Expression Of V6 Research Articles

SFPQ Promotes Lung Cancer Malignancy via Regulation of CD44 v6 Expression.

Mesenchymal stem cells (MSCs) contribute to tumor pathogenesis and elicit antitumor immune responses in tumor microenvironments. Nuclear proteins might be the main players in these processes. In the current study, combining spatial proteomics with ingenuity pathway analysis (IPA) in lung non-small cell (NSC) cancer MSCs, we identify a key nuclear protein regulator, SFPQ (Splicing Factor Proline and Glutamine Rich), which is overexpressed in lung cancer MSCs and functions to promote MSCs proliferation, chemical resistance, and invasion. Mechanistically, the knockdown of SFPQ reduces CD44v6 expression to inhibit lung cancer MSCs stemness, proliferation in vitro, and metastasis in vivo. The data indicates that SFPQ may be a potential therapeutic target for limiting growth, chemotherapy resistance, and metastasis of lung cancer.

Abstract 1165: Differential expression of CD44 variants drive the progression, invasion, drug-resistance and stemness characteristics in human oral squamous cell carcinoma

Abstract Purpose: Cluster of differentiation 44 (CD44) is a cell-surface glycoprotein and plays role in the progression and severity of oral squamous cell carcinoma (OSCC). Here we report differential function of CD44 variants in OSCC progression. Materials and methods: The expression of CD44 standard (CD44s) and variants (CD44v4, CD44v6); the activation of signaling pathways (MAPK, PI3K); the cell viability; and the MMP-9/-2 activity were assessed using RT-PCR, immunohistochemistry, Western blotting, MTT assay and gelatin zymography. These experiments were carried out using fresh human OSCC tissue specimens, including adjacent normal, noninvasive (N0), invasive tumor samples (N1-3) and chemo-radiation resistant tumor samples (RCRT). We also used OSCC cell lines, including parental (SCC9/SCC4) and Cisplatin-resistant (CisR-SCC9/-SCC4) to confirm our observation with human tissues. Knock down of CD44 variants or inactivation of MAPK/PI3K pathways was achieved for in vitro analysis in OSCC cells. Summary: Differential expressions of CD44 variants (v4and v6) were associated with overall oral cancer aggressiveness. CD44v4 expression was positively correlated with the activation of MAPK pathway causing chemoresistance. Conversely, CD44v6 expression and activation of PI3K-Akt caused invasiveness of OSCC. Finally, an overlapping role of two major signal transduction pathways such as MAPK and PI3K, that impinge on the variants/ isoforms expressions of a single gene i.e. CD44 that leads to the OSCC aggressiveness. Conclusion: Collectively, these results established that CD44 variants (v4and v6) expression driven by MAPK/PI3K are associated with the overall progression, drug-resistance, invasion, and stemness characteristics leading to the aggressiveness of OSCC. Hence targeting these pathways may be exploited for treating OSCC. Keywords: OSCC, CD44s, CD44v4/6, MAPK, PI3K, Notch, Chemoresistance, Invasion/migration, stemness, aggressiveness of OSCC. Note: This abstract was not presented at the meeting. Citation Format: Tanushree Kashyap, Siddavaram Nagini, Ajay Rana, Rajakishore Mishra. Differential expression of CD44 variants drive the progression, invasion, drug-resistance and stemness characteristics in human oral squamous cell carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 1165.

Chronic Fibrosing Vasculitis in the Setting of Still Disease

Chronic Fibrosing Vasculitis in the Setting of Still Disease

CD147 and Ki-67 overexpression confers poor prognosis in squamous cell carcinoma of oral tongue: A tissue microarray study

Squamous cell carcinoma of the oral tongue (SCCOT) exhibits high risk for recurrence and regional metastasis even after surgical resection. We assessed the clinicopathologic and prognostic significance of a group of functionally related biomarkers. We used a tissue microarray consisting of SCCOT from 32 patients for this study. These patients were treated at the University of Texas MD Anderson Cancer Center from 1995 to 2008. Biomarker expression levels were examined by immunohistochemistry and graded semiquantitatively to determine their prognostic significance. CD147 and Tp63 expressions were significantly associated with a higher T stage and Ki-67 labeling index, as well as a shorter overall survival (OS) rate. Expression of Tp63 associated positively with poorly differentiated histology. There was significant association of Tp63 with the expression levels of CD147 and Glut-1. Glut-1 overexpression was marginally associated with a higher T stage. There was no prognostic significance of CD44 v6 expression in SCCOT. SCCOT with CD147 overexpression in combination with high Ki-67 labeling index had poor OS. CD147 and Ki-67 overexpression is associated with aggressive disease with poor prognosis in SCCOT.

