The location and role of mTOR in the cell.
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Understanding Breast Cancer
With an Emphasis on Triple Negative Breast Cancer
Compiled by John G. Connor, M.Ac., L.Ac.
June 2010
Table of Contents
· Triple Negative Breast Cancers (TNBCs) are those which are ER (Estrogen Receptor) negative, PR (Progesterone Receptor) negative and HER2 negative. These tumors intrinsically have a poor prognosis relative to those with endocrine positive disease.
· Triple-negative tumors are synonymous with basal-like tumors, and can be identified by immunohistochemistry. Based on gene-expression profiling, basal-like tumors are still heterogeneous and can be subdivided into at least five distinct subgroups. The development of distant metastasis in basal-like tumors is associated with the presence of central fibrosis and a small amount of lymphocytic infiltrate. (Kreike et al 2007)
· TNBC has associations with loss of expression of the androgen receptor and E-cadherin and P-cadherin, and with positive expression of basal cytokeratins CK5 and CK17 (basal phenotype), p53, vimentin, a high MIB1 labeling index, VEGF and in addition appears to be strongly EGFR-driven.
· The majority of BRCA1 associated breast cancers are TN and basal-like. Although sensitive to chemotherapy, early relapse is common and a predilection for visceral metastasis, including brain metastasis, is seen. Targeted agents, including EGFR, VEGF and poly (ADP-ribose) polymerase (PARP) inhibitors*, are currently in clinical trials and hold promise in the treatment of this aggressive disease. (Anders & Carey, 2008) * See glossary for more information about PARP inhibitors.
· Of the estimated 1 million cases of breast cancer diagnosed annually worldwide, it is estimated that over 170,000 will harbor the triple-negative (estrogen receptor/progesterone receptor/HER2-negative) phenotype. Most, though not all, triple-negative breast cancers will be basal-like on gene expression micorarrays. When BRCA1 mutation carriers develop breast cancer, it is usually basal-like; given the central role of BRCA1 in DNA repair, this could have profound therapeutic implications. The armamentarium of "targeted therapeutics" for triple-negative breast cancer is evolving and includes strategies to inhibit angiogenesis, epidermal growth factor receptor, and other kinases. Finally, the positive association between triple-negative breast cancer and BRCA mutations makes inhibition of poly(adenosine diphosphate-ribose) polymerase-1 an attractive therapeutic strategy that is in active study. (Anders & Carey 2009)
Tissue Pathology and other Tests to Perform on Breast Cancer Cells
Important Tissue Pathology Tests to Perform:
o Histocological grade: Mitotic index, S-phase expression
o DNA index: Aneuploid (unfavorable) versus diploid (favorable)
o ER/PR (CD 24 over-expression predicts tamoxifen resistance)
o KI-67 (<10% favorable)
o Cathepsin-D (also G)
o Mutant proteins: p53, Bcl-2, PTEN, p21, p27, p63, nm23
o Her 2 neu (Co Her II and ER expression predicts tamoxifen resistance)
o Her I (EGFr)
o PDGF/c-KIT (CD117)
o Cytokeratins (CK) – detects circulating tumor cells CK 19 andCK 7 (micrometastasis)
o CK-19 – low levels associated with poor outcome in triple negative breast cancer
o E-Cadherin (lack of = poor prognosis)
o Survivin (over-expression associated with poor prognosis)
o TGF-β1: Primes BC for lung and bone metastasis co-expressed with AP-1 and NF-κB, stimulating IL-8
Important Blood Tumor Markers to Assess:
o CA 27.29 <38
o CA 15-3 (MUC-1) <35
o CA 125
o Prolactin
o Carbonic Anhydrase (CA)-9 Preferred range = Low (angiogenesis/hypoxia)
o Bcl-2 – a strong predictor of chemo-resistance (Breast Cancer Res. 2008;10(2):R30.Epub 2008 Apr1)
Tests relating to cancer in the bone:
o Bone specific alkaline Phosphatase
o Ki-67
Blood marker tests to test for hypercoagulation, angiogenesis and metastasis:
o Fibrinogen <375
o D-dimer <.40
o PAI-1
(Elevated levels of any one of these three above -- but even more importantly when they are all elevated -- is strongly predictive of forming life-threatening blood clots (thromboembolisms). Many chemotherapeutic drugs significantly increase this risk. These drugs should either not be given until levels are normalized or should be given with targeted agents that inhibit hypercoagulation.)
o LDH (lactate dehydrogenase)
o ESR (erythrocyte sedimentation rate)
o Serum insulin (fasting) and IGF-1
o Serum zinc
o Ceruloplasmin
o Serum copper
o Vitamin D 25-HYDROXY
o C-reactive protein (CRP) (Donnie likes it to be below 0.8)
o VEGF
Rationale and Studies Supporting the Use of the Above Markers in Breast Cancer
· Bcl-2 – an NF-κB regulated gene that functions by blocking the apoptosis pathway, thus immortalizing cancer cells. It has been suggested that Bcl-2 over expression results in the up regulation of VEGF expression with increased neoangiogensis in human cancer xenografts.
· CA 125 – Sensitivity of this test to ovarian cancer is 75%-80%. Laboratory Tests and Diagnostic Procedures, p. 273 (2008)
· CA 125 – This test is especially useful in monitoring ovarian carcinoma and breast cancer. Although CA 125 cannot distinguish benign from malignant tumors, levels >65 U/ml are associated with malignancy in 90% of pelvic masses. Patients with very high CA 125 levels (>450 U/ml) have poor median survival, whereas patients with levels less than 55 U/ml tend to do somewhat better. Laboratory Tests and Diagnostic Procedures, p. 273 (2008)
· CA 15-3 (MUC-1) - an important blood tumor marker to assess breast, pancreatic & ovarian cancers.
· CA 27.29 - an important blood tumor marker to assess breast cancer.
· cathepsin D - We have focused our research on cathepsin D as a potential target, since it is often overexpressed in aggressive human breast cancers, including ER-negative tumors, and rarely associated with HER2-Neu amplification. Validated assays in the primary tumor of molecular markers such as ER, HER2-Neu and cathepsin D should help to predict which targeted therapy should be applied to cure breast cancer patients. (Rochefort et al, 2003)
· Cathepsin D – (Cath D) – is an aspartyl lysosomal protease expressed in all tissues that might play a role in antigen processing, cell proliferation and tissue renewal, and activation of different pro hormones. It is suggested that CathD expression was stronger in malignant than in benign breast disease, the positivity being present in both epithelial neoplastic and stromal cells. (Brujan et al, 2009)
· cathepsin D - The expression of ER, PR, c-erB-2 and cathepsin D were evaluated in 55 breast cancer patients. Metastases were found to occur in bone (24.4%), lungs (20.8%) and liver (10.7%). Histopathological type of invasive ductal carcinoma was associated with the higher incidence of bone metastasis. (Irawan, et al 2008)
· cathepsin G - Using a mouse model it was found that the major source of cathepsin G at the tumor-bone interface seems to be osteoclasts that up-regulate production of cathepsin G via interaction with tumor cells. It was also found that in vitro osteoclastogenesis is reduced by inhibition of cathepsin G in a coculture model and that in vivo inhibition of cathepsin G reduces mammary tumor-induced osteolysis. This suggests that cathepsin G is a potentially novel therapeutic target in the treatment of breast cancer bone metastasis. (Wilson et al, 2008)
· CD24 has been reported to have a role in metastasis of several malignancies including breast cancer. We investigated the effect of CD24 expression on biologic features in human breast cancer cells, and found that CD24 expression is associated with more rapid growth and enhanced ability of adhesion and invasion in MCF-7. The proportion of cells was higher in synthetic phase and lower in arrestic phase for CD24(high) than for CD24(low/-) cells. Cyclin D1 and p27 had stronger expressions in CD24(low/-) cells than CD24(high) cells. These suggested one possible pathway for CD24 effect on proliferation. (Kim et al 2007)
· Ceruloplasmin – is used to test for cancer (breast), cardiovascular disease, cirrhosis, diabetes mellitus, rheumatoid arthritis. Ceruloplasmin is an alpha2-globulin transport protein that transports copper and aids in mobilizing iron stores. It is an acute-phase reactant that becomes elevated during stress, pregnancy, and infection. Laboratory Tests and Diagnostic Procedures, p. 322 (2008) *
Copper – High serum levels of copper correlate with certain cancers and high copper appears to play a role in cancer promotion. The angiogenic activity of bFGF (basic fibroblast growth factor), VEGF, TNF-α (Tumor necrosis factor alpha) and IL-1 (Interleukin-1) were found to be copper dependent.
* Serum copper and ceruloplasmin levels were estimated in 20 patients each of prostate and colon cancer. Although copper to ceruloplasmin ratio was not significantly altered, copper and ceruloplasmin levels were increased significantly in the cancer patients as compared to controls. (Navak et al 2003)
· CK-19 negativity in basal-like carcinoma - Basal-like carcinoma of the breast is associated with genetic instability and aggressive behavior. In this study, we evaluated the luminal cytokeratin marker CK-19 in young women with breast cancer treated with conservative surgery and radiation therapy (CS+RT. Positive expression of ER, PR, HER-2/neu, CK19, and p53 were 33.1%, 34.5%, 10.0%, 79.5%, and 20.9%, respectively. With 20 local relapses and 38 distant metastases, the 10-year overall, breast relapse-free, and distant relapse-free survival were 79.65%, 87.29%, and 67.35%, respectively. Tumor stage and nodal status were associated with distant relapse-free and overall survival. In multivariate analysis, CK19 negativity was a predictor of poor local (RR, 3.54; 95% CI, 1.87-7.65; p < 0.01) distant (RR, 1.44; 95% CI, 0.86-2.70; p = 0.17), and overall survival (RR, 1.89; 95% CI, 1.04-3.55; p = 0.03). CONCLUSIONS: Lack of CK19 expression identifies a subset of patients with a significantly higher risk of local relapse. Distant relapse and overall survival rates also correlated with CK19 negativity. Further evaluation of the prognostic significance of basal and luminal cytokeratins in young women with breast cancer is warranted. (Parikh et al 2008)
· CK-19 - Detection of CK-19 mRna-positive CTCs before adjuvant chemotherapy predicts poor clinical outcome mainly in patients with ER-negative, triple-negative, and HER2-positive early-stage breast cancer. (Ignatiadis et al, 2007)
· CK-19 - The detection of CK-19 mRNA–positive CTCs in the blood of early breast cancer patients after adjuvant chemotherapy is an independent risk factor indicating the presence of chemotherapy-resistant residual disease. (Xenidis et al 2009)
· C-kit -- an immunohistochemical assay to detect c-kit protein in biopsy tissue for use as an aid in diagnosing gastrointestinal stromal tumors and determining patient eligibility for imatinib mesylate therapy, a hemoglobin A 1C test for home or office use to assess glycemic control in diabetic patients, and an immunoassay system capable of simultaneously detecting six autoantibodies in blood serum for use as an aid in the diagnosis of certain autoimmune disorders. (July 22, 2005 — The U.S. Food and Drug Administration (FDA)
· C-Reactive Protein - Because it disappears rapidly when inflammation subsides, its detection signifies the presence of a current inflammatory process. Laboratory Tests and Diagnostic Procedures, p. 393 (2008)
· CTCs (Circulating Tumor Cells) - Biocharacterization of CTCs in the peripheral blood of advanced breast cancer patients may represent a real-time tumor biopsy. This study suggests that a subset of patients with HER-2 negative primary tumors develops HER-2 positive CTCs during disease progression. (Pestrin et al, 2009)
· CTCs (Circulating Tumor Cells) - Of all prognostic markers, the levels of CTCs at baseline and at the first follow-up visit were the most significant predictors of progression-free and overall survival in patients with metastatic breast cancer. (Cristofanilli et al 2004)
· DNA index - see glossary of terms.
· E-cadherin - The loss of E-cadherin expression or function in epithelial carcinomas has long been thought as a primary reason for disruption of tight epithelial cell-cell contacts and release of invasive tumor cells from the primary tumor. (Hazan 2004)
· E-Cadherin – E and N cadherin are calcium dependent cell adhesion molecules that mediate cell-cell adhesion and also modulate cell migration and tumor invasiveness. The loss of E-cadherin-mediated adhesion has been shown to play an important role in the transition of epithelial tumors from a benign to an invasive state. (Hazan 2000)
· E-cadherin and AR - One recent study concluded that E-cadherin and AR (androgen receptor) could be candidate predictive factors for chemotherapy response in triple negative breast cancer. (Koo 2009)
· EGFR – epidermal growth factor receptor - participates in cellular signaling pathways. It is expressed in many human tumor types.
· EGFR - There are now several trials exploring the potential of EGFR inhibitors in triple negative disease in the metastatic breast cancer setting and evaluating a combination of EGFR-inhibitors + platinum agent.
· EGFR (serum) and serum HER-2neu are useful predictive and prognostic markers in metastatic breast cancer patients treated with metronomic chemotherapy. (Sandri et al 2007)
· EGFR over-expression in triple negative and basal-like breast carcinoma is now well-established, as is therefore the therapeutic value of EGFR-inhibition.
