Production of Thrombopoietin by Human Carcinomas and Its Novel Isoforms

Abstract

Thrombocytosis is occasionally seen in patients with carcinomas and has been assumed to be attributable to interleukin-6 or granulocyte-macrophage colony-stimulating factor produced by carcinoma cells. In this study, we clarified whether thrombopoietin (TPO) is involved in carcinoma-associated thrombocytosis. Expression of TPO mRNA was observed in the majority of 27 carcinoma cell lines as determined by reverse transcriptase-polymerase chain reaction (RT-PCR). There were 6 PCR products differing in size; sequence analysis showed the full-length TPO mRNA (TPO-1), 12- and 116-bp deleted variants (TPO-2 and TPO-3, respectively), and 3 novel isoforms (197- and 128-bp deleted forms and a 60-bp insert form of TPO-3; named TPO-4, TPO-5, and TPO-6, respectively). Of 27 lines, 24 expressed TPO-1 mRNA with various other isoforms. Culture supernatants of COS-1 cells transfected with TPO-5 or TPO-6 cDNA did not promote the proliferation of TPO-responsive cells, whereas Western blot analysis on the cell lysates demonstrated TPO-5 but not TPO-6 protein, suggesting poor extracellular secretion (TPO-5) or poor protein synthesis (TPO-6). TPO protein was detected in 10-fold concentrated culture supernatants of cells of these carcinoma lines, with a median concentration of 0.38 fmol/mL as evaluated by enzyme-linked immunosorbent assay. High blood TPO levels were observed with a median value of 3.46 fmol/mL (range, 0.34 to 8.67 fmol/mL) in patients with advanced carcinomas associated with thrombocytosis. These results indicate that thrombocytosis in patients with carcinomas might be caused, at least in part, by TPO produced by carcinoma cells.

Lack of pathogenic mutations in the 5′‐untranslated region of the thrombopoietin gene in patients with non‐familial essential thrombocythaemia

“…At least six different human isoforms have been reported in addition to the two produced by alternative promoter usage. Some of them have been widely observed (23, 30, 32): others have been demonstrated in carcinoma cell lines (33). Functional studies of the isoform lacking exon 2 has shown that it can be more efficiently translated, although under normal circumstances it comprises only 2% of liver TPO transcripts (23).…”

Platelets in cancer

“…This risk prediction model, which has been validated in several large cohorts of patients in various clinical settings, provided evidence that thrombocytosis is an important biomarker of cancerassociated thrombosis, indicating a role for platelets in this process (7). Cancer-mediated thrombocytosis may be explained by the ability of several tumor cells to produce and regulate thrombopoietin (TPO), (17,18), a key cytokine which stimulates megakaryocyte formation and platelet production. In a multicenter study involving 619 patients with epithelial ovary cancer, thrombocytosis was found to be associated with high plasma level of TPO and interleukin-6 (IL-6), and linked to an advanced disease and poor survival (17).…”

Involvement of Platelets in Cancers

“…28 Other authors have reported the releasing of TPO by tumor cells in ovarian and hepatocellular cancers. 29,30 The elevation of TPO production promotes increased megakaryocyte growth differentiation and platelet production. [27][28][29][30][31] Tumor Cell-Induced Platelet Activation-Released Factors…”