Vitamin B12 enhances the antitumor activity of 1,25-dihydroxyvitamin D3 via activation of caspases and targeting actin cytoskeleton.

BACKGROUND: 1,25-dihydroxyvitamin D3 (1,25(OH)2D3) is an effective anticancer agent, and when combined with other agents it shows superior activities. Vitamin B12 has been shown to contribute to increasing the effectiveness of anticancer drugs when used in combination. Thus, the current study aimed at investigating the anticancer potential of the combination of 1,25(OH)2D3 and vitamin B12. METHODS: MTT assay was used to determine the cytotoxic activity of combining 1,25(OH)2D3 and vitamin B12 against six different cancer cell lines and one normal cell line. The surviving fraction after clonogenic assay was measured, and the effects of 1,25(OH)2D3/B12 combination on the activity of different caspases, cell adhesion, actin cytoskeleton, cell morphology, and percentage of polarized cells were evaluated. RESULTS: Vitamin B12 did not cause cytotoxicity, however, it enhanced the cytotoxicity of 1,25(OH)2D3 against cancer cells. The cytotoxic effects of 1,25(OH)2D3 and its combination with vitamin B12 was not evident in the normal mammary MCF10A cell line indicating cancer cell-specificity. The cytotoxic effects of 1,25(OH)2D3/B12 combination occurred in a dose-dependent manner and was attributed to apoptosis induction which was mediated by caspase 4 and 8. Moreover, 1,25(OH)2D3/B12-treated cells showed enhanced inhibition of clonogenic tumor growth, reduced cell adhesion, reduced cell area, reduced percentage of cell polarization, and disorganized actin cytoskeleton resulting in reduced migratory phenotype when compared to cells treated with 1,25(OH)2D3 alone. CONCLUSION: 1,25(OH)2D3 and vitamin B12 exhibited synergistic anticancer effects against different cancer cell lines. The combination therapy of 1,25(OH)2D3 and vitamin B12 may provide a potential adjunctive treatment option for some cancer types.

Association of Leptin Receptor Q223R Gene Polymorphism and Breast Cancer Patients: A Case Control Study.

INTRODUCTION: Leptin is a hormone secreted from adipocytes that regulates metabolism and energy homeostasis through the leptin receptor (LEPR). The aim of this study was to investigate the association of leptin receptor gene Q223R gene polymorphism, and plasma leptin level among obese breast cancer females. MATERIALS AND METHODS: The study enrolled 160 breast cancer patients and 160 healthy control females. LEPR Q223R polymorphism was determined by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP). Serum leptin was determined using enzyme-linked immunosorbent assay human leptin kit.  Immunohistochemical tests from paraffin blocks were carried out for estrogen and progesterone staging using the precise antibodies. RESULTS: An association was found between LEPR gene Q223R gene polymorphism among obese breast cancer females. Statistical difference was found between GG (60.6%) Arg/Arg genotype (OR=2.986; 95%CI=1.540 to 5.789; p= 0.001) compared to AA (33.1%) Gln/Gln genotype. GG Q223R LEPR polymorphism showed statistically significant difference among obese breast cancer patients (BMI more than 25) compared to control (P < 0.0001). GG genotype of Q223R LEPR polymorphism showed statistically significant increased leptin level (p-value =0.0001) among obese patients (mean± SD; 23.39±4.32) compared to control (17.83±5.67). CONCLUSIONS: Q223R LEPR polymorphism GG genotype was associated with increased leptin profile among obese breast cancer females.

Ischemia modified albumin: an oxidative stress marker in ß-thalassemia major.

BACKGROUND: Ischemia modified albumin (IMA) is an altered type of serum albumin that forms under conditions of oxidative stress. This study reports on the levels and clinical significance of IMA in patients with ß-thalassemia major. METHODS: Blood specimens were collected from 166 subjects (101 ß-thalassemia major patients and 65 healthy controls). Serum levels of IMA, ferritin, malondialdehyde (MDA), ferroxidase, transaminases, total protein, and albumin were determined using conventional methods. RESULTS: Serum levels of IMA (ABSU) were significantly higher in patients than in controls (0.725±0.155 vs 0.554±0.154, p=0.000). Similarly, higher levels were also observed for ferritin, MDA, ferroxidase, and transaminases. No significant differences were observed between patients and controls with respect to total protein and albumin. Spearman univariate analysis demonstrated significant correlation between IMA and ferritin, MDA, ferroxidase, and transaminases. Multiple linear regression analysis revealed significant association of IMA with ferritin and ferroxidase after adjusting for the other variables (r=0.343, p=0.002; r=0.228, p=0.029 respectively). MDA however, correlated significantly with ferritin only (r=0.654, p=0.000). CONCLUSION: Our findings suggest that increased levels of IMA in thalassemic patients are likely to be a result of iron-induced oxidative stress and hence its potential significance as a new marker of oxidative stress in such patients.

