Using a monoclonal antibody (MAb1.1ASML) raised against a surface glycoprotein of the metastasizing rat pancreatic carcinoma cell line BSp73ASML, cDNA clones have been isolated that encode glycoproteins with partial homology to CD44, a presumed adhesion molecule. In one of the clones, pMeta-1, the epitope marks an additional extracellular domain of 162 amino acids inserted into the rat CD44 protein between amino acid positions 223 and 247 (by analogy to human and murine CD44). The new variants are expressed only in the metastasizing cell lines of two rat tumors, the pancreatic carcinoma BSp73 and the mammary adenocarcinoma 13762NF; they are not expressed in the non-metastasizing tumor cell lines nor in most normal rat tissues. Overexpression of pMeta-1 in the nonmetastasizing BSp73AS cells suffices to establish full metastatic behavior.

Abstract. CD44 is a ubiquitous surface molecule that exists as a number of isoforms, generated by alternative splicing of 10 "variant" exons. Little is known about the expression and function of the variant isoforms, except that certain isoforms may play a role in cancer metastasis. We produced mAbs against CD44 variant regions encoded by exons 4v, 6v, and 9v, by immunizing mice with a fusion protein spanning variant exons 3v to 10v. A comprehensive analysis of human tissues revealed that CD44 variant isoforms were expressed widely throughout the body, principally by epithelial cells. However there was differential expression of CD44 variant exons by different epithelia. Most epithelia expressed exon 9v, but much fewer expressed 6v or 4v. The regions of epithelia that expressed the highest levels of the variant isoforms were the generative cells, particularly the basal cells of stratified squamous epithelium, and of glandular epithelium. CD44 variant isoforms were also expressed differentially by leukocytes, with CD44-9v expressed at very low levels and CD44-6v and 4v virtually absent. However, CIM4-9v and CD44-6v were the main variants that were transiently upregulated on T cells after mitogenic stimulation and on myelomonocytic cell lines by TNFot and IFN~ treatment. Some epithelial cell lines could preferentially upregulate CIM4-6v upon IFN3, incubation. These results show that CD44 variant isoforms are expressed much more widely than first appreciated, and that expression of the variant isoforms on some cell types can be modulated by particular cytokines.C D44 is a widely expressed cell surface glycoprotein that serves as an adhesion molecule in cell-substrate and cell-cell interactions, including lymphocyte homing, hemopoiesis, cell migration, and metastasis (for reviews see Haynes et al., 1991; Underhill, 1992; Gtinthert, 1993;Lesley et al., 1993). CD44 also has other functions that relate to lymphocyte activation (Haynes et al., 1989) and the binding of certain cytokines to endothelium (Tanaka et al., 1993). CD44 is a proteoglycan with an NH2-terminal region that is structurally related to several hyaluronate binding proteins (Underhill, 1992). CD44 is known to bind hyaluronate and collagen (Carter and Wayner, 1988;Aruffo et al., 1990;Culty et al., 1990;Miyake et al., 1990) and a chondroitin sulfated form of CD44 binds fibronectin (Jalkanen and Jalkanen, 1992). Additional ligands for CD44 may well exist. The numerous functions and molecular interactions of CD44 probably relate to its complex structure. In addition to the "standard" 85-95-kD form (CD44s), severalThe Basel Institute for Immunology was founded and is supported by E Hoffman-La Roche Ltd., Basel, Switzerland.Address all correspondence to Dr. Charles R. Mackay, Leukosite Inc., 800 Huntington Ave., Boston, MA 02115, or Dr. Ursula Giinthert, The Basel Institute for Immunology, Grenzaeherstrasse 487, CH-4005 Basel, Switzerland.larger "variant" isoforms exist (CD44v) t that are generated by alternative splicing of at least 10 exons (Screaton et al....

Abstract. Plakoglobin (~-catenin), a member of the armadillo family of proteins, is a constituent of the cytoplasmic plaque of desmosomes, as well as of other adhering cell junctions, and is involved in anchorage of cytoskeletal filaments to specific cadherins. We have generated a null mutation of the plakoglobin gene in mice. Homozygous -/-mutant animals die between days 12-16 of embryogenesis due to defects in heart function. Often, heart ventricles burst and blood floods the pericard. This tissue instability correlates with the absence of desmosomes in heart, but not in epithelial organs. Instead, extended adherens junctions are formed in the heart, which contain desmosomal proteins, i.e., desmoplakin. Thus, plakoglobin is an essential component of myocardiac desmosomes and seems to play a crucial role in the sorting out of desmosomal and adherens junction components, and consequently in the architecture of intercalated discs and the stabilization of heart tissue.

SummaryWe have examined the cell surface molecule CD44, which is attracting interest because of reports that isoforms are associated with metastasis. The prognostic value of CD44 expression has yet to be assessed for a solid tumour.Benign (59) and malignant (primary 61, metastatic 59) gastric tissues were examined with antibodies directed at epitopes common to known CD44 isoforms. Normal mucosa was CD44 negative. In atrophic gastritis and intestinal metaplasia expression was restricted to the epithelial cells of the basal glands and was positively correlated with an increased leucocyte infiltrate and with the expression of HU\ DR by mucosal cells. These observations suggest a role for chronic inflammation in the induction of CD44 expression on benign mucosa. No such association was observed between inflammatory infiltrate and CD44 expression on gastric tumours.CD44 expression, observed in only 49% of primary tumours, was associated with distant metastases at time of diagnosis and, among 31 curatively resected patients, with tumour recurrence (p~ 0-0014) and increased mortality (p = 0-001) during follow-up averaging 17 months. When we used an antibody directed against the CD44 variant exon 9v, we found a good correlation between the expression of total CD44 and of exon 9v containing isoforms, and 9v expression in primary tumours was significantly and positively associated with tumour recurrence and mortality. Lancet 1993; 3 4 2 :1 0 1 9 -2 2

Experimental colitis in mice is characterized by infiltration of activated T helper (Th) cells and macrophages into the lamina propria. Particularly, these cells expressed CD44 variant exon 7 (CD44v7)-containing isoforms. Upregulation of CD44v7 isoforms was induced by CD40 ligation, an inflammation-driving interaction between activated Th cells and macrophages. To define the role of CD44v7 in colitis, mice bearing a targeted deletion for exon v7 were generated. In trinitrobenzene sulfonic acid–induced colitis, wild-type mice developed severe signs of persistent inflammation. Mice lacking CD44v7 initially showed unspecific inflammation, then recovered completely. The pathogenic origin was shown to reside in bone marrow–derived CD44v7+ cells, because adoptive transfer experiments demonstrated an absolute requirement for CD44v7 on hematopoietic cells for maintenance of colitis. Interleukin (IL)-10–deficient mice, which develop a chronic Th1-driven enterocolitis, were crossbred with CD44v6/v7 null mice. In IL-10 × CD44v6/v7 double deficient mice, intestinal inflammation developed only weakly and at an older age. Analysis of cell death in the inflamed lesions revealed that mononuclear cells in the CD44v7 null infiltrates had higher rates of apoptosis than those from wild-type mice. Thus, the region encoded by CD44v7 appears to be essential for survival of effector lymphocytes, resulting in persistence of inflammation.