Anti-IL-8 monoclonal antibodies inhibits the autophagic activity and cancer stem cells maintenance within breast cancer tumor microenvironment.

BACKGROUND: Breast cancer tumor microenvironment (TME) is a promising target for immunotherapy. Autophagy, and cancer stem cells (CSCs) maintenance are essential processes involved in tumorigenesis, tumor survival, invasion, and treatment resistance. Overexpression of angiogenic chemokine interleukin-8 (IL-8) in breast cancer TME is associated with oncogenic signaling pathways, increased tumor growth, metastasis, and poor prognosis. OBJECTIVE: Thus, we aimed to investigate the possible anti-tumor effect of neutralizing antibodies against IL-8 by evaluating its efficacy on autophagic activity and breast CSC maintenance. METHODS: IL-8 monoclonal antibody supplemented tumor tissue culture systems from 15 females undergoing mastectomy were used to evaluate the expression of LC3B as a specific biomarker of autophagy and CD44, CD24 as cell surface markers of breast CSCs using immunofluorescence technique. RESULTS: Our results revealed that anti-IL-8 mAb significantly decreased the level of LC3B in the cultured tumor tissues compared to its non-significant decrease in the normal breast tissues.Anti-IL-8 mAb also significantly decreased the CD44 expression in either breast tumors or normal cultured tissues. While it caused a non-significant decrease in CD24 expression in cultured breast tumor tissue and a significant decrease in its expression in the corresponding normal ones. CONCLUSIONS: Anti-IL-8 monoclonal antibody exhibits promising immunotherapeutic properties through targeting both autophagy and CSCs maintenance within breast cancer TME.

The effect of Fas/FasL pathway blocking on apoptosis and stemness within breast cancer tumor microenvironment (preclinical study).

Evasion of the immune system is the tumor's key strategy for its maintenance and progression. Thus, targeting the tumor microenvironment (TME) is considered one of the most promising approaches for fighting cancer, where immune cells within the TME play a vital role in immune surveillance and cancer elimination.FasL is one of the most important death ligands expressed by tumor-infiltrating lymphocytes (TILs) and plays a vital role in eliminating Fas-expressing cancer cells via Fas/FasL pathway-induced apoptosis. However, tumor cells can express elevated levels of FasL inducing apoptosis to TILs. Fas/FasL expression is linked to the maintenance of cancer stem cells (CSCs) within the TME, contributing to tumor aggressiveness, metastasis, recurrence, and chemoresistance.This study is considered the first study designed to block the overexpressed FasL on the tumor cells within TME mimicking tissue culture system using rFas molecules and supplementing the Fas enriched tissue culture system with blocked Fas - peripheral blood mononuclear cells PBMCs (using anti-Fas mAb) to protect them from tumor counterattack and augment their ability to induce tumor cell apoptosis and stemness inhibition.A significantly increased level of apoptosis and decreased expression of CD 44 (CSCs marker) was observed within the east tumor tissue culture system enriched with Fas molecules and anti-Fas treated PBMCs and the one enriched with Fas molecules only compared to the breast tumor tissues cultured alone (p < 0.001). Accordingly, we can consider the current study as a promising proposed immunotherapeutic strategy for breast cancer.

Angiogenic activities of interleukin-8, vascular endothelial growth factor and matrix metalloproteinase-9 in breast cancer.

Angiogenesis is a major contributor to tumor growth and metastasis within breast cancer tumor microenvironment in which different proangiogenic factors have been identified and associated with tumor progression, metastasis and poor prognosis. The aim of the current study was to evaluate the angiogenesis among breast cancer patients through ex vivo assessment of the angiogenic factors interleukin 8 (IL-8) and vascular endothelial growth factor (VEGF)-A expressions in excised tumor tissues as well as matrix metalloproteinase 9 (MMP-9) serum levels as well as the prognostic value of MMP-9. Our study included 28 invasive ductal carcinoma female patients who were scheduled for modified radical mastectomy at Medical Research Institute, Alexandria University, Egypt and 10 control subjects. Both IL-8 and VEGF-A expressions were immunohistochemically detected in tumor tissues and serum MMP-9 was determined by ELISA. Although no significant correlations were found between each of IL-8, VEGF-A, MMP-9 levels, and patients' clinicopathological parameters, a significant positive correlation was found between these angiogenic factors each other suggesting their synergistic roles in proceeding angiogenesis. Higher serum MMP-9 level was detected in breast cancer patients compared to the control group, indicating that it can be used as a prognostic biomarker in breast cancer patients.

