3732

CD82

CD82 molecule;CD82;CD82 molecule

C33 antigen|CD82 antigen|inducible membrane protein R2|kangai 1 (suppression of tumorigenicity 6, prostate; CD82 antigen (R2 leukocyte antigen, antigen detected by monoclonal and antibody IA4))|metastasis suppressor Kangai-1|tetraspanin-27|tspan-27

11p11.2

protein-coding

BACKGROUND: The transgenic adenocarcinoma of the mouse prostate (TRAMP) model closely mimics PC-progression as it occurs in humans. However, the timing of disease incidence and progression (especially late stage) makes it logistically difficult to conduct experiments synchronously and economically. The development and characterization of androgen depletion independent (ADI) TRAMP sublines are reported. METHODS: Sublines were derived from androgen-sensitive TRAMP-C1 and TRAMP-C2 cell lines by androgen deprivation in vitro and in vivo. Epithelial origin (cytokeratin) and expression of late stage biomarkers (E-cadherin and KAI-1) were evaluated using immunohistochemistry. Androgen receptor (AR) status was assessed through quantitative real time PCR, Western blotting, and immunohistochemistry. Coexpression of AR and E-cadherin was also evaluated. Clonogenicity and invasive potential were measured by soft agar and matrigel invasion assays. Proliferation/survival of sublines in response to androgen was assessed by WST-1 assay. In vivo growth of subcutaneous tumors was assessed in castrated and sham-castrated C57BL/6 mice. RESULTS: The sublines were epithelial and displayed ADI in vitro and in vivo. Compared to the parental lines, these showed (1) significantly faster growth rates in vitro and in vivo independent of androgen depletion, (2) greater tumorigenic, and invasive potential in vitro. All showed substantial downregulation in expression levels of tumor suppressor, E-cadherin, and metastatis suppressor, KAI-1. Interestingly, the percentage of cells expressing AR with downregulated E-cadherin was higher in ADI cells, suggesting a possible interaction between the two pathways. CONCLUSIONS: The TRAMP model now encompasses ADI sublines potentially representing different phenotypes with increased tumorigenicity and invasiveness.

KAI1 is a metastasis suppressor gene located on human chromosome 11p11.2. It belongs to a structurally distinct family of cell surface glycoproteins. Decreased KAI1 expression has been observed in several common solid epithelial tumors, including prostatic, pancreatic, lung, hepatic, colorectal, ovarian, and esophageal cancers. A recent study also observed frequent loss of KAI1 expression in a number of squamous cell carcinomas of the cervix by immunohistochemical technique. To further confirm whether this gene is altered in this malignancy, we analyzed KAI1 expression in various stages of cervical carcinoma by a molecular method. Total cellular RNA was extracted from 84 primary invasive cervical carcinomas and 6 metastatic or recurrent lesions. cDNA was synthesized and was used for real-time quantitative polymerase chain reaction analysis. The level of KAI1 expression was obtained as the value of threshold cycle (Ct) and was quantitated with a comparative Ct method. In addition, paraffin blocks of the tumors were selected and prepared for immunohistochemical study with an anti-KAI1 polyclonal antibody, C-16. Both the real-time quantitative polymerase chain reaction method and immunohistochemical study revealed a frequent decrease in KAI1 expression in invasive cervical cancers and metastatic or recurrent lesions. However, the reduction in KAI1 was not related to progression of the disease. When tumor cell differentiation was analyzed, poorly differentiated tumors showed a greater decrease in KAI1 expression than well or moderately differentiated tumors (P < 0.001). Histologically, KAI1 loss was observed equally in both squamous cell carcinoma and adeno-/adenosquamous carcinoma. Since down-regulation of KAI1 occurs in both early and late stages of cervical cancer, we suggest that its involvement in the progression of this malignancy is an early event.

The molecular mechanisms responsible for metastasis are not fully understood. Recently, expression of the KAI1 gene on human chromosome 11p11.2 was found to be down-regulated in metastatic prostate cancer cell lines compared with normal human prostate, suggesting that KAI1 may be a metastasis suppressor gene. The aim of this study was to investigate whether there is reduced expression of KAI1 in late-stage bladder cancer. Sixty-six paraffin-embedded bladder tissue sections were analyzed for KAI1 mRNA by in situ hybridization. Nineteen of these were from patients with no histological evidence of bladder cancer, and 47 were from papillary transitional cell carcinomas (TCCs); of these, 16 were highly invasive. KAI1 mRNA was highly expressed in the specimens of normal bladder (11 of 11; 100%), inflammatory bladder (5 of 8; 63%), and noninvasive papillary TCCs of grades 1 and 2 (15 of 24; 63%), compared to grade 3 papillary TCCs (1 of 7; 14%) or invasive TCCs (1 of 16; 6%). The differences in expression between local and invasive disease were statistically significant (P cancer and that KAI1 may represent an invasion/metastasis suppressor gene in bladder cancer.

expression of a newly described inhibitor of tumour metastasis, KAI1, was examined in bladder cancer progression and compared with the expression of p53 and pRb, which are markers of advanced disease. KAI1 mRNA (by in situ hybridization) and protein levels (by immunohistochemistry) were examined in 135 paraffin-embedded bladder tissue sections. Significant decreases in KAI1 mRNA and protein levels were detected between normal and tumour tissue (p<0.001 and p=0.026, respectively), and between non-invasive and invasive tumours (p=0.046 and p<0.001, respectively). Loss of KAI1 protein expression was accompanied by a shift in staining pattern from a uniform distribution to a weaker, membranous or heterogeneous pattern. Normal tissue and low-grade tumours showed little p53 protein staining. High level staining (indicative of mutant p53) was associated with increased grade in non-invasive tumours (p=0.031) but was not significantly higher in invasive tumours. Whilst p53 protein staining increased with malignant progression and KAI1 mRNA expression decreased, there was no significant correlation between the two patterns (p=0.33, adjusted for group, p=0.18) or when only cancer samples were analysed (p=0.065, adjusted for group, p=0.26), even when taking into account overexpression of MDM-2 protein as a pathway for inactivation of p53. There was no correlation between loss of KAI1 mRNA expression and gain of abnormal pRb staining (p=0. 30, or adjusted for tumour samples only, p=0.59). These results suggest that loss of KAI1 expression is associated with invasive bladder cancer, but is not related to mutation of p53 or to loss of normal pRb expression.

We have recently characterized a human bladder cancer cell line T24 and a more aggressive lineage related variant of it, T24T. To gain further insights, we have studied their metastatic ability in an in vivo model system. Results show that T24 forms significantly fewer [4/12 (1/11) mice had metastases with 1-2 lesions/mouse] metastasis in SCID/bg mice than T24T [14/14 (6/6) mice had metastases with a mean of 24-28 lesions/mouse]. To begin exploring the mechanisms underlying this difference, we evaluated the mRNA and protein expression levels of metastasis-suppressor genes, known to be important in the progression of other cancers, in our model of bladder cancer progression. A higher mRNA expression of BRMS1, a metastasis suppressor in breast cancer, was observed in T24 cells. In addition, RhoGDI2 mRNA expression was only observed in T24 when compared to T24T, suggesting that Rho activation might play a significant role in the metastatic cascade. However, a basal level mRNA expression of KISS1, described as metastasis suppressor in melanoma and breast, was observed in both the lines and had slightly higher expression in T24T. No difference of Nm23-H1, KAI1, MKK4/SEK1 and E-Cadherin protein levels were noted between these two lines. In summary, it appears that the T24/T24T paired cell lines constitute a useful model for the study of human bladder cancer metastasis that will allow both the discovery and mechanistic evaluation of genes potentially involved in this process.

The molecular basis for the loss of KAI1 expression in invasive and metastatic tumors and tumor cell lines is not understood. Recently, identification of a sequence with homology to the consensus p53-binding motif in the promoter of the KAI1 metastasis suppressor gene, has led to a proposal that transcriptional regulation by p53 controls expression of KAI1, and that a dramatic down-regulation of KAI1 mRNA levels in invasive tumors and many tumor cell lines, is directly due to loss of p53 function. We have tested this hypothesis by assessing KAI1 mRNA levels in a series of 22 cell lines derived from bladder and prostate cancers, in which we confirmed the p53 gene sequence and characterized the functional status of the endogenous p53 protein. We anticipated that cell lines expressing p53 capable of transactivation should express high levels of KAI1 mRNA compared with cell lines expressing defective p53, or which were p53-null. KAI1 mRNA levels were determined by northern analysis using a full-length KAI1 cDNA probe, and varied widely between cell lines examined. However, there was no association between these levels and p53 status. Furthermore, transfection of representative cell lines with wild-type p53, or exposure to DNA damaging agents, had no effect on KAI1 mRNA levels. Our data suggest that p53 is not a major factor regulating levels of KAI1 mRNA in bladder and prostate cancer cell lines.

OBJECTIVE: To examine the expression of the KAI1 metastasis suppressor gene and to evaluate its relationship with tumor recurrence in primary pTa and pT1 urothelial bladder carcinoma. METHODS: Samples were obtained from 87 patients after transurethral resection (TUR). tumor stage and grade were reviewed in 33 patients with pTa and in 54 patients with pT1, with a mean follow-up time of 47.4 +/- 30.1 months. The KAI1 protein immunohistochemical assay was performed. Prognosis was analyzed using the Kaplan-Meier method and Cox's proportional hazards model. Correlation between KAI1 expression and recurrence according to each clinicopathological factor was comparatively evaluated using the chi-squared test. RESULTS: Decreased expression of KAI1 protein failed to reach statistical significance for stage (P = 0.25) or morphology of tumor stem (P = 0.19), but it was significantly related to tumor size (P = 0.016). The recurrence-free 5-year survival rates of the group with decreased KAI1 expression was 69.7%, which was significantly higher than the 22.2% for the KAI1-positive group (P < 0.0001). In univariate and multivariate analyses, decreased expression of KAI1 protein, stage pT1, tumor size >3 cm and sessile tumors were independent prognosis factors of recurrence. Despite the lower recurrence rate expected by considering only the clinicopathological factors, decreased KAI1 expression was able to identify the group with a high risk of recurrence. CONCLUSIONS: Downregulated KAI1 expression in bladder tumors tends to relate to stage and morphology of the tumor stem and was significantly correlated to tumor size. Decreased expression of KAI1 was associated with the degree of invasiveness and progression of the cancer and was an independent prognostic factor of recurrence in primary pTa and pT1 urothelial bladder carcinoma.

KITENIN is a newly identified binding partner of the KAI1/CD82 metastasis suppressor. Recent studies using a mouse model of colon cancer, have suggested that KITENIN might be a metastasis enhancer whose functions are modulated by an interaction with KAI1/CD82. To begin exploration of the possible importance of KITENIN to human cancer, we examined KITENIN mRNA (by RT-PCR) and protein expression (by Western blotting) in a large series of bladder cancer cell lines, and then compared these levels to the expression of KAI1/CD82 and of previously determined in vitro invasive behaviour of these same cancer cell lines. We report that KITENIN was uniformly expressed in all cancer cell lines, but those lines in which KAI1/CD82 was not detected, had a higher in vitro invasive ability and altered actin organisation (as determined by fluorescence microscopy), than those lines in which KAI1/CD82 was present. Our data suggest that the relationship between KITENIN and KAI1/CD82 may be an important determinant of tumour cell behaviour.

Levels of the KAI1 metastasis suppressor are reduced in advanced stages of many human cancers, leading to the loss of KAI1 function. A recent study has suggested that the loss of KAI1 function may also occur via alternative splicing of KAI1 mRNA which deletes an exon encoding a critical 28 amino acids from the protein. Using PCR, 20 bladder tumours at differing stage and grade, a non-tumourigenic urothelial cell line (SV-HUC-1) and 17 bladder cancer cell lines were examined for expression of this alternatively spliced (AS) KAI1 mRNA. Full-length KAI1 mRNA was present in all tumour samples and low levels of AS KAI1 mRNA in 15/20 samples. There was no association between the presence or absence of AS mRNA and clinicopathological characteristics of these tumours. Low levels of AS KAI1 mRNA were present in SV-HUC-1 and 14/17 bladder cancer cell lines. There was no association between the presence or absence of AS KAI1 mRNA and tumourigenicity, or in vivo invasive abilities of these cell lines. In all cell lines expressing AS KAI1 mRNA, levels were 3- to 5-fold lower than levels of wild-type mRNA, irrespective of wild-type mRNA levels. Low levels of an alternatively spliced form of KAI1 mRNA are present in most bladder cancer tumours and tumour cell lines, but are not associated with invasive behaviour.

OBJECTIVE: To investigate the relationship between the expression of the cancer metastasis suppressor gene KAI1 and MMP-2 and MMP-9 in human bladder cancer cell lines that express variable levels of KAI1. MATERIALS AND METHODS: Five bladder cancer cell lines (BL-28/0, BL-13/0, BL-17/0/x1, B10, and D2) were grown in standard culture conditions. Gelatinase activities in serum-free conditioned medium were assessed using gelatin zymography. Whole cell lysates were prepared and Western blotting used to detect the protein expression of MMP-9, MMP-2, TIMP-1, TIMP-2, and KAI1. Semiquantitative RT-PCR was performed to analyze the mRNA expression level of MMP-2, MMP-9, TIMP-1, TIMP-2, and KAI1. RESULTS: Western blotting analysis confirmed that KAI1 was expressed in BL-28/0 and Bl-13/0 but not in D2, B10 and BL-17/0/x1 cell lines. This was consistent with in vitro invasion assays reported previously which showed that cell lines lacking KAI1 expression were 2x to 10x more invasive than cell lines that expressed KAI1. MMP-2 protein was detected in BL-28/0, BL-13/0. and BL-17/0/x1 only. Very low levels of MMP-9 were present in BL-28/0, BL-13/0, B10, and BL-17/0/x1 but not D2, whilst very low levels of TIMP-1 were present in all cell lines. No TIMP-2 was detected. Gelatin zymography showed detectable MMP-2 expression in conditioned medium from BL-28/0 and BL-13/0. Very weak MMP-9 was detected in BL-28/0 conditioned medium only. mRNA expression of MMP-2 was only detectable in BL-28/0 and BL-13/0 cell lines. MMP-9 mRNA levels were extremely low in all lines and not detectable in D2 cells. TIMP-1 and TIMP-2 mRNA were detected in all lines. CONCLUSION: We found that KAI1 expression in bladder cancer cell lines is related to a poor invasive potential and expression of latent MMP-2 but not MMP-9. These results are unexpected given other studies showing high levels of MMP-2 and MMP-9 protein expression in patients with invasive bladder cancer. This may reflect differences in the regulation and secretion of MMP-2 and MMP-9 in vitro compared with the in vivo situation, where tumor cells interact with the surrounding environment.

