WO1995007717A9 - Procede pour inhiber la fixation de reticulocytes a l'endothelium par interference avec les interactions vla-4/vcam-1 - Google Patents

Procede pour inhiber la fixation de reticulocytes a l'endothelium par interference avec les interactions vla-4/vcam-1

Info

Publication number
WO1995007717A9
WO1995007717A9 PCT/US1994/010395 US9410395W WO9507717A9 WO 1995007717 A9 WO1995007717 A9 WO 1995007717A9 US 9410395 W US9410395 W US 9410395W WO 9507717 A9 WO9507717 A9 WO 9507717A9
Authority
WO
WIPO (PCT)
Prior art keywords
reticulocytes
vla
sickle
vcam
binding
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Ceased
Application number
PCT/US1994/010395
Other languages
English (en)
Other versions
WO1995007717A1 (fr
Filing date
Publication date
Application filed filed Critical
Priority to AU78352/94A priority Critical patent/AU7835294A/en
Priority to EP94929205A priority patent/EP0720486A1/fr
Publication of WO1995007717A1 publication Critical patent/WO1995007717A1/fr
Publication of WO1995007717A9 publication Critical patent/WO1995007717A9/fr
Anticipated expiration legal-status Critical
Ceased legal-status Critical Current

Links

Definitions

  • the invention relates to the inhibition of interactions between sickle reticulocytes and activated endothelial cells. More particularly, the invention relates to treating human diseases associated with abnormal reticulocyte-endothelium interactions, by inhibiting these interactions.
  • Sickle cell anemia is a disease that is associated with various debilitating complications, including hemolytic anemia, increased infection, ischemic organ damage and episodes of severe pain. Pauling et al., Science 110:543 (1949); and Ingram, Nature 180:326 (1957) teach that this disease and its complications are manifestations of homozygosity for a single amino acid substitution of valine for glutamic acid and in the B6 position of the globin chain.
  • Various researchers including Hahn and Gillespie. Arch. Intern. Med. 39:233 (1927); Harris et al., Arch. Intern. Med. 9_Z:145 (1956); Charache and Conley, Blood 42:25 (1964); and Ham and Castle, Trans. Assoc.
  • the primary pathologic event may be formation of mechanically rigid "sickled" cells with deoxygenation.
  • the sickled cells which cannot normally traverse the microvasculature, occlude small and large blood vessels, causing ischemic organ damage and episodes of pain.
  • others including Hofrichter et al., Proc. Natl. Acad. Sci. USA 71:4864 (1974); Eaton et al., Blood 47:621 (1976); Eaton and Hofrichter, Blood 70:1245 (1987); Mozzarelli et al., Science 237:500 (1987); and Ferrone, Ann. Ny Acad. Sci.
  • capillary obstruction is an uncommon occurrence because hemoglobin S polymerization does not occur immediately after deoxygenation and most erythrocytes traverse the capillary bed before sickling occurs.
  • the fraction of erythrocytes that sickle in the microvasculature and cause obstruction may be determined by other factors that increase capillary transit time or decrease the delay in polymerization.
  • VLA-4 receptor may mediate binding to the inducible endothelial cell adhesion molecule VCAM-1 on cytokine-activated endothelial cells in vitro.
  • VCAM-1 the inducible endothelial cell adhesion molecule
  • the invention demonstrates the mechanism by which a subpopulation of SSRBC bind to activated endothelial cells, thereby leading to increase in transit time and resulting in vascular occlusion in sickle cell disease.
  • the invention further provides a method for inhibiting such binding, thus preventing such vascular occlusion and its resulting complications.
  • This method comprises the step of administering a blocking agent of VLA-4 antigens on the surface of reticulocytes that express VLA-4, including sickle reticulocytes.
  • Various agents can be used to mediate such blocking, including anti-VLA-4 or anti-VCAM-1 antibodies which may optionally be single chain, humanized or chimeric, Fab.
  • the invention satisfies these objects by providing a method for blocking the binding interactions between VLA-4 expressing reticulocytes, including sickle reticulocytes and activated endothelial cells by administering a blocking agent of VLA-4 antigen on the surface of reticulocytes.
  • the invention clearly demonstrates, for the first time, that administration of such blocking agents is sufficient to prevent binding between sickle reticulocytes and activated endothelial cells.