Immunotherapeutic Approach to Breast Cancer: The Anti-Tumor Effect of Anti-IL-6 Monoclonal Antibodies in Malignant Mammary Tumor Implanted-Mice.

It has been established that suppression of apoptosis during carcinogenesis is the main cause of development and progression of breast cancer. Breast cancer patients have higher circulating levels of IL-6 protecting cancer cells from apoptosis and positively correlated with poor prognosis of the disease. The current work is carried out to fulfill one of our in vivo preclinical studies' for approaching a novel breast cancer immunotherapy through induction of tumor cell apoptosis. The study aims at investigating the potential of anti-IL-6 monoclonal antibodies (mAbs) to suppress IL-6 anti-apoptotic activities in tumor microenvironment of malignant mammary tumor implanted-mice. To achieve this goal, 4 groups of mice were used, group I: served as control, group II: mice implanted with Ehrlich ascites carcinoma cell lines (EAC), through intramuscular injection till tumor inoculation, group III: injected intratumorally with10 µl saline for 3 successive days, and group IV: mice were injected intratumorally one day after tumor inoculation with a dose of 1.5 mg / kg of recombinent anti-IL-6 monoclonal antibodies in10 µl saline for 3 successive days. Apoptosis was evaluated in tumor samples from anti-IL-6 treated tumor implanted mice and compared with controls. Levels of apoptosis in tumor tissue samples of tumor implanted mice treated with anti-IL-6 were significantly (P=0.009) higher than untreated ones. In conclusion, anti-IL-6 monoclonal antibodies have the potential to suppress the anti-apoptotic effect of interleukin-6 (IL-6) within the tumor microenvironment of tumor implanted in mice.

Effects of chronic ethanol and vitamin C administration on production of tumor necrosis factor-alpha and interleukin-6 in rats.

Chronic alcoholism complicated by alcoholic liver disease (ALD) is characterized by activation of inflammatory responses. Alcohol intake increases gut permeability allowing substances such as lipopolysaccharides (LPS) which are strong inducers of proinflammatory cytokines such as tumor necrosis factor-alpha (TNF-alpha) and interleukin-6 (IL-6) to enter the circulation. Vitamin C is an antioxidant with many cellular activities seemed to protect cells against alcohol-induced peroxidation. In present study, serum levels of TNF-alpha and IL-6 were measured by ELISA method in four groups of albino rats, each group consists of 10 rats. Group (I) was untreated group (control), group (II) was treated with ethanol, group (III) was treated with ascorbic acid and group (IV) was treated with ethanol + ascorbic acid. Results revealed that both TNF-alpha and IL-6 serum levels were very highly significantly increased in group (II) and (IV) than control group (1) (P < 0.001). Group (III) showed significantly (P < 0.001) decreased TNF-alpha serum level than group (II) and (IV) while it showed significantly (P < 0.001) increased IL-6 serum level than control group (I) and also significantly decreased IL-6 serum level than group (IV). Serum IL-6 level was significantly (P < 0.01) decreased in group (III) than (II). These results indicate that serum levels of the proinflammatory cytokines TNF-alpha and IL-6 may serve as predictive biomarkers for progression of ALD. In addition, using TNF-alpha neutralizing agent (or its antagonist)/or IL-6 as an anti-apoptotic factor could be useful as a treatment strategy of ALD.

Interleukin-2 regulatory effect on P-selectin and interleukin-8 production in patients with chronic renal failure.

Patients with chronic renal failure (CRF) show a clinical state of immunodysfunction that occurs in both humoral and cellular immunity as well as inflammatory response. In this study, we investigated the mononuclear-endothelial cells (MCs/ECs) interaction and the possible protective role of IL-2 as the main T lymphocyte activator in CRF patients. The levels of soluble P-selectin (sP-selectin) and interleukin-8 (IL-8) as the two main mediators of MCs/ECs interaction were measured in IL-2 supplemented and non-supplemented peripheral blood mononuclear cells (PBMCs) supernatant of CRF patients. The obtained results were correlated with those of sex and age matched controls. Significantly higher levels of sP-selectin and IL-8 were detected in both IL-2 supplemented and non-supplemented PBMCs culture supernatant of CRF patients than controls (P = 0.000). Those levels were significantly lower in IL-2 supplemented PBMCs culture supernatant than non-supplemented ones of both CRF (P = 0.000) (for both mediators) and normal control groups (P = 0.01, P= 0.04 for sP-selectin and IL-8 respectively). The higher sP-selectin in CRF indicates impairment of MCs/ECs interaction that may be resulted from blockade of P-selectin receptors on PBMCs by P-selectin molecules shedded from ECs to plasma and bind to PBMCs in vivo. The elevated IL-8 level in PBMCs of CRF reflect the imbalance of Thl/Th2 ratio and subsequent impairment of cellular immunity in those patients. The lower level of both sP-selectin and IL-8 in IL-2 supplemented PBMCs supernatant than in non-supplemented one seemed to be due to the IL-2 induced proliferation of Th1 lymphocytes yielding newly in vitro formed T cells which do not carry P-selectin as well as relative increase of Th1/Th2 ratio in both normal and CRF groups. Thus, IL-2 may improve the MCs/ECs interaction and correct the Th1/Th2 ratio in CRF providing a novel promising therapeutic approach to improve the immuno-pathological condition of those patients.