We have recently characterized a human bladder cancer cell line T24 and a more aggressive lineage related variant of it, T24T. To gain further insights, we have studied their metastatic ability in an in vivo model system. Results show that T24 forms significantly fewer [4/12 (1/11) mice had metastases with 1-2 lesions/mouse] metastasis in SCID/bg mice than T24T [14/14 (6/6) mice had metastases with a mean of 24-28 lesions/mouse]. To begin exploring the mechanisms underlying this difference, we evaluated the mRNA and protein expression levels of metastasis-suppressor genes, known to be important in the progression of other cancers, in our model of bladder cancer progression. A higher mRNA expression of BRMS1, a metastasis suppressor in breast cancer, was observed in T24 cells. In addition, RhoGDI2 mRNA expression was only observed in T24 when compared to T24T, suggesting that Rho activation might play a significant role in the metastatic cascade. However, a basal level mRNA expression of KISS1, described as metastasis suppressor in melanoma and breast, was observed in both the lines and had slightly higher expression in T24T. No difference of Nm23-H1, KAI1, MKK4/SEK1 and E-Cadherin protein levels were noted between these two lines. In summary, it appears that the T24/T24T paired cell lines constitute a useful model for the study of human bladder cancer metastasis that will allow both the discovery and mechanistic evaluation of genes potentially involved in this process.

PURPOSE: Brain metastases are an increasingly common complication in breast cancer patients. The metastasis Suppressor Genes (MSG) Nm23, KISS1, KAI1, BRMS1, and Mkk4 have been associated with the metastatic potential of breast cancer in vitro and in vivo. METHODS: The mRNA expression of Nm23, KISS1, KAI1, BRMS1, and Mkk4 in fresh frozen tissue samples of brain metastases from ductal invasive breast cancer specimens was examined in relation to primary tumors. In a first step, mRNA expression screening was carried out using a semi-quantitative RT-PCR approach, in a second step quantitative real-time RT-PCR was performed on selected specimens. By immunohistochemical staining, gene products were visualized on the protein level. RESULTS: Semi-quantitative RT-PCR revealed reduced mRNA expression of Nm23, KISS1, KAI1, BRMS, and Mkk4 in brain metastases. Results for KISS1, KAI1, BRMS, and Mkk4 were confirmed by real-time RT-PCR. In detail, mRNA expression reduction in breast cancer brain metastases was tenfold. expression of MSG could be confirmed by immunohistochemical staining on protein level. CONCLUSIONS: Our investigations revealed significantly reduced mRNA expression of metastases suppressor genes KISS1, KAI1, BRMS1, and Mkk4 in breast cancer brain metastasis. Particularly, in the case of KISS1 and Mkk4, an important role for future treatment of patients with breast cancer brain metastatic lesions can be assumed.

KAI1, also known as CD82, is a candidate metastasis suppressor gene and has been indicated in the disease progression of certain solid tumours, including those of breast cancer. The present study aimed to investigate the importance of KAI1 as a potential metastasis suppressor in breast cancer cells. MDA-MB-231 and MCF-7 sublines with different patterns of KAI1 expression were created by way of anti-KAI1 transgene or transfection of KAI1 expression construct. Cell adhesion was markedly increased in cancer cells showing increased expression of KAI1 (MCF-7(KAI1EXP), p=0.021 vs. control cells), while it was significantly reduced in the KAI1 knockout subline, MDA-MB-231(KAI1KO) (p=0.002 and 0.0004, respectively). Significant increase of cell migration of MCF-7(KAI1EXP) cells (p=0.024 vs. control) and restricted motility of MDA-MB-231(KAI1KO) cells (p=0.003) were observed. Furthermore, MCF-7(KAI1EXP) cells also showed reduced cell invasion (p=0.022), while MDA-MB-231(KAI1KO) cell line showed a significant increase in invasion (p=0.0063 and p=0.007, respectively). KAI1 did not affect cell growth. It is concluded therefore that KAI1 plays an important role in cell adhesion, invasion and migration of breast cancer cells, in vitro, and is a potential metastasis suppressor gene in breast cancer.

Downregulation of KAI1 metastasis suppressor protein is associated with dismal prognosis in a variety of cancers. mutation of p53 was suggested to be involved in KAI1-downregulation. In cervical cancer, p53 is inactivated by human papillomavirus (HPV) oncoprotein E6 with the grade of inactivation depending on the HPV type. KAI1-expression was immunohistochemically determined in 67 specimens of cervical cancer, HPV-typing was performed using polymerase chain reaction (PCR), cloning, and sequencing. KAI1-downregulation was found in 68.1% of patients, HPV-infection in 91%. There was no association of KAI1-downregulation and infection with a particular HPV type. KAI1-downregulation in cervical cancer seems independent of HPV-E6 induced p53 inactivation.

OBJECTIVES: Loss of KAI1 protein is a frequent event in various types of cancer and has been reported to be associated with increased rate of metastasis and shorter survival time. METHODS: expression of KAI1 and p53 was immunohistochemically determined in 75 biopsy specimens from patients with primary irradiated, invasive squamous cell cervical cancer and 30 cases with CIN I-III. RESULTS: In invasive cancer, 22 cases showed strong expression, 42 cases down-regulation, and 11 cases no expression of KAI1. All specimens with CIN I showed strong KAI1 expression, while in CIN II and III down-regulation was observed in 45% of cases. There was no significant correlation of KAI1 expression with clinical and histopathological parameters. No influence of KAI1 expression on prognosis was observed. CONCLUSIONS: KAI1-down-regulation is a frequent event in cervical cancer occurring early in carcinogenesis. KAI1 expression did not influence the prognosis of patients with primary irradiated cervical cancer.

KAI1 is a metastasis suppressor gene located on human chromosome 11p11.2. It belongs to a structurally distinct family of cell surface glycoproteins. Decreased KAI1 expression has been observed in several common solid epithelial tumors, including prostatic, pancreatic, lung, hepatic, colorectal, ovarian, and esophageal cancers. A recent study also observed frequent loss of KAI1 expression in a number of squamous cell carcinomas of the cervix by immunohistochemical technique. To further confirm whether this gene is altered in this malignancy, we analyzed KAI1 expression in various stages of cervical carcinoma by a molecular method. Total cellular RNA was extracted from 84 primary invasive cervical carcinomas and 6 metastatic or recurrent lesions. cDNA was synthesized and was used for real-time quantitative polymerase chain reaction analysis. The level of KAI1 expression was obtained as the value of threshold cycle (Ct) and was quantitated with a comparative Ct method. In addition, paraffin blocks of the tumors were selected and prepared for immunohistochemical study with an anti-KAI1 polyclonal antibody, C-16. Both the real-time quantitative polymerase chain reaction method and immunohistochemical study revealed a frequent decrease in KAI1 expression in invasive cervical cancers and metastatic or recurrent lesions. However, the reduction in KAI1 was not related to progression of the disease. When tumor cell differentiation was analyzed, poorly differentiated tumors showed a greater decrease in KAI1 expression than well or moderately differentiated tumors (P < 0.001). Histologically, KAI1 loss was observed equally in both squamous cell carcinoma and adeno-/adenosquamous carcinoma. Since down-regulation of KAI1 occurs in both early and late stages of cervical cancer, we suggest that its involvement in the progression of this malignancy is an early event.

BACKGROUND & OBJECTIVE: metastasis suppressor gene KAI1/CD82 plays an important role in infiltration and metastasis of several types of human cancers, while the researches on its relation with cervical carcinoma are far from adequate now. Therefore, immunohistochemical techniques were employed in this study to determine the expression of KAI1 gene, and explore its clinical significance in cervical squamous cell carcinoma. METHODS: SP immunohistochemistry was used to detect the expression of KAI1 gene in 99 specimens of cervical squamous cell carcinoma, 25 specimens of cervical intraepithelial neoplasm (CIN) II-III, and 18 specimens of normal cervix. Correlations of expression of KAI1 gene to clinicopathologic factors, and prognosis of cervical squamous cell carcinoma were statistically analyzed. RESULTS: The rates of negative, weak, moderate, and strong expression of KAI1 in cervical squamous cell carcinoma were 52.5% (52/99), 16.2% (16/99), 15.2% (15/99), and 16.2% (16/99), respectively, significantly lower than those in normal cervix, and CIN II-III (P=0.000). expression of KAI1 has no correlation with FIGO stage, age, pelvic lymph node metastasis, tumor histological grade, depth of cervical infiltration, serum squamous cell carcinoma antigen (SCC) level, tumor size, and gross type of cervical lesion (P>0.05). Both univariate and multivariate analyses showed expression of KAI1 has no correlation with prognosis of cervical squamous cell carcinoma (P>0.05). CONCLUSION: KAI1 gene may be an early event in development of cervical cancer, and has no correlation with prognosis of cervical squamous cell carcinoma.

OBJECTIVE: To explore the effect of metastasis suppressor gene KAI1 on the proliferation and invasive ability of cervical cancer cell line CaSki. METHODS: pCMV-KAI1 cDNA plasmid was transferred into cervical carcinoma cell line CaSki by liposome, which had low level of endogenous KAI1 expression. The expressions of KAI1 protein and mRNA were determined by immunohistochemistry and real-time fluorescence quantitative PCR (RT-PCR), the proliferation of KAI1-transfected CaSki cells was investigated by MTT assay and the invasive ability of these cells was evaluated by in vitro invasion assays. RESULTS: After the transfection of pCMV-KAI1 cDNA, the level of KAI1 mRNA and protein expression in CaSki cell were increased (P < 0.05), while the cell proliferation was suppresssed, and the migrative ability of passing through the membrane filte also decreased evidently (P < 0.05). CONCLUSION: The KAI1 metastasis suppressor gene suppressed the ability of proliferation and invasion of cervical cancer cell CaSki in vitro.

KAI1 is a potential metastatic suppressor gene for prostate cancer. We found by Northern blot analysis that six of ten (60%) gastric and colon cancer cell lines exhibited undetectable or very low expression level of KAI1 mRNA. The effects of KAI1 on the adhesion, motility and invasiveness of colon cancer cells was therefore investigated by using two kinds of stable transfectants, i.e., antisense transfectants of BM314 cells whose KAI1 mRNA expression was suppressed by transfer of antisense KAI1 cDNA and sense transfectants of DLD-1 cells with the enhanced KAI1 mRNA by sense cDNA transfer. The following results were obtained: (1) KAI1 gene expression had no significant effect on in vitro cell growth rate of colon cancer BM314 and DLD-1 cells; (2) Cell aggregation assay showed that KAI1 enhanced the Ca++-independent aggregatability of those colon cancer cells; (3) It was revealed by cell motility and invasion assays that KAI1 suppressed both the motility and in vitro invasiveness of those cells and (4) Furthermore, both the binding to fibronectin and the migration on fibronectin-coated plates of those cells were inhibited by KAI1 expression. These suggest that reduced KAI1 gene expression may contribute to the invasiveness and metastatic ability of colon cancer cells.

BACKGROUND: The expression of the KAI1 gene and its gene product were studied in metastatic and non-metastatic human colorectal cancer to evaluate its role in the metastatic process. METHODS: KAI1 mRNA and protein expression was examined in 36 primary colorectal carcinomas and 6 liver metastasis using Northern blot and Western blot analyses. Forty-six normal colonic tissue samples served as controls. The exact site of KAI1 expression was analyzed by in situ hybridization and by immunohistochemistry in primary tumors, in the corresponding normal tissues, in lymph node metastases and liver metastases. RESULTS: Densitometric analysis of Northern blots revealed overexpression of KAI1 mRNA in 87% of colonic cancer tissues in comparison with the corresponding normal colonic tissues. This increase was 9.1-fold in median (P < .001). KAI1 mRNA expression was strongly dependent on tumor stage. Colorectal cancer at stages II and III revealed significantly higher KAI1 mRNA levels than stage IV tumors (P < .03 and P < .015, respectively) or normal controls. In addition, liver metastases showed reduced KAI1 mRNA expression when compared with their corresponding primary tumor. In situ hybridization confirmed the stage-dependent expression results obtained by Northern blots, in which the KAI1 mRNA signal was exhibited almost exclusively in the epithelial cells. Lymph node and liver metastases were largely devoid of KAI1 mRNA. Western blot analysis showed a highly significant increase of KAI1 protein level in stage II cancers in comparison with the normal colon (P < .001) but also in comparison with the more advanced tumor stages III and IV (P < .03) and P < .02, respectively), when metastases were already present. In accordance, KAI1 immunostaining decreased successively with the advance of the tumor stage and was absent in lymph node and liver metastases. CONCLUSIONS: These data demonstrate that the KAI1 mRNA expression and the KAI1 protein level increase in an earlier tumor stage of colorectal cancer, decrease in advanced stages, and are lost in metastases. The loss of KAI1 might favor the ability of colorectal cancer cells to metastasize.

