Cluster of Differentiation 44 (CD44)
The adhesion molecule CD44 is a transmembrane glycoprotein that plays a role in lymphocyte activation, recirculation, and balance, extracellular matrix adhesion, angiogenesis, cell proliferation, cell differentiation and cell migration and as a receptor for hyaluronic acid. These various biological properties are very useful for the physiological activity of normal cells, but are also associated with the pathological activity of tumor cells. Increased expression of CD44 is associated with poor prognosis of several malignancies such as lung cancer, ovarian cancer, breast cancer, colorectal cancer, gastrointestinal neuroendocrine tumors, and others. In recent years, scientists have focused more on the relationship between CD44 and blood malignancies.
They assumed that CD44 plays an important role in normal myelopoiesis because anti-CD44 antibodies are fundamentally altered in in vitro myelopoiesis in long-term bone marrow culture. In the context of leukemia, studies have shown that it is possible to stop differentiation in some leukemic cells by binding (ligation) to CD44 with specific antibodies, indicating the possibility of recent advances in targeted therapy for CD44 differentiation in leukemia therapy.
CD44 structure and function
The human CD44 gene is located in the forearm on chromosome 11 which consists of at least 20 exons that rotate 50 kilobases of the DNA chain. This gene consists of 2 groups of exons, consisting of exons 1-5 and 16-20 which are expressed simultaneously in all cell types and are standard forms. 10 exon variables (6-15) can be randomly connected via standard exons at the insertion site between exons 5 and 16. Transcription for this gene is via alternative linkage complexes that are generated in functionally distinct isoforms. This type of isoform is divided into peptide units consisting of the extracellular region of the protein, according to the alternative splicing theory that will result in more than 1000 variations of the CD44 molecule overall.
The smallest CD44 molecule, which lacks an intact structure, is standard CD44 (CD44s) which is mainly expressed by basic lymphohematopoietic cells. CD44 is also known as hematopoietic (CD44H). CD44s are formed from the distal extracellular region (including the “ligand-binding sites”), the proximal membrane region, the transmembrane ring region and the cytoplasmic end (figure 1). Including multiple glycosylated sites and chondroitic acid binding sites, the extracellular regions can bind different extracellular matrices. The N-terminal is the region primarily responsible for hyaluronic acid binding.
The transmembrane region is very characteristic of most single-membrane glycoproteins including sites that can interact with hexadecanoic acid. The cytoplasmic sequence can be phosphorylated as a substrate for protein kinase C. As a GTP binding protein, CD44 can bind to GDP substrate and has GTP enzymatic activity so that it can increase the interaction between CD44 and ankirins.2,3,4
The CD44 variant (CD44v) is predominantly expressed on epithelial cells, characterized by a broadly glycosylated amino acid site and a chondroitic acid binding site. Connected continuously or in a septal direction, the exon combinations have different variables and different CD44 molecules. Currently, there are more than 10 kinds of CD44v in several cell lines detected by polymerase chain reaction. 1.2
Alternative splicing is the basis for distinguishing the structure and function of these proteins and may be related to tumor metastasis. After immunological activation, CD44v on T lymphocytes and other leukocytes is transiently regulated. The CD44 isoform includes the last 3 exon products in the region variable (CD44v8-v10), also known as epithelial CD44 or CD44E which is specifically expressed on epithelial cells. The longest CD44 isoform is expressed by a fusion of 8 exons of the variable region (CD44v3-v10) present in keratinocytes. pMeta-1 (CD44v4-v7) and pMeta-2 (CD44v6,v7) are referred to as metastatic CD44 because their cDNA causes transfection, the possibility of metastasis in nonmetastatic tumor cells in mice.
Figure 2 shows the CD44 exon arrangement consisting of constant-region exons and exon variations. Figure 1) epithelial CD44 (CD44v8-v10), 2) keratinocyte CD44 (CD44v3-v10), 3) pMeta-1 (CD44v4-v7), pMeta-2 (CD44v6, v7).