  • Figure 1 shows the results of assays for ⁇ ] integrin expression by cells in venous blood, as described in Example 1.
  • Figure 2 shows the results of assays for reticulocyte counts and ⁇ ] integrin expression by cells in venous blood, as described in Example 2.
  • Figure 3 shows the results of assays for alpha and beta integrin chain by SSRBC. as described in Example 3.
  • Figure 4 shows the results of assays for ⁇ j or ⁇ .4 integrin and glycophorin A expression by venous blood cells, as described in Example 4.
  • Figure 5 shows the results of assays for binding of erythrocytes to TNF- ⁇ - stimulated endothelial cells, as described in Example 5.
  • Figure 6 shows anti-VLA-4 and anti-VCAM-1 antibody inhibition of binding of SSRBC to activated endothelial cells, as described in Example 6.
  • Figure 7 shows the nucleotide sequences encoding the variable regions of the heavy and light chains of anti-VLA-4 murine monoclonal antibody HP 1/2.
  • Panel A is the nucleotide sequence encoding the variable heavy region, with the first nucleotide representing the beginning of the first codon.
  • Panel B is the nucleotide sequence encoding the variable light region, with the first nucleotide representing the beginning of the first codon.
  • Figure 8 shows the nucleotide sequences of Vj-j and VK -encoding regions having CDR-encoding sequences from murine HP 1/2 transplanted therein.
  • Panel A shows the transplanted Vj-j sequence.
  • Panel B shows the transplanted VK sequence.
  • Figure 9 shows the nucleotide sequences encoding the variable regions of the heavy and light chains of the humanized anti-VLA-4 antibody hHPl/2.
  • Panel A is the nucleotide sequence encoding the V j region.
  • Panel B is the nucleotide sequence encoding the V region.
  • Figure 10 shows the expression of adhesion proteins on erythrocytes from an infant with necrotizing enterocolitis.
  • Figure 11 shows the expression of adhesion proteins on erythrocytes from an infant with no known risk of necrotizing enterocolitis.
  • the invention relates to the inhibition of binding interactions between VLA-4 expressing reticulocytes and activated endothelial cells. More particularly, the invention relates to the inhibition of such binding interactions that are involved in the complications associated with human disease, especially sickle cell disease.
  • the invention provides an experimental model for studying the role of increased capillary transit time of sickle reticulocytes in common complications of sickle cell disease, such as vascular occlusion, hemolytic anemia, increased infection, ischemic organ damage and episodes of severe pain.
  • the invention further provides a method for preventing such complications in patients suffering from sickle cell disease.
  • the invention provides an experimental model for studying the role of increased capillary transit time of sickle reticulocytes in the complications associated with sickle cell disease.
  • the mechanism of such increased transit time has been poorly understood.
  • unclear or conflicting reports on the molecules involved in the binding interactions leading to increased transit time have not facilitated the discovery of means for blocking such binding.
  • the invention provides the necessary information for producing an experimental model for studying the involvement of increased capillary transit time in complications arising from sickle cell disease.
  • erythrocyte integrin expression is found in normal controls with reticulocyte counts less than 5%, although a small population of ⁇ ] expressing reticulocytes is detected in blood from non-sickle patients with reticulocyte counts greater than 5%, particularly in neonates with elevated reticulocyte counts.
  • Reticulocyte counts in 25 samples obtained from 22 sickle cell disease patients range from 4% to 45% with a mean and standard deviation of 18 ⁇ 11%. Since ⁇ j expression is limited to the reticulocyte fraction, the total
  • erythrocytes from patients with sickle cell anemia clearly express the 0:4 integrin chain, and this expression is limited to the reticulocyte fraction.
  • the per centage of 0:4 expressing erythrocytes is essentially identical to the per centage of ⁇ ] expressing cells.
  • Expression of ⁇ 1 , cr?, 0:3, 0C5, oc6 or ⁇ v integrin chains is not detected on SSRBC by flow cytometry. Double staining with both anti- ⁇ 4 and anti- ⁇ j shows that these integrins are co-expressed by the same cells.
  • Tumor necrosis factor (TNF) stimulation of endothelial cell monolayers which is associated with VCAM-1 induction, results in markedly increased binding of SSRBC to the monolayers, relative to unstimulated endothelial cell monolayer controls, with only a small number of SSRBC or normal RBC binding to unstimulated human umbilical vein endothelial cells.