· Estradiol Receptor and Progesterone Receptor in Breast Cancer – This diagnostic test is used to predict response to hormonal therapy in patients with breast cancer. Estrogen receptors and progesterone receptors are intracellular proteins that specifically bind estrogens and progesterones. The establishment of receptor status in patients with breast cancer is crucial because the receptors are the most predictive factor for the response to hormonal therapies for primary and metastatic breast cancer. Although receptor status is important in determining which patients are likely to benefit from endocrine therapy, estrogen and progesterone receptor status is only a weak predictor of long-term relapse and mortalities and is not to be used alone to assign a patient to a particular prognostic grouping. It should be noted that breast cancers that are initially hormone dependent might progress to a hormone-independent form, despite the continued expression of the receptor. This may limit the long-term usefulness of the hormonal therapies. Laboratory Tests and Diagnostic Procedures, p. 486-487 ( 2008)
· HER-2/neu Oncogene – Is overexpressed in brain cancer, breast cancer (predictive of poor short-term prognosis, but can benefit from Herceptin therapy), colorectal cancer (predicts poor survival), esophageal cancer, malignant mesothelioma, non-small-cell lung cancer, ovarian cancer, pancreatic adenocarcinomas, endometrial carcinoma, synovial sarcomas and uterine serous papillary carcinoma. HER2/neu is a gene that codes for a transmembrane tyrosine kinase. Amplification of the gene product is noted in up to 30% of breast, ovarian and endometrial carcinomas. High levels of the gene product are associated with low estrogen and progesterone receptor concentrations and patients have little or no response to hormonal therapy. Breast cancer patients with overexpression of HER-2/neu have a better response to doxorubicin HCl (Adriamycin)-based chemotherapy regimens. Testing for HER-2/neu overexpression is useful when one is predicting which patients will have a response to monoclonal antibodies directed against HER-2/neu. Laboratory Tests and Diagnostic Procedures, p. 632-633 ( 2008)
· HER-2/neu protein is a glycoprotein cell surface receptor that is composed of an extracellular domain, a transmembrance domain, and an intracellular domain and is over expressed by many adenocarcinomas including breast adenocarcinoma.
· Homocysteine – is created when the body uses the amino acid, methionine, for methylation. Methylation is an important reaction in the body, which leaves homocysteine as a by-product. Normally homocysteine is converted back to methionine, or used to create cysteine and other useful substances. If these conversions are blocked, however, homocysteine accumulates, which can lead to a host of negative reactions. Abnormal metabolism and elevated blood levels of homocysteine create a condition that is highly toxic to both cellular and fibro-elastic components of the vascular wall. Homocysteine can damage blood vessels and nerves, and has been linked to heart attacks, strokes, cancer (particularly colon, breast, and prostate), Alzheimer’s disease, osteoporosis and other neurological diseases, depression, birth defects, gout, cervical dysplasia, and rheumatoid arthritis.
· Ki-67 – A strong correlation exists between the proliferation rate of tumors and clinical outcome, and there is an association between a high proliferation rate and the degree of tumor aggressiveness. The Ki-67 immunostaining, along with histopathologic evaluation, has been shown to be beneficial in correlating the proliferation state of cancers such as breast, lymphatic, lung and brain with clinical prognosis. Laboratory Tests and Diagnostic Procedures, p. 690 (2008)
· Ki-67 - In this study cytological grading, together with Ki-67 and c-kit indices, proved to be helpful in flat epithelia atypia and columnar cell lesions evaluations on core breast biopsy. (Tomasino, et al, 2009)
· Ki-67 - The average Ki-67 index was lowest in luminal A (15.8%), intermediate for ERBB2 (27.8%) and highest for triple negative tumors (>50%). (Bhargava, et al, 2009)
· Ki-67 – The Ki-67 monoclonal antibody is used to determine the proliferation state of breast cancer, bone cancer, brain tumors, cervical cancer, colorectal cancer, endometrial cancer, liver tumors, lymphomas. Laboratory Tests and Diagnostic Procedures, pp.689-690 (2008)
· Mitotic index - see glossary of terms.
· nm23 protein expression appears widely expressed in normal breast, early and advanced breast cancer stages. Interestingly our study found that strong staining intensity and nuclear localization of nm23 protein may prove to be a useful marker of breast cancer progression. (Ismail et al 2008)
· p21 -- an important gene-protein involved in maintaining and regulating cell behavior. High levels of p21 are associated with cancer protection while the lost expression of p21 is associated with cancer progression.
· p21- The expression of p21, a transcriptional target of p53, increases in cells where p53 is stabilized due to exposure to genotoxic agents. (from RnDSytems web site)
· P27 – a cyclin-dependent kinase inhibitor. A cancer marker.
· p53 –– a tumor suppressing gene located on chromosome 17p.
· p53 – Breast cancers with p53 mutations appear to be relatively resistant to ionized radiation and seem to benefit significantly from CMF chemotherapy. Laboratory Tests and Diagnostic Procedures, p. 361 (2008)
· p53 - Negative estrogen and progesterone receptor status (ER/PR) was positively associated with both p53 protein overexpression (OR = 2.6, 95% CI = 1.7-4.0), and p53 mutation in breast cancer. (Rossner et al, 2008)
· p53 - Proapoptotic proteins directly induced by p53 include among others, Bax, Apaf 1, PUMA. P53AIP1, PIDD and NOXA. (Gasco 2002)
· p53 – The detection of mutations in tumor-suppressor genes is associated with approximately half of human cancers, including colorectal, breast, bladder, esophageal, liver, lung, ovarian and brain (p53) and pancreas cancers; leukemias; and male germ cell cancers (DCC). Laboratory Tests and Diagnostic Procedures, p. 361 (2008)
· p53 functions to eliminate and inhibit the proliferation of abnormal cells, thereby preventing neoplastic development.
· p53 mutation - Although the overall frequency of p53 mutation in breast cancer is approx. 20%, strikingly, in typical medullary breast carcinomas, p53 mutation occurs in 100% of cases. This is of particular interest, since it is now well recognized that medullary breast cancers share clinicopathological similarities with BRCA1-associated cases. Indeed, methylation-dependent silence of BRCA1 expression occurs commonly in medullary breast cancers.(Gasco 2002)
· p53 mutation correlates with poor response to DOX (Doxorubicin) in breast cancer.
· p53, P-cadherin and E-cadherin - Another study supported the assessment that basal cytokeratins and androgen receptors, in addition to the traditional pathologic parameters (tumor size and nodal status), in order to provide prognostic information in the group of tumors with triple-negative phenotype. Assessment of p53, P-cadherin and E-cadherin did not add significant prognostic information in this class of tumors. In the lymph node-negative subgroup, basal phenotype can provide powerful prognostic information independent of other well-established markers and can indentify a specific subgroup of patients that may benefit from a more aggressive approach to adjuvant therapy. These researchers emphasized the importance of routine staining of TNBC with androgen receptors and basal cytokeratins. (Rakha 2006)
· p63 - There are several markers relatively frequently utilized in the immunohistochemistry in the differential diagnosis of breast lesions. Myoepithelial markers (p63, alpha-SMA, smooth muscle myosin heavy chain, and others) are useful to highlight myoepithelial cells. They are employed to verify myoepithelial cell lining in intraductal papillary lesions, or to recognise peripheral myoepithelial cells for non-invasive carcinoma, although their staining results are not always excellent. (Moriya, et al, 2009)
· PDGF - Inhibition of PDGF has shown to suppress metastatic cancer.
· PDGF - platelet-derived growth factor – Expression of PDGF, and activation (by autophosphorylation) of its receptor (PDGFR), a tyrosine kinase, is associated with the growth of metastasis in several forms of cancer including kidney, breast, prostate, sarcomas, cervical and in head and neck cancer.
· Plasminogen activator inhibitor-1 (PAI-1) inactivates PTEN.
· Prolactin – A review article by Dumitdrescu and Cotarla (2005) cites a low-to-moderate increased risk for breast cancer in women with elevated prolactin levels. Laboratory Tests and Diagnostic Procedures, p. 907 (2008)
· Prolactin is an important blood tumor marker to assess in breast and prostate cancers.
· PTEN - The inactivation of PTEN – phosphatase and tensin homologue – an important tumor suppressing gene -- was found in about one third of all breast cancers. The reduced expression of PTEN protein correlated with lymph node metastases and a worse prognosis in the patients with breast cancer.
· PTEN loss in metastases is predictive of resistance to EGFR-drugs and chemotherapy. The combination of PTEN and KRAS mutational analysis could help to identify a subgroup of patients who have higher chances of benefiting from EGFR inhibition. (J Clin Oncol. 2009 Apr 27)
· PTEN- Mutations or deletions of the PTEN gene are increasingly being reported in human malignancies, including breast cancer.
· PTEN mutations, or lost expression of, relate to the activation of NF-κB, COX-2 and EGFR. They also feature of poor prognosis and resistance to chemotherapy, hormone therapy and herceptin therapy.
· S-phase expression - see glossary of terms.
· Survivin – an NF-kB regulated gene that functions by blocking the apoptosis (cell suicide) pathway, thus immortalizing cancer cells.
· Survivin - The expression of survivin, a member of the inhibitor-of-apoptosis (IAP) family, is elevated in various human cancers originating in the breast, lung, prostate, colon, pancreas and stomach and is the result of p53 mutation.
· VEGF - OBJECTIVES: The aim of this work was to explore the value of serum vascular endothelial growth factor-A (VEGF-A) in patients with metastatic triple negative breast cancer (TNBC) treated with chemotherapy. The primary end point was overall survival (OS). Secondary end points were response rate (RR), progression-free survival (PFS) and VEGF-A level at baseline, mid-therapy and at the end of therapy. DESIGN AND METHODS: Female patients aged 18 years or above with histologically proven metastatic TNBC were included. Serum VEFG-A levels were measured at baseline, after the 3rd and 6th cycles of FAC chemotherapy regimen (Fluorourcil, Adriamycin, and Cyclophamide). RESULTS: The overall RR was 57%. The median PFS and OS were 7 and 11.2 months, respectively (95% CI: 4.3-9.7 and 3.8-18.5 months, respectively). Patients whose disease progressed despite therapy had a significantly higher baseline VEGF-A level than those who did not progress. VEGF-A level did not drop with continuation of therapy. Patients with high VEGF-A level had a significantly lower PFS but not OS than patients with low levels. CONCLUSION: The outcome of metastatic TNBC is poor with FAC chemotherapy regimen. Alternative chemotherapeutic regimens and novel therapeutic approaches including targeting of VEGF and/or its receptors are warranted. (Taha et al 2009)
Breast Cancer and Bone Metastasis
· In spite of advances in treatment strategies, about 25%-40% of patients with breast cancer still eventually develop metastatic disease. The choice of the optimal therapeutic strategy for patients with metastatic disease is largely influenced by prior exposure to adjuvant therapies. (Guarneri and Conte, 2009)
· Blood Test relating to cancer in the bone: Bone specific alkaline Phosphatase.
· Bone specific alkaline phosphatase - Biochemical markers of bone resorption and bone formation are abnormally raised in the blood and urine of patients with metastatic bone disease. Markers of bone collagen breakdown, such as N-telopeptide, as well as markers of osteoblast function, such as bone specific alkaline phosphatase, appear to be of use in assessing and managing patients with malignancies that metastasize to bone. In this study, both NTx and BAP showed a significant correlation with both the presence of bone metastases and the extent of skeletal involvement. Biochemical markers of bone remodeling can also be used to guide decision making regarding the treatment of metastatic bone disease and to determine the effectiveness of therapy for patients with cancer to bone whose broad-based symptoms make it difficult to discern true response to therapy. (Demers et al, 2000)
· Ki-67 – The Ki-67 monoclonal antibody is used to determine the proliferation state of breast cancer, bone cancer, brain tumors, cervical cancer, colorectal cancer, endometrial cancer, liver tumors, lymphomas. Laboratory Tests and Diagnostic Procedures, pp.689-690 (2008)
· More recent data have suggested that IL-8 plays a major role in the predilection of breast cancer for metastasis to bone and that IL-8 expression in breast cancer relates to estrogen receptor negativity. (Benoy, et al, 2004)
· Increased levels of IGF-1 lead to activation of NF-κB, which in turn decreases apoptosis, increases mitosis, and increases the proliferation of cancer in bone.
· Cathepsin G – is a serine protease which plays a vital role in the bone microenvironment by modulating tumor-stromal interaction in a manner that favors tumor establishment and regulates chemotaxis* of monocytes, a subset of which has the potential to differentiate into osteoclasts. Cathepsin G-induced chemotaxis* of monocytes is mediated by proteolytic activation of protease-activated receptor-1 (PAR-1). Attenuation of PAR-1 activation abrogates cathepsin G-mediated induction of monocyte chemotaxis. Therapeutic targeting of both PAR-1 signaling in osteoclast precursors as well as cathepsin G at the tumor-bone interface has the potential to reduce osteolysis by inhibiting the recruitment, differentiation and activation of osteoclast precursors. (Wilson et al, 2009)
* Chemotaxis = The characteristic movement or orientation of an organism or cell along a chemical concentration gradient either toward or away from the chemical stimulus. Bacteria exhibit chemotaxis when they move toward a source of nutrients.
