Prostate cancer invasion is influenced more by expression of a CD44 isoform including variant 9 than by Muc18.
The standard form of cell adhesion glycoprotein CD44 is a metastasis suppressor in prostate cancer. However, we previously showed by RT-PCR and Western blotting that cancer overexpresses unique CD44 variant v7-v10 isoforms. Muc18 is another cell adhesion marker reportedly overexpressed by prostate cancer. Matched frozen section-confirmed tumor and benign tissues were harvested from 10 prostatectomy specimens and tumor was microdissected from two lymph node metastases. Tissues were homogenized for RNA preparations, and RT-PCR was performed for the CD44v7-v10 sequence. In cultured prostate cancer cells, we caused RNA interference against CD44v9 and/or Muc18. We used PC3M cells and a derivative cell line called G(s)alpha, that constitutively expresses this G-protein and is more invasive. Lipofection was performed for a green fluorescent protein plasmid and for two 22-mer DNA fragments, cloned into a plasmid expression vector to generate hairpin, interfering dsRNA. Assays for invasion into Matrigel, a basement membrane matrix, were performed in 4-5 experiments. RT-PCR demonstrated expression of a 608 bp band representing CD44v7-v10 or a 638 bp band of CD44v6-v10 in prostate cancer tissues and metastases but not benign tissue. Cultured G(s)alpha cells overexpressed CD44v9 by comparison with PC3M cells. At 90 h after 6-hour lipofection, protein silencing was evident by Western blots. Silencing the CD44v9 expression reduced invasiveness into Matrigel to 21.6+/-7.0% in PC3M cells (P<0.001) and 31.2+/-18.3% in G(s)alpha cells (P=0.001), compared to cells exposed to transfection vehicle alone. Silencing Muc18 expression reduced invasiveness to 76.9+/-13.5% of the control value in PC3M cells (P<0.05) and 84.8+/-29.9% in G(s)alpha cells (P=0.18). Prostate cancer invasion is facilitated more by its overexpression of CD44 variant 9 than by Muc18. Its relative overexpression by G(s)alpha cells is a novel finding, suggesting a link between signal transduction and cell adhesion marker expression.
Omara-Opyene
Archangel Levi
AL
Department of Pathology, Immunology, and Laboratory Medicine, The University of Florida, Gainesville, FL, USA.
Qiu
Jingxin
J
Shah
Girish V
GV
Iczkowski
Kenneth A
KA
D015703:Antigens, CD; D051929:CD146 Antigen; D045744:Cell Line, Tumor; D006801:Humans; D018960:Hyaluronan Receptors / Q000032:analysis / Q000502:physiology*; D007150:Immunohistochemistry; D008297:Male; D008562:Membrane Glycoproteins / Q000032:analysis / Q000502:physiology*; D009361:Neoplasm Invasiveness; D019006:Neural Cell Adhesion Molecules; D011471:Prostatic Neoplasms / Q000473:pathology*; D034622:RNA Interference; D020133:Reverse Transcriptase Polymerase Chain Reaction
D015703:Antigens, CD; D051929:CD146 Antigen; C117722:CD44v10 antigen; C448906:CD44v9 antigen; D018960:Hyaluronan Receptors; C497902:MCAM protein, human; D008562:Membrane Glycoproteins; D019006:Neural Cell Adhesion Molecules