL-18 receptor by IL-12. Binding of IL-18 produced by IL-12 stimulated monocytes to its receptor on T cells leading to activation of kinases and IFN- production and T helper type I immune response.
Also, IL-12 administration increases the circulating as well as liver associated levels of IL-18 in wild type mice. IL-12 and IL-18 regulate each other’s production.
Our work finds that there is no significant difference between the mean values of IL-18 and different clincopathological characteristics in non-metastatic patients with breast cancer before stimulation with IL-12 as well as in metastatic patients after stimulation with IL-12.
However, after IL-12 addition, there is significant decrease in the mean values of IL-18 in non-metastatic patients in grade III than in grade II.
Regarding IFN- levels in different clincopathological parameters, our study revealed a significant decrease in the mean values of IIFN- in non-metastatic patients with age >50 years than those <50 years and also with tumor size >5 cm before supplementation with IL-12, whereas after supplementation, there is significant decrease in negative estrogen and progesterone receptors (ER/PR) patients than patients with positive ER and PR.
As for the metastatic patients, our work indicated a significant decrease in the mean values of IFN- patients in grade III than that with grade II and, after addition of IL-12 there is significant difference in the mean values with different ER and PR status.
These data clearly indicated that levels of IL-18 and IFN- were significantly decreased in breast cancer patients who have a biologically aggressive tumor.
These obtained findings produce considerable evidence that the supplementation with IL-12 plays an important role in the production of IFN- by peripheral blood mononuclear cells of patients with breast cancer.
Our work revealed that IL-12 has a role in the rejection of established tumors, which dependant on the induction of a T-helper type I response producing IFN- that acts on host cells.
IL-12 can also up-regulates Fas expression, Fas (CD95-APO-1) a cell surface molecule capable of inducing ligand-mediated apoptosis on human breast cancer cells which plays a role in the elimination metastatic tumor activity.
IL-12 also inhibits angiogenesis and primes macrophages for nitric oxide production which is involved in controlling tumor growth in animal model.
from the data collected form our work, we suggest that using IL-12 as immuno-regulator for patients with breast cancer may be of a value in improvement of immuno-suppressive state associated with malignancy.
A better understanding of IL-12 biology, indications side effects, profiles and toxicity management will aid in its optimal application in the management of cancer patients ad can be used as effective weapon against cancer.
Many studies, found that IL-12 treatment of a murine model of breast cancer leads to tumor regression and systemic anti-tumor immunity, as it is a potent stimulant of immune response and effient element in production of IFN- by synergy with IL-18.
As this work shows that cytokines play important role in immune response in cancer patients.
Therefore, the characterization of different cytokines production in cancer patients is important for investigating the impaired cellular immunity and application of immuno-therapeutic principles to the treatment of breast cancer.
Out work provide a new insight into the regulatory mechanisms controlling the immuno-suppression in breast cancer patients.
Our result indicate the importance of regulatory role of IL-12 on increasing the production of IL-18 and IFN- in females with breast cancer to induce immune response to produce anti-tumor reaction.