Correlation Between Expression of Cell Adhesion Molecules CD44v6 and E-cadherin and Lymphatic Metastasis in Non-small Cell Lung Cancer

To explore the relationship between expressions of cell adhesion molecules CD44 v6 and E-cadherin (E-cad) and lymphatic metastasis in non-small cell lung cancer (NSCLC). Eighty- seven tissue samples obtained from patients with primary NSCLC were collected in our hospital from Dec., 2007 to Dec., 2012, and the expressions of CD44 v6 and E-cad gene proteins in these samples were detected by immunohistochemical method. In the tissue without lymphatic metastasis, the positive expression rate of CD44 v6 was significantly lower, whereas the normal expression rate of E-cad was notably higher than that with lymphatic metastasis (55.6% vs. 78.4%, 47.2% vs. 21.6%), and both differences had statistical significance (P<0.05). Besides, CD44 v6 and E-cad expressions had a significant correlation in the NSCLC tissue with lymphatic metastasis (P<0.05). The positive expression of CD44 v6 and abnormal expression of E-cad may play a very important role in promoting lymphatic metastasis of NSCLC, with synergistic effect. Hence, detection of CD44 v6 and E-cad expressions is conductive to judging the lymphatic metastasis in NSCLC.

THAP11, a novel binding protein of PCBP1, negatively regulates CD44 alternative splicing and cell invasion in a human hepatoma cell line

THAP11 is an essential factor involved in ES cell pluripotency and cell growth. Here, we identified THAP11 as a novel physiological binding partner of PCBP1. In HepG2 cells, THAP11 overexpression inhibited CD44 v6 expression and cell invasion. However, when deleting the binding domain with PCBP1 or endogenous PCBP1 was knocked down, THAP11 failed to inhibit CD44 v6 expression, indicating that THAP11 regulates CD44 v6 expression through interacting with PCBP1. In HCC patients, the expression of THAP11 mRNA significantly correlated with PCBP1 mRNA expression. Our results suggest a novel role of THAP11 in CD44 alternative splicing and hepatoma invasion.Structured summary of protein interactions:THAP11 physically interacts with PCBP1 by anti bait coimmunoprecipitation (View interaction) THAP11 physically interacts with PCBP1 by anti tag coimmunoprecipitation (View Interaction: 1, 2) THAP11 and PCBP1 colocalize by fluorescence microscopy (View interaction)

Abstract 3480: CD44 V6 and V9 isoforms act in resistance to Fas-mediated apoptosis in breast cancer initiating cells

Abstract Breast cancer initiating cells (BCIC) are CD44+/CD24 low/ESA+ cells derived from breast tumors that highly tumorigenic and resistant to apoptosis and to chemotherapy. To determine anti-apoptotic mediator (s) to both Fas death receptor signaling and chemotherapeutic agents induced apoptosis in BCIC, we detest the expression of V5, V6 and V9 of CD44 isoforms in BCIC derived from breast cancer cell lines MCF-7, MDA-MB-231, SUM159 and SUM1315, and assess their effects on both Fas death receptor signaling and chemotherapeutic agents induced apoptosis. Methods: 1). We measured relative expression of the various CD44 isoforms in parental and BCIC populations using quantitative real time-PCR. 2).We then determined which CD44 isoforms play a role in BCIC's resistance to apoptosis. SUM159 was chosen to use due to the overexpression of all CD44 isoforms. 3). After isolation of cells expressing the BCIC markers from SUM159 cells (CD44+/CD24 low/ESA+), we knocked down CD44 V5, V6 and V9 by siRNA in SUM159 BCIC cells and challenged them with increasing doses of the chemotherapeutic agent doxorubicin and FasL. 4).Tissue microarrays of human breast cancer (www.tissue-array.com) were also employed to assess association between the CD44 V9 expression and clinical parameters such as nodal metastasis, histological status, ER status, PgR status, HER2 status and survival in 40 cases of primary breast cancers. Results: 1).CD44 isoforms V5, V6 and V9 are differentially expressed in BCIC compared to their parental cell lines. V9 is markedly overexpressed in all 4 BCIC tested. 2). BCIC cells treated with siRNA targeting isoforms V5, V6 and V9 demonstrated no significant effect on doxorubicin IC50 when compared to controls. The existence of CD44 V9 and CD44 V6 in breast cancer cells confer resistance to apoptosis induced by FasL, knockdown of CD44 V6 resulted in 39% increase in caspase 3/7 activity when compared to controls while knockdown of CD44V9 caused a 49% increase. Further study showed that this is through the inhibition of the extrinsic apoptotic pathway by suppressing caspases-8 activation, thereby blocking downstream apoptotic events. 3). 40 cases of primary breast cancers provided assessable cores paired with corresponding uninvolved tissue from the same patient. The analysis showed a tendency between CD44V6 expression and HER2 status. We found that approximately 72.8% of HER2-positive patients express CD44V9 compared to 48.3% of HER2-negative patients expressing CD44V9. It did not reach statistically significance probably because of small size of samples. Conclusions: Loss of expression of the V6 and V9 but not V5 isoforms in SUM159 BCIC resulted in an increased sensitivity to apoptosis in response to the Fas L but not to the chemotherapeutic agent doxorubicin. These findings establish a role for CD44 V6 and V9 isoforms in aptoptotic resistance of BCIC and suggest that they may serve as targets for therapeutic intervention. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 3480. doi:1538-7445.AM2012-3480

