Expression of Purinergic Receptors in Non-melanoma Skin Cancers and Their Functional Roles in A431 Cells

Abstract

We investigated the use of purinergic receptors as a new treatment modality for nonmelanoma skin cancers. Purinergic receptors, which bind adenosine 5'-tri-phosphate, are expressed on human cutaneous keratinocytes. Previous work in rat and human epidermis suggested functional roles for purinergic receptors in the regulation of proliferation, differentiation, and apoptosis. Immunohistochemical analysis of frozen sections in human basal cell carcinomas and squamous cell carcinomas for P2X5, P2X7, P2Y1, P2Y2, and P2Y4 receptors was performed, accompanied by detailed analysis of archive material of tumor subtypes in paraffin sections. Functional studies were performed using a human cutaneous squamous cell carcinoma cell line (A431), where purinergic receptor subtype agonists were applied to cells and changes in cell number were quantified via a colorimetric assay. Immunostaining in paraffin sections was essentially the same as that in frozen sections, although more detail of the subcellular composition was visible. P2X5 and P2Y2 receptors were heavily expressed in basal cell carcinomas and squamous cell carcinomas. P2X7 receptors were expressed in the necrotic center of nodular basal cell carcinomas and in apoptotic cells in superficial multifocal and infiltrative basal cell carcinomas, and squamous cell carcinomas. P2Y1 receptors were only expressed in the stroma surrounding tumors. P2Y4 receptors were found in basal cell carcinomas but not in squamous cell carcinomas. P2X5 receptors appear to be associated with differentiation. The P2X7 receptor agonist benzoylbenzoyl-adenosine 5'-triphosphate and high concentrations of adenosine 5'-triphosphate (1000-5000 microM) caused a significant reduction in A431 cell number (p<0.001), whereas the P2Y2 receptor agonist uridine 5'-triphosphate caused a significant amount of proliferation (p<0.001). We have demonstrated that non-melanoma skin cancers express functional purinergic receptors and that P2X7 receptor agonists significantly reduce cell numbers in vitro.

UV-B induced alteration in purinergic receptors and signaling on HaCaT keratinocytes

“…One minute long, 40 mJ/cm 2 UVB irradiation - concentrations of ATP and BzATP (specific agonist of the P2X 7 receptor) caused a significant decrease in A431 cell number [30]. In our experiments, after irradiation, the P2X 7 receptors were diminished in the cells, and we detected less P2X 7 receptor mRNA.…”

“…Mock treated samples were handled the same way as irradiated ones. Untreated controls cells were taken up at the time of the experiment, while UV-B irradiated and mock treated cells were supplied with 37 °C culture media and were used for experiments at 8,12,16,24,30,36 and 48 h as indicated.…”

P2X7: a growth-promoting receptor—implications for cancer

“…So far, no extensive screening of P2X 7 expression in malignant tumours has been performed; however, different laboratories have reported increased expression of this receptor in prostate [30], breast and skin cancers [31,32], neuroblastoma [9], leukemia [6] and thyroid papillary carcinoma [33]. In most cases, these studies simply report histological evidence of receptor overexpression in tissue slices, with no analysis of functional activity.…”

Targeting of the P2X7 receptor in pancreatic cancer and stellate cells

“…In the cancer field, P2X7R is believed to play multiple roles. First, it can be an anti‐tumour receptor inducing cancer cell death 16, 17. Second, P2X7R can also be a procancer receptor, as it supports cancer cell proliferation, migration and invasion, both in vitro 18, 19 and in vivo 20.…”