BACKGROUND: The importance of KAI1 to colorectal cancer (CRC) progression is unclear, with conflicting data regarding changes in KAI1 expression during tumour progression. MATERIALS AND METHODS: In situ hybridisation and immunohistochemistry, with a semiquantitative scoring system, were used to assess KAI1 mRNA and protein levels, respectively, in normal colon samples (43), and non-metastatic (24) or metastatic (33) primary cancers, and liver metastases (48). RESULTS: There was significant loss of KAI1 mRNA and protein staining in non-metastatic primary tumours versus normal tissues (Bonferroni, p < 0.05) but levels recovered to near normal in metastatic primary tumours and liver metastases. Increased KAI1 expression significantly correlated with distant metastases (Mann-Whitney, p = 0.0001, mRNA; p = 0.033, protein), cancer-specific survival (Cox regression analysis p = 0.0002, mRNA; 0.0493, protein) and overall patient survival (p = 0.0001, mRNA). Multivariate analysis (Cox proportional hazards model) confirmed high KAI1 mRNA expression was an independent prognostic indicator of distant metastasis (p < 0.0001), cancer-specific survival (p < 0.0001) and overall patient survival (p < 0.0001). CONCLUSION: These data indicate a complex relationship between KAI1 and progression of human CRC, in which expression is reduced in localised primary tumours, but regained in disease associated with metastasis.

BACKGROUND & OBJECTIVE: The study of tumor metastasis suppressor gene KAI1 in colorectal cancer (CRC) is limited nowadays. There are some controversies about the expression of KAI1 and its relationship with pathologic classification of CRC, and up today, the reports of the correlation between KAI1 and the prognosis of CRC still be few. This study was designed to investigate the role of KAI1 in development of CRC and its value in predicting the prognosis of CRC. METHODS: The expression of KAI1 in 91 cases of primary CRC and 25 cases of lymph node metastases were determined using immunohistochemistry. RESULTS: The positive cases of KAI1 in primary CRC were 54 (59.3%), the weak positive cases were 22 (24.2%), the negative cases were 15 (16.5%). KAI1 expression reduced significantly in the cases with low differentiation, lymph node metastasis and distant metastatic tumor (P< 0.01). The positive expression rate of KAI1 in the patients who survived more than 5 years (67.14%) was higher than that in the patients who survived less than 5 years (33.33%) (P< 0.05). The 5-year survival rates of the patients with positive, weak positive, and negative expression of KAI1 decreased in turn (87.04%, 63.34%, 60.00%; P< 0.05). KAI1 expression in lymph node metastases was lower than that in primary locus (P< 0.05). CONCLUSION: The abnormal expression of KAI1 participates in malignant progression of CRC. Detecting the expression of KAI1 probably possesses clinical significance in evaluating the differentiation, lymph node metastasis,and distant metastasis of CRC, and predicting the prognosis of CRC.

AIM: To investigate KAI1 gene expression in the progression of human colonic carcinoma and its clinical significances. METHODS: KAI1 expression was detected by in situ hybridization and immunohistochemistry in the 4 established cell lines of colorectal carcinoma with different metastatic potentials, and in 80 specimens of colonic carcinoma, 21 colonic carcinoma specimens with lymphatic metastasis and 20 controls of normal colonic mucosa. RESULTS: The expressions of KAI1 in HT29 and SW480 cell lines were higher than those in LoVo and SW620. The expression of KAI1 gene was significantly higher in colorectal carcinoma compared with normal colonic mucosa and lymphatic metastasis (chi(2)=46.838, P<0.01). The expression of KAI1 gene had no relationship with histological grade. The KAI1 expressions in Dukes A and B carcinoma were higher at both mRNA and protein levels compared to Dukes C carcinoma (chi(2)=16.061, P<0.05). The expression of KAI1 in colonic carcinoma specimens with lymphatic metastasis was almost lost. The results of in situ hybridization were in concordance with immunohistochemistry. CONCLUSION: KAI1 is highly related to the metastasis of colonic carcinoma and may be a useful indicator of metastasis in colonic carcinoma.

The KAI1 gene is identified as a tumor metastasis suppressor gene in many types of cancer. We examined KAI1 gene and its protein KAI1/CD82 expression by in situ hybridization and immunohistochemical analysis, and found that KAI1 mRNA and protein expression were inversely correlated with lymph node and distant metastasis in digestive tract carcinomas, but not with age and gender of the patient, or with tumor differentiation. Moreover, KAI1/CD82 protein expression positively reflected the survival outcome of patients. Western blot analysis showed that VP-16 increased KAI1/CD82 protein expression obviously in various cancer cell lines, especially in those that were highly metastatic. This increased KAI1/CD82 expression was associated with its translocation from the cytomembrane to the nucleus, in which it interacted with nuclear p53 protein, forming a strong complex, observed by confocal microscopy and co-immunoprecipitation, respectively. In nude mice, after feeding with VP-16, the number of tumors metastasized from spleen to liver was obviously reduced, and KAI1/CD82 protein expression became stronger in those metastatic tumors. Accordingly, this demonstrated that KAI1 might be used as an indicator for predicting the clinical outcome, and VP-16 may be clinically considered as a promising candidate for anti-metastasis with regard to its potential to upregulate KAI1 expression.

PURPOSE: KAI1 is a metastasis suppressor gene located on human chromosome 11p11.2. It is a member of the structurally distinct family of cell surface glycoprotein, transmembrane 4 protein superfamily. KAI1 was initially isolated as a gene that suppressed metastasis of rat prostate tumor cells. Decreased KAI1 expression has been observed recently in various human cancers, including pancreatic, lung, hepatic, colorectal, breast, ovarian, esophageal, and cervical cancers. Frequent down-regulation of the KAI1 protein was also observed in endometrial cancer cell lines. The aim of this study was to determine whether this gene is altered in human endometrial carcinoma. In addition, its prognostic significance in this tumor was also evaluated. EXPERIMENTAL DESIGN: tumor specimens from 18 cases with various degrees of endometrial hyperplasia, 97 primary endometrial carcinomas with various stages, and 28 metastatic lesions of this cancer were examined in this study. Using the method of immunohistochemistry, we characterized the KAI1 protein expression in the 143 endometrial tumors. expression of KAI1 at RNA level was also examined in 35 of the 143 samples using a real-time quantitative PCR method. The data from immunohistochemical analysis were correlated with various clinicopathological factors. RESULTS: High levels of KAI1 protein expression were detected in almost all of the specimens with endometrial hyperplasia (17 of 18). In contrast, loss of KAI1 expression occurred in an increasing frequency (27.8-71.4%) from early stages of primary endometrial carcinomas to metastatic tumors (P < 0.001). In addition, more poorly differentiated tumors demonstrated significantly lower KAI1 expression as compared with the well-differentiated tumors (P < 0.001). It was also found that patients with KAI1-negative tumors had a lower survival rate than those with KAI1-decreased or positive tumors (P = 0.0042 and 0.0286, respectively). However, in multivariate analysis, the prognostic significance of KAI1 expression was inferior to tumor stage. CONCLUSION: These data suggest that KAI1 expression is down-regulated in advanced endometrial cancer. Clinically it may be a useful indicator of the tumor progression and may provide prognostic information on the outcome of this disease.

Kangai (KAI)-1 (CD82) is a metastasis suppressor gene, which belongs to the family of tetraspanin proteins. A loss of KAI-1 expression is associated with the advanced stages of many human malignancies. The present study was designed to investigate the expression pattern of KAI-1 in the normal endometrium and uterine tumors and to correlate it with the expression of tumor suppressor protein p53. KAI-1 could be found in the normal endometrium throughout the menstrual cycle. Thirteen of 42 endometrial carcinomas demonstrated moderate KAI-1 expression, but low expression of p53. Twenty-nine of 42 endometrial carcinomas showed reduced or absent KAI-1 expression, which correlated with strong expression of p53 (p < 0.001). There were significant correlations between KAI-1 expression and histological type, e.g., 93% of endometrioid carcinomas displayed a low or moderate immunostaining for KAI-1, whereas nearly all of the serous/clear cell carcinomas were KAI-1 negative (p < 0.001); tumor grading, e.g., 73% of high grade tumors showed no KAI-1 expression (p < 0.001). Most of the investigated uterine sarcomas were negative for KAI-1, whereas they displayed a strong immunostaining for p53. In conclusion, KAI-1 and p53 show inverse expression. The reduced KAI-1 expression may be the result of dysregulated p53 function and could be an important step in the endometrial carcinogenesis.

BACKGROUND: KAI1/CD82, a tumor metastasis suppressor gene, is correlated inversely with the progression and invasion of several tumors. It also has been reported that the KAI1 gene is related to the tumor suppressor gene p53. This study was performed to clarify the correlation between KAI1/CD82 expression and clinicopathologic characteristics and p53 expression in patients with esophageal squamous cell carcinoma (ESCC). The authors also investigated mutation of the KAI1 gene coding region to determine whether this may reduce KAI1 expression in ESCC. METHODS: Using immunohistochemistry with anti-KAI1 polyclonal antibody and monoclonal antibody against p53, KAI1/CD82 and p53 expression were detected in 55 patients with ESCC who had undergone surgery. The authors examined the KAI1 gene mutation in 22 patients with ESCC by polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP) analysis and DNA sequencing. RESULTS: KAI1/CD82 expression was positive in 36 of 55 patients (65.5%). There was a significant inverse correlation between KAI1/CD82 expression and regional lymph node metastasis (P = 0.0045), distant metastasis (P = 0.0092), the number of lymph node metastases (P = 0.0019), and pathologic stage (P = 0.0046). The survival rates of KAI1/CD82 negative patients were poorer than those of positive patients (P = 0. 024). The correlation between KAI1 positive and p53 positive tumors was not statistically significant. None of the 22 patients with ESCC showed mutation of the KAI1 gene by PCR-SSCP. In one patient, there was polymorphism in the SSCP assay and DNA sequencing. CONCLUSIONS: The authors demonstrated immunohistochemically that the expression of KAI1 protein appeared to be correlated with lymph node metastasis. mutation does not seem to be a mechanism for dysregulation of the KAI1 protein in ESCC.

KAI1 is a metastasis suppressor gene located on human chromosome 11p11.2. It belongs to a structurally distinct family of cell surface glycoproteins. Decreased KAI1 expression has been observed in several common solid epithelial tumors, including prostatic, pancreatic, lung, hepatic, colorectal, ovarian, and esophageal cancers. A recent study also observed frequent loss of KAI1 expression in a number of squamous cell carcinomas of the cervix by immunohistochemical technique. To further confirm whether this gene is altered in this malignancy, we analyzed KAI1 expression in various stages of cervical carcinoma by a molecular method. Total cellular RNA was extracted from 84 primary invasive cervical carcinomas and 6 metastatic or recurrent lesions. cDNA was synthesized and was used for real-time quantitative polymerase chain reaction analysis. The level of KAI1 expression was obtained as the value of threshold cycle (Ct) and was quantitated with a comparative Ct method. In addition, paraffin blocks of the tumors were selected and prepared for immunohistochemical study with an anti-KAI1 polyclonal antibody, C-16. Both the real-time quantitative polymerase chain reaction method and immunohistochemical study revealed a frequent decrease in KAI1 expression in invasive cervical cancers and metastatic or recurrent lesions. However, the reduction in KAI1 was not related to progression of the disease. When tumor cell differentiation was analyzed, poorly differentiated tumors showed a greater decrease in KAI1 expression than well or moderately differentiated tumors (P < 0.001). Histologically, KAI1 loss was observed equally in both squamous cell carcinoma and adeno-/adenosquamous carcinoma. Since down-regulation of KAI1 occurs in both early and late stages of cervical cancer, we suggest that its involvement in the progression of this malignancy is an early event.

OBJECTIVE: To investigate the expression of two tumor metastasis suppressor genes nm23 and KAI1 in gallbladder adenocarcinoma, and their clinicopathological significance. METHODS: Specimens and clinical data from 31 gallbladder adenocarcinoma patients were collected. Histopathological grading and the expression of nm23 and KAI1 were detected by HE and immunohistochemical staining, respectively. All cases were followed up for at least three years. RESULTS: Immunohistochemical staining showed that the positive rate of nm23 and KAI1 proteins was 71.0% (22/31) and 61.3% (19/31), respectively. The positive expression rates of nm23 and KAI1 proteins in the early stage carcinomas were significantly higher than those in the moderate and advanced stage ones (P exact = 0.0051 and P exact = 0.0084), and both had an negative correlation with clinicopathologic stage (P trend = 0.0047 and P trend = 0.0058). There was a significant difference in the expression of nm23 and KAI1 proteins among well, moderately and poorly differentiated carcinomas (P exact = 0.0328 and P exact = 0.0020). The expression of nm23 and KAI1 was positively correlated with histopathological grade (P trend = 0.0086 and P trend = 0.0006). There was also a significant difference in the expression of nm23 and KAI1 proteins between 17 survival and 14 dead patients (P exact = 0.0038 and P exact = 0.0001 ). A synergistic effect of nm23 and KAI1 protein on the survival was observed , and seemed to be more important than any individual gene alone (P exact = 0.0005). CONCLUSION: The expressions of nm23 and KAI1 proteins are negatively correlated with clinical stage, but positively with histopathological grade in gallbladder adenocarcinoma. These two tumor metastasis suppressor genes may act synergistically to inhibit the tumor metastasis.

KAI1 was immunohistochemically detected in the lower part of fundic glands and intestinal metaplasia in non-cancerous gastric tissues, while only 11.0% (8/73) of cancerous tissues were shown to be positive. The relationship between various clinicopathological characteristics of patients with gastric cancer and KAI1 expression in their tumors was then analyzed to reveal that the incidence of positivity was 16.6% (8/48) of patients with stage Ia-IIIa and 0% (0/25) of patients with stage IIIb-IVb (P < 0.05) and that the incidence of peritoneal or hepatic metastasis tended to be lower in KAI1-positive tumors than in negative ones. These results suggest that decreased KAI1 expression may be related to the progression of gastric cancer.

