Using biologic markers to optimize therapies

5-Fluorouracil (5FU) has been the main treatment of choice in colorectal cancer (CRC) in both the advanced and adjuvant settings for the past 40 years, with response rates of only 10% to 20% in the advanced setting. The combination of 5FU with newer therapies such as oxaliplatin and irinotecan has significantly improved response rates to 40% to 50%. Despite these improvements more than half of advanced CRC patients derive no benefit from treatment, which is due to the development of drug resistance. This review highlights the current prognostic and predictive markers that have been identified for CRC to date and the limitations to their usefulness. Furthermore, we discuss novel targeted therapies for endothelial growth factor receptor and vascular endothelial growth factor. Finally, we focus on the development of microarray analysis to identify panels of predictive markers for CRC, which ultimately aim to tailor treatment according to an individual patient and tumor profile.     

A quantitative approach to determining disease response during therapy using multiple biologic markers: application to carcinoma of the breast.

This analytical study was undertaken in an effort to develop a model for a quantitative approach to the evaluation of multiple biological marker levels in blood and urine as a means for determining tumor changes during treatment of patients with malignant disease. The potential biologic markers measured in patients with carcinoma of the breast consist of three urinary polyamines (putrescine, spermidine and spermine), three urinary nucleosides (pseudouridine, N2, N2-dimethylguanosine and 1-methylinosine), and plasma carcinoembryonic antigen (CEA). The distribution patterns of the seven markers measured pretreatment and five weeks after initiating therapy were examined by grouping the patients into the three categories of progression, stable, or regression based on their clinical response to treatment. In addition to the individual marker measurements, the pretreatment and posttreatment values of the ratios of the polyamine levels (spermine/putrescine, spermine/spermidine, and spermidine/putrescine) and the nucleoside levels (N2, N2-dimethylguanosine/pseudouridine, 1-methylinosine/pseudouridine, and 1-methylinosine/N2, N2-dimethylguanosine) were also evaluated. In the pretreatment measurements, CEA levels were elevated for 76% of the patients and the three nucleosides were elevated for 36% of the patients and the three nucleosides were elevated for 36% to 37% of the patients. Urinary spermidine and spermine levels were abnormal for 27% and 24%, respectively, while putrescine levels were elevated for 7% of the patients. When all 14 marker measurements and the 12 ratios of these measurements were considered, the multiple regression equation evaluated the treatment results with a multiple correlation coefficient (R = 0.891; P less than 0.100) about 2.4 times higher than with the most sensitive single marker variable, N2, N2-dimethylguanosine/pseudouridine (R = 0.377; P less than 0.05), alone. Stepwise regression analysis revealed that the minimum number of multiple marker measurements and their ratios required to achieve the maximum value of the multiple correlation coefficient (R = 0.653; p = 0.010) was fifteen. These include the pre and posttreatment measurements of CEA, spermine, N2, N2-dimethylguanosine and 1-methylinosine, as well as two ratios of the polyamines and three ratios of the nucleosides in the post-treatment of the polyamines and three ratios of the nucleosides in the post-treatment measurements. These data suggest that the utilization of regression analysis to evaluate the monitoring utility of multiple marker measurements may be of clinical value.     

New biologic markers in non-Hodgkin's lymphomas

Many of the tumor markers mentioned in this article may seem to be of only research or theoretic interest. However, many of those mentioned are likely to be of great clinical utility in the future. In particular, the strength of these immunohistochemical findings have been compared with standard clinical prognostic features (e.g., stage, age, B symptoms) and have been modeled to show independent prognostic relevance in both intermediate-grade and low-grade lymphomas. Beyond their predictive value, these markers have identified phenotypes that may serve as new therapeutic targets: (1) restoration of HLA loss with alpha and gamma interferons; and (2) restoration of tumor-infiltrating T-cell lymphocytes with interleukin-2. New phenotype-directed cytokine therapies seem compelling with this new knowledge. We await future prospective trials of phenotyping to further refine this new exciting prospect.     

Potential biologic markers in Burkitt's lymphoma.