Role of Hyaluronan-Mediated CD44 Signaling in Head and Neck Squamous Cell Carcinoma Progression and Chemoresistance

Head and neck squamous cell carcinoma (HNSCC) is an aggressive malignancy that may involve the oral cavity, pharynx, larynx, and paranasal sinuses. The mechanisms of tumor progression underlying the clinical behavior of HNSCC remain unclear. CD44 comprises a family of transmembrane receptors that can give rise to multiple CD44 variant isoforms. Hyaluronan (HA), a major extracellular matrix component is the primary ligand for CD44 receptors. HA and CD44 signaling play an important role in HNSCC progression. Several CD44 variant isoforms (including v3-, v6-, and v10-containing isoforms) are associated with advanced disease, possibly through unique growth factor interactions with binding domains in the inserted variant regions of the cytoplasmic domain of CD44. In HNSCC, HA mediates the formation of a complex including CD44 and the epidermal growth factor receptor (EGFR) which is overexpressed in a large proportion of HNSCCs. Downstream effectors under EGFR regulation are activated, promoting promote cell growth and tumor survival. The leukemia-associated Rho-guanine nucleotide exchange factor (LARG) also associates with CD44 and EGFR to promote several Ras and RhoA pathway effectors, leading to cell migration, growth, and tumor survival. The secretion of matrix metalloproteinases, necessary for tumor cell invasion, is also regulated by these HA/CD44-mediated pathways. Finally, EGFR-mediated pathways play major roles in the HA/CD44 promotion of chemoresistance in HNSCC. Understanding HA/CD44-mediated signaling pathways may lead to improved treatment of HNSCC.

Quantitative assessment of expression of cell adhesion molecule (CD44) splice variants: CD44 standard (CD44s) and v5, v6 isoforms in oral leukoplakias: An immunohistochemical study

The aim of this study was to semiquantitatively analyze the immunohistochemical expression pattern of CD44 standard (CD44s) and variant (CD44v) isoforms in leukoplakias using a panel of monoclonal antibodies recognizing epitopes of CD44s and of the variant exons v5 and v6. To evaluate the efficacy of CD44s and CD44 v5, v6 immunoexpression as possible molecular markers in detecting high-risk leukoplakias when screening for this oral precancer. Samples of oral leukoplakia (40 cases) and of normal mucosa (10 cases) were evaluated. Oral leukoplakia was graded into: hyperkeratosis without dysplastic change (8 cases), mild dysplasia (13 cases), moderate dysplasia (10 cases), and severe dysplasias (9 cases). Expression of CD44s,v5, v6 was analyzed by immunohistochemistry in a semiquantitative manner. Three areas of epithelium were scored B, S, and C, i.e., stratum basale, stratum corneum, and stratum spinosum, respectively in leukoplakias. Scoring of all specimens followed a two-parameter system, which implemented percentage of positive cells and staining intensities. Statistical analyses for each parameter of all groups and normals, mean, and standard deviation were calculated by using computer software package EPISTAT. In normal epithelium CD44s, CD44v5, and CD44v6 were expressed as membranous proteins localized on the surface of epithelial cells. Both basal and spinous layer of epithelia expressed strong positive staining of CD44s, v5, v6 which then gradually faded into the negative staining of the superficial keratin layer. Profile of CD44s and v5 revealed that the mean levels of stratum B, S, and C in normal cases were comparable to the study cases and by Student 't' test P>0.05 not significant. There was, however, a statistically significant decrease in the expression of v6 with increasing grades of dysplasias when compared with normal mucosa. Among CD44s and its variant isoforms,v5, v6, in this study, variant isoform v6 may serve as a marker in detecting high-risk leukoplakias.

