So far, NSCLC is the most thoroughly studied cancer with respect to Sulf-2. Despite the development of new therapies that target specific signaling pathways, the overall prognosis for this disease is poor [65
], and new therapies are being vigorously sought. Is Sulf-2 a worthwhile therapeutic target to consider for this disease? There is very limited Sulf-2 expression in normal lung () [31
]. However, in lung squamous cell carcinoma, Sulf-2 is clearly expressed in tumor cells and in stromal elements surrounding tumor () [31
]. In lung adenocarcinoma, Sulf-2 is strongly induced in tumor stroma. Thus, Sulf-2 satisfies the criterion for a cancer target in that it is selectively expressed in the cancer cells/tumor stroma relative to normal tissue. The minimal phenotypes in Sulf-2 null mice also bolster confidence that off-target effects from Sulf-2 therapeutics may be acceptable. The evidence that Sulf-2 promotes carcinogenesis, rather than just being a correlate of carcinogenesis, is based on the use of NSCLC cell lines [31
]. The xenograft experiments with Sulf-2 knockdown constitute the strongest evidence. However, although widely employed in pre-clinical studies of cancer therapeutics, there are limitations in the ability of xenograft models in nude mice to predict efficacy of targeted therapies in the clinic. These limitations include the artificial nature of tumor cell lines passaged in culture for many generations, the species differences between tumor and stromal cells, the ectopic location of the xenograft, and the lack of an adaptive immune response in the immunodeficient host mouse. Some of these concerns can be overcome by introducing actual fragments of human tumors into nude mice to study the response of the tumors to anti-cancer therapies. Another approach to overcome these limitations would be to exploit murine tumor models driven by expression of oncogenes and/or loss of tumor suppressors. Murine models of NSCLC have been described in which expression of mutated forms of K-ras, Egfr
, or BRaf
in the lung leads to the development of lung adenocarcinomas or pre-malignant lesions [73
]. It will be important to determine whether Sulf-2 is upregulated in these models and if so whether it is expressed in early (adenomas) or late stage (overt carcinoma) tumors. The contribution of Sulf-2 to the development of lung carcinomas could be evaluated by a Sulf-2 inhibitor (e.g., antibody) or by crossing the conditional mouse onto a Sulf-2 null background and determining the impact of the absence of Sulf-2 on tumor initiation and growth.
Sulf-2 is an extracellular enzyme and would therefore offer considerable advantages as a therapeutic target. It is potentially amenable to inhibition by either small molecules or antibodies. Function-perturbing antibodies have not yet been reported for either Sulf. However, a small molecule inhibitor of the Sulfs has already been identified in the form of PI-88. This agent consists of a mixture of chemically sulfated yeast oligosaccharides with a molecular weight range of 1400-3100 daltons [75
]. PI-88 was originally identified as an inhibitor of heparanase, an endoglycosidase of HS chains which is broadly upregulated in human tumors [76
]. PI-88 is a non-cleavable heparin mimetic which inhibits heparanase because it competes with HS native substrates. In addition to inhibiting heparanase, PI-88 can bind to wide range of growth factors (FGF-1, FGF-2, HGF, and VEGF) [75
]. PI-88 has recently been shown to inhibit the enzymatic activity of both Sulfs at comparable potencies found for heparanase [77
]. It inhibits Sulf activity against 4-MUS, heparin, and HSPGs. Progen Pharmaceuticals is currently testing PI-88 in clinical trials for advanced melanoma (phase II) and post-resection liver cancer (phase III)(http://www.progen.com.au/pipeline/
). Clinical trials in lung cancer and prostrate cancer have also been carried out [78
]. A recurring problem with PI-88 is the development of immune-mediated thrombocytopenia in a significant number of recipients [78
]. Because of the broad range of activities of PI-88, it is difficult to interpret either beneficial or adverse effects of the drug. In principle, specific Sulf-2 drugs could be identified by screening for inhibitors of its enzymatic activity. Selectivity of drug candidates could be evaluated by testing for inhibition of heparanase and a range of protein-HS interactions.
Lung cancer is a very heterogeneous disease with multiple genetic and epigenetic alterations at the level of individual tumors [65
]. Dysregulation of signaling through receptor tyrosine kinases (RTKs) is commonly found in NSCLC [79
]. In particular, a number of studies have found a correlation between the activation/overexpression of EGFR, c-Met, and IGF-1R and enhanced risk of development, progression and poorer prognosis of NSCLC. Activation of these pathways results in cell proliferation, differentiation, migration, adhesion, and protection from apoptosis. Genitifib and erlortinib, which are inhibitors of the kinase activity of EGFR, are FDA approved for the treatment of locally advanced or metastatic NSCLC (www.cancer.gov
). Cetuximab, an antibody directed against EGFR, is in clinical trials for patients with advanced non-small-cell lung cancer (www.clinicalresearch.nih.gov
An important future question to address is whether there is cross-talk between Sulf-2 and EFGR. Some EGFR ligands (i.e., HB-EGF and amphiregulin) are heparin-binding [80
], which raises the possibility of Sulf-2 regulation of their interactions with HSPGs. Alternatively, Wnt signaling, which is Sulf-2 dependent in these cells (see above), could transactivate EGFR, as has been demonstrated in breast cancer cell lines [81
]. Interestingly, one of the NSCLC cell lines (H292 cells) whose transformed phenotype is Sulf-2 dependent [31
]is also EFGR dependent [82
]. Cross-talk between Sulf-2 and the other critical RTKs in NSCLC (e.g., c-Met and IGF-1R) also deserves investigation. Strategies for combination treatments of NSCLC, based on inhibition of both Sulf-2 and one of the critical RTKs, could emerge from such investigations.
There is a strong need for diagnostic and prognostic biomarkers for NSCLC. Irrespective of whether Sulf-2 turns out to be a significant driver in this disease, it deserves consideration as a potential biomarker for this cancer and the others in which it is overexpressed. Sulf-2 is secreted and could potentially accumulate in blood or other body fluids. Its enzymatic activity could become the basis for a highly sensitive assay for its detection in fluids or for its in situ localization in tumors.