PURPOSE: metastasis remains an incurable common complication in patients with gastric cancer. A variety of theories have been proposed to explain the inefficiency of the metastatic process. To compare protein expression of metastasis-related genes (nm23, KISS1, KAI1 and p53) between primary tumours and metastatic tumours may be useful in illustrating these theories. METHODS: metastasis-related tissue microarrays (including normal tissues, primary tumours, nodal metastases and liver metastases) were constructed. The protein expression of nm23, KISS1, KAI1 and p53 in lymph node and liver metastases from advanced gastric cancer specimens was mainly examined by immunohistochemical staining in relation to primary tumours. RESULTS: Immunohistochemical staining showed reduced protein expression of nm23, KISS1 and KAI1 in lymph node and liver metastases compared with primary tumours. Results for p53 were to the contrary. CONCLUSIONS: Our investigations revealed a tendency of reduced protein expression of metastasis suppressor genes nm23, KISS1 and KAI1 in gastric cancer with the progress of metastasis. This means that the progression theory is an important determinant of metastatic efficiency.

OBJECTIVE: To investigate the mRNA levels of Kiss-1 and KAI-1 metastasis suppressor genes in gastric cancer, and to explore its clinical value. METHODS: In situ hybridization was used on routinely paraffin-embedded sections of resected specimens of 49 cases with gastric cancer and 20 cases with pericancerous tissue. RESULTS: The positive rates and the scores of Kiss-1 and KAI-1 mRNA in gastric cancer tissue were significantly lower than those in pericancerous tissue (P<0.01). The positive rates and scores in normal to mild-atypical hyperplasia cases were significantly higher than those in middle to severe-atypical hyperplasia of pericancerous tissue (P<0.05,P<0.01). The positive rates and scores of Kiss-1 and KAI-1 mRNA in patients with infiltrating depth T1~T2, without lymph node metastasis, with only first group of lymph node metastasis, and without distant organ metastasis were significantly higher than those in patients with infiltrating depth T3~T4, with lymph node metastasis, with second or third group of lymph node metastasis and with distant organ metastasis. A strong positive correlation was found between the expressive scores of Kiss-1 and KAI-1 mRNA in gastric cancer (r=0.53, P<0.01). CONCLUSION: The expression of Kiss-1 and/or KAI-1 mRNA may be important biological markers of reflecting invasive and metastatic potential and prognosis in gastric cancer. The assays of Kiss-1 and/or KAI-1 mRNA expression level in benign lesions of stomach may have important clinical value for the protection and early-stage finding of gastric cancer.

BACKGROUND: The aim of this study was to clarify the clinical significance of TM4SF members CD9, CD63 and CD82 in human gastric carcinoma. METHODS: By employing RT-PCR and immunohistochemistry, we studied the expression of CD9, CD63 and CD82 in 49 paired tissue specimens of normal gastric mucosa and carcinoma. All tissues were obtained from patients who underwent curative surgery. RESULTS: All normal gastric epithelium and gastric ulcer tissues strongly expressed transcripts and proteins of CD9, CD63 and CD82 as compared with corresponding controls. We found a significant correlation between CD63 mRNA level and different pM statuses (P = 0.036). Carcinomas in M0 stage revealed a stronger expression of CD63 than carcinomas in M1 stage. expression of CD9 protein was found significantly stronger in pN0, pM0 than in advanced pN stages (P = 0.03), pM1 (P = 0.013), respectively. We found the relationship between CD63 expression, gender (p = 0.09) and nodal status (p = 0.028), respectively. Additionally, advanced and metastasized tumor tissues revealed significantly down-regulated CD82 protein expression (p = 0.033 and p = 0, respectively), which correlated with the tumor pTNM stage (p = 0.001). CONCLUSION: The reduction of CD9, CD63 and CD82 expression are indicators for the metastatic potential of gastric carcinoma cells. Unlike their expression in other tumor types, the constitutive expression of CD63 may indicate that this factor does play a direct role in human gastric carcinogenesis.

PURPOSE: The similar appearance of renal tumor histological subtypes can complicate differential diagnoses. This problem is most notable for the chromophobe subtype of renal cell carcinoma, which can be histologically indistinguishable from oncocytoma with investigational molecular markers failing to provide reliable differentiation. KAI1 is a metastasis suppressor gene whose expression correlates inversely with the metastatic potential of most solid tumor cancer types. We tested the hypothesis that KAI1 is differentially expressed among renal tumor histological subtypes. MATERIALS AND METHODS: Immunohistochemical staining for KAI1 protein was performed in 152 nephrectomy specimens, including 48 clear cell, 35 papillary and 31 chromophobe renal cell carcinoma samples, 28 oncocytomas and 10 tumor-free kidneys. Staining was scored as none/minimal, low, moderate or high. KAI1 mRNA levels were compared by quantitative reverse transcriptase-polymerase chain reaction in an additional 22 chromophobe renal cell carcinoma and oncocytoma samples. RESULTS: In all 10 tumor-free kidneys KAI1 protein was detected exclusively in distal tubule cell membranes. Of the tumor specimens KAI1 protein was absent in all papillary renal cell carcinoma specimens. It was present in only 1 of 48 clear cell renal cell carcinomas (2%) and 2 of 28 oncocytomas (7%) but only at low levels. In contrast, 27 of 31 chromophobe renal cell carcinoma specimens (87%) expressed KAI1 protein, most at moderate or high levels. The diagnostic accuracy of KAI1 immunostaining for discerning chromophobe renal cell carcinoma from oncocytoma was 90% with similar results observed at the RNA level. CONCLUSIONS: KAI1 is an accurate biomarker for chromophobe renal cell carcinoma that may aid in the diagnostic differentiation of chromophobe renal cell carcinoma from oncocytoma. It remains to be determined whether KAI1 expression contributes to the low metastatic potential of chromophobe renal cell carcinoma.

OBJECTIVE: To investigate the expression of the metastasis suppressor gene KAI1 in laryngeal squamous cell carcinoma and the relationship between expression of KAI1 and it's role in tumorgenesis and development. METHODS: The sections of 84 samples of primary laryngeal squamous cell carcinoma (PLSCC), 27 samples of laryngeal precancerous lesion (LPL), 10 samples of vocal cords polyp (VCP) and 10 samples of normal laryngeal tissues (NLT) were examined by in situ hybridization for KAI1mRNA. RESULTS: It was found that 10 samples of NLT and 10 samples of VCP showed strong expression of KAI1, the Integrated A Total of KIA1 in these two types of tissues were 136,206.8 +/- 36,675.536 and 133,674.505 +/- 42,858.456, respectively, with no statistically significant difference between them, and high prevalence of decreased expression of KAI1 in LPL and PLSCC compared with that in VCP and in NLT (P < 0.01). There was a significant relationship between down-regulation of KAI1 expression with pathological grade, lymph node metastases, T stage and clinical stage (P < 0.01) and did not correlate with gender. CONCLUSION: KAI1 may play an important role on tumor genesis, development, invasion and metastasis of PLSCC, and become one of the molecular markers for early diagnosis and predicting of aggressive and metastasis potential to assess the clinical stage of PLSCC.

OBJECTIVE: To observe the levels of KAI1/CD82 in carcinoma of larynx and the relationship between it and their clinicopathological features,in association with the results of metastasis. METHOD: The expressions of KAI1/CD82 in 61 patients with carcinoma of larynx were detected by immunohistochemistry. The samples of tissue cutting from the edge of carcinoma of larynx (control group) were also studied,and the results were compared. RESULT: The expression of KAI1/CD82 protein in carcinoma of larynx was lower than that in control group (P < 0.05). Statistic difference about the expression of KAI1/CD82 between primary lesions and metastatic lesions was found (P < 0.05); The expression of KAI1/CD82 were closely correlated with lymph node metastasis (P < 0.05), but their expressions were not correlated with age, clinical staging, pathological differentiation (P > 0.05). CONCLUSION: KAI1/CD82 may play an important regulating role in metastasis of carcinoma of larynx, and they may be used as potential markers for tumor metastasis.

OBJECTIVE: This study aimed to investigate the expressions and significance of KAI1/CD82 and cyclin D1 in laryngeal squamous cell carcinoma (LSCC). METHODS: Real-time quantitative PCR (Q-PCR) and Western blot assay were employed to detect the expressions of KAI1/CD82 and cyclin D1 in the laryngeal tissues of 86 LSCC patients, 32 patients with laryngeal polyp and 38 patients with laryngeal leukoplakia, and the influence of both proteins on the clinicopathological features and survival of LSCC patients. RESULTS: The changes in mRNA and protein expressions of KAI1/CD82 and cyclin D1 were consistent in three groups, and the expressions of KAI1/CD82 and cyclin D1 were significantly different among three groups (P<0.01 or <0.05). The KAI1/CD82 expression in patients with TNM stage III-IV LSCC, poorly differentiated LSCC, clinical stage III-IV LSCC or lymph node metastasis was markedly lower than that in those with TNM stage I-II LSCC, well differentiated LSCC, clinical stage I-II LSCC or no lymph node metastasis (P<0.01 or <0.05). However, there was no marked difference in KAI1/CD82 expression between males and females and among patients in different age groups (P>0.05). In LSCC patients positive for KAI1/CD82 protein expression, the median survival time was 76 months, which was significantly longer than that in LSCC patients negative for KAI1/CD82 protein expression (48 months; X(2)=16.293, P=0.000). The Cyclin D1 expression in patients with TNM stage III-IV LSCC, poorly differentiated LSCC, or clinical stage III-IV LSCC was dramatically higher than that in patients with TNM stage I-II LSCC, well differentiated LSCC, or clinical stage I-II LSCC (P<0.01 or <0.05). However, no marked difference was noted in cyclin D1 expression between males and females, among patients in different age groups and between patients with and without lymph node metastasis (P>0.05). In LSCC patients positive for cyclin D1 protein expression, the median survival time was 40 months, which was markedly shorter than that in LSCC patients negative for cyclin D1 protein expression (X(2)=9.517, P=0.02). In LSCC patients, there was a negative correlation between KAI1/CD82 expression and cyclin D1 expression (X(2)=7.86, P<0.01). CONCLUSION: KAI1/CD82 affects cell cycle. Both KAI1/CD82 and cyclin D1 are involved in the occurrence and development of LSCC, and may provide clinical information for evaluation of invasiveness, metastasis and prognosis of LSCC. Thus, KAI1/CD82 and cyclin D1 may serve as markers for determination of invasiveness, metastasis and prognosis of LSCC.

Down-regulation of KAI1 expression has been shown to be associated with formation of metastases or disease progression in prostate and pancreatic cancer. In the present study we analyzed the expression pattern of KAI1 in metastatic and nonmetastatic hepatocellular carcinomas (HCCs) in comparison with normal livers to evaluate whether alteration of KAI1 also facilitates the metastatic ability in this malignancy. Thirty-nine primary HCCs and 10 normal liver tissue samples were studied for KAI1 messenger RNA (mRNA) expression with use of Northern blot analysis and in situ hybridization. By Northern blot analysis, moderate to strong KAI1 mRNA expression was present in normal liver samples. In contrast, KAI1 mRNA expression in tissue samples of primary HCCs was markedly decreased compared with normal controls. The normal/tumor ratio of KAI1 mRNA expression was 2.6:1 (P <.01). Primary HCCs that gave rise to metastasis showed significantly lower KAI1 mRNA levels than nonmetastasized HCCs (P <. 05). As seen by in situ hybridization, moderate to strong cytoplasmic KAI1 mRNA staining was present in almost all normal hepatocytes. Bile ducts, blood vessels, and connective tissue showed no or only faint KAI1 mRNA expression in the normal liver samples. In nonmetastatic HCCs, the cancer cells exhibited in situ hybridization signals that were similar to the normal controls. In contrast, most of the primary HCC cells in samples with metastases showed only faint or moderate KAI1 mRNA expression predominantly in the perinuclear regions. When KAI1 mRNA expression of primary hepatocellular cancer cells was compared with metastasized cancer cells in lymph nodes, with intrahepatic satellite metastasis, or with peritoneal metastasis in the same patients, significantly lower (P <.01) KAI1 mRNA levels were present in the metastasized HCC cells. Reduced KAI1 mRNA in HCC cells seems to influence their metastatic ability and thereby enhances the malignant potential of HCC.

KAI1 gene is a novel metastasis suppressor gene. Its transcription level has been reported to be related with metastases in several kinds of tumors. The Authors used RT-PCR method to investigate the expression levels of KAI1 gene in hepatocellular carcinoma (HCC) tissues from 42 patients. The results demonstrated that 57.1% of tumor tissues were positive in KAI1 expression. There was no statistically significant difference on KAI1 gene expression rate between large (>5 cm) and small (<=5 cm) tumors, encapsulated and non-capsulated tumors, as well as alpha-fetoprotein (AFP) positive (>20 ug/ml) and negative (<=20 ug/ml) tumors; however, the expression rate was higher in tumors without intrahepatic metastasis than in those with intrahepatic metastasis (63.9% vs. 16.7%, P<0.05). It is suggested that KAI1 gene expression may be related to the metastases of hepatocellular carcinoma.