Characterization of the expression of variant and standard CD44 in prostate cancer cells: Identification of the possible molecular mechanism of CD44/MMP9 complex formation on the cell surface

CD44 is a glycosylated adhesion molecule and osteopontin is one of its ligand. CD44 undergoes alternative splicing to produce variant isoforms. Our recent studies have shown an increase in the surface expression of CD44 isoforms (sCD44 and v4-v10 variant CD44) in prostate cancer cells over-expressing osteopontin (PC3/OPN). Formation of CD44/MMP9 complex on the cell surface is indispensable for MMP9 activity. In this study, we have characterized the expression of variant CD44 using RT-PCR, surface labeling with NHS-biotin, and immunoblotting. Expression of variant CD44 encompassing v4-v10 and sCD44 at mRNA and protein levels are of the same levels in PC3 and PC3/OPN cells. However, an increase in the surface expression of v6, v10, and sCD44 in PC3/OPN cells suggest that OPN may be a ligand for these isoforms. We then proceeded to determine the role of sCD44 in MMP9 activation. Based on our previous studies in osteoclasts, we hypothesized that phosphorylation of CD44 has a role on its surface expression and subsequent activation of MMP9. We have prepared TAT-fused CD44 peptides comprising unphosphorylated and constitutively phosphorylated serine residues at positions Ser323 and Ser325. Transduction of phosphopeptides at Ser323 and Ser323/325 into PC3 cells reduced the surface levels of CD44, MMP9 activity, and cell migration; but had no effect on the membrane localization of MMP9. However, MMP9 knock-down PC3 cells showed reduced CD44 at cellular and surface levels. Thus we conclude that surface expression of CD44 and activation of MMP9 on the cell surface are interdependent.

R425 – Association of CD44 v6 and v10 Isoforms In HNSCC Progression

Problem The CD44 family of receptors includes multiple variant isoforms, several of which have been linked to tumor progression. The objective of this study was to investigate the role of CD44 v6- and v10-containing isoforms in head and neck squamous cell carcinoma (HNSCC) progression. Methods A laboratory investigation utilizing a panel of 13 HNSCC cell lines and clinical tissue specimens from a series of 82 patients with HNSCC was performed. Results Immunohistochemical analysis of 82 primary HNSCC tumors and 24 metastatic lymph nodes revealed expression of CD44 v6 and v10 variants. A greater proportion of metastatic lymph nodes vs. primary tumors demonstrated strong expression of CD44 v6 (18/24 vs. 26/82) and CD44 v10 (14/24 vs. 16/82), while expression of standard CD44 was not significantly different in metastatic lymph nodes and primary tumors (10/24 vs 60/82). In addition, greater than expected expression of CD44 v10 was found among 14 specimens obtained from patients previously treated with chemoradiation. To further analyze the role of these CD44 variants in HNSCC progression, HNSCC cell lines were selected on the basis of their CD44 v6 and v10 expression. A HNSCC cell line without CD44 expression and cells of the same variant-expressing cell lines transfected with short-hairpin RNA for CD44 were used as controls. Compared to controls, HNSCC cells expressing CD44 v6-containing isoforms demonstrated increased hyaluronan-mediated tumor cell growth and migration. Furthermore, HNSCC cells expressing CD44 v10-containing isoforms demonstrated increased hyaluronan-mediated cisplatin resistance. Conclusion CD44 v6- and v10-containing isoforms are associated with HNSCC progression. Significance CD44 v6- and v10-containing isoforms may be useful tumor markers and targets for HNSCC therapy. Support This work was supported by an AAO-HNS/AHNS Young Investigator Award.