· The expression of ER, PR, c-erB-2 and cathepsin D were evaluated in 55 breast cancer patients. Metastases were found to occur in bone (24.4%), lungs (20.8%) and liver (10.7%). Histopathological type of invasive ductal carcinoma was associated with the higher incidence of bone metastasis. (Irawan, et al 2008)
· It was found that transforming growth factor beta receptor 1 (TGF-βR1) is a commonly upregulated gene at the tumor bone interface. These studies demonstrate that inhibition of TGF-βR1 signaling at the tumor bone interface will be a therapeutic target in the treatment of breast cancer-induced osteolysis. (Futakuchi, et al, 2009)
· Expression of PDGF, and activation (by autophosphorylation) of its receptor (PDGFR), a tyrosine kinase, is associated with the growth of metastasis in several forms of cancer including kidney, breast, prostate, sarcomas, cervical and in head an neck cancer.
· PDGF – Platelet-derived Growth Factor – Inhibition of PDGF has shown to suppress metastatic cancer.
· Inhibiting PDGFR phosphorylation may, especially in combination with agents such as Taxane-rich Yew extract, Artemisinin with a whole plant extract, produce substantial therapeutic effects against bone metastasis.
· Curcumin and EGCG can also interfere with the expression of VEGF by processes other than hypoxia, such as transforming growth factor (TGF)-β release, COX-2 over-expression, hydrogen peroxide release from bone cells, constitutive and aberrant EGFR and Src signaling, and most importantly, by aberrant NF-κB signaling in established cancers.
· Boswellia is a LOX-5 inhibitor and will help prevent cancer from going to the bone. (Donnie – Meds of the Earth 2009)
· I-3-C demonstrated an ability to inhibit breast cancer bone metastasis by down-regulating MMP-9 & NF-κB. (Mol Cancer Ther 2006)
· Ursolic acid possesses anabolic activity in the bone (Pharm Res. Vol 58 Issue 5-6 pp. 290-296) making it useful for strengthening bone against cancer.
· Chaste tree – has cancer suppressing effects and improves bone health.
· Reishi has shown radiation and chemotherapy-protective ability due to its stimulating effects on bone marrow.
· In one study it was shown that resveratrol’s antiproliferative effects on osteosarcoma cells are mediated by the activation of the ERKs/p53 signaling pathway and therefore identifies new targets for strategies to treat and/or prevent osteosarcoma. (Alkhalaf and Jaffal, 2006)
· Matrix metalloproteinase MMP-7, in human breast-to-bone metastases. We propose that the solubilization of RANKL by MMP-7 is a potential mechanism through which MMP-7 mediates mammary tumor–induced osteolysis. Our studies indicate that the selective inhibition of MMP-7 in the tumor-bone microenvironment may be of benefit for the treatment of lytic breast-to-bone metastases. (Thiolloy et al, 2009)
· In mouse models of "triple-negative" or basal-like breast cancer, treatment with TGFbeta neutralizing antibodies or receptor kinase inhibitors strongly inhibits development of lung- and bone metastases. These TGFbeta antagonists do not significantly affect tumor cell proliferation or apoptosis. Rather, they de-repress anti-tumor immunity, inhibit angiogenesis and reverse the mesenchymal, motile, invasive phenotype characteristic of basal-like and HER2-positive breast cancer cells. Patterns of TGFbeta target genes upregulation in human breast cancers suggest that TGFbeta may drive tumor progression in estrogen-independent cancer, while it mediates a suppressive host cell response in estrogen-dependent luminal cancers. Consequently, TGFbeta antagonists may convert basal-like or HER2-positive cancers to a more epithelioid, non-proliferating (and, perhaps, non-metastatic) phenotype. Conversely, these agents might antagonize the therapeutic effects of anti-estrogens in estrogen-dependent luminal cancers. These predictions need to be addressed prospectively in clinical trials and should inform the selection of patient populations most likely to benefit from this novel anti-metastatic therapeutic approach. (Tan, Alexe and Reiss, 2009)
· An upcoming G&D paper reveals how two specific matrix metalloproteinase (MMP) proteins contribute to bone metastasis in advanced breast cancer - lending important new insight into the design of clinically useful small molecule inhibitors.
"More than 70% of late stage breast cancer patients have skeletal complications," explains Dr. Yibin Kang. "It is important to uncover molecular mechanism of bone metastasis in order to come up with better treatments to reduce the pain and suffering from bone metastasis."
MMP1 and ADAMTS1 are upregulated in breast cancer cell lines with an enhanced ability to metastasize to bone. Dr. Kang and colleagues demonstrated that MMP1 and ADAMTS1 enzymatically cleave and release EGF-like growth factors from tumor cells to stimulate EGFR signaling in the bone-building osteoblasts. The researchers went on to show that such signaling reduces the production of OPG, a suppressor of the bone-resorbing osteoclasts, eventually leading to hyperactivity of osteoclasts, bone destruction and subsequent expansion of bone metastasis.
This paper supports a rationale for the therapeutic targeting of MMP1 and ADAMTS1, and suggests that inhibition of EGFR signaling in bone stromal cells to block osteoclast activity may represent a viable method of mitigating bone complications in advanced metastatic breast cancers. (The study was led by Dr. Yibin Kang in Princeton University in close collaboration with Dr. Joan Massague at MSKCC and Dr. Michael Reiss at the Cancer Institute of New Jersey. It will be published online ahead of print at www.genesdev.org/cgi/doi/10.1101/gad.1824809.)
Herceptin, Tamoxifen and Aromatase Resistance
· Herceptin – a drug that inhibits HER-2 neu. Using botanical agents to enhance the immune system has been shown to improve the effectiveness of monoclonal antibodies, including herceptin, against HER-2 neu.
· Estrogen-related receptor alpha (ERRalpha) and ERRgamma status may be predictive of sensitivity to hormjonal blockade therapy, and ERRalpha status may also be predictive of ErbB2-based therapy such as Herceptin. Moreover, ERRalpha and ERRgamma are candidate targets for therapeutic development. (Ariazi et al 2002)
· PTEN – phosphatase and tensin homologue is an important tumor suppressing gene. PTEN mutations, or lost expression of, relate to poor prognosis and resistance to chemotherapy, hormone therapy and herceptin therapy.
· Many clinical studies have found an association between HER2 overexpression and Tamoxifen failure, and recently a meta-analysis of seven studies concluded that metastatic breast cancer over expressing HER2 was resistant to tamoxifen. (Read et al, 1990) (Pietras et al, 1995) (De Laurentiis M, er al, 2000)
· Tamoxifen – Hormone-blocking drugs, such as tamoxifen, are used to treat and prevent recurrences of breast cancer. Estrogen stimulates cancer growth because it binds to a nuclear receptor that contains a heat-shock protein (HSP) molecule. Upon binding the HSP disassociates itself and allows changes in the receptors to occur. This leads to activation of DNA transcription. Hormone-blocking drugs work by binding to specific estrogen receptors on the surface of tumor cells where hormones would normally bind, inhibiting the cell from growing and reproducing. Tamoxifen works by competing with the estrogen receptors, thus inhibiting the effects of estrogen on target cells. Tamoxifen resistance, however, may allow the proliferation of a more aggressive form of cancer that can be difficult to treat. This resistance can develop to the extent that tamoxifen itself actually stimulates cancer growth. Herbal Medicine, Healing & Cancer (1999)
· Tamoxifen -- is the endocrine therapy for all stages of estrogen receptor positive (ER) breast cancer (BC). Although tamoxifen has shown improved overall 5 year survival for ER positive BC it virtually has had no benefit on ER negative BC. In contrast to the consistent response to estrogen in normal target tissue (e.g. uterus and vagina), only half of the so-called ER-positive tumors respond to adjunctive tamoxifen; and of the half that do respond, many mutate over time and become resistant to tamoxifen. Thus some intrinsic mechanism is already present in half of the ER-positive breast tumors that subverts tamoxifen’s action.
· Resistance to Tamoxifen - AIB1 (amplified in breast cancer 1), also called SRC-3 and NCoA-3, is a member of the p160 nuclear receptor co-activator family and is considered an important oncogene in breast cancer. Increased AIB1 levels in human breast cancer have been correlated with poor clinical prognosis. Overexpression of AIB1 in conjunction with members of the epidermal growth factor receptor (EGF/HER) tyrosine kinase family, such as HER2, is associated with resistance to tamoxifen therapy and decreased disease-free survival. This review summarizes the possible mechanisms of how dysregulation of AIB1 at multiple levels can lead to the initiation and progression of breast cancer as well as its role as a predictor of response to breast cancer therapy, and as a possible therapeutic target. (Lahusen et al 2009)
· Resistance to Tamoxifen - Patients receiving adjuvant tamoxifen whose tumors express high levels of both HER2/neu (HER2) and the estrogen receptor (ER) coactivator AIB1 often develop tamoxifen resistance. (Shou et al 2004)
· Women who take tamoxifen for 5 years probably have a 2 to 4 times increased risk of developing other cancers including endometrial cancer, uterine sarcoma, a non-estrogen dependent more aggressive breast cancer, and a substantially increased risk of thromboembolic complications.
· Women with breast cancer who use tamoxifen appear to be at increased risk of developing estrogen-receptor negative contralateral tumors, researchers from the Fred Hutchinson Cancer Research Center in Seattle report.
· Tamoxifen induces cellular oxidative stress leading to the activation of activating protein-1 (AP-1), which in turn causes not only tamoxifen resistant breast cancer to develop but leads to tamoxifen activated breast cancer. (Jour. of Nat’l Cancer Inst. Dec. 6, 2000)
· Tamoxifen and p53 expression - In another study it was found that tamoxifen (Tam) use in maintenance therapy of breast cancer may be associated with an increased risk of endometrial cancer. This preliminary evaluation indicates that there is a higher incidence of p53 expression in Tam treated patients.” (Barakat RR., Adhikari D., Federici M., Saigo PE., Bertino JR., Banerjee D. Bcl-2 and c-Ki-ras status in endometrial cancers occurring after breast cancer with or without tamoxifen treatment. Evaluation of p53, Memorial Sloan Kettering Cancer Center, New York, NY.)
