Glycoengineering in cancer therapeutics: a review with fucose-depleted trastuzumab as the model.

Experimentally modified trastuzumab antibodies show increased cytotoxic potency when used with human effector cells against HER2-overexpressing human breast cancer cells in vitro and ex vivo. Furthermore, the superior efficacy of 'glycoengineered' trastuzumab has been confirmed in vivo utilizing a preclinical xenograft model of human HER2-amplified, trastuzumab-resistant human breast cancer. The increased cytotoxic potency coupled with other improvements are achieved by a seemingly modest change in trastuzumab's structure, that is, depletion of two α-L-fucose residues from trastuzumab's heavy chains. Fucose-free trastuzumab binds with much greater affinity to human natural killer cells. This improved binding induces much greater antibody-dependent cellular cytotoxicity against HER2-overexpressing cells. The pharmaceutical industry has recognized the advantages of fucose-free therapeutic antibodies and has developed technologies that aim to mass produce such antibodies for human use. Here, we summarize data from multiple academic and pharmaceutical laboratories highlighting fucose depletion of antibodies as a key strategy of glycoengineering in cancer therapeutics. We use fucose-depleted trastuzumab as a model to show the advantages of this new class of anticancer agents. We predict that these advantages will translate clinically into improved therapeutics for many patients including those with HER2-overexpressing neoplasms.

The emerging importance of α-L-fucose in human breast cancer: a review.

Breast cancer cells incorporate the simple sugar alpha-L-fucose (fucose) into glycoproteins and glycolipids which, in turn, are expressed as part of the malignant phenotype. We have noted that fucose is not simply a bystander molecule, but, in fact, contributes to many of the fundamental oncologic properties of breast cancer cells. Here, we summarize the evidence from us and others that fucose is necessary for key functions of neoplastic progression including hematogenous metastasis, tumor invasion through extracellular matrices including basement membranes and up-regulation of the Notch signaling system, with implications for epithelial-to-mesenchymal transition and activation of breast cancer stem cells. Additionally, certain breast cancer biomarkers are fucose-rich while a well-known marker of breast cancer progression, soluble E-selectin, is a known counter-receptor of fucosylated selectin ligands. We provide illustrative examples and supportive evidence drawn from work with human breast cancer cell lines in vitro as well as clinical studies with human pathologic material. And finally, we discuss evidence that fucose (or its absence) is central to the mechanisms of action of several experimental targeted therapies which may prove useful in breast cancer treatment. We propose that alpha-L-fucose is essential in order to construct first, the malignant and then the metastatic phenotype of many human breast cancers. This knowledge may inform the search for novel treatment approaches in breast cancer.

Role of MRI in breast cancer management.

In breast cancer, different situations call for different imaging tests. Mammography is the test of choice for screening women with no signs or symptoms of breast cancer. For diagnosis, tailored mammographic views and ultrasonography are the norm. Magnetic resonance imaging (MRI) is highly sensitive for cancer staging, problem-solving, posttreatment surveillance, and other indications. It can detect primary breast cancers and additional foci of cancer that are occult to standard imaging. Continued improvements in technology and studies to assess outcomes will help to better define MRI's role in breast cancer.

Cell surface associated alpha-L-fucose moieties modulate human breast cancer neoplastic progression.

Glycosylation drives critical processes important for mammalian cell-cell and cell-matrix interactions. Alpha-L-fucose (alpha-L-f) is a key monosaccharide component of oligosaccharides that has been found to be overexpressed during tumor progression. Modification of cell surface fucosylation, we hypothesized, alters tumor cell phenotype and function at the end of the neoplastic progression cascade including tumor invasion. Alpha-L-fucosidase (alpha-L-fase) is a glycosidase that specifically removes (alpha-L-f) from oligosaccharide sites. We first verified the effectiveness of the alpha-L-fase to specifically decrease the level of alpha-L-f on the cell surface of several human breast cancer cell lines and also examined the recovery time for these cells to repopulate their surfaces. To investigate the potential effect of defucosylation on tumor functions, we studied the proliferation, and invasion in vitro of human breast cancer MDA-MB-231 cells as the representative cell model. We further examined several fucose-associated molecules previously shown to be involved in tumor progression, including CD44 and CD15 (Lewis X antigen). We found that alpha-L: -fase pretreatment significantly decreased the invasive capability of breast cancer cells. Deoxyfuconojirimycin (DFJ), a specific alpha-L: -fase inhibitor, reversed this effect. After fucosidase treatment, the level of both CD15 and CD44 were found to be reduced as measured by flow cytometry. alpha-L-fase treatment, further, did not affect tumor cell proliferation in vitro under identical experimental conditions. Gelatin zymography of conditioned media from tumor cells treated with alpha-L-fase demonstrated no change in MMP-2 activity while MMP-9 was significantly reduced. In summary, fucose containing glycans were found widely distributed on the cell surface of breast cancer cells and could be effectively removed by alpha-L-fase treatment. This decreased fucosylation, in turn, was seen to impair the interaction between tumor cells and extracellular matrices, and thus affected key cell functions modulating tumor invasion. Further elucidation of the molecular pathways involved in the inhibition of tumor cell invasion may suggest a rationale for the use of glycobiologic therapeutics to deter tumor progression.

