Kefah Mokbel

Targeting Matrilysin and Its Impact on Tumor Growth In vivo: The Potential Implications in Breast Cancer Therapy

Introduction: Matrilysin (MMP-7) is a metalloproteinase that is involved in the degradation of extracellular matrix, invasion, and tumor progression. The current study examined if targeting matrilysin using retroviral ribozyme transgenes may have an impact on breast cancer cells and may have clinical implications. Experimental Design: Retroviral hammerhead ribozyme transgenes were designed to specifically target human matrilysin mRNA. The breast cancer cell MDA-MB-231 was transfected with either a retroviral matrilysin transgene or a control retroviral transgene. Stably transfected cells were tested for their invasiveness and migratory properties in vitro. The cells were also used in creating a tumor model in athymic nude mice in which the growth of tumors and levels of matrilysin were assessed. In addition, levels of both protein and mRNA of matrilysin were investigated in a cohort of human breast tumors. Results: Expression of matrilysin in MDA-MB-231 was successfully eliminated by the retroviral hammerhead ribozyme transgene for matrilysin as revealed by reverse transcription-PCR. Matrilysin transgene–transduced cancer cells (MDA-MB-231ΔMatrilysin) exhibited a significantly lower degree of invasion (number of invading cells 16.0 ± 2.5) compared with wild type (MDA-MB-231WT; 26.2 ± 6.2, P < 0.05) or control transgene-transduced cancer cells (MDA-MB-231pRevTRE; 25.3 ± 4.2, P < 0.01). However, the rate of growth of the cells in vitro was not significantly affected. In the in vivo tumor model, MDA-MB-231ΔMatrilysin tumors, which had very low levels of immunoreactive matrilysin, grew at a significantly lower rate (0.24 ± 0.03 cm3, 4 weeks after inoculation) compared with the wild-type MDA-MB-231WT (1.46 ± 0.04 cm3) and MDA-MB-231pRevTRE (1.12 ± 1.0 cm3) tumors. In human breast tumors, breast cancer cells stained matrilysin at a significantly higher density, compared with normal mammary epithelium. The highest level of matrilysin was seen in high-grade tumors and that from patients with moderate and poor prognosis. Finally, high levels of matrilysin were significantly linked with a poor long-term survival (P = 0.0143). Conclusion: Matrilysin, which is aberrantly expressed in human breast tumors, can be effectively eliminated from breast cancer cells by way of hammerhead ribozyme transgene. Elimination of matrilysin is associated with low invasiveness and slow tumor growth. Taken together, the study suggests that targeting matrilysin may have important therapeutic implications.

Oestrogen producing enzymes and mammary carcinogenesis: a review.

There is a large and compelling body of epidemiological and experimental evidence that oestrogens are instrumental in the aetiology of breast cancer. Their mechanisms of action are varied, including stimulation of cellular proliferation through receptor-mediated hormonal activity, increasing genetic mutation rates through cytochrome P450-mediated metabolic activation, and induction of aneuploidy. The local biosynthesis of oestrogens especially in postmenopausal women is believed to play a very important role in the pathogenesis and development of hormone dependent breast carcinoma and the over-expression of regulatory enzymes seems to be associated with the development of a more aggressive disease and associated with poor outcome and increased local and distant recurrences. In this article we highlight the role of CYP19 gene expression and aromatase activity in mammary carcinogenesis. Other oestrogen producing (17-β-hydroxysteroid dehydrogenase and steroid sulphatase) and catalyzing enzymes (3-β-hydroxysteroid dehydrogenase, Oestrogen sulfotransferase, CYP1A1, CYP1B1, and CYP3A4) are also discussed in some detail. Understanding the mechanisms that regulate these enzymes is crucial to the development of new endocrine therapies in post-menopausal females with hormone dependant breast cancer. Currently, third generation aromatase inhibitors has revolutionized the treatment of oestrogen dependant breast cancer. However, the important role of both STS and 17-β-HSD type 1 in local oestrogen production provides novel potential targets for endocrine therapy. Such endocrine therapy is currently being explored and the development of STS inhibitors, combined aromatase/steroid sulfatase inhibitors and 17-β-HSD type 1 inhibitors is underway with promising initial results.