Association of haptoglobin phenotypes with ceruloplasmin ferroxidase activity in ß-thalassemia major.

BACKGROUND: Haptoglobin (Hp) and ceruloplasmin (CP) are 2 plasma antioxidants playing a role in preventing iron-induced oxidative damage. This study presents data related to Hp phenotypes and ceruloplasmin ferroxidase activity in relation to iron store markers in patients with ß-thalassemia major. METHODS: Blood specimens were collected from 196 subjects (124 ß-thalassemia major patients and 72 healthy controls). Serum levels of iron, total iron binding capacity (TIBC), ferritin, high sensitivity C-reactive protein (hs-CRP), ceruloplasmin, and ferroxidase activity were determined using conventional methods. Haptoglobin phenotypes were determined by polyacrylamide gel electrophoresis. RESULTS: As expected, the mean levels of iron store markers, except TIBC, were significantly higher in patients than in controls. Ceruloplasmin concentrations (mg/dl) and its ferroxidase activity (U/l) were significantly higher in patients than in controls (57.9±18.8 vs 46.9±14.2 and 159.9±47.8 vs 95.3±20.9; p<0.001, for CP and Hp, respectively). As for Hp phenotypes, no significant differences were observed between iron store markers and ferroxidase activity among the control group. In the patients group however, significantly higher concentrations of ceruloplasmin and its ferroxidase activity were observed among patients with Hp2-2 phenotype as compared to patients with the other phenotypes. Additionally, correlations according to Hp phenotypes revealed strong association between ceruloplasmin ferroxidase activity and serum ferritin in patients with Hp 2-2 phenotype and not in the others (r=0.331, p<0.05). CONCLUSION: Thalassemia patients with Hp 2-2 phenotype are under greater iron-driven oxidative stress than patients with other phenotypes.

Identification of rare and novel Cryptosporidium GP60 subtypes in human isolates from Jordan.

Little is known about the epidemiology of Cryptosporidium in Jordan and no genotyping studies have been conducted on Cryptosporidium isolates from humans or animals from Jordan. Genotyping of 44 Cryptosporidium isolates from Jordanian children at the 18S rRNA locus and a unique diagnostic locus identified four Cryptosporidium species; C. parvum (22), C. hominis (20), C. meleagridis (1) and C. canis (1). Sub-genotype analysis of 29 isolates at the 60-kDa glycoprotein (GP60) locus identified three C. parvum, two C. hominis subtype families and one C. meleagridis subtype. Several rare and novel subtypes were identified indicating unique endemicity and transmission of Cryptosporidium in Jordan.

Amifostine exerts anti-angiogenic activity and suppresses vascular endothelial growth factor secreted by hemopoietic stem/progenitor cells.

OBJECTIVE: To assess, in vitro, the effect of Amifostine (AMF, WR-2721) on angiogenesis and levels of vascular endothelial growth factor (VEGF) secreted from hemopoietic stem/progenitor cell populations. METHODS: We conducted the study in the research laboratories of the Hashemite University, Jordan between September 2003 and January 2005 where we took samples were from Myelodysplastic syndrome (MDS) patients and healthy donors attending Al-Hussein Cancer Center and We determined the proliferation of human umbilical vein endothelial cells (HUVECs) in cultures supplemented with media conditioned with AMF-treated and AMF-untreated pure hemopoietic cells [CD34+ cells, and erythroid, myeloid and megakaryocytic progenitors]. Furthermore, in the same conditioned media, we evaluated levels of elaborated VEGF by a sensitive enzyme linked immunosorbent assay. RESULTS: Biologically, media conditioned with AMF-treated cells reduced proliferation of HUVECs compared to media conditioned with untreated control cells (p<0.05). In cultures of AMF-untreated cells, elaboration of VEGF was higher (p<0.05) in media conditioned with cells from MDS patients compared to healthy donors. A 30 minutes pre-exposure of cells to AMF (500 mM) suppressed levels of VEGF secreted within 24 hours in 63 of 89 evaluated cultures. The percentage of reduction of VEGF in AMF-sensitive cultures was comparable in cultures of MDS cells (18%, 2-37%; median, range) and normal cells (12%, 2-45%). CONCLUSION: The results showed that AMF exerts an anti-angiogenic activity and suppresses the secretion of VEGF in hemopoietic stem/progenitor cells obtained from both healthy individuals and patients with MDS.

Lifestyle related risk factors for breast cancer in Jordanian females.