CD44 is a Cell Adhesion Molecules (CAMs) which is very important to maintain the stability of the tissue structure. In dynamic situations, cells alter their cell and cell matrix interactions through the properties of CAMs that function as expression modifiers. CAM expression is normally tightly regulated, by controlling cell proliferation, mobility, differentiation, and survival. As an adhesion molecule (binder), CD44 plays a role in many ways for example having a role in maintaining the balance of lymphocyte production, T lymphocyte activation, assisting the attachment between fibroblasts, lymphocytes and extracellular material (ECM) such as hyaluronic acid, chondroitin sulfatase, fibronectin, laminin and collagen. . It also plays a role in signal transmission, updating the composition of interstitial tissue, helping drug absorption and drug sensitivity. In addition, it also plays a role in forming pseudopods and plays a role in cell migration
Lymphocyte extravasation at the site of inflammation consists of several steps, namely recognition and rolling, adhesion, diapedesis, and migration. Rolling of T lymphocytes on the endothelium can be mediated by CD44. The interaction between CD44 on T lymphocytes and hyaluronan on the endothelial surface is sufficient to initiate the rolling process required for extravasation to tissues. This rolling interaction together with chemokines facilitates adhesion, mediated by integrins and their receptors on endothelial cells that trigger diapedesis and migration to sites of inflammation and infection. 2
Hyaluronan synthase produces and secretes hyaluronan. This release of hyaluronan interacts multivalently with CD44 to induce or stabilize signaling domains in the plasma membrane. This signaling domain contains receptor tyrosine kinases (ErbB2 and EGFR), other receptors such as TGFβR1, and non-receptor kinases (Scr family), which are oncogenic pathways such as cell proliferation by MAP kinase and P13 kinase. adapter proteins mediate the interaction of CD44 with surface effectors such as RhoA, Rac1, and Ras. Heparan sulfate activates the receptor tyrosine kinase, the c-Met receptor which induces cytoskeletal changes to promote cell motility and invasion, by means of actin filaments joining the CD44 tail via the ERM family or ankirin. All of these activities were influenced by hyaluronan produced by tumor cells (figure 4).
Most epithelial cells, hematopoietic cells, and non-epithelial cells generally produce CD44s. The standard CD44 isoform is produced by all mature blood cell types, mostly by bone marrow precursor mononuclear cells and all CD34+ HPC. The level of each expression varies depending on the origin of the hematopoietic cells and the degree of differentiation. For example, CD44 was high in monocytic cells, moderate levels in polymorphonuclear (PMN) and CD34+ HPC and low in erythrocytes and platelets. CD44-6v and CD44-9v isoforms have been detected in monocytes, macrophages, lymphocytes, and dendritic cells.2,3
CD44 is produced at very high levels in many tumor types, and is associated with the biologic properties of tumors including tumorogenesis, growth, metastasis, and prognosis. It is a definite indicator of tumor severity and disease activity and is also called a metastasis-associated protein. In fact, the spread of metastases involves interactions between tumor cells and endothelial cells, which suggests that CD44 may be involved in the process of metastatic expansion. There are several opinions regarding the relationship between lymphocyte activation and tumor cell metastases including strong invasion, reversible adhesion to cell migration, accumulation and proliferation in lymph nodes and sometimes release into the circulatory system and peripheral tissues. Seiter et al assumed that this similarity might be in view of the general effects of CD44v6, suggesting that the mechanism of CD44v6 in tumor metastasis is similar to that of lymphocyte activation. Tumor cells can cause hidden lymphocytes because overexpression of CD44v6 is eliminated through recognition and killed by the immune system, so tumors can invade lymph nodes and metastasize more easily.2,3
CD44 expression in hematological malignancies
Recent findings have shown that CD44 is overexpressed by hematopoietic cells and is involved in interactions between the bone marrow stromal layer and cells of hematopoietic origin and this overexpression is associated with poor prognosis of some haematological malignancies.