BACKGROUND & OBJECTIVE: tumor metastasis-suppressor gene KiSS-1 is related to the metastasis of malignancies. However, its correlation to the metastasis, especially the formation of portal vein tumor thrombus (PVTT), of hepatocellular carcinoma (HCC) has seldom been reported. This study was to investigate the function of KiSS-1 gene in the formation of PVTT of HCC, and explore the machanism. METHODS: The expression of KiSS-1 and matrix metalloproteinase-9 (MMP-9) in 50 specimens of HCC (31 cases with PVTT and 19 cases without PVTT) and 11 specimens of PVTT was detected by reverse transcription-polymerase chain reaction (RT-PCR) and immunohistochemistry (IHC). RESULTS: The positive rates of KiSS-1 mRNA and protein were significantly lower in PVTT group and HCC with PVTT group than in HCC without PVTT group (18.2% and 16.1 % vs. 63.2%, 0% and 12.9% vs. 47.4%, P < 0.05)u the positive rates of MMP-9 mRNA and protein were significantly higher in PVTT group and HCC with PVTT group than in HCC without PVTT group (72.7% and 77.4% vs. 31.6%, 81.8% and 83.9% vs. 42.1%, P < 0.05). There was no significant difference between PVTT group and HCC with PVTT group (P > 0.05). KiSS-1 expression was negatively related to MMP-9 expression (r=-0.362 for mRNA, and r=-0.473 for protein, each P < 0.05). CONCLUSIONS: KiSS-1 and MMP-9 are closely related to the formation of PVTT. KiSS-1 gene might suppress the formation of PVTT by suppressing MMP-9 synthesis.

BACKGROUND & OBJECTIVE: metastasis suppressor gene KAI1 can inhibit the metastasis potential of many malignant tumors. This study was to explore the effect of KAI1 gene on the adhesion of hepatocellular carcinoma cell line MHCC97-H with high metastatic potential, and to find the possible mechanism. METHODS: The plasmid containing KAI1 gene was transfected into MHCC97-H cells. The growth status of the cells was observed. The changes of adhesion factors, soluble intercellular adhesive molecule-1 (sICAM-1) and E-cadherin, in the cells were detected by ELASA and Western blot. The character of cell adhesion was determined through plate colony formation test and cell adhesion test. RESULTS: The morphology of MHCC97-H cells did not change markedly after transfection of KAI1 gene, but black granules in cytoplasm were increased. Four days after transfection, the expression of sICAM-1 and E-cadherin were decreased by 24.28% and 26.02%. The adhesion rate was decreased by 11.34% at 3 h after transfection, and by 24.00% at 4 h after transfection, but the cloning efficiency did not change much. CONCLUSION: KAI1 gene could affect the growth pattern and proliferation of MHCC97-H cells, suppress sICAM-1 secretion and E-cadherin production, and inhibit adhesion of MHCC97-H cells.

BACKGROUND: Downregulation of KAI1 gene expression has been found in many types of cancer cells and is closely related to cancer invasion and metastasis. This study was aimed at investigating the effects and possible underlying mechanisms of KAI1 gene on invasion and metastasis of human hepatocellular carcinoma (HCC). METHODS: The invasive ability, visco-elastic properties and cell adhesion forces were analysed in different HCC cells originating from the MHCC97-H cell line transfected with either the sense or the antisense KAI1 expression plasmid. Tumuorigenicity, metastatic abilities, extracellular matrix (ECM) and intercellular adhesion molecule-1 (ICAM-1) expression were also evaluated in the nude mouse models of the xenografted and orthotopic liver cancer cells. RESULTS: Compared with their parental cells, in the HCC cells transfected with the sense KAI1 gene, the invasive ability in vitro was significantly decreased (P<0.01); the cellular elastic coefficients K(1), K(2) and mu were significantly higher (P<0.05); the cells adhesion forces to fibronectin were significantly lower (P<0.01). The sense KAI1 gene transfection into the cancer cells also inhibited their invasion and lung metastasis in the orthotopic liver cancer nude mice. However, the opposite changes were observed in the HCC cells transfected with the antisense KAI1 gene. KAI1 gene transfection also affected ECM and ICAM-1 expression in the transplanted liver cancer. CONCLUSION: The KAI1 gene plays an important role in the invasion and metastasis of human HCC and its upregulation in HCC cells suppresses their invasive and metastatic abilities. KAI1 gene functioned as a metastasis inhibitor by regulating the HCC cell biophysical behaviours including aggregation, adhesion, motility and visco-elastic properties.

KAI1, a metastasis suppressor gene of prostate cancer, is located on human chromosome 11p11.2. Down-regulation of KAI1 mRNA during tumor progression and metastasis has been reported for several kinds of cancer, but the mechanism of this down-regulation is not known. In the present study, our aim was to ascertain the relationship between down-regulation of KAI1 mRNA expression and KAI1 gene alterations in lung cancer. Forty-nine cases of adenocarcinoma of the lung were studied by reverse-transcriptase polymerase chain reaction (RT-PCR) assay of KAI1 mRNA and by immunohistochemical detection of KAI1 protein. In addition, markers of the microsatellite loci D11S1344 and D11S1326 were used to investigate loss of heterozygosity (LOH) and replication errors (RERs) of the KAI1 gene region. The RT-PCR assay showed that there was no correlation between KAI1 mRNA expression and either the age of the patients or tumor size. By contrast, KAI1 mRNA expression was significantly correlated with gender (P=0.047), metastasis to the lymph nodes or other organs (P=0.004), the histological grade of the tumor (P=0.036) and the pathological stage (P=0.049). Immunohistochemical staining showed that in one case without metastasis, loss of KAI1 mRNA was associated with invasion of the stroma by KAI1 protein-negative cancer cells. The numbers of informative cases by microsatellite analysis were 14 (28.6%) of 49 at D11S1344 and 27 (55.1%) of 49 at D11S1326; none of 49 adenocarcinomas showed LOH or RERs at these loci. These results suggest that down-regulation of KAI1 mRNA expression rarely if ever involves LOH or RERs of the KAI1 gene region in primary lung adenocarcinoma.

KAI1/CD82 has been shown to be a metastasis suppressor for several human cancers, and a recent study revealed that wild-type tumor suppressor p53 can directly activate KAI1/CD82 gene expression. However, the response of KAI1/CD82 expression in cancer cells to exogenous stimulants has not been investigated. The present study examined whether tumor necrosis factor (TNF), which mediates many of the cellular responses associated with inflammatory reactions or cancer progression, can affect the KAI1/CD82 expression in lung cancer cells and, if so, whether nuclear factor (NF)-kappaB, a key molecule in TNF-mediated gene expression, is involved in the mechanism of KAI1/CD82 induction. Our results demonstrated that expression of KAI1/CD82 in PC-14 cells expressing mutant p53 could be augmented by TNF-alpha, and that transfer of the gene for a specific inhibitor of NF-kappaB, IkappaB alphaSR (mutant IkappaB alpha; NF-kappaB super-repressor), into PC-14 cells could inhibit this augmentation. The amount of NF-kappaB in the nucleus of PC-14/IkappaB alphaSR cells correlated well with KAI1/CD82 mRNA and protein expression. In addition, IkappaB alphaSR gene transfer inhibited the spontaneous expression of KAI1/CD82 protein in KAI1/CD82-high-expressing RERF-LC-OK cells, which contain a mutant-type p53. These observations indicate that NF-kappaB activation may play a role in the regulation of KAI1/CD82 expression in lung cancer cells independently of wild-type p53, and suggest that KAI1/CD82 expression may be regulated by interaction with the host microenvironment.

KAI1, which is identical to CD82, was initially identified as a metastasis-suppressor gene for human prostate cancer, and its expression is reported to be a favorable prognostic factor for operable human lung cancer. In this study, we examined the functional role of KAI1/CD82 in the late phase of metastatic spread of human lung-cancer cells. For this, KAI1/CD82 cDNA was introduced into KAI1/CD82 low-expressing human lung-cancer cell lines, SBC-3 and PC-14, and then the metastatic potential of the transformants was analyzed by i.v. inoculation of KAI1/CD82-transduced cells, SBC-3/KAI1 and PC-14/KAI1, into NK cell-depleted SCID mice. Contrary to our expectations, KAI1/CD82 gene transfer promoted multiorgan metastasis of i.v.-inoculated human lung-cancer cells, while s.c. tumor growth was unaffected. cancer cells from metastatic tumors of NK cell-depleted SCID mice injected i.v. with SBC-3/KAI1 expressed appreciable cell-surface KAI1/CD82, and cells not expressing KAI1/CD82 (revertants) were not detected in the tumors. Our findings indicate that under conditions where the host's natural cytotoxicity is suppressed, KAI1/CD82 may enhance the formation of tumors by circulating lung-cancer cells at metastatic sites.

OBJECTIVE: To investigate the correlation of a newly-defined tumor metastasis suppressor gene KAI1/CD82 and metastatic potential of human cancer cell lines. METHODS: expressions of KAI1/CD82 in eight human cancer cell lines of prostate, lung and melanoma with different metastasis potential were studied by Northern blot hybridization and the mutation were detected by PCR-SSCP. RESULTS: expressions of KAI1 mRNA decreased in human prostate cancer cell lines PC3 and PC3M, and lung cancer cell line PG which were metastatic cancer cell lines (integral light density values: 0.0319, 0.0266 and 0.0549, respectively). expression of KAI1 mRNA showed high level in PAa, a human lung cancer cell line with no metastasis (integral light density value: 0.7313). In human melanoma cell lines with different metastasis potential (WM35, WM1341b, WM983a and WM451), expression of KAI1 showed moderate expression (integral light density values are 0.1798, 0.1582, 0.1501, 0.1800 respectively). PCR-SSCP analysis indicated the existence of band shift in exon 7 of KAI1 gene in PC3, PG and PC3M. CONCLUSION: Decreased KAI1/CD82 expression may be associated with enhanced metastatic potential in prostate cancer cell lines and lung cancer cell line other than melanoma cell lines. The decreased expression may be partially nelaled with mutation of KAI1 gene.

Although the KAI1/CD82 protein has been reported to inhibit cell metastasis in many studies, its mechanism of action has not yet been fully elucidated. In the present study, we investigated the possible effects of KAI1/CD82 on the metastatic phenotype in H1299 lung carcinoma cells. These studies were based on the pivotal role that the acquisition of motile phenotype plays on the initial steps of metastasis. KAI1/CD82-mediated morphological changes were observed using phase contrast microscopy. We report here, that a KAI1/CD82-induced phenotypic change was involved in the decrease of Rac1 expression and GTPase activity. However, we found that KAI1/CD82 did not regulate Rac1 mRNA levels. This suggests the existence of another regulatory mechanism of Rac1 protein maturation or activation. To identify the signaling pathway of Rac1 regulation, we investigated the PI3K/Akt/mTOR pathway, since the PI3K/Akt pathway regulates Rac1 activation and mTOR is known to play a regulatory role in protein translation. H1299/CD82-transfectants showed lower mTOR expression and cell growth than the control group. The data obtained from this study suggested that KAI1/CD82 decreased the metastatic phenotype of H1299 lung carcinoma cells by down-regulating Rac1 expression through the PI3K/Akt/mTOR pathway.

KAI1/CD82, a metastasis suppressor gene of prostate cancer, is located on the human chromosome 11p11.2. Down-regulation of KAI1/CD82 during tumor progression and metastasis has been reported in several cancers, but the mechanism of this down-regulation remains unknown. The relationship between down-regulation of KAI1/CD82 mRNA expression and KAI1/CD82 gene alterations in human melanoma cell lines were investigated. The promoter methylation status was examined after a 331-bp GC-rich fragment of the promoter region was amplified in G361, SK-MEL-24 and SK-MEL-28 cell lines treated with bisulfite. In order to detect methylated CpGs in all three cell lines, 331-bp fragments were sequenced. To examine the restoration of KAI1/CD82 mRNA and protein expression, the cells were exposed to methylase inhibitor, 5-aza-2'-deoxycytidine (5-AzaC). Bisulfite-sequencing data showed no methylation in G361 and SK-MEL-24 cells, and slight methylation in SK-MEL-28 cells at CpG sites 23-26 in the promoter. Real-time PCR and flow cytometry analysis showed that 5-AzaC-treated cells restored KAI1/CD82 mRNA and protein expression in SK-MEL-24 and SK-MEL-28 cells, compared to the controls. The restoration of KAI1/CD82 mRNA and protein expression detected no significant difference between SK-MEL-24 and SK-MEL-28 cells. This means that 5-AzaC did not affect the methylated cells only. Loss of heterozygosity (LOH) at polymorphic microsatellite loci on the human chromosome 11 in the human melanoma cells was also examined. Microsatellite analysis showed LOH at D11S1344 in SK-MEL-24 and SK-MEL-28 cells, and G361 showed allelic imbalance. In conclusion, this study suggests that down-regulation of KAI1/CD82 mRNA expression in human melanoma cell lines is related to LOH or allelic imbalance, but not to methylation of the KAI1/CD82 gene region.

Introduction. Uveal melanoma (UM) is an intraocular tumor that leads to metastatic disease in approximately 50% of afflicted patients. There is no efficacious treatment for metastatic disease in this cancer. Identification of markers that can offer prognostic and therapeutic value is a major focus in this field at present. KAI1 is a metastasis suppressor gene that has been reported to play a role in various human malignancies, although it has not previously been evaluated in UM. Purpose. To investigate the expression of KAI1 in UM and its potential value as a prognostic marker. Materials and Methods. 18 cases of human primary UM were collected and immunostained for KAI1 expression. A pathologist evaluated staining intensity and distribution semiquantitatively. Each case was categorized as group 1 (low staining) or group 2 (high staining). Results. In group 2, two of the 12 cases presented with metastasis. Conversely, in group 1, five out of 6 cases had metastasis. The mean follow-up of patients who did not develop metastasis was 81.81 months (median: 75 months) versus 42.14 months (median: 44 months) for patients with metastasis. Conclusions. KAI1 is a promising candidate marker that may offer prognostic value in UM; it may also represent a therapeutic target in metastatic disease.

metastasis is the primary cause of mortality from cancer, but the mechanisms leading to metastasis are poorly understood. In particular, relatively little is known about metastasis in cancers of mesenchymal origins, which are known as sarcomas. Approximately ten proteins have been characterized as 'metastasis suppressors', but how these proteins function and are regulated is, in general, not well understood. Gp78 (also known as AMFR or RNF45) is a RING finger E3 ubiquitin ligase that is integral to the endoplasmic reticulum (ER) and involved in ER-associated degradation (ERAD) of diverse substrates. Here we report that expression of gp78 has a causal role in the metastasis of an aggressive human sarcoma and that this prometastatic activity requires the E3 activity of gp78. Further, gp78 associates with and targets the transmembrane metastasis suppressor, KAI1 (also known as CD82), for degradation. Suppression of gp78 increases KAI1 abundance and reduces the metastatic potential of tumor cells, an effect that is largely blocked by concomitant suppression of KAI1. An inverse relationship between these proteins was confirmed in a human sarcoma tissue microarray. Whereas most previous efforts have focused on genetic mechanisms for the loss of metastasis suppressor genes, our results provide new evidence for post-translational downregulation of a metastasis suppressor by its ubiquitin ligase, resulting in abrogation of its metastasis-suppressing effects.