OBJECTIVE: To compare the lifestyle related risk factors for breast cancer such as physical activity, cigarette smoking, the use of contraceptive pills and increased body weight between non-familial and familial breast cancer females in Jordan. METHODS: This study was carried out in the Kingdom of Jordan during the period 2000 through to 2002. A questionnaire was used to collect information from 99 females who were histologically and pathologically diagnosed for breast cancer. Data of the questionnaire was entered and analyzed using statistical package for social sciences. RESULTS: This study showed no significant difference between familial, non-familial breast cancer females and controls in the following risk factors: physical activity, contraceptive methods, and smoking. On the other hand, a statistically significant difference in weight was found between the familial breast cancer females, the total breast cancer females and the controls. In addition, the highest percentage of overweight and obese was found among postmenopausal breast cancer females. CONCLUSION: Postmenopausal obesity is a significant risk factor among Jordanian breast cancer females.

Haptoglobin polymorphism in breast cancer patients form Jordan.

BACKGROUND: Previous reports regarding the occurrence of breast cancer and its association with Hp polymorphism are conflicting. The possible role of family history as a factor in determining the degree of association between the disease and Hp polymorphism has not been reported before. In this study, the distribution of haptoglobin phenotype among patients with familial and nonfamilial breast cancer was investigated. METHODS: Haptoglobin phenotypes were determined in serum of 128 breast cancer patients (familial, n=42; nonfamilial, n=86) and in controls (n=200) by vertical polyacrylamide gel electrophoresis. RESULTS: No significant difference of Hp phenotype distribution was observed between patients as a combined group when compared with the control group. In the familial group, the frequency of Hp1-1 and Hp2-1 phenotype distribution was higher and Hp2-2 was lower than that in the nonfamilial and the control groups. Similar but inversed Hp distribution pattern was observed in the nonfamilial group when compared with that in the other groups. An appreciable finding is the observation that Hp2-2 phenotype frequency in the nonfamilial group was significantly higher than that in the familial group (p=0.0365). CONCLUSIONS: Results of this study demonstrate that the pattern of Hp phenotype distribution in breast cancer patients is family history-dependent. Hp1 and Hp2 allele frequencies were over-represented in patients with familial and nonfamilial breast cancer, respectively. The pattern is probably attributed to genetic and oxidative stress mechanisms.

Mutation analysis of the breast cancer gene BRCA1 among breast cancer Jordanian females.

OBJECTIVE: To screen mutations of the tumor suppressor breast cancer susceptibility gene 1 (BRCA1) within 3 exons among Jordanian breast cancer females. METHODS: A total of 135 Jordanian breast cancer females were genetically analyzed by denaturing gradient electrophoresis (DGGE) for mutation detection in 3 BRCA1 exons (2, 11 and 20) between 2000-2002 in Al-Basheer Hospital, Amman, Jordan. RESULTS: Of the studied patients 50 had a family history of breast cancer, 28 had a family history of cancer other than breast cancer, and 57 had no family history of any cancer. Five germline mutations were detected among breast cancer females with a family history of breast cancers (one in exon 2 and 4 mutations in exon 11). Another germline mutation (within exon 11) was detected among breast cancer females with family history of cancer other than breast cancer, and no mutation was detected among breast cancer females with no family history of any cancer or among normal control females. CONCLUSION: Screening mutations within exon 2, exon 11 and exon 20 showed that most screened mutations were within BRCA1 exon 11 among breast cancer Jordanian families with a family history of breast cancer.

Comparison of some risk factors between non-familial and familial breast cancer females in Jordan.

OBJECTIVE: To compare the risk factors such as age, menopause, menarche, age at the first pregnancy, number of pregnancies and breast feeding period between the familial and non-familial breast cancer females in Jordan. METHODS: This study was carried out in Al-Basheer Hospital, Amman, Jordan during the period 2000 and 2002. A questionnaire was used to collect information from 99 females who were histologically and pathologically diagnosed with breast cancer. Data of the questionnaire were entered and analyzed using statistical package for social sciences. RESULTS: The highest percentage of non-familial and familial breast cancer occurrence was among age group 51-60 years. The age of the first pregnancy is another risk factor of which the highest percentage of breast cancer was reported for both familial (57.1%) and non-familial (65.4) breast cancer females who have their first pregnancy while they were 20-years-old and above. More than 4 pregnancies also represents a risk factor for both non-familial (67.9%) and familial (68.6%) breast cancer patients. In this study, there are no statistical differences between menopause and menarche age among the breast cancer females. Finally, an inverse relationship was shown between breast feeding period (equal or more than 24 months) and the occurrence of breast cancer in both non-familial (2%) and familial (0%) breast cancer. CONCLUSION: This study found that age 51-60 years and the increase number of pregnancies (more than 4) in the age of 20 years or more are risk factors for both types of breast cancer. On the other hand, longer period of breast feeding (more than 24 months) decreases the risk of breast cancer in both types.