· After five years, the incidence of gallstone formation was 37.4% in tamoxifen-treated patients compared with 2.0% in patients who did not receive tamoxifen. Adjuvant tamoxifen therapy leads to gallstone formation in postmenopausal breast cancer patients and is most apparent after 3 years of treatment. The risk of gallstone formation increased with longer duration of hormone use for current postmenopausal patients and with higher doses of estrogen used (in another study). (Barclay 2003)
· The lysosomal protease Cathepsin D (Cath D) is associated with increased invasiveness and metastasis in breast cancer. Immunofluorescence showed that estradiol located Cath D to the cell surface, while tamoxifen accumulated Cath D to dense lysosomes in perinuclear regions. Moreover, tamoxifen increased the intracellular transporter of Cath D, the mannose 6-phosphate/IGF-II receptor (M6P/IGF2R). In contrast, estradiol decreased the levels of this receptor. Thus secretion of Cath D is hormone dependent and may be mediated by altered expression of the M6P/IGF2R. (Dabrosin et al 2004)
· Tamoxifen contributes to hypothyroidism through thyroid peroxidase, inducing oxidative stress. (Koc et al 2000) (Beyssen et al 2000) (Cutuli et al 2000) (Giani et al 1996) (Iino et al 1996)
· X-box-binding protein-1, or the XPB1 gene is an alternatively spliced transcription factor that participates in a stress-signaling pathway to protect cells from damage. Over-expression of the spliced variant of the gene in estrogen receptor-positive breast cancer cells led to reduced sensitivity to Tamoxifen and Faslodex. (Clark et al 2007)
· Gamma linolenic acid accelerates response to Tamoxifen in breast cancer. (Robertson et al, 2000)
· Cell culture work has shown that human breast cancer cells have a remarkable ability to adapt to the environment in which they are grown and to circumvent any form of imposed growth inhibition. Growth of estrogen responsive human breast cancer cells in the long-term absence of estrogen results in an initial phase of slow growth but this is followed by up regulation of growth such that the cells eventually grow at the same rate in the absence of estrogen as they originally grew only in the presence of estrogen. Only one report has documented loss of ER in this process. In most models, ER are not lost and remain functional in that estrogen-regulated genes such as pS2 or progesterone receptor are still increased by the re-addition of estradiol. (Shaw et al 2005)
· Tamoxifen (TAM) has estrogenic activity on liver and endometrium causing severe oxidative stress with various biochemical derangements. Coenzyme Q(10), Riboflavin and Niacin (CoRN) are well-known potent antioxidants and protective agents against many diseases including cancer. In this context, this study was undertaken to find if co-administration of TAM along with CoRN could alleviate the sole TAM-induced biochemical derangements in postmenopausal women with breast cancer. A statistically significant alteration in various blood chemistry parameters, such as serum total bilirubin (S. BIL), serum glutamate oxaloacetate transaminase (SGOT), serum glutamate pyruvate transaminase (SGPT), gamma glutamyl transpeptidase (gamma-GT), uric acid (UA), lipoprotein lipase (LPL), lecithin: cholesterol acyl transferases (LCAT), potassium, calcium and Na(+), K(+)-ATPase in sole TAM-treated group, was favorably reverted back to near normal levels on combinatorial therapy with CoRN. CONCLUSION: TAM on co-administration with CoRN has a favorable impact on various blood chemistry profiles. However, large scale randomized studies over a longer time span are required to ascertain the safety and efficacy of co-administrating antioxidants with conventional chemotherapy. (Yuvaraj et al 2008)
· Tamoxifen-induced hepatic steatosis - Oral administration of tamoxifen, an endocrine therapy for breast cancer, often induces hepatic steatosis (THS, tamoxifen-induced hepatic steatosis) as a complication, which can progress to non-alcoholic steatohepatitis (NASH). The development of this complication is strongly associated with three clinical risk factors; specifically, insulin resistance, central obesity, and hypertriglyceridemia, however a genetic predisposition to THS has yet to be investigated. Our study provides the first evidence that CYP17 polymorphism participates in the development of THS, and sheds light on the genetic causes of this side effect and genetic differences between tamoxifen-treated individuals. (Ohnishi et al 2005)
· Tamoxifen and CYP2D6 - The clinical outcomes of breast cancer patients treated with tamoxifen may be influenced by the activity of cytochrome P450 2D6 (CYP2D6) enzyme. Among 67 patients examined, those homozygous for the CYP2D6*10 alleles revealed a significantly higher incidence of recurrence within 10 years after the operation (P = 0.0057; odds ratio, 16.63; 95% confidence interval, 1.75-158.12), compared with those homozygous for the wild-type CYP2D6*1 alleles. Patients with the CYP2D6*10/*10 genotype showed a significantly shorter recurrence-free survival period (P = 0.036; adjusted hazard ratio, 10.04; 95% confidence interval, 1.17-86.27) compared to patients with CYP2D6*1/*1 after adjustment of other prognosis factors. The present study suggests that the CYP2D6 genotype should be considered when selecting adjuvant hormonal therapy for breast cancer patients. (Kiyotani et al 2008)
· Tamoxifen and CYP2D6 - Tamoxifen has been the mainstay adjuvant hormonal treatment for breast cancer for many years. Conversion of tamoxifen to its active metabolite, endoxifen, is reduced by low activity of the cytochrome P450 enzyme, CYP2D6. We examined the effect of reduced CYP2D6 activity on the response to tamoxifen in patients with familial early-onset breast cancer. Poor metabolizer status for CYP2D6 predicts worse overall survival in patients with familial breast cancer. Therefore, CYP2D6 inhibitor drugs should not be prescribed concomitantly with tamoxifen. Prospective studies should be undertaken to establish the effect of CYP2D6 status on outcome in familial breast cancer patients treated with tamoxifen. (Newman et al, 2008)
· Tamoxifen and CYP2D6 - Although prospective data are lacking, the balance of evidence available currently suggests that a single nucleotide polymorphism in the CYP2D6 gene, particularly the presence of 2 null alleles, predicts for reduced tamoxifen metabolism and possibly poorer outcome than expected in patients with a wild-type genotype. (Higgins et al 2009)
· Tamoxifen and CYP2D6 - Adjuvant endocrine treatment with aromatase inhibitors improves disease-free survival compared with tamoxifen in postmenopausal women with estrogen receptor-positive breast cancer. This difference could be due to differences in tamoxifen metabolism because levels of endoxifen, the active tamoxifen metabolite, vary with the number of mutant alleles, including the *4 allele, of the gene encoding cytochrome P450 2D6 (CYP2D6). CONCLUSIONS: Modeling suggests that among patients who are wild type for CYP2D6, 5-year disease-free survival outcomes are similar to or perhaps even superior with tamoxifen than with aromatase inhibitors. Endocrine therapy tailored to CYP2D6 genotype could be considered for women who are newly diagnosed with breast cancer, particularly those who have with concerns about either the relative toxicity or the increased cost of aromatase inhibitors. (Punglia et al 2008)
· Tamoxifen and CYP2D6 - At the least, however, women who are taking tamoxifen should avoid the concomitant use of drugs that inhibit CYP2D6 activity if possible. Ironically, some of the most potent inhibitors of CYP2D6 are the selective serotonin uptake inhibitors and the selective serotonin norepinephrine inhibitors (SSNRIs), which are frequently used for treatment of hot flashes. Examples of these inhibitors include paroxetine and fluoxetine. However, other SSNRIs, such as venlafaxine, do not inhibit CYP2D6 yet are quite effective for treatment of hot flashes, and as such would be the preferred treatment for these patients. Regardless, we believe that these studies have brought the field of pharmacogenetics onto the radar screen of clinicians caring for breast cancer patients. The model reported by Punglia et al. suggests that selection of endocrine therapy based on CYP2D6 genotype might make tamoxifen more than just another choice, but actually the preferred choice, for women with ER-positive breast cancer who are wild type for the CYP2D6 gene. (Hayes et al 2008)
· Soy foods have been a staple in Asia for centuries but the consumption of this food in the West is recent. Intake of soy among women at high risk for or with breast cancer has become a public health concern because genistein, a major component of soy, has weak estrogenic effects on breast epithelium, and has been found to negate the benefit of tamoxifen in some animal and in vitro studies. We found no evidence that soy intake adversely affected levels of tamoxifen or its metabolites. However, age, menopausal status, BMI, and use of hypertensive medications significantly influenced circulating levels of tamoxifen and its metabolites in this population. (Wu et al, 2007)
· Several groups have reported that long-term culture of MCF7 cells in tamoxifen or 4-hydroxythamioxifen results in the emergence of clones of cells resistant to the growth of inhibitory action of these anti-estrogens, and this was not associated with loss of ER content or ER function. (Shaw et al 2005)
· Aromatase – is an enzyme complex involving cytochrome P450, which mediates the conversion of androgens to estrogens. . The production of estrogens from androgens is mediated by the aromatase enzyme, the aberrant expression of which plays a critical role in the disease process in other tissues, most notably the breast.
· Arimidex and Femara are aromatase inhibitors that reduce circulating estrogen. Breast cancer that is driven by estrogen expresses aromatase activity. This is particularly true once tamoxifen resistance has occurred and the cancer that was previously driven by estrogen is now able to covert androgens into estrogens to aid in further proliferation of the cancer. Herbal Medicine, Healing & Cancer (1999)
· HER-2/neu and response to the aromatase inhibitor letrozole versus tamoxifen.
Patients with normal serum HER-2/neu receiving letrozole demonstrated a significantly greater ORR (objective response rate) and CB (clinical benefit) and longer TTP (time to progression) and TTF (time to treatment failure) than patients receiving tamoxifen (an anti-estrogen). Although in patients with elevated serum HER-2/neu there was no significant difference between letrozole and tamoxifen in ORR or CB, there was a strong trend favoring longer TTP and significantly longer TTF with letrozole. (Lipton A, et al 2003)
· Tamoxifen and Aromatase Inhibitors - Several studies have now shown that for postmenopausal women with ER-positive breast cancer, estrogen depletion with an aromatase inhibitor is generally more effective than tamoxifen in both the adjuvant and metastatic settings. A panel convened by the American Society of Clinical Oncology has recommended the use of an aromatase inhibitor at some time in the course of treatment for all postmenopausal women with ER-positive breast cancer. However, despite this enthusiastic endorsement, aromatase inhibitors are not ideal for all women because they are associated with substantially more common and often more severe musculoskeletal complaints that often lead to nonadherence, as well as with higher long-term risks for osteoporosis and fractures. Moreover, because aromatase inhibitors are contraindicated in premenopausal women, tamoxifen is currently the treatment of choice for these patients. Tamoxifen and the related selective ER modulator raloxifene are approved for chemoprevention, whereas aromatase inhibitors are not. Finally, tamoxifen is also effective in the metastatic setting, and as more women receive adjuvant aromatase inhibitor therapy, those who do relapse will be treated with tamoxifen. Therefore, in the foreseeable future, most premenopausal women and many postmenopausal women at risk for or with ER-positive breast cancer may consider tamoxifen at some point in their treatment. (Hayes et al 2008)
For more information on this subject please see our article on Tamoxifen.
Other Markers for Consideration in Breast Cancer
· AP-1 – a transcription factor that controls the expression of genes – the protein products of which participate in complex signal transduction, and thus cell transformation.
· A number of studies suggest that breast cancers associated with BRCA1 mutations are likely to be Triple Negative and the majority of these are also the basal phenotype. Basal-like tumors express certain cytokeratins characteristic of the ’basal’ layer myoepithelial cells lining the terminal duct lobular unit (namely cytokeratins 5, 6, 14 and 17). Basal-like tumours are usually high-grade, exhibit comedeonecrosis, pushing borders and an inflammatory lymphocytic infiltrate. (Young et al 2009)
· BRCA1 – mutations tend to exhibit a very similar histological phenotype to basal-like tumors. The fact that basal-like tumors tend to have very high expression levels of VEGF suggests molecularly targeting VEGF should return special benefit in triple negative disease from the anti-VEGF agent Avastin.
· BRCA-1 – A gene on chromosome 17 that normally helps to suppress cell growth. A person who inherits certain mutations in a BRCA1 gene has a higher risk of getting breast, ovarian, prostate, and other types of cancer. It is a breast cancer susceptibility gene identified on chromosome 17. BRCA-2 is on chromosome 13. They participate in repairing radiation-induced breaks in double-stranded DNA. Mutations in BRCA1 and BRCA2 might disable this mechanism leading to more errors in DNA replication and ultimately to cancerous growth.
· BRCA1 and BRCA2 – are inherited breast cancer tumor suppressor genes that can be recognized on the human genome. BRCA1 and BRCA2 are found on chromosomes 17 and 13 respectively. Inherited mutations of these genes have been found to be associated with an increased risk of familial breast and ovarian cancer occurrences. Carriers of BRCA1 mutations have an increased risk of ovarian cancer (epithelial or transitional cell) and microglandular adenosis, and carriers of BRCA2 are at increased risk for Fanconi’s anemia, pancreatic cancer and prostate cancer. Persons with BRCA1 and BRCA2 mutations have an increased risk of breast cancer. Laboratory Tests and Diagnostic Procedures, p. 255 (2008)
· Another transcriptional coactivator for p53 is BRCA-1. It is, of course, well known that germ-line mutations in BRCA1 predispose carriers to breast and ovarian cancer. (Gasco 2002)
· Triple negative breast cancers have an aggressive clinical course, and EGFR and BRCA1 might be candidate therapeutic targets in this disease. (Toyama, 2008)
· Data from preclinical and clinical studies indicate that both BRCA1 and triple negative tumors have unique sensitivities to platinum agents such as cisplatin and carboplatin, as well as to the genotoxic biological agents, the PARP inhibitors.
· List of triple negative sensitive genotoxic agents: Cytoxan, paraplatin, platinol, adriamycin, ellence, mitomycin/mutamycin, radiotherapy and PARP inhibitors.
· CA-9 (Carbonic Anhydrase-9) - an important blood marker to assess angiogenesis/hypoxia. You want it to be low.
· CAV-1 is highly upregulated in triple negative breast cancer. (Donald Yance – Medicines of the Earth 2009) A recent study by Pinilla et al9 showed that caveolin-1 (CAV1) expression is associated with a triple negative phenotype in sporadic and hereditary breast cancer. ABI-007 (Abraxane) is a novel, biologically interactive, nanometer-sized albumin-bound paclitaxel particle initially developed to avoid the toxicities associated with polyethylated castor oil. In their phase III study, Gradishar et al10 compared albumin-bound nanoparticle paclitaxel, ABI-007 with polyethylated castor oil-based paclitaxel in women with metastatic breast cancer. This study showed greater efficacy and a favorable safety profile of ABI-007 though no subgroup analysis of molecular phenotypes for differential efficacy of the treatment was performed. After the incorporation of ABI-007 with albumin in blood circulation, ABI-007 is preferentially transported from blood to tumor site in two ways. One way is through leaky junction of endothelial cells that are highly pronounced around the tumor tissue by induction of angiogenesis. The second way and perhaps more prominent way is acting through receptor-mediated transcytosis of this albumin-bound ABI-007.11 This second mechanism is mediated by CAV-1. Breast cancer patients with higher CAV-1 expression, as in the cases of breast cancer with triple negative phenotype, may show better efficacy with more favorable safety profile if they receive ABI-007.