OBJECTIVE: To investigate the relationship between the expressions of KAI1, nm23, ETS-1, vascular endothelial growth factor (VEGF) and microvascular density (MVD) and lymph node metastasis and prognosis in nasopharyngeal carcinoma (NPC). METHODS: The Envision immunohistochemical method was used to detect the expressions of KAI1, nm23, ETS-1 and VEGF in 50 cases of non-keratinizing carcinoma (NKC) with cervical lymph node metastasis, 30 cases of NKC without cervical lymph node metastasis at the primary diagnoses and 30 cases of non-tumor nasopharyngeal tissues (NP). The microvascular density was counted by immunostaining with CD34. RESULTS: (1) The expression rates of KAI1 and nm23 protein in NKC with cervical lymph node metastasis group and without cervical lymph node metastasis group and NP group increased successively , the difference being significant (P < 0.05); The expression rates of ETS-1 and VEGF protein in NKC with cervical lymph node metastasis group and without cervical lymph node metastasis group and NP group increased successively, the difference being significant (P < 0.05). (2) In 80 NKC cases, the MVD was respectively lower in KAI1 and nm23 protein positive groups than those in the negative groups (P < 0.05); the MVD was respectively higher in ETS-1 and VEGF protein positive groups than those in the negative groups (P < 0.05 ). (3) There was significant difference between the MVD, the number of NKC without cervical lymph node metastasis cases in the single expression of KAI1 or nm23 protein and in common expression of KAI1 and nm23 protein (P < 0.05), in the same as between the single expression of ETS-1 or VEGF protein and in common expressions of ETS-1 and VEGF protein (P < 0.05). (4) There was positive correlation between the expressions of KAI1 and nm23 protein (P < 0.01), as well as between the expressions of ETS-1 and VEGF protein (P < 0.01). (5) the 5-year survival rates of the patients correlated with cervical lymph node metastasis and the expressions of KAI1, nm23, ETS-1 and VEGF proteins in NKC (P < 0.05). CONCLUSIONS: The expressions of KAI1, nm23, ETS-1 and VEGF proteins were highly related to MVD in NPC,cervical lymph node metastasis and prognosis. They might be considered to be reference indicator for evaluating the cervical lymph node metastasis and prognosis of NPC.

BACKGROUND & OBJECTIVE: KAI1/CD82 was recently detected as a tumor metastasis suppressor gene. Its silencing contributes to progression and infiltration of some tumors. Our study was designed to investigate the expression of KAI1/CD82 in neuroblastoma, and explore its correlation to clinicopathologic characteristics and prognosis of patients with neuroblastoma. METHODS: The EnVision immunohistochemistry was used to detect the expression of KAI1/CD82 in 90 specimens of neuroblastoma (28 specimens of ganglioneuroblastoma and 62 specimens of neuroblastoma). Clinical data and follow-up data of the 90 patients were analyzed. RESULTS: Positive rate of KAI1/CD82 was significantly higher in ganglioneuroblastoma than in neuroblastoma (39.3% vs. 14.5%, P=0.014). Its expression was negatively correlated to clinical stage of neuroblastoma (P=0.003). CONCLUSIONS: The change of KAI1/CD82 expression is an early event in tumorigenesis of neuroblastoma. Its down-regulation may be considered as a potential indicator to judge the differentiation and metastasis of neuroblastoma, which can serve as one of the combined indexes to clinical assessment of prognosis.

The KAI1 gene maps to chromosome 11p11.2, is a metastasis-suppressor gene for human prostate cancer and also is involved in the progression of human pancreatic and non-small cell lung cancer. Recently, we showed that introduction of a single copy of normal, neomycin-tagged human chromosome 11 into highly metastatic MDA-MB-435 breast cancer cells suppressed breast cancer metastasis. Concomitantly, KAI1 levels were higher in chromosome 11/MDA-MB-435 cell clones. The purpose of this study was to test whether KAI1 expression is indicative of breast cancer metastasis using a panel of immortalized breast epithelial and breast cancer cell lines that represent multiple stages of breast cancer progression. metastatic cell clones isolated from the parental mixed, wild-type population of MDA-MB-435 cells expressed the lowest levels of KAI1 mRNA and chromosome 11 containing MDA-MB-435 (neo11/MDA-MB-435.A3 and neo11/MDA-MB-435.B1) cells had approximately twice as much KAI1 mRNA than the parental clones. MCF-10A, an immortalized normal-like non-tumorigenic mammary epithelial cell line, had the highest level of KAI1 mRNA. We compared the metastatic propensity and invasive ability of a continuum of breast cancer cells with varying degrees of progression toward malignancy and found that these parameters tended to correlate inversely with KAI1 mRNA expression. These data suggest that, in addition to its role in human prostate, pancreatic and non-small cell lung cancer, KAI1 may also be a useful marker for staging human breast disease.

The multistep process of carcinogenesis implies the accumulation of multiple molecular defects. Alteration of tumor suppressor and metastasis suppressor genes are the important steps. Increasing experimental evidence indicates that decreased expression of tumor-metastasis/suppressor genes and gene products are involved in the progression of a variety of human malignancies. In the present study, we have extended this analysis to non-small cell lung carcinomas (NSCLC). The expression and prognostic significance of the tumor suppressor gene PTEN and metastasis suppressor genes nm23-H1 and KAI-1 was evaluated in NSCLCs. Immunohistochemical staining was performed on formalin-fixed, paraffin-embedded tissues from 53 bronchogenic adenocarcinomas and 51 squamous cell carcinomas using monoclonal antibodies against PTEN, nm23H-1, and KAI-1 proteins. Immunohistochemical results were correlated with tumor stage, grade, lymph nodes positivity, metastasis, and patient survival. Significant co-expression of PTEN, nm23-H1 and KAI-1 was observed in NSCLC (P<.001 to .002). The immunohistochemical expression of these proteins was significantly higher in stages 1 and 2 compared with stages 3 and 4 (P=.04 for PTEN and KAI-1, P=.039 for nm23-H1). When all stages were considered together, loss of immunoreactivity for PTEN, nm23-H1 and KAI-1 was found in advanced NCSCLs (P=.015 for PTEN, P=.001 for KAI-1, P=.004 for nm23-H1), which is suggestive of co-downregulation of these proteins in the process of tumor progression. On multivariate analysis, negative staining for PTEN (P=.014), KAI-1 (P=.034), and nm23-H1 (a trend toward association for nm23-H1 reached near significance P=.08) correlated with disease-related death. Positive lymph node status was associated with negative immunostaining for PTEN (P=.007) but no correlation was observed for nm23-H1 and KAI-1. Loss of expression was linked to distant metastasis (P=.006 for PTEN, P=.002 for nm23H1, P=.001 for KAI-1). On multivariate analysis, co-downregulation of PTEN (P=.009), KAI-1 (P=.02), and nm23-H1 (P=.011) independently predicted shortened survival in NSCLC. Although NSCLC exhibits strong co-expression of PTEN, nm23-H1 and KAI-1, there is a loss of these proteins in high-stage tumors. Co-downregulation of PTEN, KAI-1, and nm23-H1 significantly correlates with distant metastasis and predicts shortened survival. Our study supports a role of these tumor suppressor and metastasis suppressor genes in the evolution and progression of NSCLC.

We conducted a study on the mechanism of KAI1/CD82-mediated suppression of tumor invasiveness and metastasis, and examined its effect on MMP-9 activity and the TIMP1 levels in H1299 human non-small cell lung carcinoma cells. The H1299 human lung carcinoma cells were transfected with pcDNA3.1-CD82 and stable transfectant clones that had a high KAI1/CD82 expression were obtained. We performed Western blot analysis, cell invasion assay, gelatin zymography, and RT-PCR to assess the KAI1/CD82 expression and tumor invasiveness, the MMP-9 activity, the MMP-9 mRNA and protein levels, and the TIMP1 levels in the H1299/CD82 transfectant cells and compared the results with those of the control groups. The H1299/CD82 transfectants exhibited significant suppression of cell invasion, reduced MMP9 enzyme activity, elevated MMP9 mRNA and MMP-9 protein levels, and elevated TIMP1 levels. It may be postulated that KAI1/CD82 over-expression in the H1299 non-small cell lung carcinoma cells suppresses the tumor invasiveness and metastatic potential by inducing MMP9 inactivation via the up-regulation of TIMP1.

BACKGROUND AND OBJECTIVE: Vasculogenic mimicry (VM), found in many high invasive ability tumors, is associated with tumor invasion and metastasis. Many genes exhibit abnormal levels of expression in these tumors. This study aims to find good markers for predicting the invasion and metastasis of non-small cell lung cancer (NSCLC). METHODS: VM and expression of CD82/KAI1 and HIF-1alpha were examined via immunohistochemistry and histochemistry of 160 NSCLC and 20 normal lung tissue specimens. RESULTS: In NSCLC, positive rates of 37.5%, 48.8%, and 36.9% were obtained for CD82/KAI1, HIF-1alpha, and VM, respectively. In normal lung tissue, positive rates of 95.0%, 0, and 0 were obtained for CD82/KAI1, HIF-1alpha, and VM, respectively. A significant difference was found between the NSCLC and normal lung groups (P<0.01). VM and the expression levels of CD82/KAI1 and HIF-1alpha were significantly related to tumor differentiation, lymph node metastasis, clinical staging, and postoperative survival time (P<0.01 for all). A negative correlation was found between the expression levels of CD82/KAI1 and HIF-1alpha; a similar relationship was observed between CD82/KAI1 and VM. A positive relationship between the expression of HIF-1alpha and VM was revealed; there was a significant relationship between microvessel density (MVD) and the expression of CD82/KAI1 or HIF-1alpha or VM. VM and overexpression of HIF-1alpha were related to poor prognosis: the survival rates were significantly lower in positive patients than in negative patients (both P<0.01). The survival rates of the CD82/KAI1-positive and CD82/KAI1-negative groups were significantly different (P<0.01). The five-year survival rate was significantly different between the group with MVD>/=22 and the group with MVD<22 (P<0.01). pTNM stage, positive expressions of CD82/KAI1 and HIF-1alpha, and VM were independent prognostic factors of NSCLC (P<0.01). CONCLUSIONS: VM and the expressions of CD82/KAI1 and HIF-1alpha in NSCLC are related to differentiation, lymph node metastasis, clinical staging, and prognosis. The combined detection of CD82/KAI1, HIF-1alpha, and VM has an important role in predicting the progression and prognosis of NSCLC.

BACKGROUND: This study investigates the expression of CD82/KAI1 and epithelial-cadherin (E-cad) in non-small cell lung cancer (NSCLC). METHODS: Tissues of resected primary NSCLC and normal lung tissue were investigated. Protein expression was detected with immunohistochemical staining. mRNA expression levels of CD82/KAI1 and E-cad were determined by semiquantitative reverse transcriptase polymerase chain reaction (RT-PCR) after mRNA extraction. RESULTS: The expression of CD82/KAI1 and E-cad was significantly lower in NSCLC compared to normal lung tissue (P < 0.01). CD82/KAI1 and E-cad mRNA and protein expression were found to be in close relationship with the grade of differentiation, lymph node metastasis and pathologic tumor-node-metastasis (pTNM) stages, and survival in NSCLC (P < 0.01), but no relationship was observed with gender, diameter, type, histological type and age (P > 0.05). expression of mRNA CD82/KAI1 and E-cad were consistent with their proteins (P < 0.01), and there was a significant relationship between expression of CD82/KAI1 and E-cad. The survival rate of the CD82/KAI1-positive and CD82/KAI1-negative groups was significantly different (P < 0.01). In addition, the survival rate was significantly different between the E-cad-positive and E-cad-negative groups. pTNM stages and positive expression of CD82/KAI1 and E-cad were independent prognostic factors of NSCLC (P < 0.05). CONCLUSION: Lower expression of CD82/KAI1 and E-cad was found in NSCLC compared to normal lung tissue. Decreased expression of CD82/KAI1 and E-cad was closely related to cellular differentiation, pTNM stages, invasion and metastasis.