  • the increase in SSRBC binding parallels VCAM-1 induction.
  • VLA-4 expressed by sickle reticulocytes in the binding of these cells to activated endothelium is supported by monoclonal antibody (mAb) blocking studies.
  • mAb monoclonal antibody
  • Preincubation of TNF ⁇ -stimulated endothelial cells with anti-VCAM-1 mAb, or of SSRBC with anti-oq. integrin mAb causes up to 75% inhibition of binding of SSRBC to endothelial cells.
  • a control mAb that recognizes intercellular adhesion molecule 1 (ICAM-1) does not inhibit SSRBC binding to activated endothelial cells.
  • the invention provides a method for preventing increased capillary transit time and the resultant vascular occlusion in patients suffering from sickle cell anemia.
  • the irreversibly sickled cell was initially believed to play a central role in the development of vascular complications, more recent clinical studies suggest that the frequency and severity of vascular complications is inversely related to the per centage of these cells and instead is closely correlated with the per centage of young deformable cells (see e ⁇ , Ballas et al., Blood 22:1216 (1988)). Furthermore, gx.
  • the present demonstration that VLA-4/VC AM- 1 interactions are responsible for binding of sickle reticulocytes to activated endothelial cells, and that administration of blocking agents of VLA-4 antigen on the surface of sickle reticulocytes can prevent this binding indicates that the method according to this aspect of the invention can prevent vascular occlusion in patients suffering from sickle cell anemia.
  • the method according to this aspect of the invention comprises the step of administering to a patient suffering form sickle cell anemia a blocking agent of VLA-4 on the surface of sickle reticulocytes.
  • blocking agent of VLA-4 antigens is intended to mean an agent that is capable of interfering with interactions between VLA-4 antigens and ligands on endothelial cells or in the extracellular matrix, including but not limited to either VCAM-1 or fibronectin on the surface of activated endothelial cells.
  • VCAM-1 a monoclonal antibody against VLA-4 as a blocking agent.
  • any agent that can block VLA-4 antigens can be successfully used in the method of the invention.
  • any agent capable of blocking VLA-4 antigens on the surface of sickle reticulocytes is considered to be an equivalent of the monoclonal antibody used in the examples herein.
  • the invention contemplates as equivalents at least peptides, peptide mimetics, carbohydrates and small molecules capable of blocking VLA-4 antigens on the surface of sickle reticulocytes.
  • the blocking agent that is used in the method of the invention to block VLA-4 antigens on the surface of sickle reticulocytes is a monoclonal antibody or antibody derivative.
  • Preferred antibody derivatives include humanized antibodies, chimeric antibodies, single chain antibodies.
  • Monoclonal antibodies against VLA-4 are a preferred blocking agent in the method according to the invention.
  • Human monoclonal antibodies against VLA- 4 are another preferred blocking agent in the method according to the invention. These can be prepared using in vitro-primed human splenocytes, as described by Boerner et al., J. Immunol. 147:86-95 (1991).
  • Another preferred blocking agent in the method of the invention is a chimeric antibody having anti-VLA-4 specificity and a human antibody constant region.
  • These preferred blocking agents can be prepared according to art-recognized techniques, as exemplified in U.S. Patent No. 4.816.397 and in Morrison et al., Proc. Natl. Acad. Sci. USA £:6851-6855 (1984).
  • Yet another preferred blocking agent in the method of the invention is a humanized antibody having anti-VLA-4 specificity.
  • Humanized antibodies can be prepared according to art-recognized techniques, as exemplified in Jones et al.. Nature 321:522 (1986); Riechmann. Nature 332:323 (1988); Queen et al.. Proc. Natl. Acad. Sci. USA £6:10029 (1989): and Orlandi et al. Proc. Natl. Acad. Sci. USA 86:3833 (1989).
  • Those skilled in the art will be able to produce all of these preferred blocking agents, based upon the nucleotide sequence encoding the heavy and light chain variable regions of HP1/2 [SEQ. ID. NOS.
  • Two other preferred blocking agents are single chain antibodies, which can be prepared as described in U.S. Patent No. 4.946,778. the teachings of which are hereby incorporated by reference: and biosynthetic antibody binding sites, which can be prepared as described in U.S. Patent No. 5,091.513, the teachings of which are hereby incorporated by reference.