Interleukin-8, ferritin and soluble transferrin receptors in type II diabetes mellitus.

Type II diabetes mellitus (DM) is the most common form of diabetes that constitutes the majority of cases worldwide including Egypt. Chronic elevated glucose level in DM increases monocyte adhesion to aortic endothelial cells (ECs) which is mediated primarily through induction of interleukin-8 (IL-8). This study aimed to investigate the possible role of IL-8 as a potent chemoattractant, pro-inflammatory cytokine in the immuno-inflammatory response of type II diabetic patients in correlation to ferritin and sTFR as markers of glucose homeostasis that characterizes the disease. The current work was conducted on 20 diabetic females and 10 healthy age and sex matching subjects as a group of control. Serum levels of IL-8, ferritin and sTFR were measured in all study subjects under investigation. Results revealed that both serum levels of IL-8 and ferritin were significantly elevated in type II diabetic patients (P = 0.0029 and 0.03 respectively) compared with those of control group while no significant difference was detected between sTFR levels in diabetic patient and control groups. In addition, a significant positive correlation was detected (P = 0.032) between serum levels of IL-8 and sTFR of the studied diabetic patient group. In conclusion, quantitative determination of IL-8, ferritin and sTFR could help in predicting type II diabetes-associated immuno-inflammatory manifestations characterize the micro-and macrovascular disease complications, particularly for high risk populations.

Study of P53 in peripheral blood and synovial mononuclear cells of rheumatoid arthritis and osteoarthritis patients and its relation to the degree of disease activity.

Over expression of P53 has been described in many inflammatory conditions including rheumatoid arthritis (RA) and osteoarthritis (OA) as a protective mechanism to induce apoptosis of synovial cells. Lack of P53 function through mutation in human synoviocytes increases the development of normal synovial fibroblasts into transformed aggressive synovial fibroblasts. P53 levels were determined in supernatant of cultured mononuclear cells (MCs) isolated from peripheral blood (PBMCs) of patients with RA (n = 10) and OA (n = 10) as well as 10 normal healthy controls (C). P53 levels were also determined in supernatants of MCs isolated from synovial fluid (SFMCs) of RA and OA patients. Results of this work revealed that P53 level was significantly higher in PBMCs supernatant of RA group than those of both (C) and (OA) groups (P = 0.022). P53 level was non-significantly higher in SFMCs supernatant of RA than OA group. Significantly higher levels of P53 was detected in SFMCs culture supernatant than that of PBMCs within each RA (P = 0.003) and OA (P = 0.001) group. Results also showed a significantly positive correlation between P53 levels (in both PBMCs and SFMCs) and the disease activity score (DAS) in RA group (P = 0.01, P = 0.02 respectively) while insignificantly positive correlations between P53 level (in both PBMCs and SFMCs) and radiological grading of OA group were obtained. These results indicate that mutations and consequent dysfunction of P53 gene may result in chronic inflammation and hyperplasia in RA patients. In conclusion, gene therapy targeting P53-dependent pathway could be a promising therapy for RA and OA diseases.

Antibody profile, interferon-gamma and nutritional status in cryptosporidial infection among school children.

Cryptosporidium parvum is more common in children than in adults. The host responses that induce protective immunity against Cryptosporidium is poorly understood. The present work studied the antibody profile, interferon gamma (IFN-gamma) serum level and nutritional status among infected school children, aged eleven years in a rural school in Abis 8 village. Fecal examination showed nineteen cases only infected with Cryptosporidium and ten children free from parasites (controls). The IgA, IgG and IgM serum levels in infected cases were within normal level, without significant differences as compared to controls. The IgE serum level was significantly high in infected children. The cytokine IFN-gamma was significantly lower in the infected cases as compared to controls. Malnutrition was diagnostic among infected children. It was concluded that malnutrition is the important finding related to Cryptosporidium infection. Malnutrition decreased IFN-gamma the recognized factor in protection from infection and elevated serum IgE level.