A Complex of EpCAM, Claudin-7, CD44 Variant Isoforms, and Tetraspanins Promotes Colorectal Cancer Progression

High expression of EpCAM and the tetraspanin CO-029 has been associated with colorectal cancer progression. However, opposing results have been reported on CD44 variant isoform v6 (CD44v6) expression. We recently noted in rat gastrointestinal tumors that EpCAM, claudin-7, CO-029, and CD44v6 were frequently coexpressed and could form a complex. This finding suggested the possibly that the complex, rather than the individual molecules, could support tumor progression. The expression of EpCAM, claudin-7, CO-029, and CD44v6 expression was evaluated in colorectal cancer (n = 104), liver metastasis (n = 66), and tumor-free colon and liver tissue. Coexpression and complex formation of the molecules was correlated with clinical variables and apoptosis resistance. EpCAM, claudin-7, CO-029, and CD44v6 expression was up-regulated in colon cancer and liver metastasis. Expression of the four molecules did not correlate with tumor staging and grading. However, coexpression inversely correlated with disease-free survival. Coexpression was accompanied by complex formation and recruitment into tetraspanin-enriched membrane microdomains (TEM). Claudin-7 contributes to complex formation inasmuch as in the absence of claudin-7, EpCAM hardly associates with CO-029 and CD44v6 and is not recruited into TEMs. Notably, colorectal cancer lines that expressed the EpCAM/claudin-7/CO-029/CD44v6 complex displayed a higher degree of apoptosis resistance than lines devoid of any one of the four molecules. Expression of EpCAM, claudin-7, CO-029, and CD44v6 by themselves cannot be considered as prognostic markers in colorectal cancer. However, claudin-7-associated EpCAM is recruited into TEM and forms a complex with CO-029 and CD44v6 that facilitates metastasis formation.

Evaluation of CD44 variant 6 expression and clinicopathological factors in pulmonary metastases from colon carcinoma

Although relatively little is known about the molecular mechanisms underlying tumor progression, recently CD44 glycoproteins and the c-Met receptor tyrosine kinase have been identified as potentially important components of the metastatic cascade. CD44 is a family of transmembrane receptors generated from a single gene by alternative splicing and differential glycosylation. Important biological processes involving CD44 glycoproteins include cell adhesion, lymphocyte homing, hematopoiesis, tumor progression and metastasis. The precise mechanism via which CD44 promotes tumorigenesis have not yet been elucidated. We evaluated the expression of adhesion molecule CD44 variant 6 in pulmonary metastases from colorectal carcinomas and its correlation with clinicopathological parameters. Twenty patients were randomly selected from the patients who had undergone a resection of pulmonary metastasis from colorectal cancer. Formalin-fixed, paraffin-embedded archival specimens of tumor tissues and adjacent normal mucosa from these patients were the subjects of the present study. Immunoreactivity for CD44 was quantified. Specimens were considered positive if almost 25% of the neoplastic cells were stained. CD44 v6 expression was related to the interval between colon resection and metastases diagnosis, the number of pulmonary metastases, and the survival after lung resection. No statistical correlation was found between CD44 v6 positivity and disease-free interval after colon resection, number of metastases or 2-year survival after lung resection. Probably CD44 v6 is necessary and sufficient to confer metastatic potential to carcinoma cells increasing the migration capacity and participating in invasion via changes in adhesion to the extracellular ligands, but is not necessary to modify the clinical history of the metastases. Therefore the evaluation of CD44 v6 expression in lung metastases does not influence the therapeutic scheme.

Expression of CD44v5 and -v6 in Barrett's Carcinoma Is Not Increased Compared to That in Nondysplastic Barrett's Mucosa

Barrett's adenocarcinoma currently shows the highest increase in the incidence of all malignant tumors. Reliable molecular markers to identify Barrett's patients at risk are still missing. Our own results demonstrate that the expression of CD44v6 correlates with the development of dysplasia in colorectal neoplasms. Therefore, we examined the expression of CD44 variants v5 and v6 in normal esophageal mucosa, nondysplastic Barrett's mucosa, and Barrett's carcinoma. mRNA from biopsy specimens of patients with Barrett's esophagus ( n = 19) or Barrett's carcinoma ( n = 15) and patients without esophageal diseases (controls; n = 9) were extracted and used as templates for cDNA synthesis. CD44 variants were detected by RT–PCR with primers hybridizing with CD44 sequences up- and downstream of variable exons. CD44v6 expression was found in 36 of 56 biopsy specimens (64%) of nondysplastic Barrett's mucosa, in 100% of squamous epithelium, and in none of the gastric mucosa specimens. Eleven of 15 specimens (73%) of Barrett's carcinoma tested positive for v6 expression. The identification of v5 expression did not give additional information. There was no correlation between CD44v5 or -v6 expression and staging or grading of the tumors. Expression of CD44v5 and -v6 seems to be independent of the development of cancer in Barrett's mucosa.

