SPA‐1 controls the invasion and metastasis of human prostate cancer

Abstract

Recent studies suggest that SIPA1 encoding a Rap GTPase-activating protein SPA-1 is a candidate metastasis efficiency-modifying gene in human breast cancer. In this study, we investigated the expression and function of SPA-1 in human prostate cancer (CaP). Immunohistochemical studies of tumor specimens from CaP patients revealed a positive correlation of SPA-1 expression with disease progression and metastasis. The correlation was recapitulated in human CaP cell lines; LNCaP that rarely showed metastasis in SCID mice expressed an undetectable level of SPA-1, whereas highly metastatic PC3 showed abundant SPA-1 expression. Moreover, SIPA1 transduction in LNCaP caused prominent abdominal lymph node metastasis without affecting primary tumor size, whereas shRNA-mediated SIPA1 knockdown or expression of a dominant-active Rap1 mutant (Rap1V12) in PC3 suppressed metastasis. LNCaP transduced with SPA-1 (LNCaP/SPA-1) showed attenuated adhesion to the precoated extracellular matrices (ECM) including collagens and fibronectin, due to defective ECM-medicated Rap1 activation. In addition, LNCaP/SPA-1 showed a diminished level of nuclear Brd4, which is known to bind SPA-1, resulting in reduced expression of a series of ECM-related genes. These results suggest that SPA-1 plays an important role in controlling metastasis efficiency of human CaP by regulating the expression of and interaction with ECM in the primary sites. (Cancer Sci 2011; 102: 828-836) P rostate cancer (CaP) is the most common male malignancy in the Western world.(1) Data from the Surveillance, Epidemiology, and End Results program of the National Cancer Institute between 1999 and 2006 indicated that 16% of patients presented regional lymph node involvement or distant metastasis at the time of diagnosis. Importantly, those with distant metastasis showed poor prognosis with a 5-year survival rate of 30.2% (http://www.seer.cancer.gov/). Thus, clarification of molecular mechanisms of metastasis (2) is a crucial step towards establishing a new therapeutic strategy.Cancer metastasis consists of a series of complex processes, and multiple genetic factors may be involved in the metastatic efficiency, such as mutations accumulated during primary tumor evolution, genetic features intrinsic to the original tumor cell clones and host genetic backgrounds. (3,4) One of the initial steps of metastasis is invasion and detachment of tumor cells from the primary tissues, in which the interaction of tumor cells with ECM plays an important role. (5,6) In agreement, among numerous metastasis-predictive genes in various tumors, the common molecular signature of metastasis involves those related to ECM interactions. (7)(8)(9) Accumulating evidence indicates an important role of Rap signaling in controlling cell-ECM and cell-cell adhesion, in part via regulation of integrins and cadherins.(10,11) Rap signaling is controlled by multiple regulatory factors, including Rap GDP ⁄ GTP exchange factors and Rap GTPase-activating proteins (GAP), (12) and the deregulation may profoundly af...

p53 expression controls prostate cancer sensitivity to chemotherapy and the MDM2 inhibitor Nutlin-3

“…With a deleted TP53 gene on one allele and a frame-shift mutation on the other producing a severely truncated non-functional p53 protein, the p53-null PC3 line is a highly metastatic prostate cancer cell line (Table 1). 43,53,54 Stably p53WT transfected PC3 cells (PC3/p53WT) were also employed in this set of experiments to investigate p53's signaling (100 nM) or vehicle control (DMSO) along with serial dilutions of doxorubicin. As seen in Figure 4, we were able to increase the sensitivity of LNCaP cells approximately 100-fold to the effects of doxorubicin when p53 activity was WT.…”

Suppression of SIPA-1 expression may reduce bladder cancer invasion and metastasis via the downregulation of E-cadherin and ZO-1

“…The control vector consisted of the following sequence: 5'-GAT CCG TAC AGC GGT CCA ATC ATA GTA GTG CTC CTG GTT GCT ATG ATT GGA CCG CTG TAC TTT TTT AT-3'. SIPA-1 knockdown in T24 cells was achieved by transfecting the cells with a pSINsi-hU6 vector containing short hairpin (sh)RNA targeting SIPA1, or a control vector (16). The sequence of the shRNA for SIPA-1 was as follows: 5'-GAT CCG CTA CTT GCA ACA CCA TTC TTA GTG CTC CTG GTT GAG AAT GGT GTT GCA AGT AGC TTT TTT AT-3'.…”

“…BIU-87 cells were transfected with a pcDNA3.1 vector containing the SIPA gene, or a control vector using opti-MEM (Life Technologies) and Lipofectamine 2000 (Life Technologies), and were selected using G418 reagent (Life Technologies) (16). The control vector consisted of the following sequence: 5'-GAT CCG TAC AGC GGT CCA ATC ATA GTA GTG CTC CTG GTT GCT ATG ATT GGA CCG CTG TAC TTT TTT AT-3'.…”

“…Recent studies have indicated that SIPA-1 is associated with the motility of cancer cells (14)(15)(16). In addition, nuclear SIPA-1 activates the integrin β1 promoter and promotes the invasion of breast cancer cells (15).…”

“…A previous study found that knockdown of SIPA-1 in colorectal cancer cells resulted in reduced cell growth in vitro; however, similar knockdown exhibited a contrasting effect on invasion and migration, which were increased in SIPA1-knockdown cells compared with the control cells (13). Furthermore, SIPA-1 knockdown regulated the invasion and metastasis of human prostate cancer (16). The Ras family includes Ras, Rap and Ral proteins, of which Ras and Ral have been demonstrated to have an association with bladder cancer (17,18).…”