Specific biochemical molecules used as potential biologic markers, including modified nucleosides, polyamines, and pyrimidine catabolic end-products, were quantitatively measured in the urine of seven patients with Burkitt's lymphoma before, during, and after one or more courses of therapy. The results of this preliminary study demonstrated that patients with this disease frequently excrete significantly increased amounts oof modified nuceleosides (considered to be derived primarily from transfer ribonucleic acid), polyamines, and beta-aminoisobutyric acid during the course of their disease. With successful treatment and rapid destruction of tumor cells, a concomitant rise in these molecules occurs. Elevations were observed prior to chemotherapy and changes in levels associated with treatment or tumor progression appeared to correlate with disease status and to aid in assessing antitumor response. Periodic follow-up analysis of these molecules may be helfful in appraising relapse or recurrence of the malignancy prior to overt evidence of tumor by existing clincial means.     

Precancerous lesions and biologic markers in esophageal cancer.

Neoplastic development in the esophagus is characterized by abnormal activity in the basal cell proliferative compartment. Malignancy starts with mild dysplasia. Barrett's esophagus is associated with columnar epithelial dysplasia and presents increased risk for adenocarcinoma. Esophagitis, especially with chronicity, may constitute a predisposing characteristic in high incidence areas. This paper briefly describes the main features of precancerous lesions in the esophagus. It then focuses on a discussion of the biological markers of malignancy in the oral cavity and esophagus that are currently under study. Such biomarkers include promising differentiation markers such as keratins, involucrin, particulate transglutaminase, as well as growth factors, and most studied but nonspecific onco-developmental markers, e.g., carcinoembryonic antigen, alpha 1-fetaprotein, hormone/enzyme markers, e.g., human chorionic gonadotropin and placental lactogen, and a number of other miscellaneous markers.     

Identification of candidate molecular markers predicting sensitivity in solid tumors to dasatinib: rationale for patient selection

Dasatinib is a multitargeted kinase inhibitor that was recently approved for the treatment of chronic myelogenous leukemia and Philadelphia chromosome-positive acute lymphoblastic leukemia with resistance or intolerance to prior therapy. It is also in clinical trials for treating patients with solid tumors. The identification of molecular markers predictive of response to dasatinib could assist in clinical development by selecting patients most likely to derive clinical benefit. Using baseline gene expression profiling of a panel of 23 breast cancer cell lines, we identified genomic signatures highly correlated with in vitro sensitivity to dasatinib. The ability of these signatures to predict dasatinib sensitivity was further confirmed and validated in independent test cell lines. A six-gene model was used to correctly predict dasatinib sensitivity in 11 out of 12 (92%) additional breast and 19 out of 23 (83%) lung cancer cell lines. Quantitative real-time PCR and immunohistochemical assays further confirmed the differential expression pattern of selected markers. Finally, these gene signatures were observed in a subset of primary breast, lung, and ovarian tumors suggesting potential utility in patient selection. The subset of breast cancer patients expressing the dasatinib-sensitive signature includes a distinct clinical and molecular subgroup: the so-called "triple negative" (i.e., estrogen receptor-negative, progesterone receptor-negative, and HER2-negative) or "basal" breast cancer subtype. This patient population has a poor prognosis and currently has few effective treatment options. Our results implicate that dasatinib may represent a valuable treatment option in this difficult-to-treat population. To test this hypothesis, clinical studies are now under way to determine the activity of dasatinib in these patients.     

Proteomic analysis of nipple aspirate fluid to detect biologic markers of breast cancer

The early detection of breast cancer is the best means to minimise disease-related mortality. Current screening techniques have limited sensitivity and specificity. Breast nipple aspirate fluid can be obtained noninvasively and contains proteins secreted from ductal and lobular epithelia. Nipple aspirate fluid proteins are breast specific and generally more concentrated than corresponding blood levels. Proteomic analysis of 1 microl of diluted nipple aspirate fluid over a 5-40 kDa range from 20 subjects with breast cancer and 13 with nondiseased breasts identified five differentially expressed proteins. The most sensitive and specific proteins were 6500 and 15 940 Da, found in 75-84% of samples from women with cancer but in only 0-9% of samples from normal women. These findings suggest that (1) differential expression of nipple aspirate fluid proteins exists between women with normal and diseased breasts, and (2) analysis of these proteins may predict the presence of breast cancer.     