Expression of CD44 and its isoforms in the fetal neuroblasts.

CD44 is a polymorphic transmembrane glycoprotein that exists as multiple isoforms resulting from alternative splicing and posttranslational modifications. Enhanced expression of CD44 has been correlated to the tumorigenicity and metastatic behavior in different malignant tumors. In contrast, human neuroblastomas exhibit an inverse correlation between CD44 expression and tumor malignancy. To determine whether there is a CD44 silencing in sympatho-adrenal precursors as a possible explanation for the down-regulation of CD44 in neuroblastomas, the expression of standard CD44H and v6, v7, v7v8, or v10 isoforms was analyzed by immunohistochemistry in human adrenal glands of 14- to 20-week-old gestational age fetuses. All of the fetal neuroblasts localized in the adrenal gland parenchyma and migrating from the sympathetic nerve chain into the fetal adrenal cortex strongly expressed CD44H but none of the CD44 isoforms could be detected in these cells. In contrast, a strong expression of CD44v7 and v6 was detected in the fetal adrenal cells. These results indicate that, as for many other cell types, the CD44H expressed by fetal neuroblasts may contribute to controlling their migration into the adrenal medulla and that the down-regulation of CD44H in neuroblastoma cells should be explained by mechanisms other than the fetal suppression of CD44H expression in their normal counterparts.

Overexpression of CD44 Variant 6 in Human Endometrial Cancer and Its Prognostic Significance

Objective. We evaluated the prognostic significance of CD44 isoform CD44 V6 in endometrial cancer.Method. Immunohistochemistry was used to determine the expression of CD44 V6 in 78 randomly selected endometrial cancer specimens.Results. CD44 V6 was detected in 53% (41/78) of the tumor samples. Clinicopathological evaluation revealed an inverse correlation with CD44 V6 expression and conventional poor prognosticators such as lower segment involvement, nuclear and structural grade, myometrial invasion, serosal involvement, lymph-vascular space invasion, and adnexal metastasis. Furthermore, the CD44 V6-negative group was found to be associated have a higher recurrence rate and a shorter disease-free survival.Conclusion. These findings indicate that absence of CD44 V6 expression in cases of endometrial cancer might be coupled with a more aggressive course.

Expression of CD44 isoforms in infiltrating lobular carcinoma of the breast.

Expression of CD44 has been shown to correlate with the progression and prognosis of some malignant tumors. The aim of this study was to evaluate the expression of CD44 isoforms in infiltrating lobular carcinomas and analyse their potential as prognostic indicators. A panel of 39 tumors were examined for their expression of membranous and cytoplasmic CD44s, v3, v5, v6, v7 and v3-10 in the infiltrating cells, by immunohistochemical staining. The protein positive tumors showed membranous and/or cytoplasmic staining with all antibodies used except for CD44v7, which only displayed cytoplasmic staining. Cytoplasmic expression of CD44v3 (P = 0.014) and membranous expression of v6 (P = 0.039) were significantly associated with alveolar, classical/alveolar carcinomas and mucinous/alveolar carcinomas. Furthermore, in alveolar, classical/alveolar and mucinous/alveolar carcinomas, cytoplasmic staining of CD44v5 was correlated with lymph node negative patients (P = 0.048), whereas membranous v5 was correlated with lymph positive patients (P = 0.048). In classical, classical/trabecular and trabecular carcinomas expression of membranous CD44s was significantly correlated with lymph node status (P = 0.042).