· CA 72-4 – Cancer Antigen 72-4 – This test is more sensitive than CA 125 for ovarian cancer. Positive tests may occur in 85% of invasive ductal breast carcinomas and in ovarian and endometrial adenocarcinomas. Specificity can be improved with concurrent testing for other tumor markers. Laboratory Tests and Diagnostic Procedures, p. 272 (2008)
· CD44-4 – is a major E-Electin Ligand that mediates breast cancer cell transendothelial migration.
· CK-5 – Antibodies against stratified-epithelial CKs, such as CK5, identify normal basal/myoepithelial cells and thus may be helpful to recognize invasiveness and malignancy in prostatic and breast lesions, particularly when combined with p63.
· COX-2 may be a molecular target for treating HER-2 over-expressing breast cancer. Breast cancers that demonstrate elevated COX-2 expression were more predictive of poor survival in patients with estrogen receptor (ER)-positive tumors than in those with ER-negative tumors, according to a 2002 report by Finnish researchers. Similarly elevated expression was of greater prognostic value in patients with no abnormalities of the p53 gene and the HER-2 gene. Most common cancers with altered (amplified) COX-2 expression include: prostate, colon, breast, cervical brain, gastric, pancreatic, lung, head and neck, kidney and bladder.
· D-Dimer test – will tell you if you might have a clot somewhere in your body, or risk forming a clot. 0-1 is good and indicates no clot indicated. The higher above the 1 you get, the more the likelihood of a clot. The range Donald Yance likes to see is below 0.40. Detectable fibrin degradation, as measured by plasma D-dimer, is a clinically important marker for lymphovascular invasion and early tumor metastasis in operable breast cancer. Plasma D-dimer levels have been shown to be increased in patients with various solid tumors including lung, prostate, cervical, ovarian, breast and colon cancer.
· HDCT – high-dose chemotherapy. Triple negative disease has been found significantly responsive to HDCT.
· HIF – hypoxia inducible factor. HIF-1 –– a transcription factor that controls the expression of genes – the protein products of which participate in complex signal transduction, and thus cell transformation.
· HSP (Heat Shock Proteins) A small HSP called alpha-basic crystalline is commonly expressed in triple negative tumors and this HSP overexpression increased cell migration and invasion, among other molecular activity, via the MEK/ERK pathway, suggesting that inhibition of the underling MEK/ERK pathway may be an effective therapy for these types of basal-like breast tumors.
· Human protease-activated receptor 1 (hPAR1) is a bona fide receptor of the hemostatic protease thrombin, and has a central function in tumor progression. PAR1 function is attenuated by p53. (Salah, et al, 2008)
· High mammographic breast density known to predict increased breast cancer risk is associated with higher concentrations of circulating IGF-I and insulin-like growth factor-I (IGF-I), which also plays a critical role in carcinogenesis and tumorigenesis. These considerations would help to account for the antitumor effect of caloric restriction via mTOR inhibition, as caloric restriction may involve underlying insulin and IGF pathways, and suggest that both caloric restriction and glucose / insulin control may play specific beneficial functions in triple negative disease via the newfound contribution of mTOR inhibition, and add another item of defense to the growing arsenal deployable against triple negative breast carcinoma.
· IL-8 - A pro-inflammatory cytokine. An important blood marker to assess angiogenesis/hypoxia. You want it to be low.
· The MIB-1 labeling index for non-proliferative disorders, proliferative disorders, and breast cancer was 3.4±1.9%, 8.9±6.2% and 20±12%, respectively. The MIB-1 labeling index and tumor size, lymph node metastasis status, and clinical stage according to the TNM classification correlated significantly. Survival rate was inversely correlated with the MIB-1 labeling index. No patient with an MIB-1 labeling index of less than 10% had lymph node metastases, and all are alive without recurrence. Patients with an MIB-1 labeling index of over 30% had an extremely poor prognosis. Conclusions The MIB-1 labeling index is very useful for predicting both either extremely good or extremely poor prognosis, and axillary lymph node metastasis. (Nakagomi et al 2008)
· mTOR – (mammalian target of rapamycin). The mTOR kinase is downstream of the PI3K/Akt pathway, an important regulator of cell proliferation and survival, and shown to also affect VEGF production at multiple levels, and breast cancers with mTOR overexpression showed a three times greater risk for disease recurrence.
o mTOR: A Central Regulator Of Cell Proliferation, Angiogenesis, and Cell Metabolism
o mTOR is a kinase protein predominantly found in the cytoplasm of the cell. It acts as a central regulator of many biological processes that are essential for cell proliferation, angiogenesis, and cell metabolism.1-3 mTOR exerts its effects primarily by turning on and off the cell's translational machinery, which includes the ribosomes, and is responsible for protein synthesis.1
The location and
role of mTOR in the cell.
o mTOR is a key intracellular point of convergence for a number of cellular signaling pathways. mTOR performs its regulatory function in response to activating or inhibitory signals transmitted through these pathways, which are located upstream from mTOR in the cell. These diverse signaling pathways are activated by a variety of growth factors (including vascular endothelial growth factors (VEGFs), platelet-derived growth factor (PDGF), epidermal growth factor (EGF), insulin-like growth factor 1 (IGF-1)), hormones (estrogen, progesterone), and the presence or absence of nutrients (glucose, amino acids) or oxygen.4,5
o One or more of these signaling pathways may be abnormally activated in patients with many different types of cancer, resulting in deregulated cell proliferation, tumor angiogenesis, and abnormal cell metabolism.1,4,5
o mTOR is an important therapeutic target because:
o It is a key intracellular point of convergence for a number of signaling pathways that are abnormally activated in many types of cancer1,4
o It appears to be a stable target that does not mutate1,5,6
o Inhibiting mTOR:
o May inhibit abnormal cell proliferation, tumor angiogenesis, and abnormal cell metabolism
o May potentially enhance the efficacy of other cancer treatments1, 5, 7
· NFκB – Nuclear Factor Kappa Beta
o NFκB -These results identify the NFκB-mediated signaling pathway and a target gene for MIS action and suggest a putative role for the MIS ligand and its downstream interactors in the treatment of ER-positive as well as negative breast cancers. (Segev et al 2000)
o NFκB is up-regulated in practically all cancers. A transcription factor that controls the expression of genes – the protein products of which participate in complex signal transduction, and thus cell transformation.
o NFκB upregulates telomerase in the cancer cell which makes the cancer cell immortal. It upregulates inflammation, angiogenesis, metastases and tumor promotion. It stimulates COX-2 and all these other inflammatory pathways as well.
o NF-κB up-regulation has been found to be associated with almost every kind of cancer. NF-κB and p53 pathways together play crucial roles in most human cancers in which hyper-activation of NFκB and inactivation of p53 is a common occurrence. Inhibition of NF-κB and activation of p53 promotes apoptosis in cancer cells.
o NFκB is hyperactive in many human cancers raising the resistance of cancer cells to chemotherapy drugs and chemoradiation.
· PAR-1 – protease activated receptor-1 – is identified as a survival factor that protects cells from undergoing apoptosis. Whereas PAR1 gene expression correlates with tumor progression its neutralization effectively initiates an apoptotic pathway leading at least in part to significantly reduced tumor formation. (Salah et al 2007)
· PAR-1 expression correlates with tumour invasiveness, as well as with cancer cell survival. In this study it was shown that hypoxia enhances PAR-1 expression in MDAMB231 breast cancer cells. Evidence was also shown for a novel role of PAR-1 in protecting hypoxic breast cancer against cell death. The overall results identify for the first time a functional role for PAR-1 in the cellular responses of breast cancer to a hypoxic microenvironment. (Naldini et al, 2009)
· PARP (poly(ADP-ribose)polymerase) - Data from preclinical and clinical studies indicate that both BRCA1 and triple negative tumors have unique sensitivities to platinum agents such as cisplatin and carboplatin, as well as to the genotoxic biological agents, the PARP inhibitors.
· Platinum-based chemotherapy achieves increased response rates for TN tumors, with a trend towards worse survival in early breast cancer through an improved survival in advanced disease. (Sirohi 2008)
· Serum IL-8 levels are increased in 67% of patients with advanced breast cancer. . .The IL-8 levels increase significantly in patients with more advanced disease. An elevated serum IL-8 is related to an accelerated clinical course, a higher tumor load, and the presence of liver or lymph node involvement. . . These results further expand the concept that inflammation and inflammatory cytokines are critical components of tumor progression. (Benoy, et al, 2004)
· Serum copper levels correlate with tumor incidence, tumor burden, malignant progression in a variety of human cancers – Hodgkin’s and non-Hodgkin’s lymphoma, sarcomas, leukemias and cancer of the cervix, breast, liver and lung as well as brain tumors. High serum levels of copper correlate with certain cancers and high copper appears to play a role in cancer promotion.
· SPARC (secreted protein acidic and rich in cystein), also known as osteonectin and BM40, is a 32 kDa secreted glycoprotein that interacts with extracellular matrix (ECM) proteins to promote adhesion of cells from the matrix, thereby inducing a biological state conducive to cell migration. SPARC is also thought to play an important role in tissue remodelling, angiogenesis, embryonic development and tumourigenesis. The study concluded that SPARC plays a crucial role in tumour development in breast cancer and as such has a significant bearing on patient prognosis and long-term survival. (Watkins et al 2005)
· t-PA - tissue-type plasminogen activator is associated with the ability to break down fibrin. Fibrin formation and platelet aggregation are important steps to tumor formation.
o One study showed a significant relationship between high levels of t-PA and a favorable prognosis in breast cancer.
· TRAIL - Prognostic significance of tumour necrosis factor-related apoptosis-inducing ligand (TRAIL) receptor expression in patients with breast cancer. TNF-related apoptosis-inducing ligand (TRAIL) induces apoptosis upon binding to TRAIL receptors 1 and 2 (TRAIL-R1/DR4 and TRAIL-R2/DR5). Determination of the TRAIL receptor expression profile may aid in defining which breast cancer patients have a higher risk of lymph node metastasis and worse overall survival and on the other hand will help to guide TRAIL-based tumour therapy. (Ganten et al 2009)
· Vimentin - In this study we examined 198 sarcomas, 38 carcinomas, 13 'tumours with a spindle cell component' and 22 malignant melanomas with a commercial monoclonal vimentin antibody. All histopathological material was formalin fixed and paraffin embedded. The results show this antibody to be a sensitive and specific marker of mesenchymal derivation or differentiation. It is a useful tool in separating sarcomas from most carcinomas, and in separating malignant melanomas from carcinomas. When used in combination with a cytokeratin antibody it identifies carcinosarcomas and synovial sarcomas. (Leader et al 2007)
· Weisenthal Test - The Weisenthal Cancer Group today announced that clinical data published at the annual meeting of the American Society of Clinical Oncology (ASCO) show that a new laboratory test it has developed accurately identified patients who would benefit from treatment with the molecularly-targeted anti-cancer therapies gefitinib (Iressa, AstraZeneca) and erlotinib (Tarceva, Genentech). The new test, called the EGFRx assay, predicted accurately for the survival of patients treated with the targeted drugs. The finding is important because the EGFRx test, which can also be applied to many emerging targeted cancer drugs, could help to help to solve the growing problem of knowing which patients should receive costly, new treatments that can have harmful side-effects and which work for some but not all cancer patients who receive them.
Larry Weisenthal, M.D., Ph.D., a medical oncologist and developer of the EGFRx assay explains that the new test relies upon what he calls “Whole Cell Profiling” in which living tumor cells are removed from an individual cancer patient and exposed in the laboratory to the new drugs. A variety of metabolic and apoptotic measurements are then used to determine if a specific drug was successful at killing the patient’s cancer cells. The whole cell profiling method differs from other tests in that it assesses the activity of a drug upon combined effect of all cellular processes, using several metabolic and morphologic endpoints. Other tests, such as those which identify DNA or RNA sequences or expression of individual proteins often examine only one component of a much larger, interactive process.
According to Dr. Weisenthal, this may explain why EGFRx whole cell profiling is the only test to date to demonstrate a statistically significant association between prospectively reported test results and patient survival. Using the EGFRx assay and the whole cell profiling method, Dr. Weisenthal’s group correlated test results, which were obtained by his lab and reported to physicians prior to patient treatment, with significantly longer or shorter overall patient survival depending upon whether the drug was found to be effective or ineffective at killing the patient’s tumor cells in the laboratory. Patients prospectively identified by Dr. Weisenthal as favorable candidates averaged 485 days of life after treatment with the targeted therapy drugs. In contrast, patients identified as unfavorable candidates for the drugs averaged 75 days survival after receiving the drugs. This compares to 76 days average survival among patients identified as unfavorable candidates and who did not receive a targeted therapy drug. Survival among patients identified by Dr. Weisenthal as unfavorable candidates was therefore similar regardless of whether or not they received the targeted drugs.
Comparing the whole cell profiling approach with other types of tests Dr. Weisenthal states, “Over the past few years, researchers have put enormous efforts into genetic profiling as a way of predicting patient response to targeted therapies. However, no gene-based test as been described that can discriminate differing levels of anti-tumor activity occurring among different targeted therapy drugs. Nor can an available gene-based test identify situations in which it is advantageous to combine a targeted drug with other types of cancer drugs. So far, only whole profiling has demonstrated this critical ability to distinguish between the activities of different targeted agents. The reason this is critical is because there is a growing array targeted drugs to choose from. Also, most patients today are treated not with a targeted therapy drug alone but rather with a combination of chemotherapy drugs. Therefore, the existing DNA and RNA tests do not reflect the way cancer medicine actually is practiced today.”