  • antibody and antibody derivative blocking agents can also act in the method of the invention by binding the receptor for VLA-4, thus acting as agents for blocking the VLA-4 antigen on the surface of sickle reticulocytes, within the meaning of this term for purposes of the invention.
  • antibody and antibody derivative blocking agents according to the invention include embodiments having binding specificity for VCAM-1 or fibronectin, since these molecules are likely to be important in the adhesion between sickle reticulocytes and activated endothelial cells.
  • the blocking agents used in the method according to the invention are not antibodies or antibody derivatives, but rather are soluble forms of the natural binding proteins for VLA-4.
  • These blocking agents include soluble VCAM-1, bifunctional VCAM-1/Ig fusion proteins, or VCAM-1 peptides as well as fibronectin, fibronectin having an alternatively spliced non-type III connecting segment. These blocking agents will act by competing with the activated endothelial cell-bound binding protein for VLA-4 on the surface of sickle reticulocytes.
  • blocking agents are preferably administered parenterally.
  • the blocking agents are preferably administered as a sterile pharmaceutical composition containing a pharmaceutically acceptable carrier, which may be any of the numerous well known carriers, such as water, saline, phosphate buffered saline, dextrose, glycerol, ethanol, and the like, or combinations thereof.
  • a pharmaceutically acceptable carrier such as water, saline, phosphate buffered saline, dextrose, glycerol, ethanol, and the like, or combinations thereof.
  • the blocking agent if an antibody or antibody derivative, will be administered at a dose between about 0.1 mg/kg body weight/day and about 10 mg/kg body weight/day. If non-antibody or antibody analogs are used, administration should be at a molar equivalency to that of the antibody binding sites at the above-described concentrations.
  • the invention provides an experimental model for studying other diseases involving increased reticulocyte counts and/or vaso-occlusion.
  • a condition is severe hereditary spherocytosis. which results in a poorly understood occurrence of leg ulceration and dermatitis.
  • leg ulceration and dermatitis These complications have previously been attributed to increased blood viscosity and reduced tissue oxygen delivery caused by rigid microspherocytes.
  • these attributions are called into question by the observation that the leg ulcers improve or disappear after splenectomy.
  • Splenectomy predictably results in increased erythrocyte counts and circulating microspherocytes. which should aggravate leg ulcers if increased blood viscosity alone is responsible for ulceration and dermatitis.
  • VLA-4 expressing reticulocytes plays a role in the pathophysiology of skin complications in sever hereditary spherocytosis, since splenectomy invariably decreases reticulocyte counts.
  • the invention thus provides a method for studying the potential role of VLA-4 expression by reticulocytes in the complications by administering to the cells blocking agents of VLA-4 antigen on the surface of reticulocytes in in vitro and/or in vivo studies.
  • erythrocytes parasitized by Plasmodium falciparum bind to endothelial cells, and vascular occlusive complications characteristic of malaria are thought to be mediated by this binding (see MacPherson et al., Am. J. Pathol. 119:385 (1985); Oo et al., J. Neuropathol. Exp. Neurol. 46:223 (1987); Pongponrantn et al., Am. J. Trop. Med. Hyg. 44:168 (1991)).
  • the factors that predispose certain individuals to the most catastrophic complication of cerebral malaria are not well understood.
  • Necrotizing enterocolitis in infants is another pathologic condition in which abnormal interactions between endothelial cells and erythrocytes have been pathophysiologically implicated. Abnormal binding of red cells to bowel endothelial cells results in vascular compromise, occlusion, and a clinical syndrome marked by colitis and bowel infarction.
  • Example 1 Expression of ⁇ j ⁇ Integrin by a Subpopulationof Reticulocytes
  • Venous blood samples anticoagulated with EDTA were collected from patients with sickle cell anemia during routine follow up visits to the Grady Memorial Hospital Sickle Cell Center or from normal controls after approval of and according to the guidelines of the Emory University. Georgia Institute of Technology, and Grady Memorial Hospital Institutional Review Boards. A sample of blood with an elevated reticulocyte count was identified and collected at the Grady Memorial Hospital hematology laboratories. The identity and medical history of this patient was not available, although this sample was identified as newborn blood. 10 ⁇ l of whole blood was washed in PBS with 5% fetal calf serum and 0.1% azide (FACS buffer), pelleted and stained with 20 ⁇ l of mAb OKT-5 (anti- CD8. ascites 1 :100 in FACS buffer. ATCC.