Isoform Expression of CD44 Adhesion Molecules, Bcl-2, p53 and Ki-67 Proteins in Lung Cancer

CD44, belongs to the cell adhesion molecule family and is expressed on cell surfaces in several isoforms which are generated by alternative splicing of messenger RNA. These splice variants have been shown in several cancer cell types and are thought to be involved in tumor progression. The aim of the current study was to evaluate the expression of selected CD44 variants on lung cancer cells of various histology and to compare these with other markers of tumor spread. Surgical samples of primary lung carcinoma of various histology were subjected to alkaline phosphatase-anti-alkaline phosphatase complex immunohistochemistry using a panel of monoclonal antibodies: anti-CD44 v5, v6, v7/8, v10, anti-Ki-67, anti-Bcl-2 and anti-p53. Positive cells were scored in a semiquantitative way. The patients were subdivided into groups with and without metastases, as found during surgery. All CD44 variants tested could be demonstrated on lung cancer cells, but the incidence of particular isoforms varied, depending on lung cancer histology. In general, CD44 expression was highest in squamous cell tumors and lowest in anaplastic small cell carcinomas. Squamous cell cancers had high expression of v5 and v6 variants, while in anaplastic large cell and small cell carcinomas v10 was abundant. When Ki-67, Bcl-2 and p53 protein expression was compared to the incidence of CD44 variants, coincidence was found for v10 only. Most of the cases positive for v10 were also Ki-67 positive (p = 0.0146). In 12 cases with metastases, tumor cells had high v6 and Ki-67 expression, but these data were not significant compared to cases without metastases. Overall, these data suggest that v5 and v6 variants are of significance in squamous cell lung carcinoma, presumably in the promotion of metastasis, while in anaplastic small cell or large cell cancers only v10 expression seems to correlate with proteins associated with tumor growth and progression.

Reduced expression of CD44v3 variant isoform is associated with unfavorable outcome in non–small cell lung carcinoma

The expression of CD44 standard form (CD44s) and variant isoforms v3 and v6 was analyzed in 233 resected non–small cell lung carcinoma (NSCLC) specimens by immunohistochemistry (IHC), and the mRNA status of CD44v3 and CD44v6 in a cohort of samples was determined by in situ hybridization (ISH) and further confirmed by reverse transcriptase polymerase chain reaction (RT-PCR). The expression of CD44s, CD44v3, and CD44v6 was correlated with clinicopathologic variables and survival. The expression of CD44v3 and v6 was reduced in 97% and 90% of the adenocarcinomas and in 86% and 74% of the large cell/anaplastic carcinomas, respectively, as compared with squamous cell carcinomas, where they were reduced in 53% and 51% of the cases ( P = .0001 and P = .004 for v3 and v6). The corresponding values for CD44s were 92%, 70%, and 51%, respectively ( P = .011). The reduced CD44s and CD44v6 expression was associated with lymph node metastases ( P = .03 and P = .005, respectively) and the reduced expression of CD44s also with advanced stage ( P = .04). Recurrences during the follow-up were more often found within the tumors showing reduced expression of CD44v3 ( P = .04). Combining ISH and IHC results showed that CD44v3 and v6 mRNA were not always processed into protein, suggesting a regulation disturbance posttranscriptionally since malignant transformation of cells has occurred. In survival analyses, the reduced expression of CD44s and CD44v3 was associated with a shortened disease-free survival ( P = .04 and P = .01, respectively). In multivariate analysis, CD44v3 retained its independent prognostic value ( P = .03). These results emphasize the value of CD44, and especially the v3 variant isoform in the behavior of NSCLC. H UM P ATHOL 31:1088-1095.

Immunohistochemical expression of the CD44 variant 6 in colorectal adenocarcinoma.

The CD44 variant 6 (CD44 v6) has been postulated to be involved in both carcinogenesis and tumor progression. In the present study, CD44 v6 was stained immunohistochemically in 63 colorectal cancer tissues to assess significance of CD44 v6 in the carcinogenesis and progression of colorectal carcinoma. None of the normal colonocytes showed an expression of CD44 v6. CD44 v6 expression was very strong in 24, moderate in 13, and negative in 26 tumor tissues. A negative or moderate expression of CD44 v6 was significantly associated with a larger tumor size (P < 0.05) and invasion through the bowel wall (P < 0.01). There was no correlation between the expression of CD44 v6 and gross type, histologic differentiation, lymph node involvement, liver metastasis, and clinical stage of the disease. The present study showed that the expression of CD44 v6 was characteristic of neoplastic changes in the colonocytes and that a diminished expression of CD44 v6 was associated with the penetration of colorectal cancer through the bowel wall, but not with either lymph node or distant metastasis.