Role of anastrozole across the breast cancer continuum: from advanced to early disease and prevention

Breast tumorigenesis is a continuum from preinvasive lesions to early breast cancer and advanced disease. In this article the data supporting the use of the aromatase inhibitor anastrozole in postmenopausal women across this continuum are reviewed. In advanced disease, anastrozole has a significant survival benefit and tolerability advantage compared with megestrol acetate when used as second-line treatment. As first-line therapy, anastrozole significantly prolongs time to progression compared with tamoxifen in women with hormone receptor-positive (HR+) disease. In the adjuvant setting, anastrozole has superior efficacy and tolerability compared with tamoxifen in newly diagnosed patients and those who have already received 2-3 years' prior adjuvant tamoxifen. Therefore, anastrozole should be considered a preferred treatment option for postmenopausal women with HR+ early breast cancer. Furthermore, anastrozole has preoperative efficacy in HR+ large or locally advanced tumors. Finally, anastrozole substantially reduces the incidence of contralateral breast cancer compared with tamoxifen in women with HR+ early breast cancer and, therefore, is a potential chemopreventive agent. Anastrozole is thus positioned to become the standard care for postmenopausal women with HR+ disease across the whole breast cancer continuum. Additional data from ongoing studies will further clarify the role of anastrozole across the continuum and answer outstanding questions regarding the optimal timing and duration of treatment.     

Role of ultrasonography in treatment selection

In spite of difficulties impacting objectivity and reproducibility due to its dependence on the technical and diagnostic ability of each examiner, ultrasonography (US) in general can evaluate very well soft tissue lesions, especially small lesions of 1 cm or less in size. US can reliably delineate the nature of several lesions, and can detect dilated mammary ducts and lymph nodes. On the other hand, US may miss ductal spread or lymph node metastasis of breast cancer, especially if the lesions are too minute to be detected by the instrument. US should be used together with X-ray or MR mammography to prevent false negative cases. US is useful for evaluating the local nature and characteristics of cancerous lesions such as size, invasiveness, histological type, intraductal spread etc, and can evaluate regional lymph node and liver metastasis for pre-treatment staging. In addition, US-guided techniques are essential for preoperative pathological diagnosis by FNAC or CNB and for marking the marginal line for partial mastectomy. US should be more actively used to monitor intra-mammary recurrence after breast conserving surgery.     

The biologic basis for the use of retinoids in cancer prevention and treatment.

Retinoids (vitamin A and related molecules) are biologic agents that have demonstrated, in preclinical and clinical models, potent activity in the prevention and treatment of a variety of malignancies. Presented in this article is a review of recent clinical studies and correlative laboratory findings that advance our understanding of the biologic basis for the use of retinoids in cancer prevention and treatment.     

Role of Epstein-Barr virus DNA load monitoring in prevention and early detection of post-transplant lymphoproliferative disease

Posttransplant lymphoproliferative disease (PTLD) is a severe and life-threatening complication after stem cell or solid-organ transplantation, virtually always associated with presence of Epstein-Barr virus (EBV) in the proliferating cells. PTLD is probably caused by the iatrogenically impaired T-cell response allowing outgrowth of EBV-positive B-cells. Quantitative EBV DNA load monitoring is a minimally invasive technique increasingly recognized as a valuable tool in posttransplant patient management. In this review, we focus on the clinical utility of EBV DNA load monitoring in the peripheral blood of transplant recipients using PCR and we discuss the currently most-widely used techniques and their value and limitations in predicting and diagnosing PTLD. Options for EBV DNA load-guided pre-emptive therapy and application of monitoring EBV DNA load dynamics in the prediction of clinical response after therapy are described. Origins of elevated EBV DNA loads in immunosuppressed patients and recent insights in the EBV life cycle in the immuncompromised host are discussed. Finally, a standardization of methodology, clinical specimen type, and cut-off values is proposed. This is essential for comparisons between different institutes and more adequate patient management.     

