The 5-HT(2B) receptor plays a key regulatory role in both neuroendocrine tumor cell proliferation and the modulation of the fibroblast component of the neoplastic microenvironment.
BACKGROUND
Fibrosis is a cardinal feature of small intestinal neuroendocrine tumors (SI-NETs) both in local peritumoral tissue and systemic sites (cardiac). 5-HT, a commonly secreted NET amine, is a known inducer of fibrosis, although the mechanistic basis for it and growth factors regulating fibrosis and proliferation in the tumor microenvironment are unclear. We hypothesized that targeting 5-HT(2B) receptors on tumor cells would inhibit SI-NET 5-HT release and, thereby, fibroblast activation in the tumor microenvironment.


METHODS
We studied the 5-HT(2B) receptor antagonist PRX-08066 in NET cell lines (KRJ-I, H720) and in the coculture system (KRJ-I cells: fibroblastic HEK293 cells) using real time polymerase chain reaction, ELISA, Ki67 immunostaining, and flow cytometry-based caspase 3 assays to assess antiproliferative and profibrotic signaling pathways.


RESULTS
In the 5-HT(2B) expressing SI-NET cell line, KRJ-I, PRX-08066 inhibited proliferation (IC(50) 4.6 x 10(-9)M) and 5-HT secretion (6.9 x 10(-9)M) and decreased ERK1/2 phosphorylation and profibrotic growth factor synthesis and secretion (transforming growth factor beta 1 [TGFbeta1], connective tissue growth factor [CTGF] and fibroblast growth factor [FGF2]). In the KRJ-I:HEK293 coculture system, PRX-08066 significantly decreased 5-HT release (>60%), Ki67 (transcript and immunostaining: 20%-80%), TGFbeta1, CTGF, and FGF2 transcription (20%-50%) in the KRJ-I cell line. 5-HT itself stimulated HEK293 proliferation (25%) and synthesis of TGFbeta1, CTGF and FGF2. PRX-08066 inhibition of KRJ-I function reversed these effects in the coculture system.


CONCLUSIONS
Targeting the 5-HT(2B) receptor may be an effective antiproliferative and antifibrotic strategy for SI-NETs because it inhibits tumor microenvironment fibroblasts as well as NET cells. Fibrosis and proliferation appear to be biologically interfaced neuroendocrine neoplasia domains.
Svejda
Bernhard
B
Institute of Pathophysiology and Immunology, Centre for Molecular Medicine, Graz, Austria.
Kidd
Mark
M
Giovinazzo
Francesco
F
Eltawil
Karim
K
Gustafsson
Bjorn I
BI
Pfragner
Roswitha
R
Modlin
Irvin M
IM
D053148:Caspase 3 / Q000378:metabolism; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D018920:Coculture Techniques; D005347:Fibroblasts / Q000187:drug effects* / Q000473:pathology; D006801:Humans; D036341:Intercellular Signaling Peptides and Proteins / Q000378:metabolism; D019394:Ki-67 Antigen / Q000378:metabolism; D018358:Neuroendocrine Tumors / Q000378:metabolism* / Q000473:pathology*; D044405:Receptor, Serotonin, 5-HT2C / Q000378:metabolism*; D012701:Serotonin / Q000378:metabolism; D058830:Serotonin 5-HT2 Receptor Antagonists; D015398:Signal Transduction / Q000187:drug effects
D036341:Intercellular Signaling Peptides and Proteins; D019394:Ki-67 Antigen; D044405:Receptor, Serotonin, 5-HT2C; D058830:Serotonin 5-HT2 Receptor Antagonists; D012701:Serotonin; D053148:Caspase 3