Current management of DCIS: a review

Ductal carcinoma in-situ (DCIS) is a heterogeneous disease, in terms of its radiological characteristics, histological morphology and molecular attributes. This diversity is reflected in its natural history and influences optimal treatment strategy. A significant proportion of DCIS lesions behave in a clinically benign fashion and do not progress to invasive disease. Reliable identification of these patients could allow treatment to be less radical or safely omitted. Management should be tailored to the individual within the context of a multidisciplinary team. Approaches such as biological profiling and molecular analysis represent an opportunity to improve our understanding of the tumour biology of this condition and rationalise its treatment. This article reviews the management strategies for DCIS in the context of recent randomized trials, including the role of sentinel lymph node biopsy, adjuvant radiotherapy and tamoxifen.

Brain-derived neurotrophic factor expression predicts adverse pathological & clinical outcomes in human breast cancer

IntroductionBrain-derived neurotrophic factor (BDNF) has established physiological roles in the development and function of the vertebrate nervous system. BDNF has also been implicated in several human malignancies, including breast cancer (BC). However, the precise biological role of BDNF and its utility as a novel biomarker have yet to be determined. The objective of this study was to determine the mRNA and protein expression of BDNF in a cohort of women with BC. Expression levels were compared with normal background tissues and evaluated against established pathological parameters and clinical outcome over a 10 year follow-up period.MethodsBC tissues (n = 127) and normal tissues (n = 33) underwent RNA extraction and reverse transcription, BDNF transcript levels were determined using real-time quantitative PCR. BDNF protein expression in mammary tissues was assessed with standard immuno-histochemical methodology. Expression levels were analyzed against tumour size, grade, nodal involvement, TNM stage, Nottingham Prognostic Index (NPI) and clinical outcome over a 10 year follow-up period.ResultsImmuno-histochemical staining revealed substantially greater BDNF expression within neoplastic cells, compared to normal mammary epithelial cells. Significantly higher mRNA transcript levels were found in the BC specimens compared to background tissues (p = 0.007). The expression of BDNF mRNA was demonstrated to increase with increasing NPI; NPI-1 vs. NPI-2 (p = 0.009). Increased BDNF transcript levels were found to be significantly associated with nodal positivity (p = 0.047). Compared to patients who remained disease free, higher BDNF expression was significantly associated with local recurrence (LR) (p = 0.0014), death from BC (p = 0.018) and poor prognosis overall (p = 0.013). After a median follow up of 10 years, higher BDNF expression levels were significantly associated with reduced overall survival (OS) (106 vs. 136 months, p = 0.006). BDNF emerged as an independent prognostic variable in multivariate analysis for disease free survival (DFS) (p = 0.026) and approached significance for OS (p = 0.055).ConclusionBDNF expression was found to be significantly higher in BC specimens compared to normal tissue. Higher transcript levels were significantly associated with unfavourable pathological parameters including nodal positivity and increasing NPI; and adverse clinical outcomes including LR, death from BC, poor prognosis, reduced DFS and OS. BDNF offers utility as a prognostic marker and potential for targeted therapeutic strategies.

The mRNA expression of hTERT in human breast carcinomas correlates with VEGF expression

Background Telomerase is a ribonucleoprotein enzyme that synthesises telomeres after cell division and maintains chromosomal stability leading to cellular immortalisation. hTERT (human telomerase reverse transcriptase) is the rate-limiting determinant of telomerase reactivation. Telomerase has been associated with negative prognostic indicators in some studies. The present study aims to detect any correlation between hTERT and the negative prognostic indicators VEGF and PCNA by quantitatively measuring the mRNA expression of these genes in human breast cancer and in adjacent non-cancerous tissue (ANCT). Materials and methods RNA was extracted from 38 breast carcinomas and 40 ANCT. hTERT and VEGF165, VEGF189 and PCNA mRNA expressions were estimated by reverse transcriptase-PCR (RT-PCR) and Taqman methodology. Results The level of expression of VEGF-165 and PCNA was significantly higher in carcinoma tissue than ANCT (p = 0.02). The ratio of VEGF165/189 expression was significantly higher in breast carcinoma than ANCT (p = 0.025). hTERT mRNA expression correlated with VEGF-189 mRNA (p = 0.008) and VEGF165 (p = 0.07). Conclusions hTERT mRNA expression is associated with the expression of the VEGF189 and 165 isoforms. This could explain the poorer prognosis reported in breast tumours expressing high levels of hTERT. The relative expression of the VEGF isoforms is significantly different in breast tumour to ANCT, and this may be important in breast carcinogenesis.