Several new targeted drugs have been introduced during the last few years and dozens more are on the horizon. These so-called “smart drugs” focus their effects on specific, identifiable processes occurring within cancer cells. The new drugs are highly promising in that they sometimes provide benefit to patients who have failed traditional therapies. However, they do not work for everyone, they often have unwanted side effects, and they are all extremely expensive: some cost patients and insurance carriers $5,000 to $7,000 or more per month of treatment. Patients, physicians, insurance carriers, and the FDA are all calling for the discovery of predictive tests that allow for rational and cost-effective use of these drugs.
“Not only is this an important predictive test that is available today”, says Dr. Weisenthal, “but it is also a unique tool that can help to identify newer and better drugs, evaluate promising drug combinations, and serve as a ‘gold standard’ correlative model with which to develop new DNA, RNA, and protein-based tests that better predict for drug activity.”(Clinical Study Results Published at American Society of Clinical Oncology (ASCO) Meeting, 2006)
Cancer-preventive Botanicals & Nutrients
· Natural Compounds that Down-Regulate COX-2
o EPA/DHA in n-3 fatty acids from fish oils
o Baicalein, from Chinese skullcap (ICII >95%)
o Curcumin
o Transresveratrol
o Pterostilbene
o Quercetin
o Salicin (Corydalis 30%) Willow bark extract (WBE)
o WBE inhibits the cell growth and promotes apoptosis in human colon and lung cancer both through COX-selectivity and nonCOX-2. Other synergistic compounds with WBE include other salicyl alcohol derivates, flavonoids, proanthocyanidins.
· Several foods and common culinary herbs contain particular compounds that reduce inflammation, in part, by acting as COX-2 inhibitors. These include grapes (leaf and skin), curcumin longo (turmeric), Ocimun sanctum (basil), fish, flax, oregano, rosmarinus (rosemary) and licorice root (glycyrrhizin glabra).
· Natural COX inhibitory curcuminoid components of curcumin are active in the regulation of COX-2, EGFR, VEGF, PI3K/Akt, MEK/ERK, p53, c-Myc, NF-kappaB, Bcl-2, e-cadherin, and apoptotic pathways all known to be critically involved in breast carcinomas in general and in triple negative disease in particular, as well as HER2 (ErbB2), and some of which are also regulated by the activity of the EGCG (epigallocatechin-3 gallate) component of green tea. (Kaniklidis 2007)
· Natural compounds shown to block EGF receptor activation and its downstream effectors include:
o Resveratrol
o Vitamin D-3
o Licorice
o Quercetin (inhibits both EGF and HER2-neu expression).
o Cysteine (un-denatured whey protein concentrate)
o Selenium
o Curcumin
o I-3-C/DIM
o I-3-C (indole-3-carbinol) – induced cell death in breast cancer cell lines via the mitochondrial apoptotic pathway. I3C significantly reduced levels of EGFR in breast cancer cell lines.
· Natural compounds that normalize p53
o Curucumin
o Quercetin
o Resveratrol
o OPCs
o 6-Gingerol
o EGCG
o Oridonin (Rabdosia)
o Paw paw seed
o Withanone (Ashwagandha extract)
o Folate, Tocotrienols and Vit E succinate
o Note: Diets rich in refined sugars and starches, as well as rich in red meat promote p53 mutation.
· Natural compounds that suppress PDGF (platelet-derived growth factor)
o Curcumin causes an interruption of the PDGF and EGF signaling pathways by stimulating gene expression of PPARγ
o Curcumin, also decreases the proportion of S phase cells after PDGF stimulation
o Up-regulation of 12-LOX pathway of AA metabolism activates PDGF
o Baicalein, in Chinese skullcap, one of the most powerful anti-cancer agents induces apoptosis of cancer cells. One known mechanism is by down-regulating 12-LOX – reducing PDGF
o EGCG, from GTE, inhibits PDGF-induced VEGF expression via blocking PDGF receptor and Erk-1/2.
· Herbs and Compounds that Inhibit VEGF
o Curcuma longa (turmeric) – 95% curcumin
o Magnolia seed cones – 90% honokiol
o Camellia sinensis (green tea extract) -50% EGCG, plus other compounds.
o Vitus vinifera (grape seed extract) – 95% OPCs
o Angelica sinensis (dong quai) – 4-hydroxzyderricin
o Taxus breviflora (Pacific yew) – taxol and other related taxins
o Scutellaria baicalensis (Chinese Baical skullcap) – 95% baicalin and other compounds, mostly flavonoids
o Polygonum cuspidatum (Japanese knotweed) – 20% resveratrol
o Artemisia annua (Chinese wormwood) – 95% artemisinin and other related terpenes and flavonoids
o Silybum marianum (milk thistle) – 80% Silymarin
o Coriolus versocolor – 15% polysaccharides
o Viscum album (Mistletoe)
o Ginkgo biloba – 27% Flavones and 7% terpenes
o Rabdosia rubescens (Rabdosia) – 12:1
o Ocimum spp. (Basil) Ursolic acid.
· Natural Compounds that Reduce Bcl-2
o Curcumin
o EGCG and Theophylline - Green tea
o Baicalin and baicalein - Scutellaria baicalensis
o Hibiscus protocatechuic acid (PCA) - Hibiscus s.
o Carnosol - Rosemary
o Gingerol – Ginger
o Echinocystic acid )EA) – Panax ginseng
o OPCs – grape seed extract
o Parthenolide – feverfew
o Andrographolide – Andrographis, diterpenoid lactone
o Beta-lapachone – Lapacho
o EPA – from fish oil
o Chelidonium alkaloids – Chelidonine
o Casticin – Yarrow (Achillea millefolium)
· Raising Vitamin D levels may prevent up to half of all breast and two thirds of colorectal cancer cases in the US. The investigators recommend a daily intake of 2000 IU of Vit. D3 and when possible moderate sun exposure.
· Known activators of PTEN and/or inhibitors of PTEN mutation include: honokiol, from Magnolia grandifloria/officinalis, quercetin, resveratrol, sulforaphane and isoflavones (red clover).
· A diet rich in fruits and vegetables, protects a woman from the BRCA gene becoming activated.
· Astragalus suppressed Basal-like breast cancer: down-regulating EGF and p53.
· Both caloric restriction and glucose / insulin control may play specific beneficial functions in triple negative disease via the newfound contribution of mTOR inhibition, and add another item of defense to the growing arsenal deployable against triple negative breast carcinoma.
· mTOR - The natural agent curcumin's anticancer activity appears to operate primarily by blocking mTOR-mediated signaling pathways in the tumor cells, also inducing apoptosis and inhibiting the basal or type I insulin-like growth factor-induced motility of the cells, also inhibiting at high concentrations the phosphorylation of Akt in tumor cells.
· Curcumin and genistein inhibit EGF.
· Olive oil has been shown to suppress Her-2/neu in Her-2/neu positive breast cancer.
· Ginkgo suppresses breast cancer: Terpenoid constituents, ginkgolikde B, inhibited the proliferation of a highly aggressive human breast cancer. (Anticancer Res. 2006 Jan-Feb;26(IA):9-22)
· DIM can induce apoptosis in breast cancer cells independent of estrogen receptor status by a process that is mediated by the modulated expression of the Bax/Bcl-2 family of apoptotic regulatory factors, and NF-κB pathways.
· Silymarin exerts exceptional anticarcinogenic effects against breast cancer through an apparent down-regulatory effect on certain breast cancer promoting enzymes, namely cyclin-dependent kinases (CDKs) in the G1 phase of the cell cycle.
· Oral administration of large doses of Calcium-D-glucarate have been shown to lower serum estrogen levels in animals by 23% and breast cancer by 70%.
· In addition to the antimutagenic activity, U. tomentosa extracts and fractions exert a direct antiproliferative activity on MCF7 breast cancer.
· Reishi mushroom powdered extract (RPE) -- suppressed cell adhesion and cell migration of highly invasive breast and prostate cancer cells, suggesting its potency to reduce tumor invasiveness.
· Rabdosia (Rabdosia rubescens Hora) – contains oridonin which effectively inhibited the proliferation of a wide variety of cancer cells including those from prostate (LNCaP, DU145, PC3), breast (MCF-7, MDA-MB231), non-small cell lung (NSCL) (NCI-H520, NCI-H460, NCI-H1299) cancers, acute promyelocytic leukemia (NB4) and glioblastoma multiforme (U118, U138).
· Sulforaphane blocked the formation of mammary tumors in rats treated with a potent carcinogen.
· One study showed that sulforaphane can block late stages of the cancer process by disrupting components of the cell called microtubules, and may help to block the growth of breast cancer cell growth.
· Sulforaphane inhibits human mcf-7 mammary cancer cell mitotic progression and tubulin polymerization.
· Chaste tree (Vitex agnus-castus) berry has been shown to reduce prolactin levels in a study of women with hyperprolactinemia. Laboratory Tests and Diagnostic Procedures, p. 907 (2008)
· Artemisinin killed virtually all human breast cancer cells exposed to it within 16 hours.
· Pacific Yew – (Taxus brevifolia) – The yew tree contains a group of unique alkaloids referred to as taxanes that have been shown to exhibit significant antineoplastic actions. Taxanes represent the most important class of antitumor agents introduced in cancer therapy in the last decade. The first member of the family isolated from the yew tree was paclitaxel (taxol). Paclitaxel and docetaxel are two of the most effective chemotherapeutic drugs used today. They are used to treat many types of cancers including ovarian, breast and non-small cell lung cancer.
· Coriolus hirsutus/versicolor – Coriolus versicolor extract (CVE) significantly extended survival at five years or beyond in cancers of the stomach, colon-rectum, esophagus, nasopharynx and lung (non-small cell types, and in HLA B40-positive breast cancer subset.
· Curcumin and EGCG can also interfere with the expression of VEGF by processes other than hypoxia, such as transforming growth factor (TGF)-β release, COX-2 over-expression, hydrogen peroxide release from bone cells, constitutive and aberrant EGFR and Src signaling, and most importantly, by aberrant NF-κB signaling in established cancers.
· Aspirin is known to itself be a potent MEK/ERK inhibitor suggesting a potential role in triple negative disease if further confirmed (as demonstrated early in the research of Zhongyan Wang and Peter Brecher14 at Boston University, Nina Vartiainen15 in Finland, among many others following). In addition, along with aspirin, NSAIDs like ibuprofen, and COX inhibitors are independently of benefit in breast cancer risk reduction 16, 17, 18, 19, a benefit that may be shared by natural COX inhibitory curcuminoid components of curcumin, which is active in the regulation of COX-2, EGFR, VEGF, PI3K/Akt, MEK/ERK, p53, c-Myc, NF-kappaB, Bcl-2, e-cadherin, and apoptotic pathways all known to be critically involved in breast carcinomas in general and in triple negative disease in particular, as well as HER2 (ErbB2) 20 – 36, and some of which are also regulated by the activity of the EGCG (epigallocatechin-3 gallate) component of green tea.
· Flaxseeds – supplemented in the diet of men with prostate cancer reduced both the proliferation rate and in creased apoptosis. Flaxseeds have potent antiestrogenic effects on estrogen receptor positive breast cancer, and recently were found to down-regulate cancer-induced VEGF.
· Folate and B6 – Higher levels of folate and B6 reduce the risk of developing breast cancer.
· Honokiol inhibits in vitro and in vivo growth of breast cancer through induction of apoptosis and cell cycle arrest.
· Silymarin (milk thistle) has also demonstrated a synergistic effect when combined with cisplatin and doxirubin in treatment of ovarian cancer.
· Silymarin exerts exceptional anticarcinogenic effects against breast cancer through an apparent down-regulatory effect on certain breast cancer promoting enzymes, namely cyclin-dependent kinases (CDKs) in the G1 phase of the cell cycle.
· A silybin/phosphatidylcholine combination when used as a single agent against human ovarian cancer inhibited the angiogenic growth factor called VEGF. The silybin agent was significantly active in inhibiting ovarian tumor growth.
· A German study showed that mistletoe-treated patients survived for a mean of 4.23 yeas after inclusion in the study versus 3.05 years for the control group. A second part of the study done among 17 pairs of patients with breast cancer with axillary metastases showed a mean survival time increase from 2.41 years in the control group to 4.79 years in the mistletoe-treated group.
· Two new bioactive mono-tetrahydrofuran (THF) gamma-lactone acetogenins, asitrilobins C (1) and D (2), were isolated from the seeds of paw paw. Compounds 1 and 2 showed selective cytotoxicity comparable with adriamycin for the breast carcinoma (MCF-7) and the colon adenocarcinoma (HT-29) cell lines.