Alterations in human breast cancer adhesion-motility in response to changes in cell surface glycoproteins displaying alpha-L-fucose moieties.

Glycosylation of proteins plays multiple roles in cell-cell and cell-matrix interactions. Fucose is a monosaccharide associated with glycosylation events and is known to be over-expressed in many malignant tumors. By using alpha-L-fucosidase (alpha-L-fase), a glycosidase that specifically removes alpha-L-fucose (alpha-L-f), we have examined the potential effects of defucosylation on tumor functions, focusing on tumor progression in the context of the interaction of tumor cells with the extracellular microenvironment. In this submission, we report that alpha-L-fase treatment decreases, in static assays, tumor cell adhesion to a wide variety of ECM components including fibronectin, laminin, collagen I, hyaluronic acid and the complex human biomatrix, HuBiogel(R). By immunofluorescence, co-localization of beta1 integrin and alpha-L-f was found to decrease accordingly. Sialyl Lewis X, an alpha-L-f-containing tetrasaccharide, which modulates the rolling of leukocytes and tumor cells on endothelium, was found to be diminished on human breast cancer cells after alpha-L-fase treatment. Using a dynamic flow chamber system, we were able to determine that defucosylation impaired the rolling of mammary cancer cells on human umbilical vein endothelial cells while significantly increasing their flow speed. Further, the rolling capability of these defucosylated tumor cells was also impaired on purified E and P-selectin matrices. Based on these data, we hypothesize that decreased fucosylation impairs the interaction between tumor cells and their external milieu, which in turn, affects key cell functions modulating tumor progression. Building on our previous studies which demonstrated alpha-L-fase decreased tumor cell invasion while significantly reducing MMP-9 activity, when added to the fact that decreased adhesion on HUVEC occurs in the presence of alpha-L-fase also leads us to propose that defucosylation may modulate metastasis, and thus provides a promising additional glycobiotic target for novel therapies.

How should we screen for breast cancer? Mammography, ultrasonography, MRI.

Of the imaging techniques currently available to evaluate women for breast disease, mammography remains the mainstay of breast cancer screening, but recent guidelines have included magnetic resonance imaging (MRI) for the screening of some women at high risk. Whole-breast ultrasonography for screening has not been established as useful and so should not be offered routinely to patients.

Magnetic resonance imaging as a diagnostic tool for breast cancer in premenopausal women.

BACKGROUND: Screening mammography has led to earlier diagnosis of breast cancer; however, the increased tissue density of young women can complicate mammographic interpretation. We hypothesized that magnetic resonance imaging (MRI) has value in detection of mammographically occult breast cancers, particularly in premenopausal women for whom the sensitivity of mammography is compromised. METHODS: Data were available for 89 women with biopsy-proven breast cancer who had undergone both mammography and breast MRI. Variables evaluated included menopausal status and radiographic findings. Data were analyzed using Fisher's Exact test; P < .05 was considered significant. RESULTS: Of the 89 women in our study, 69 were perimenopausal or postmenopausal and 20 were premenopausal at the time of diagnosis. The malignant lesion was identified on mammography and MRI for a majority of patients. One third of premenopausal women had negative mammography but positive MRI findings. CONCLUSIONS: Our findings support a role for breast MRI in supplementing conventional mammography for early detection of breast cancer in premenopausal women.

Increased ipsilateral whole breast vascularity as measured by contrast-enhanced magnetic resonance imaging in patients with breast cancer.

BACKGROUND: Tumor-induced neovessel formation identified by gadolinium-enhanced magnetic resonance imaging (MRI) is a commonly used marker for breast malignancy. The purpose of this study was to assess possible differences in whole-breast vascularity as measured by contrast-enhanced MRI in the ipsilateral and contralateral breasts of patients with unilateral breast malignancies. METHODS: Gadolinium-enhanced MRI of the breast using a Siemens 1.0-T scanner with dedicated breast coil was performed on 22 consecutive patients with histologically confirmed unilateral breast carcinoma. Whole-breast vascularity of the breast containing the carcinoma was estimated as increased, decreased, or similar compared with the contralateral unaffected breast. Breast vascularity was then correlated to clinical factors including tumor size, histology, multifocality, nodal involvement, and patient age and menopausal status. RESULTS: Twenty patients had infiltrating carcinomas, and 2 patients had ductal carcinoma in situ. Four were multifocal. Fifteen of 22 patients demonstrated clear evidence of increased whole-breast vascularity in the ipsilateral breast containing the primary breast cancer compared with the contralateral breast. Although there was no clear correlation between the presence of increased whole-breast vascularity in the cancer-bearing breast with tumor size, histology, grade, mammographic appearance, or patient age and menopausal status, increased vascularity was present in 3 of 4 patients with multifocal disease and in 4 of 5 patients with metastatic disease in the axillary nodes. CONCLUSIONS: Measurable increases in whole-breast vascularity can be identified by contrast-enhanced MRI and appear with increased frequency in the cancer-bearing breast. These findings suggest that factors other than tumor size and histology may influence development of macroscopic vessels during tumor progression and may be indicative of angiogenic tumor biology.