Sentinel lymph node micrometastasis in human breast cancer: An update

INTRODUCTION The advent of sentinel lymph node biopsy (SLNB) and advances in histopathological and molecular analysis techniques have been associated with an increase in micrometastasis (MM) detection rate. However, the clinical significance of sentinel lymph node micrometastasis (SLN MM) continues to be a subject of much debate. In this article we review the literature concerning SLN MM, with particular emphasis on the prognostic significance of SLN MM. The controversies regarding histopathological assessment, clinical relevance and management implications are also discussed. METHODS Literature review facilitated by Medline and PubMed databases. Cross referencing of the obtained articles was used to identify other relevant studies. RESULTS Published studies have reported divergent and rather conflicting results regarding the clinical significance and implications of axillary lymph node (ALN) MM in general and SLN MM in particular. Some earlier studies demonstrated no associations, however most recent studies have found SLN MM to be an indicator of poorer prognosis and to be associated with non-SLN involvement. The use of adjuvant chemotherapy and/or hormonal manipulation therapy is associated with an improved survival in patients with SLN MM. Complete ALND may be safely omitted provided that adjuvant systemic therapy recommendations are equal to patients with node-positive disease. However, optimal management of SLN MM is yet to conclude. Furthermore, the identification of MM remains largely dependent on the analytical technique employed and the use of immunohistochemistry (IHC) increases the detection rate of SLN MM. Discrepancies in the histopathological interpretation of TNM classification of SLN tumour burden do exist. Published studies were non-randomized and have significant limitations including a small sample size, limited follow-up period, and lack of standardization and reproducibility of pathological examination of the SLN. CONCLUSION Patients with SLN MM have a poorer prognosis than those who are SLN negative. Therapeutic recommendations regarding patients with SLN MM should be taken in the context of multidisciplinary team setting and in selected cases of SLN MM, complete ALND may be safely omitted. A better reproducibility of pathological interpretation of the TNM classification is required so that future therapeutic guidelines can be applied without confusion.

The Evolving Role of Mammary Ductoscopy

Summary Mammary ductoscopy (MD) is an emerging technique that allows direct visualisation of the mammary duct system, and that produces sharp and clear video images and ductal washings for cytological analysis. There is a growing body of evidence that MD may have a role in the management of women with pathological nipple discharge, the guiding of breast conserving surgery for cancer, and the screening of high risk women. Further research is required to confirm these potential applications and the feasibility of its use in the rapid intervention and outpatient setting under local anaesthesia. Furthermore, the addition of molecular and genetic analysis of cells obtained by MD and the emergence of newer generations of microendoscopes are likely to enhance the use of this technique.

The Role of Suppressors of Cytokine Signalling in Human Neoplasms

Suppressors of cytokine signalling 1–7 (SOCS1–7) and cytokine-inducible SH2-containing protein (CIS) are a group of intracellular proteins that are well known as JAK-STAT and several other signalling pathways negative feedback regulators. More recently several members have been identified as tumour suppressors and dysregulation of their biological roles in controlling cytokine and growth factor signalling may contribute to the development of many solid organ and haematological malignancies. This review explores their biological functions and their possible tumour suppressing role in human neoplasms.

The evolving role of telomerase inhibitors in the treatment of cancer

SUMMARY Telomerase is a ribonucleoprotein that maintains telomeres and is essential for cellular immortality and tumour growth. The differential expression of telomerase in cancer cells makes it an attractive therapeutic target. Anti-sense oligonucleotides directed against the RNA template of hTR and small molecules that can interact and stabilise the G-quadruplex represent promising therapeutic strategies. Human trials investigating the potential role of the catalytic subunit hTERT as a universal cancer vaccine have already commenced. Alternative lengthening of telomeres (ALT) and efficacy delay remain important limitations to anti-telomerase therapy.

The Prevention of Breast Cancer: An Overview

Summary The role of lifestyle modifications, antioestrogens, cyclo-oxygenase-2 inhibitors and prophylactic mastectomy in reducing breast cancer is reviewed. It is concluded that avoiding postmenopausal obesity and regular physical activity are simple measures that seem to reduce breast cancer risk. There is no conclusive evidence that dietary modification and vitamin supplementation significantly reduce the risk of breast cancer. The evidence suggests that tamoxifen significantly reduces the risk of breast cancer in women at increased risk, but whether it reduces breast cancer mortality remains unknown. Ongoing clinical trials may prove that raloxifene is superior to tamoxifen in breast cancer prevention due to its anti-oestrogenic effects on the endometrium. Bilateral prophylactic mastectomy reduces the risk of breast cancer by 90% in high risk women.