Heparan-6-O-Endosulfatase 2 Promotes Invasiveness of Head and Neck Squamous Carcinoma Cell Lines in Co-Cultures with Cancer-Associated Fibroblasts

Abstract

Local invasiveness of head and neck squamous cell carcinoma (HNSCC) is a complex phenomenon supported by interaction of the cancer cells with the tumor microenvironment (TME). We and others have shown that cancer-associated fibroblasts (CAFs) are a component of the TME that can promote local invasion in HNSCC and other cancers. Here we report that the secretory enzyme heparan-6-O-endosulfatase 2 (Sulf-2) directly affects the CAF-supported invasion of the HNSCC cell lines SCC35 and Cal33 into Matrigel. The Sulf-2 knockout (KO) cells differ from their wild type counterparts in their spheroid growth and formation, and the Sulf-2-KO leads to decreased invasion in a spheroid co-culture model with the CAF. Next, we investigated whether a fucosylated chondroitin sulfate isolated from the sea cucumber Holothuria floridana (HfFucCS) affects the activity of the Sulf-2 enzyme. Our results show that HfFucCS not only efficiently inhibits the Sulf-2 enzymatic activity but, like the Sulf-2 knockout, inhibits Matrigel invasion of SCC35 and Cal33 cells co-cultured with primary HNSCC CAF. These findings suggest that the heparan-6-O-endosulfatases regulate local invasion and could be therapeutically targeted with the inhibitory activity of a marine glycosaminoglycan.

Galectin-3-binding protein inhibits extracellular heparan 6-O-endosulfatse Sulf-2

“…We recently reported a Sulf-2 dependent spheroid invasion model consisting of a co-culture of SCC35 cancer cells with a primary cancer associated fibroblast (HNCAF37) in Matrigel (Mukherjee et al, 2023). We have shown that Sulf-2 KO SCC35 cells invade Matrigel in this system significantly less than the wild type SCC35 cells.…”

Aspartate β-Hydroxylase Is Upregulated in Head and Neck Squamous Cell Carcinoma and Regulates Invasiveness in Cancer Cell Models

“…We have shown previously that the marine fucosylated chondroitin sulfate HfFucCS, a newly identified inhibitor of heparan 6-O-endosulfatase SULF2, is an inhibitor of HNSCC tumor cell invasion [34]. We treated the SCC35 cells with a combination of the MO-I-1151 and HfFucCS inhibitors to see if they together inhibit the invasion more efficiently (Figure 4A).…”

“…We used a Matrigel-embedded co-culture spheroid model of SCC35 tumor cells with CAF 61137, optimized previously [34], to evaluate the impact of MO-I-1151 on cell invasion (Figure 3A). The CAF-supported tumor cell invasion was recorded on day 5, and our results show that MO-I-1151 reduced the area of the co-culture spheroids by 44% (p < 0.0001) for MO-I-1151 Low and 45% (p < 0.0001) for MO-I-1151 High (Figure 3B) compared to the untreated controls.…”

“…In addition, CAFs effectively deposit and remodel the ECM in the tumor micro-environment (TME) [32], and stromal alpha-SMA is an independent prognostic factor in OSCC patients [33]. In this study, we therefore examine the expression of ASPH in HNSCC tumors, and we use a CAF-cancer cell co-culture spheroid model [34] to evaluate the impact of ASPH inhibition on cancer cell invasion.…”

“…In summary, we examined the expression of ASPH in tissues of 155 HNSCC patients and in cell models of HNSCC invasion. We tested the ability of the MO-I-1151 ASPH inhibitor alone or in combination with a recently described SULF2 inhibitor HfFucCS [34] with regard to the invasion of HNSCC tumor cells in a spheroid co-culture model with a primary HNSCC cancer-associated fibroblast (CAF 61137). Our results show that inhibition of ASPH inhibits invasion of the HNSCC cell line into Matrigel and that the effect is additive with the SULF2 inhibitor HfFucCS.…”