· Cancer occurs when the growth and differentiation of cells in a body tissue become uncontrolled and deranged. While no two cancers are genetically identical (even in the same tissue type), there are relatively few ways in which normal cell growth can go wrong. One of these is to make a gene that stimulates cell growth hyperactivity; this altered gene is known as an 'oncogene'.
· Ras is one such oncogene product that is found on chromosome 11. It is found in normal cells, where it helps to relay signals by acting as a switch. When receptors on the cell surface are stimulated (by a hormone, for example), Ras is switched on and transduces signals that tell the cell to grow. If the cell-surface receptor is not stimulated, Ras is not activated and so the pathway that results in cell growth is not initiated. In about 30% of human cancers, Ras is mutated so that it is permanently switched on, telling the cell to grow regardless of whether receptors on the cell surface are activated or not.
· Usually, a single oncogene is not enough to turn a normal cell into a cancer cell, and many mutations in a number of different genes may be required to make a cell cancerous. To help unravel the intricate network of events that lead to cancer, mice are being used to model the human disease, which will further our understanding and help to identify possible targets for new drugs and therapies.
· Transforming Growth Factor Beta (TGF-β) activates a group of transcription factors that cells have evolved to use for the regulation of many different functions depending on the cell type and on what else the cell is sensing and experiencing at the time. Tumor cells lose growth-inhibitor responses but they don’t lose the TGF-β receptors. So the cells are still capable of reacting to TGF-β with multiple responses, except that growth inhibition is no longer one of them. When tumor cells lose the ability to become growth-inhibited by TGF-β but still retain the rest of the signaling pathway they can respond to TGF-β with impunity to advance their tumorigenic behavior and form metastasis. We’ve now begun to realize that TGF-β enhances metastasis by tumor cells. (Joan Massague 2006)
· Metastasis – When tumor cells leave the primary tumor and form a metastasis, they do so with a particular profile of organ distribution. Breast cancer cells, for example, form metastasis in the lungs, bones, brain and liver. We obtain cells from patients with metastatic disease in various organs, inoculate these cells into a mouse, and ask the mouse to basically act as a cell sorter – to separate cells according to the organs that they can target and colonize. . . It allowed us to indentify breast cancer cells that metastasize to lung and bone, and then define the gene-expression signatures that correlate with the particular organ. Then we demonstrated that these genes are not just markers of organ-specific metastasis, but are also mediators of this process. They are the doers. And for this reason they constitute targets for therapy, at least in principle. In a recent paper, we reported that some of the genes that breast cancer cells use for metastatic development in the lungs are indeed already active in the primary tumor in the breast. . . because these genes are not just markers, but mediators of metastasis, we can think in terms of therapy. (Joan Massague 2006)
· Over expression of the immunosuppressive cytokine transforming growth factor beta (TGF-β) stimulates IL-17 and is another strategy that tumors have developed to evade effective immune surveillance. (Cancer Res. 2008 May 15;68(10):3915-23)
· Transforming growth factor-beta and mutant p53 conspire to induce metastasis by antagonizing p63: a (ternary) complex affair. - How and when a tumor acquires metastatic properties remain largely unknown. Recent work has uncovered an intricate new mechanism through which transforming growth factor-beta (TGFbeta) acts in concert with oncogenic Ras to antagonize p63-metastasis protective function. p63 inhibition requires the combined action of Ras-activated mutant p53 and TGFbeta-induced Smads. Mechanistically, it involves the formation of a p63-Smads-mutant p53 ternary complex. Remarkably, just two of the key downstream targets of p63 turn out to be sufficient as a prognostic tool for breast cancer metastasis. Moreover, the molecular mechanism of this inhibition points to novel therapeutic possibilities. (Marine and Berx 2009)
· Conventional chemotherapy, although directed toward certain macromolecules or enzymes, typically does not discriminate effectively between rapidly dividing normal cells (e.g., bone marrow and gastrointestinal tract) and tumor cells, thus leading to several toxic side effects. Tumor responses from cytotoxic chemotherapy are usually partial, brief, and unpredictable. In contrast, targeted therapies interfere with molecular targets that have a role in tumor growth or progression. These targets are usually located in tumor cells, although some like the antiangiogenic agents may target other cells such as endothelial cells. Thus, targeted therapies have a high specificity toward tumor cells, providing a broader therapeutic window with less toxicity. They are also often useful in combination with cytotoxic chemotherapy or radiation to produce additive or synergistic anticancer activity because their toxicity profiles often do not overlap with traditional cytotoxic chemotherapy. Thus, targeted therapies represent a new and promising approach to cancer therapy, one that is already leading to beneficial clinical effects. There are multiple types of targeted therapies available, including monoclonal antibodies, inhibitors of tyrosine kinases, and antisense inhibitors of growth factor receptors. (Arora and Scholar, 2005)
· The alternatives for treatment are constantly expanding. With the use of new effective chemotherapy, hormone therapy, and biological agents and with information regarding more effective ways to integrate systemic therapy, surgery, and radiation therapy, elaborating an appropriate treatment plan is becoming more complex. To offer better treatment with increased efficacy and low toxicity, selecting therapies based on the patient and the clinical and molecular characteristics of the tumor is necessary. After determining the stage, histological grade, and hormone receptor status, the tumor can behave in an unexpected manner, and the prognosis can vary. Other prognostic and predictive factors have been studied in an effort to explain this phenomenon, some of which are more relevant than others: HER-2/neu gene amplification and protein expression, expression of other members of the epithelial growth factor receptor family, S phase fraction, DNA ploidy, p53 gene mutations, cyclin E, p27 dysregulation, the presence of tumor cells in the circulation or bone marrow, and perineural and lymphovascular space invasion. Tamoxifen is considered the standard of care in premenopausal patients. In comparison, the aromatase inhibitor anastrozole has been proven to be superior to tamoxifen in postmenopausal patients with early-stage breast cancer. The adjuvant use of monoclonal antibodies and targeted therapies other than hormone therapy is being studied. Interestingly, some patients have an early recurrence even though they have a tumor with good prognostic features and at a favorable stage. These recurrences have been explained by the existence of certain cellular characteristics at the molecular level that make the tumor cells resistant to therapy. Selection of resistant cell clones of micrometastatic disease has also been proposed as an explanation for these events. (Gonzales-Angulo et al 2007)
· Mastectomy – might be associated with worse prognosis if conducted during the follicular phase of the menstrual cycles for patients with ER-negative tumors.
· PARP-1 is a nuclear enzyme that is involved in repairing DNA damage (called base excision repair), mediating cell death and necrosis, and regulating immune response. PARP activation occurs when cells are damaged in instances such as during chemotherapy and radiotherapy, and also in non-treatment events such as stroke, head trauma and heart ischemia. The goal of targeting PARP is to prevent tumor cells from repairing DNA themselves and developing drug resistance, which may make them more sensitive to cancer therapies.
· PARP-1 inhibitors also are attractive agents based on what seems to be not only few side effects but also a protective effect in normal tissue.
Predictive Tests for Chemotherapeutic Drug Sensitivity
BRCA1, Taxanes and breast cancer: a recent study demonstrated that a low response rate to taxane-based neoadjuvant chemotherapy is associated with BRCA1 mutations.
Class III beta-tubulin
- a predictive marker of taxanes (taxol, taxotere, abraxane)
& Navilbane
EGFR (pathology or elevated blood levels) - Gene polymorphisms active in the EGFR pathway are associated with the sensitivity of colorectal cancer patients to platinum-based chemotherapy, EGFR blocking antibodies and/or small molecule EGFR tyrosine kinase inhibitors.
Excision repair cross-complementing
factor 1(ERCC1) gene transcript - a predictive marker
of platinum-based sensitivity - cells expressing low levels of ERCC1 are
sensitive to platins. High ERCC1 transcripts predict for gemcitibine
synergy with platinum agents, unless ribonucleotide reductase expression
is also high.
Glutathione-S-transferase P1 (GSTP1) -
the variant allele GSTP1 105Val, and patients
possessing a GSTP1*B allele demonstrated notable trends toward inferior response
and survival.
K-RAS – when
positive predicts poor response to EGFR inhibiting drugs
Methylenetetrahydrofolate reductase (MTHFR)
a predictive marker resistance to 5FU, Xeloda, &
Methotrexate—lower levels correspond with better response.
MTHFR – is a predictive marker for resistance to 5FU, Xeloda and Methotrexate – lower levels correspond with better response. MTHFR = methylene tetrahydrofolate reductase. The MTHFR C7677T mutation appears to interact with folate in determining cancer risk, and there may be further interaction with riboflavin status. The effect of this mutation on cancer risk may be site specific in that individuals carrying the TT variant appear to be protected against colorectal cancer. Genetic variation in MTHFR but not TYMS may be useful for predicting toxicity from capecitabine in patients with advanced colorectal cancer.
Ribonucleotide reductase (RRM1)
- Higher levels of RRM1 observed in gemcitabine-resistant
SPARC (secreted protein acidic and rich in cystein), SPARC is thought to play an important role in tissue remodelling, angiogenesis, embryonic development and tumourigenesis. SPARC plays a crucial role in tumour development in breast cancer and as such has a significant bearing on patient prognosis and long-term survival. (Watkins et al 2005) SPARC predicts Abraxane response.
Thymidylate synthase (TS)
- could be used to decide which patients can best benefit from adjuvant
chemotherapy approaches – 5FU, Xeloda, & Methotrexate. Genotyping for the
TS polymorphism may have the potential to identify patients more likely to
respond to 5-FU or Xeloda based chemotherapy
Topoisomerase IIa (TPT2A) - Amplification of the gene TOP2A is associated with an improved breast cancer response to anthracycline and etopiside-based adjuvant chemotherapy
NOTE: Tumors that are p53 and/or Bcl-2 positive demonstrate poor response to anthracycline-based chemotherapy.
Weisenthal Tests –
Chemo agents: Etopiside, Oxiliplatin, Carboplatin, Abraxane, Nevilbine, high dose Tamoxifen
Receptor blockade drugs: Avastin, EGFR targeting drugs – Tarceva, Erbitux, and Iressa, PDGF targeting drugs Sutent and Nexavar.
Glossary of Some Breast Cancer Chemotherapeutic Drugs
· 5-FU (fluorouracil) -
· Abraxane. A newer formulation of paclitaxel. SPARC predicts Abraxane response.
· Adriamycin (Doxorubicin) -
· Anthracyclines drug class - includes doxorubicin (Adriamycin) and epirubicin (Ellence). Anthracycline-based combination regimens are often used to treat early-stage breast cancer, as well as advanced cancer.
· Aredia (pamidronate) - Bisphosphonate drugs, such as pamidronate (Aredia), are important supportive drugs for preventing fractures and reducing pain in people whose cancer has spread to the bones.
· Avastin – (Bevacizumab) is indicated for the treatment of patients who have not received chemotherapy for metastatic HER2-negative breast cancer in combination with paclitaxel. The effectiveness of Avastin in MBC is based on an improvement in progression free survival. There are no data demonstrating an improvement in disease-related symptoms or increased survival with Avastin. Avastin is not indicated for patients with breast cancer that has progressed following anthracycline and taxane chemotherapy administered for metastatic disease. Avastin is a therapeutic antibody that is believed to work by targeting and inhibiting the function of a natural protein called "vascular endothelial growth factor" (VEGF) that stimulates new blood vessel formation, a process known as angiogenesis.
VEGF – vascular endothelial growth factor is one of the most powerful stimulants of tumor angiogenesis. VEGF is a marker of angiogenesis. It is an important blood marker to assess.
o VEGF may be a clinically important marker for persistent disease and is predictive of survival in ovarian cancer patients after first-line chemotherapy.
o VEGF mRNA expression is upregulated by a wide array of oncogenes (including H-ras and K-ras, V-raf, src, PTEN, p53, Wnt, and c-jun, among others) and growth factors (including EGF, TGF-α, TGF-β, insulin-like growth factor-1, and PDGF).
· Bevacizumab - see Avastin
· Camptothecin – targets topoisomerase. (I’m not sure if this is used for breast cancer)
· Capecitabine (Xeloda) and docetaxel (Taxotere). Capecitabine is an oral drug that is chemically related to 5-FU. It is also being studied in combination with many other drugs. In 2007, the FDA approved a new type of drug, ixabepilone (Ixempra), for use in combination with capecitabine in patients with advanced breast cancer that have not responded to other types of chemotherapy.
· Carboplatin (Paraplatin) -
· Chemotherapy Regimens for early-stage breast cancer:
AC (Doxorubicin and cyclophosphamide)
AC followed by T (Doxorubicin and cylophosphamide followed by paclitaxel)
CAF (Cyclophosphamide, doxorubicin, and 5-FU)
CMF (Cyclophosphamide, methotrexate, and 5-FU)
TAC (Docetaxel, doxorubicin, and cyclophosphamide)
· Chemotherapy Regimens: For example, regimens that contain an anthracycline drug (such as doxorubicin) use the letter "A," and regimens that contain a taxane drug (such as docetaxel) use the letter "T." Cyclophosphamide (Cytoxan), fluorouracil (5-FU), and methotrexate (MTX) are standard cancer drugs used in many breast cancer chemotherapy regimens.