PURPOSE: KAI1 was originally identified in prostate cancer as a metastasis suppressor gene. Recent studies have shown a frequent down-regulation of KAI1 expression in many tumor types, whereas mutation or hypermethylation of the gene is infrequent. The aim of the present study was to examine whether loss of KAI1 expression that might be caused by genetic or epigenetic alterations could contribute to oral carcinogenesis. EXPERIMENTAL DESIGN: We analyzed mutational and methylation status of the KAI1 gene and both the mRNA and protein level in a series of oral tumors [28 precancerous lesions, 101 primary oral squamous cell carcinomas (OSCCs), and 30 metastatic OSCCs] and OSCC-derived cell lines. We also examined p53 protein expression, which has been reported to be a candidate activator for the KAI1 gene. RESULTS: With the exception of three microsatellite instabilities in the KAI1 gene, we found no mutations in the coding sequence of the KAI1 gene, no loss of heterozygosity, and no hypermethylation of the KAI1 promoter region in all samples investigated. By immunohistochemistry, however, high frequencies of KAI1 down-regulation were evident not only in the metastatic OSCCs [29 of 30 (97%)] but also in the primary OSCCs [83 of 101 (82%)] and in the precancerous lesions [13 of 28 (46%)]. There was a significant relationship between down-regulation of KAI1 protein expression and primary tumors associated with lymph node metastases (P = 0.0115), whereas there was no statistical correlation between p53 status and KAI1 expression. Taken together, reverse transcription-PCR data were consistent with the protein expression status in 16 patients from whom mRNA was available. CONCLUSIONS: Our data suggest that whereas loss of KAI1 protein expression is associated with primary tumors with lymph node metastases, the down-regulation of KAI1 is an early event in the progression of human oral cancer. The down-regulation of KAI1 is not associated with either mutation, allelic loss, methylation of the promoter, or p53 regulation.

Oral Squamous Cell Carcinoma (OSCC) is a common malignancy. Treatment failure is mainly due to loco-regional disease recurrence. KAI1 is a newly discovered metastasis suppressor gene. Fifty-seven patients with primary OSCC underwent surgery alone or surgery and adjuvant radiotherapy. Immunohistochemical evaluation of KAI1/CD82 and p53 proteins was carried out on specimen obtained at surgery. Within neoplastic fields, KAI1/CD82 expression was downregulated and negative in 42/57 (73.7%) cases. p53 expression was positive in 26/57 (45.6%) cases. No correlation was noted between KAI1/CD82 and p53 expression or clinicopathological parameters. Univariate and multivariate Cox proportional hazard models showed a correlation between KAI1/CD82 expression with disease free survival (P = 0.01, P = 0.009) and overall survival (P = 0.04, P = 0.053) respectively.

OBJECTIVE: KAI1 is a recently identified metastasis suppressor gene on human chromosome 11p11.2. It belongs to a structurally distinct family of cell surface glycoproteins. Decreased KAI1 expression seems to be involved in the progression of human prostate, lung, pancreatic, and possibly breast cancer, and recently a reduced KAI1 protein expression has been demonstrated in several ovarian carcinoma cell lines. The aim of this study is to determine whether the KAI1 gene is altered in human epithelial ovarian carcinomas. In addition, its prognostic significance in this tumor is also evaluated. METHODS: To detect KAI1 expression, 102 tumor samples from benign, borderline, primary invasive, metastatic, and recurrent epithelial ovarian tumors were prepared for immunohistochemical study with C-16, an anti-KAI1 polyclonal antibody. In addition, cellular RNA from 24 primary invasive and 7 recurrent tumors was also analyzed for KAI1 expression by using a reverse transcriptase PCR (RT-PCR) technique. The PCR single-strand conformation polymorphism method and direct DNA sequencing were used to detect KAI1 mutation in the 44 primary invasive and 8 recurrent ovarian carcinomas. RESULTS: In immunohistochemical study, decrease of KAI1 protein expression was associated with the progression of ovarian tumor. However, it had no relation to the stage of primary invasive cancers because of its frequent occurrence in early stage tumors. KAI1 expression was also frequently down-regulated in primary invasive and recurrent tumors in RT-PCR analysis. Except for a missense change at codon 241 (ATC to GTC), which causes the substitution of a valine for an isoleucine in the amino acid sequence and occurs in both normal and tumor tissues, no mutation of the KAI1 gene was found in any of the 52 carcinomas. Although there was a trend for deteriorating survival from patients with KAI1-preserved tumors to those with KAI1-decreased and -negative tumors, statistically it was not significant (P = 0.079). CONCLUSION: KAI1 may play a role in the malignant progression of epithelial ovarian carcinoma through the down-regulation of expression rather than gene mutation. Since the decreased expression presented frequently in early stage tumors, it may be an early event in the progression of this tumor and its prognostic significance needs further investigation with a larger number of cases.

OBJECTIVES: The aim of this study was to investigate the impact of downregulation of KAI1 metastasis suppressor protein in epithelial ovarian cancer. In addition, correlation of KAI1 and p53 immunostaining was investigated. METHODS: expression of KAI1 and p53 was immunohistochemically determined in 107 specimens of epithelial ovarian cancer stages I-IV. Survival of patients was investigated using uni- and multivariate analysis. RESULTS: Strong KAI1 expression was observed in 17.8% of cases, moderate in 27.1%, weak in 21.5%, and complete loss of KAI1 expression in 33.6%. Overexpression of p53 protein was observed in 45.8%. There was correlation of KAI1 expression neither with p53 expression nor with various clinical and histopathological parameters. Serous ovarian cancers showed significantly decreased staining intensity of KAI when compared to other histological types (P = 0.007). Univariate and multivariate analysis revealed that patients with strong or moderate expression of KAI1 had a significantly longer overall (P = 0.0013) and disease-free survival (P = 0.0048) when compared to those with low or absent expression. CONCLUSION: KAI1 downregulation is an independent prognostic factor in epithelial ovarian cancer, indicating dismal prognosis. Our study did not reveal a correlation between p53 status and KAI1 expression, suggesting that p53-independent mechanisms might be involved in the downregulation of KAI1.

The tetraspanin KAI1 had been described as a metastasis suppressor in many different cancer types, a function for which associations of KAI1 with adhesion and signaling receptors of the integrin superfamily likely play a role. In ovarian cancer, integrin alphavbeta3 correlates with tumor progression and its elevation in vitro provoked enhanced cell adhesion accompanied by significant increases in cell motility and proliferation in the presence of its major ligand vitronectin. In the present study, we characterized integrin alphavbeta3-mediated tumor biological effects as a function of cellular KAI1 restoration and proved for the first time that KAI1, besides its already known physical crosstalk with beta1-integrins, also colocalizes with integrin alphavbeta3. Functionally, elevated KAI1 levels drastically increased integrin alphavbeta3/vitronectin-dependent ovarian cancer cell adhesion. Since an intermediate level of cell adhesive strength is required for optimal cell migration, we next studied ovarian cancer cell motility as a function of KAI1 restoration. By time lapse video microscopy, we found impaired integrin alphavbeta3/vitronectin-mediated cell migration most probably due to strongly enhanced cellular immobilization onto the adhesion-supporting matrix. Moreover, KAI1 reexpression significantly diminished cell proliferation. These data strongly indicate that KAI1 may suppress ovarian cancer progression by inhibiting integrin alphavbeta3/vitronectin-provoked tumor cell motility and proliferation as important hallmarks of the oncogenic process.

KAI1 is a metastasis suppressor gene for prostate cancer that is located on chromosome 11p11.2-13. Using Northern blot analysis and in situ hybridization, we studied expression of KAI1 mRNA in specimens from 14 normal pancreases and 27 primary pancreatic cancers, and then correlated the findings with the clinical and histopathological parameters of the patients. Northern blot analysis showed increased steady-state levels of KAI1 mRNA expression in 24 of 27 (89%) pancreatic cancer samples. In situ hybridization showed enhanced KAI1 mRNA levels in the pancreatic cancer cells in 82% cancer tissues. The stroma surrounding the cancer mass and normal pancreatic tissue adjacent to the cancer cells exhibited very low levels of KAI1 mRNA expression. Correlation of the mRNA levels obtained by Northern blot analysis with clinical parameters of the patients revealed that KAI1 mRNA levels were significantly higher (P < 0.01) in earlier tumor stages (I, II), compared with advanced tumor stages (III, IV) in which lymph node or distant metastases were present. Furthermore, poorly differentiated tumors had significantly higher KAI1 mRNA levels than those that were moderately or well differentiated (P < 0.05). No association between KAI1 expression and survival was found. Our results indicate that KAI1 mRNA expression is reduced in patients with advanced tumor stages. This suggests that reduction of KAI1 expression might enable pancreatic cancer cells to spread in lymph nodes and to distant organs.

The KAI1 gene maps to chromosome 11p11.2, is a metastasis-suppressor gene for human prostate cancer and also is involved in the progression of human pancreatic and non-small cell lung cancer. Recently, we showed that introduction of a single copy of normal, neomycin-tagged human chromosome 11 into highly metastatic MDA-MB-435 breast cancer cells suppressed breast cancer metastasis. Concomitantly, KAI1 levels were higher in chromosome 11/MDA-MB-435 cell clones. The purpose of this study was to test whether KAI1 expression is indicative of breast cancer metastasis using a panel of immortalized breast epithelial and breast cancer cell lines that represent multiple stages of breast cancer progression. metastatic cell clones isolated from the parental mixed, wild-type population of MDA-MB-435 cells expressed the lowest levels of KAI1 mRNA and chromosome 11 containing MDA-MB-435 (neo11/MDA-MB-435.A3 and neo11/MDA-MB-435.B1) cells had approximately twice as much KAI1 mRNA than the parental clones. MCF-10A, an immortalized normal-like non-tumorigenic mammary epithelial cell line, had the highest level of KAI1 mRNA. We compared the metastatic propensity and invasive ability of a continuum of breast cancer cells with varying degrees of progression toward malignancy and found that these parameters tended to correlate inversely with KAI1 mRNA expression. These data suggest that, in addition to its role in human prostate, pancreatic and non-small cell lung cancer, KAI1 may also be a useful marker for staging human breast disease.

The human KAI1 metastasis suppressor gene encodes for a 267 amino acid plasma membrane glycoprotein, which has four transmembrane domains and one large and one small extracellular domain. Plasma membrane expression of KAI1 is downregulated during the progression of several cancers to a metastatic state, including prostate, lung, and pancreatic cancers. To elucidate the mechanisms for this downregulation, an understanding of its transcriptional regulation is critical. Therefore, a set of luciferase reporter gene plasmid constructs were generated containing various 5' flanking regions to the transcription initiation site, with or without the first exon and a portion of the first intron of the KAI1 gene. These constructs were transfected into a series of normal and malignant prostate cell lines as well as fibroblasts and the resultant luciferase activities used to define the regulatory elements for expression of the KAI1 gene. Using this approach, the minimal KAI1 promoter was found to be 0.5 kb in size and contains two regions. The first region includes the 197 bp 5' to the transcription initiation site and the second region includes the first exon and a portion of first intron (i.e., +1 to +351 bp). This combined region result in positively regulated strong, but not epithelial specific, luciferase expression. In addition to this minimal promoter element from -197 to +351 bp, there is a second regulatory element further upstream (i.e., -735 to -197 bp), which negatively regulates luciferase expression. Additional studies of Marreiros et al. (Gene 302:155, 2003) have documented that a third regulatory element even further 5' (i.e., -922 to -846 bp) encodes an enhancer for the KAI1 gene.

AIM: To investigate whether KAI1, as a metastasis suppressor gene, is associated with invasive and metastatic ability of pancreatic cancer cells. METHODS: KAI1 gene was transfected into pancreatic cancer cell line MiaPaCa II by liposomes selected with G418. expression of transfected cells was measured by Western blotting, immunofluorescence and immunocytochemistry. tumor cell invasion and metastatic ability were detected through gelatinase activity and reconstituted basement membrane (Matrigel) assay. pCMV-KAI1 was directly injected into the heterotopic human pancreatic adenocarcinoma successfully established in the groin of BALB/C nude mice, by subcutaneous injection of MiaPaCa II pancreatic cancer cells. The statistical analysis between groups was determined by Student's two tailed t test. RESULTS: By Western blotting, MiaPaCa II cells transfected by KAI1 gene indicated KAI1 expression at approximately 29.1 kDa. Cytoplasm staining was positive and uniformly spread in transfected cancer cells, using immunohistochemistry and immunofluorescence. The most obvious difference was present after 30 h (MiaPaca II 43.6 +/- 9.42, pCMV-MiaPaca II 44.8 +/- 8.56, pCMV-KAI1-MiaPaca II 22.0 +/- 4.69, P < 0.05). Gelatinolysis revealed a wider and clearer band of gelatinolytic activity in non-transfected than in transfected cells (MiaPaCa II cells 30.8 +/- 0.57, transfected cells 28.1 +/- 0.65, P < 0.05). In vivo tumor growth rates of KAI1 transfectants with KAI1-Lipofectamine 1.22 +/- 0.31 in A group were lower than control 4.61 +/- 1.98 and pCMV-KAI 11.67 +/- 0.81. Analyses of metastases with and without KAI1 transfection in mice were different in liver and lung between controls 1.62 +/- 0.39, 0.45 +/- 0.09, pCMV-KAI 1.01 +/- 0.27, 0.33 +/- 0.09 and KAI1-Lipofectamine 0.99 +/- 0.21, 0.30 +/- 0.09 respectively (P < 0.05). CONCLUSION: High expression of KAI1 gene was found in transfected MiaPaCa II human pancreatic cancer cells with lower metastatic ability. KAI1 gene plays an important role in inhibiting metastasis of pancreatic cancer after direct injection into pancreatic adenocarcinoma. These results show that the suppressed invasion and motor function of pancreatic cancer cells may be a key reason why the KAI1 gene controls pancreatic cancer cell metastasis.

KAI1 is a metastasis suppressor gene located on human chromosome 11p11.2. Previous studies have shown that the down-regulation of KAI1 mRNA and decreased expression of its gene product are significantly linked to carcinoma progression, including metastatic ability. In this study, we investigated KAI1 protein expression in thyroid neoplasms. KAI1 overexpression was observed in 64.0% of papillary carcinoma cases, and the incidence was significantly higher than in cases of follicular carcinoma (20.0%) (p = 0.0001). In papillary carcinomas, decreased KAI1 expression was frequently observed in cases invading beyond the thyroid capsule (p = 0.001), as well as in lymph node metastases (p = 0.0047) and poorly differentiated lesions (p = 0.0299). Furthermore, in anaplastic carcinoma, the incidence of KAI1 overexpression was lower than in papillary carcinoma (p < 0.0001), and only 4.2% of the cases overexpressed this gene. These results suggest that KAI1 down-regulation is significantly related to the progression of papillary carcinoma, including lymph node metastasis, and its anaplastic transformation.