CD44 and Acute Myeloid Leukemia
Bendall et al. compared the expression of the CD44 variant in normal bone marrow, peripheral blood and CD34+ cells of hematopoietic origin generated in blasts from 30 patients with acute myeloid leukemia (AML). Normal bone marrow, peripheral blood and CD34+ origin cells were negative in all variants measured using flow cytometry where exons v3,v4,v5,v6 and v7 were expressed in AML cases. RT-PCR and Southern revealed a more complex pattern of expression of exon variants in leukemic samples compared to normal hematopoietic cells. These data demonstrate a significant increase in the complexity of CD44v expression in cells in AML patients through the surface expression of several variants of the CD44 protein. They suggest that further research should be carried out directly on how the interaction of the leukemia blast with the bone marrow microenvironment changes and its diagnosis, prognosis and therapeutic opportunities. Florian et al analyzed the expression of target antigens on CD34+/CD38- cells in patients with AML, myelodysplastic syndrome, chronic myeloid leukemia, and systemic mastocytosis. Using multi-color flow cytometry, they reported that CD44 was expressed in all patients and that neoplastic stem cells in some myeloid neoplasms produced similar phenotypes including target antigens CD13, CD33 and CD44.4.
CD44 and Acute Lymphocytic Leukemia
It has been reported that high levels of the CD44 variant suggest a poor prognosis in patients with acute lymphocytic leukemia (ALL). Magyarosy et al analyzed CD44v6 expression in the bone marrow of 16 pediatric patients with ALL using immunocytochemistry. They found that the CD44v6 protein epitope was expressed by leukemic cells in 6 cases of ALL patients, particularly in the moderate or high risk group (except in 1 case). The picture is very similar to the observations made in some adult cancer patients and shows that: possible association with poor prognosis. The potential of CD44v6 expression on leukemic cells as a prognostic indicator in pediatric ALL patients should be further evaluated in larger clinical trials. Using oligonucleotide microarray analysis, Oh et al. reported that CD44 was associated with poor prognosis after analyzing tissue infiltration parameters in 86 patients with ALL. 4
The role of CD44 in chemotherapy
The role of CD44 in chemotherapy has recently been widely studied. One of them is the administration of doxorubicin. Doxorubicin binds to HNA (π-hyluronan nanocarrier) which then CD44 as a hyaluronan receptor assists the endocytosis process of the HNA-DOX complex into target cells. With a low pH environment protons and doxorubicin are released and cause cell nucleus apoptosis (figure 6).7
CD44 is a transmembrane glycoprotein that plays a role in lymphocyte activation, recirculation, and balance, extracellular matrix adhesion, angiogenesis, cell proliferation, cell differentiation and cell migration as well as a receptor for hyaluronic acid.
– This gene consists of 2 groups of exons, consisting of exons 1-5 and 16-20 which are expressed simultaneously in all cell types and are standard forms. 10 variable exons (6-15) can be connected which are variations of the shape of CD44. Alternative splicing is the basis for distinguishing the structure and function of these proteins and is associated with tumor metastasis.
– The level of each expression of CD44 varies depending on the origin of the hematopoietic cells and the degree of differentiation, but the expression of CD44 is lower in normal tissue than in tumor tissue. CD44 as a hyaluronan receptor plays an important role in chemotherapy.
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- Liu J, Jiang G. CD44 and Hematologic Malignancies. The Chinese Society of Immunology. 2006. 359-63
- Hertweck MK, Erdfelder F, Kreuzer KA. CD44 in hematological Neoplasia. University of Cologne. 2011.493-508.
- Sneath RJ, Mangham DC. The Normal Structure and function of CD44 and its role in neoplasia. Journal Mol Pathol. 1998.191-200.
- Toole BP. Hyaluronan-CD44 Interactions in Cancer: Paradoxes and Possibilities. Clin Cancer Res in aacjournals. 2009. 7462-8.
- Knudson W, Knudson CB,The Hyaluronan Receptor, CD44. Exp Cell Res. 1999.
- Jang E et al., π-hyluronan nanocarier for CD44 Targetted and pH-boosted aromatic Drug Delivery. Jurnal Mater Chem Biol. 2013.5686-5693.