· Cytoxan (cyclophosphamide) -
· Eloxatin (oxaliplatin) -
· Epirubicin (Ellence) -
· Erbitux - (cetuximab) for injection is a monoclonal antibody that targets and inhibits epidermal growth factor receptor (EGFr). EGFr is over-expressed in more than 35% of all solid malignant tumors. It is used alone or combination with other therapies for the treatment of colorectal cancer. Erbitux is indicated, in combination with irinotecan, for the treatment of EGFR-expressing, metastatic colorectal cancer in patients who are refractory to irinotecan-based chemotherapy. In addition, it is also approved for use as a single agent in the treatment of patients with EGFR-expressing, metastatic colorectal cancer who are intolerant to irinotecan-based chemotherapy.
· Etoposide-based adjuvant chemotherapy - Toposar and Vepesid - A semisynthetic derivative of podophyllotoxin, a substance extracted from the mandrake root Podophyllum peltatum. Possessing potent antineoplastic properties, etoposide binds to and inhibits topoisomerase II and its function in ligating cleaved DNA molecules, resulting in the accumulation of single- or double-strand DNA breaks, the inhibition of DNA replication and transcription, and apoptotic cell death. Etoposide acts primarily in the G2 and S phases of the cell cycle.
· FOLFOX – 5-FU + Oxaliplatin
· Gemcitabine and paclitaxel. In 2004, the Food and Drug Administration approved the antimetabolite drug gemcitabine (Gemzar) for use in combination with paclitaxel (Taxol) as a first-line treatment option for women with metastatic breast cancer.
· Herceptin – targets HER-2/neu
· Iressa - Gefitinib - Many cells, including cancer cells, have receptors on their surfaces for epidermal growth factor (EGF), a protein that is normally produced by the body and that promotes the growth and multiplication of cells. When EGF attaches to epidermal growth factor receptors (EGFRs), it causes an enzyme called tyrosine kinase to become active within the cells. Tyrosine kinase triggers chemical processes that cause the cells, including cancer cells, to grow, multiply, and spread. Gefitinib attaches to EGFRs and thereby blocks the attachment of EGF and the activation of tyrosine kinase. This mechanism for stopping cancer cells from growing and multiplying is very different from the mechanisms of chemotherapy and hormonal therapy. Gefitinib was approved by the FDA in May of 2003.
· EGFR – epidermal growth factor receptor - participates in cellular signaling pathways. It is expressed in many human tumor types.
· EGFR - There are now several trials exploring the potential of EGFR inhibitors in triple negative disease in the metastatic breast cancer setting and evaluating a combination of EGFR-inhibitors + platinum agent.
· EGFR (serum) and serum HER-2neu are useful predictive and prognostic markers in metastatic breast cancer patients treated with metronomic chemotherapy. (Sandri et al 2007)
· EGFR over-expression in triple negative and basal-like breast carcinoma is now well-established, as is therefore the therapeutic value of EGFR-inhibition.
· Lapatinib (Tykerb) was approved in 2007 for patients who have not been helped by other cancer drugs, including an anthracycline, a taxane, or trastuzumab. Lapatinib is used in combination with capecitabine (Xeloda). Research suggests it may have fewer risks for heart problems than trastuzumab. This targeted therapy drug is approved for the treatment of HER2-positive advanced breast cancer.
· Methotrexate (MTX) is a standard cancer drug used in many breast cancer chemotherapy regimens.
· Navelbine – a chemotherapy drug for the treatment of metastatic breast cancer which is a well tolerated medication with fewer and milder side effects than other chemotherapy drugs.
· Nexavar – Sorafenib – is a targeted therapy. Nexavar is classified as a tyrosine kinase inhibitor, angiogenesis inhibitor and VEGF inhibitor. It is also a PDGF inhibiting drug.
o PDGF - platelet-derived growth factor – Expression of PDGF, and activation (by autophosphorylation) of its receptor (PDGFR), a tyrosine kinase, is associated with the growth of metastasis in several forms of cancer including kidney, breast, prostate, sarcomas, cervical and in head and neck cancer.
o Inhibiting PDGFR phosphorylation may, especially in combination with agents such as Taxane-rich Yew extract, Artemisinin (with a whole plant extract), produce substantial therapeutic effects against bone metastasis.
o Inhibition of PDGF has shown to suppress metastatic cancer.
· Oxiplatin – alkylating agents.
· Paclitaxel – targets tubulin.
· Pamidronate (Aredia) -
· Platinum-based drug class - includes oxaliplatin (Eloxatin) and carboplatin (Paraplatin). These drugs may be used in combination regimens for advanced cancer or for cancers associated with BRCA genes.
· Sutent – Sunitinib - is a targeted therapy and is a receptor protein-tyrosine kinase inhibitor. It inhibits the actions of VEGF and is an angiogenesis inhibitor. It is also a PDGF targeting drug.
o PDGF - platelet-derived growth factor – Expression of PDGF, and activation (by autophosphorylation) of its receptor (PDGFR), a tyrosine kinase, is associated with the growth of metastasis in several forms of cancer including kidney, breast, prostate, sarcomas, cervical and in head and neck cancer.
o Inhibiting PDGFR phosphorylation may, especially in combination with agents such as Taxane-rich Yew extract, Artemisinin (with a whole plant extract), produce substantial therapeutic effects against bone metastasis.
o Inhibition of PDGF has shown to suppress metastatic cancer.
· Tamoxifen – a class of drugs
· Tarceva - (erlotinib) is a small molecule human epidermal growth factor type 1/epidermal growth factor receptor (HER1/EGFR) inhibitor which demonstrated, in a Phase III clinical trial, an increased survival in advanced non-small cell lung cancer (NSCLC) patients. In a Phase III trial, Tarceva has also shown an improvement in overall survival when added to gemcitabine chemotherapy as initial treatment for advanced pancreatic cancer.
· Taxanes drug class - includes paclitaxel (Taxol) and docetaxel (Taxotere). These drugs may be particularly helpful for node-positive breast cancer. A newer formulation of paclitaxel (Abraxane) is used as a secondary treatment for advanced breast cancer.
· Taxol (paclitaxel) -
· Taxotere (docetaxel) -
· Toposar - Etoposide-based adjuvant chemotherapy
· Trastuzumab (Herceptin) was approved in 1998 for treatment of metastatic breast cancer. It is used in adjuvant chemotherapy, along with drugs such as paclitaxel. This targeted therapy drug is approved for the treatment of HER2-positive advanced breast cancer.
· Trastuzumab (Herceptin). Trastuzumab is a monoclonal antibody that targets the HER2 protein on cancer cells. HER2-positive cancers account for 15 - 25% of early-stage breast cancer and are associated with more aggressive disease. Younger women tend to be most affected. In 2006, the Food and Drug Administration approved trastuzumab for treatment of HER2-positive, early-stage breast cancer (cancer confined to the breasts or lymph nodes that has been surgically removed). Trastuzumab is given along with other chemotherapy drugs following lumpectomy or mastectomy. Research indicates that trastuzumab can help prevent cancer recurrence and death among women with early-stage breast cancer, but it increases the risk of heart problems. Trastuzumab can cause heart failure. Women who have heart failure or weak heart muscle (cardiomyopathy) should not use this drug. Women who take trastuzumab need to have regular heart monitoring, especially if they have already have heart problems.
· Trastuzumab — Expression of the phosphatase and tensin homolog (PTEN) protein might help predict which breast cancer patients will be resistant to treatment with trastuzumab (Herceptin, Genentech), according to a new stud published in the August 15, 2009 issue of Cancer Research. The research uses computers to mathematically model biologic pathways that test the efficacy of a therapy. This method, known as systems biology, is relatively new and not yet clinically applicable, but has been hailed as a "great step forward" in the effort to determine which breast cancer patients will benefit from anti-HER2 therapy. Trastuzumab "has benefited thousands of women with HER2-positive breast cancer, but only a third to half of patients treated with this agent respond," said lead author Dana Faratian, MD, a clinical lecturer in pathology at the University of Edinburgh in the United Kingdom, in a press statement. "We need to know which patients will [and which patients] won't respond to treatment, and this research is a step toward realizing that aim."
· Vepesid - Etoposide-based adjuvant chemotherapy
· Xeloda (capecitabine) -
· Zoledronic acid (Zometa) and pamidronate (Aredia). Numerous chemotherapy drugs and drug combinations are being tested in clinical trials. Patients with advanced breast cancer may also receive other types of drug treatments. Bisphosphonate drugs, such as zoledronic acid (Zometa) and pamidronate (Aredia), are important supportive drugs for preventing fractures and reducing pain in people whose cancer has spread to the bones.
· Zometa (Zoledronic acid) is an intravenous bisphosphonate drug that is used to help prevent or delay bone fractures in patients with breast cancer that has spread to the bones. It is an investigational drug. Recent research suggests that the drug may also help reduce the risk for cancer recurrence in patients with early-stage breast cancer. Dosage: 4 mg. 1 x every 6 months.
· Mitotic Index and Cell Division:
Cell population growth occurs as cells pass through interphase and mitosis to complete the cell cycle. Many cells lose the capacity to divide as they mature or divide only rarely. Other cells are capable of rapid cell division. For example, as plant roots grow, cells near the tip of the root, in the apical meristem, divide rapidly to push the root through the soil. The root cap detects the pull of gravity and directs the rapid growth of cells near the tip.
By quantifying aspects of a dividing cell population, we can examine how cells differ in their capability to divide. Experimentally, we can change properties of the cell's environment and quantify the effects on cell division.
For a group of cells that rarely complete the cell cycle, we expect a high proportion of cells to be in the resting stage of the cell cycle (G1). However, in a rapidly dividing cell population, we expect a high proportion of cells to be in the stage of mitosis. One way to quantify cell division is by using the mitotic index:
The mitotic index can also be used to quantify differences in cell division when an environmental parameter is changed.
By quantifying properties of cell division, we can compare differences in cell growth among neighboring cells and in response to environmental variables. (Darbelley et al Plant Physiological Biochemistry) and (Driss-Ecole et al 1994)
· S-phase fraction: A measure of the percentage of cells in a tumor that are in the phase of the cell cycle during which DNA is synthesized. The S-phase fraction may be used with the proliferative index to give a more complete understanding of how fast a tumor is growing. See mitosis
· The DNA index represents the ratio between the mean relative DNA content of G1 cells in the sample, and the mean relative DNA content of diploid Gi cells determined as reference cells.
· The DNA index (DI), defined as the ratio of the G0/G1 channel number of tumor cells to the G0/G1 channel number of stromal cells, was analyzed in 121 aneuploid colorectal cancers. In patients with aneuploid tumors, a high DI significantly correlated with tumor penetration beyond the proper muscle layer, positive vascular invasion, and the presence of liver metastasis. This observation led us to conclude that a high DI is likely to be associated with hematogenous dissemination in aneuploid colorectal cancer. (Takayuki Yamamoto et al, 1996)
· DNA Ploidy – Is used for determining prognosis in bladder cancer (squamous), breast cancer, hepatocellular carcinoma, laryngeal squamous-cell carcinoma and ovarian cancer. Malignant cells demonstrate greater proliferation than normal cells and ten to have disordered cellular division whereby aneuploid DNA is present in individual cells. This abnormality increases with the degree of malignancy. Clinical studies indicate that the proportion of proliferating cells in a breast tumor biopsy specimen and the degree of aneuploidy have prognostic significance for breast cancer. Laboratory Tests and Diagnostic Procedures, p. 453 ( 2008)
· PARP (poly(ADP-ribose)polymerase) – PARP1 is a nuclear enzyme that is involved in repairing DNA damage (called base excision repair), mediating cell death and necrosis, and regulating immune response. PARP activation occurs when cells are damaged in instances such as during chemotherapy and radiotherapy, and also in non-treatment events such as stroke, head trauma and heart ischemia. The goal of targeting PARP is to prevent tumor cells from repairing DNA themselves and developing drug resistance, which may make them more sensitive to cancer therapies.
· RRM1 - Ribonucleotide reductase M1 - is a key enzyme involved in DNA synthesis, catalyzing the biosynthesis of deoxyribonucleotides from the corresponding ribonucleotides, and data indicate that higher levels of RRM1 are associated with chemoresistance to gemcitabine-based therapies. Up-regulation of RRM1 has been previously observed pre-clinically in different gemcitabine-resistant human cell lines and, clinically, in adenocarcinoma of the colon and, more recently, in NSCLC. (Ceppi et al 2006)
· Tyrosine kinases play a critical role in the modulation of growth factor signaling. Activated forms of these enzymes can cause increases in tumor cell proliferation and growth, induce antiapoptotic effects, and promote angiogenesis and metastasis. In addition to activation by growth factors, protein kinase activation by somatic mutation is a common mechanism of tumor genesis. Because all of these effects are initiated by receptor tyrosine kinase activation, they are key targets for inhibitors. (Arora and Scholar, 2005)
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