Application of biological markers to the study of lung cancer causation and prevention

Lung cancer is now the major cause of cancer deaths in the USA and is an increasingly significant cancer worldwide. Biological markers could be used to prevent lung cancer by allowing more timely and precise understanding of the role of environmental factors. So far, biological markers that can serve as carcinogen dosimeters have been investigated in only a small number of pilot studies of populations with current or past exposure to lung carcinogens. We describe several of our collaborative studies involving smokers, various worker populations, lung cancer cases and controls in order to illustrate the advantages and the limitations of 'molecular epidemiology'. The enzyme-linked immunosorbent assay (ELISA) with antibodies to polycyclic aromatic hydrocarbon (PAH)-DNA adducts has been used in conjunction with one or more of the following: physicochemical techniques to monitor carcinogen adducts on haemoglobin, cytogenetic methods to quantify sister chromatid exchange (SCE) and chromosomal aberrations, and Southern and western blot analyses of oncogene activation. Increased levels of markers of biologically effective doses have generally been seen in exposed and high-risk groups when compared to controls. We have also observed significant background levels of such markers and interindividual variation in levels of certain biological markers resulting from exposures to carcinogens. Thus, these methods may be particularly useful in identifying segments of the population that have received a significant effective dose and hence can be considered to be at elevated risk of cancer. Such studies are necessary to validate laboratory methods and lay the groundwork for more definitive molecular epidemiological investigations of lung cancer.     

Role of autophagy in cancer prevention

Macroautophagy (autophagy hereafter) is a catabolic process by which cells degrade intracellular components in lysosomes. This cellular garbage disposal and intracellular recycling system maintains cellular homeostasis by eliminating superfluous or damaged proteins and organelles and invading microbes and by providing substrates for energy generation and biosynthesis in stress. Autophagy thus promotes the health of cells and animals and is critical for the development, differentiation, and maintenance of cell function and for the host defense against pathogens. Deregulation of autophagy is linked to susceptibility to various disorders including degenerative diseases, metabolic syndrome, aging, infectious diseases, and cancer. Autophagic activity emerges as a critical factor in the development and progression of diseases that are associated with increased cancer risk as well as in different stages of cancer. Given that cancer is a complex process and autophagy exerts its effects in multiple ways, the role of autophagy in tumorigenesis is context-dependent. As a cytoprotective survival pathway, autophagy prevents chronic tissue damage that can lead to cancer initiation and progression. In this setting, stimulation or restoration of autophagy may prevent cancer. In contrast, once cancer occurs, many cancer cells upregulate basal autophagy and utilize autophagy to enhance fitness and survive in the hostile tumor microenvironment. These findings revealed the concept that aggressive cancers can be addicted to autophagy for survival. In this setting, autophagy inhibition is a therapeutic strategy for established cancers.     

Prognostic significance of biologic markers in node-negative breast cancer patients: a prospective study

It is generally thought that future advances in the treatment and cure of breast cancer patients will be made possible through a deeper understanding of tumor biology and an improved capability to define the prognosis of each single patient. This will lead to the formulation of new, more selective, and patient-tailored therapies. It is therefore important, when studying potential prognostic factors, to follow methodologic requirements and guidelines which involve the carrying out of prospective studies as confirmatory steps. Repeatedly or recently investigated prognostic markers (tumor size, menopausal status, ER, PgR, ³H thymidine labeling index, c-erbB-2 and p27 expression) were evaluated on a series of 286 prospectively recruited node negative breast cancer patients who underwent loco-regional treatment alone and were closely followed. The individual and relative prognostic contribution of each variable with respect to other factors, as well as their ability to identify node negative patients at risk, were assessed by univariate and multivariate analysis. At a five-year follow-up, only tumor size (p=0.021) and TLI (p=0.016) individually proved to be significant prognostic indicators of relapse-free survival. Conversely, p27 expression was not related to RFS and c-erbB-2 expression appeared to have only a short-term effect on patient prognosis. TLI and tumor size, tested in multivariate analysis along with ER and menopausal status, maintained their independent prognostic relevance. The study, performed on a large series of node-negative patients given loco-regional treatment alone, for the first time prospectively recruited, showed the prognostic relevance of TLI and its independence from other clinico-pathologic and biologic factors over a five-year period.