In penile squamous cell carcinoma (PSCC), the outcome largely depends on early detection and resection of inguinal lymph node metastases. We investigated the role of metastasis suppressor protein kang ai 1 (KAI1)/cluster of differentiation 82 (CD82), which is known to be of prognostic significance for a wide variety of cancers. Moreover, we analysed the tumours for human papillomavirus (HPV) DNA and loss of heterozygosity at the 11p11.2 locus. Tissue samples of 30 primary PSCCs were investigated immunohistochemically using an anti-KAI1/CD82 polyclonal antibody. The expression was assessed according to the degree of KAI1/CD82-positive tumour cells as positive, decreased or negative. The presence of HPV6/11, HPV16 and HPV18 DNA was analysed by polymerase chain reaction. All patients with decreased or negative expression of KAI1/CD82 in primary lesions had lymph node metastases (p = 0.0002). Patients with positive KAI1/CD82 expression showed a significant better prognosis for survival compared to the other groups (p = 0.0042). Presence of HPV DNA was associated with decreased or negative KAI1/CD82 expression. Lacking or decreased expression of metastasis suppressor gene KAI1/CD82 appears to be a prognostic parameter for the occurrence of lymph node metastases in PSCC. Our study suggests an association of decreased KAI1/CD82 expression with tumour progression, development of metastases and disease-specific death.

OBJECTIVE: To study the correlation of the expressions of CD82 and hTERT and HPV infection with the clinical pathological features of penile cancer and identify their prognostic significance in the lymphatic metastasis of the disease. METHODS: A total of 44 patients underwent partial or radical penectomy and lymph node dissection. The expressions of CD82 and hTERT were determined by immunohistochemistry, and HPV infection was detected by PCR. RESULTS: The positive rates of CD82, hTERT, and HPV DNA in penile carcinoma were 47.7%, 38.6% and 25.9%, respectively. The amplified HPV DNA was HPV-16. The pathological stage and hTERT expression were positively correlated with inguinal lymph node metastasis of penile cancer (P = 0.032, P = 0.041), and so was the pathological stage with the expression of CD82 (P = 0.045), but neither the pathological stage, nor the expression of CD82 or the positive rate of HPV DNA showed any correlation with lymph node metastasis (P = 0.627, P = 0.094, P = 0.633). CONCLUSION: The pathological grade and hTERT expression are independent prognostic factors for lymph node metastasis in penile carcinoma. These features help the prognosis and identification of the patient at the risk of nodal metastasis.

KAI1/CD82 protein is a member of the tetraspanin superfamily and has been rediscovered as a cancer metastasis suppressor. The mechanism of KAI1/CD82-mediated suppression of cancer metastasis remains to be established. In this study, we found that migration of the metastatic prostate cancer cell line Du145 was substantially inhibited when KAI1/CD82 was expressed. The expression of focal adhesion kinase (FAK) and Lyn, a Src family tyrosine kinase and substrate of FAK, was up-regulated at both RNA and protein levels upon KAI1/CD82 expression. The activation of FAK and Lyn, however, remained unchanged in Du145-KAI1/CD82 cells. As a downstream target of FAK-Lyn signaling, the p130CAS (Crk-associated substrate) protein was decreased upon the expression of KAI1/CD82. Consequently, less p130CAS-CrkII complex, which functions as a "molecular switch" in cell motility, was formed in Du145-KAI1/CD82 cells. To confirm that the p130CAS-CrkII complex is indeed important for the motility inhibition by KAI1/CD82, overexpression of p130CAS in Du145-KAI1/CD82 cells increased the formation of p130CAS-CrkII complex and largely reversed the KAI1/CD82-mediated inhibition of cell motility. Taken together, our studies indicate the following: 1) signaling of FAK-Lyn-p130CAS-CrkII pathway is altered in KAI1/CD82-expressing cells, and 2) p130CAS-CrkII coupling is required for KAI1/CD82-mediated suppression of cell motility.

cancer metastasis suppressor KAI1/CD82 belongs to the tetraspanin superfamily and inversely correlates with the metastatic potential of a variety of cancers. The mechanism of KAI1/CD82-mediated metastasis suppression remains unclear. In this study, we found a M(r) 68,00 cell-surface protein physically associated with KAI1/CD82 and named it KASP: a KAI1/CD82-associated surface protein. Distinctive from known KAI1/CD82 associations that usually occur in the context of "tetraspanin web," the KAI1/CD82-KASP association is likely to be direct because it is: (a) highly stoichiometric; (b) stabilized by chemical cross-linking; and (c) independent of cholesterol-enriched lipid rafts. Therefore, KASP is one of the major transmembrane proteins that associates with KAI1/CD82. Consistent with the wide distribution of KAI1/CD82, KASP is expressed ubiquitously in human tissues. Through peptide sequencing, KASP was identified as an immunoglobulin superfamily member called EWI2 or PGRL. Although EWI2/PGRL has been found to associate with tetraspanins CD9 and CD81, it forms distinct complexes with different tetraspanins, and its association with KAI1/CD82 could be independent of CD81 and CD9. Overexpression of EWI2/PGRL in Du145 metastatic prostate cancer cells inhibits cell migration on both fibronectin- and laminin-coated substratum, indicating that EWI2/PGRL directly regulates cell migration. Furthermore, EWI2/PGRL synergizes KAI1/CD82 in inhibiting cell migration, indicating that EWI2/PGRL is likely required for the function of KAI1/CD82. In summary, we identified a major KAI1/CD82-associated protein, EWI2/PGRL, that is important for KAI1/CD82-mediated suppression of cancer cell migration.

NDRG1 and KAI1 belong to metastasis suppressor genes, which impede the dissemination of tumor cells from primary tumors to distant organs. Previously, we identified the metastasis promoting transcription factor, ATF3, as a downstream target of NDRG1. Further analysis revealed that the KAI1 promoter contained a consensus binding motif of ATF3, suggesting a possibility that NDRG1 suppresses metastasis through inhibition of ATF3 expression followed by activation of the KAI1 gene. In this report, we found that ectopic expression of NDRG1 was able to augment endogenous KAI1 gene expression in prostate cancer cell lines, whereas silencing NDRG1 was accompanied with significant decrease in KAI1 expression in vitro and in vivo. In addition, our results of ChIP analysis indicate that ATF3 indeed bound to the promoter of the KAI1 gene. Importantly, our promoter-based analysis revealed that ATF3 modulated KAI1 transcription through cooperation with other endogenous transcription factor as co-activator (ATF3-JunB) or co-repressor (ATF3-NFkappaB). Moreover, loss of KAI1 expression significantly abrogated NDRG1-mediated metastatic suppression in vitro as well as in a spontaneous metastasis animal model, indicating that KA11 is a functional downstream target of the NDRG1 pathway. Our result of immunohistochemical analysis showed that loss of NDRG1 and KAI1 occurs in parallel as prostate cancer progresses. We also found that a combined expression status of these two genes serves as a strong independent prognostic marker to predict metastasis-free survival of prostate cancer patients. Taken together, our result revealed a novel regulatory network of two metastasis suppressor genes, NDRG1 and KAI1, which together concerted metastasis-suppressive activities through an intrinsic transcriptional cascade.

KAI1/CD82, a tetraspanin membrane protein functions as a metastasis suppressor in many types of human cancers and has been shown to regulate cell adhesion properties. In the present study, we investigated the underlying mechanism of KAI1/CD82-mediated changes in cell adhesion to the extracellular matrix using human prostate cancer cells. We found that high KAI1/CD82 expression attenuated short-term cell adhesion to uncoated- or fibronectin-coated plates. Moreover, high KAI1/CD82 expression generated an extracellular environment unfavorable for cell adhesion as compared to low KAI1/CD82 expression, suggesting KAI1/CD82-dependent regulation of extracellular matrix (ECM) molecule(s) expression and/or secretion. Among ECM components examined, fibronectin exhibited decreased expression and secretion in high KAI1/CD82-expressing cells. Furthermore, high KAI1/CD82 expression interfered with the activation of beta (1) integrin at the cell surface while total beta (1) integrin levels remained unchanged, concomitant with reduced formation of focal adhesion complex and decreased bundling of actin filaments. Finally, high KAI1/CD82 expression significantly retarded cell motility in a scratch wound assay. Taken together, our results strongly suggest that KAI1/CD82 attenuates the activation of beta (1) integrin, and thereby down-regulates outside-in signaling of beta (1) integrin, leading to the reduction of focal adhesion formation and fibronectin expression/secretion, which subsequently interferes with cell adhesion properties and motility.

OBJECTIVE: To explore the mRNA expression of KAI1 gene in laryngeal squamous-cell carcinoma and its clinical significance. METHODS: Fresh laryngeal cancer samples taken from 40 laryngeal carcinoma cases and normal control laryngeal tissues from 9 subjects were examined with semi-quantitative reverse transcription polymerase chain reaction (RT-PCR). RESULTS: Moderate, low and negative expression rates of KAI1 gene mRNA in nine normal laryngeal tissues were 33.3% (3/9), 33.3% (3/9) and 33.3% (3/9), respectively. The high, moderate, low and negative expression rates of KAI1 mRNA in 25 laryngeal cancers without lymph node metastasis were 40.0% (10/25), 28.0% (7/25), 20.0% (5/25) and 12.0% (3/25), respectively. The moderate, low and negative expression rates of KAI1 mRNA in 15 laryngeal cancers with lymph node metastasis were 20.0% (3/15), 26.7% (4/15) and 53.3% (8/15), respectively. The KAI1 mRNA expression in the laryngeal cancers without lymph node metastasis was higher than that in normal laryngeal tissues (P < 0.05). The KAI1 mRNA expression in the laryngeal cancers with lymph node metastasis was lower than that in the laryngeal cancers without lymph node metastasis (P < 0.05). The high, moderate and low expression rates of KAI1 mRNA in 10 highly differentiated laryngeal cancers were 50.0% (5/10), 30.0% (3/10) and 20.0% (2/10), respectively. The high, moderate, low and negative expression rates of KAI1 mRNA in 12 low differentiation laryngeal cancers were 8.3% (1/12), 16.7% (2/12), 16.7% (2/12) and 58.3% (7/12), respectively. The differences of KAI1 mRNA expression between high and low differentiation laryngeal cancers were statistically significant (P < 0.05). CONCLUSION: The decrease of KAI1 mRNA expression may be related to lymph node metastasis and low differentiation of laryngeal squamous-cell carcinoma.

CD82 (KAI1) and CD63 (ME491) are highly glycosylated proteins which belong to the transmembrane 4 superfamily (TM4SF). CD82 has been implicated as a possible prostate cancer metastasis suppressor gene, whereas CD63 is involved in the progression of human melanoma cancer. Down-regulation of both CD82 and CD63 expression has been associated with the metastatic potential of several solid tumors. Currently, information is lacking on the role of CD82 and CD63 during thyroid carcinogenesis. The aim of this study was to determine whether the expression of CD82 and CD63 is a useful prognostic indicator in patients with thyroid carcinoma. The expression of CD82 and CD63 was analysed by reverse transcriptase-PCR (RT-PCR) and immunohistochemistry in benign goiter (n=12) and 75 primary thyroid carcinoma tissue specimens (PTC: 33, FTC: 24, UTC: 18) out of which 36 were non-metastasized primary tumors and 39 were metastasized tumors (regional lymph node and/or distant metastases). All of the benign goiter tissues showed CD82 expression. By contrast, a significant decrease in CD82 mRNA and protein levels was detected in carcinoma tissues as compared to benign goiter tissues (p<0.001). A similar down-regulation was observed in metastasized tumor tissues when compared with non-metastasized tumors (all p<0.05). CD82 expression was correlated with pTNM status of differentiated and undifferentiated thyroid tumor and the pathologic stage of differentiated thyroid tumor. In contrast to CD82, CD63 mRNA and protein expression was unchanged in all thyroid carcinomas. Benign goiter tissues showed weak expression of CD63. There were no significant correlation between CD63 mRNA/protein expression and any clinical/pathological parameters. Our results support the hypothesis that down-regulation of CD82 expression may reflect an increased in vivo metastatic potential of thyroid cancer cells. CD82 may serve as a prognostic marker of metastasis in thyroid cancer. Constitutive expression of CD63 may indicate that this factor does not play a direct role in thyroid carcinogenesis.

Kangai (KAI)-1 (CD82) is a metastasis suppressor gene, which belongs to the family of tetraspanin proteins. A loss of KAI-1 expression is associated with the advanced stages of many human malignancies. The present study was designed to investigate the expression pattern of KAI-1 in the normal endometrium and uterine tumors and to correlate it with the expression of tumor suppressor protein p53. KAI-1 could be found in the normal endometrium throughout the menstrual cycle. Thirteen of 42 endometrial carcinomas demonstrated moderate KAI-1 expression, but low expression of p53. Twenty-nine of 42 endometrial carcinomas showed reduced or absent KAI-1 expression, which correlated with strong expression of p53 (p < 0.001). There were significant correlations between KAI-1 expression and histological type, e.g., 93% of endometrioid carcinomas displayed a low or moderate immunostaining for KAI-1, whereas nearly all of the serous/clear cell carcinomas were KAI-1 negative (p < 0.001); tumor grading, e.g., 73% of high grade tumors showed no KAI-1 expression (p < 0.001). Most of the investigated uterine sarcomas were negative for KAI-1, whereas they displayed a strong immunostaining for p53. In conclusion, KAI-1 and p53 show inverse expression. The reduced KAI-1 expression may be the result of dysregulated p53 function and could be an important step in the endometrial carcinogenesis.