  • FACS buffer 5% fetal calf serum and 0.1% azide
  • mAb 4B4 anti-Bi integrin. 100 ⁇ g/ml/ Coutler. Hialeah. FL
  • mAb AP3 anti-B3 integrin. ascites 1 : 100 in FACS buffer, Dr. Peter Neuman. Milwaukee Blood Institute. Milwaukee. WI). Cells were washed, incubated with phycoerythrin conjugated goat and mouse IgG (1 :40 in FACS buffer, Tago, Burlingame, CA) and washed again.
  • reticulocyte containing fraction In order to identify the reticulocyte containing fraction, they were then resuspended in FACS buffer containing 100 ng/ml of thiazole orange (Molecular Probes, Eugene, OR) and allowed to incubate at room temperature protected from light for 30 minutes. Erythrocytes were then examined on a FAC scan flow cytometer
  • Erythrocytes were identified by forward and side scatter and 50,000 events were collected per tube. Reticulocytes were identified by green fluorescence (FL1) and cell surface expression was detected by yellow fluorescence (FL2). The results are shown in Figure 1. Erythrocytes from the normal control (A) (reticulocyte count expressing cells 0.2%) did not express R .
  • erythrocytes from a newborn (reticulocyte count 10%, % ⁇ j expressing cells 1.5%) and from a patient with sickle cell anemia (C) (reticulocyte count 16.5%, % ⁇ - ⁇ expressing cells 4%) both expressed the ⁇ j integrin chain.
  • Example 3 Expression of VLA-4 by SSRBC Washed erythrocytes were incubated within antibody recognizing CD8 or mAb recognizing the integrin chains R> ⁇ . cq (mAb TS 2/7 Dr. Martin Hemler. Dana Farber Cancer Institute. Boston. MA), co (mAb P1E6. Telios Pharmaceuticals. San Diego. CA). 013 (mAb P1B5, Telios), 0:4 (mAb. HP2/1. AMAC. Larrabee. ME). 0:5 (mAb. P1D6. Telios). or oc6 (mAb GoH3, AMAC).
  • HUVEC Human umbilical vein endothelial cells
  • Results represent the mean number of adherent erytfirocytes/mnr ⁇ S.D.
  • the expression of cell surface VCAM-1 on HUVEC stimulated in parallel was determined in parallel by ELISA as in Example 1.
  • the results are expressed as O.D. 450 nm and represent the mean of four data points ⁇ S.D.
  • Example 6 VLA-4 and VCAM-1 Dependence of Binding ofSSRBC to Activated Endothelial Cells
  • TNF ⁇ stimulated endothelial cells were left untreated or preincubated with mAb recognizing VCAM-1 (mAb BBA 6, 10 ⁇ g/ml, R&D Systems. Minneapolis, MN) or ICAM-1 (mAb 84H10, tissue culture supernatants.
  • Sickle RBC were untreated or preincubated with anti- ⁇ 4 (50 ⁇ g/ml. mAb HP2/1. The results are shown in Figure 6.
  • CDRs complementarity determining regions
  • the complementarity determining regions (CDRs) of the light and heavy chains of the anti-VLA-4 monoclonal antibody HP 1/2 were determined according to the sequence alignment approach of Kabat et al., 1991, 5th Ed., 4 vol., Sequences of Proteins of Immunological Interest, U.S. Department of Health and Human Services, NIH, USA.
  • the CDRs of murine HP 1/2 VH correspond to the residues identified in the humanized VH sequences disclosed herein as amino acids 31-35 (CDR1), 50-66 (CDR2) and 99-110 (CDR3), which respectively correspond to amino acids 31-35, 50-65 and 95-102 in the Kabat alignment.
  • the CDRs of murine HP 1/2 VK correspond to the residues identified in the humanized VK sequences disclosed herein as amino acids 24-34 (CDR1), 50-56 (CDR2) and 89-97 (CDR3), and to the same residues in the Kabat alignment.
  • the Kabat NEWM framework was chosen to accept the heavy chain CDRs and the Kabat REI framework was chosen to accept the kappa chain CDRs.
  • Transplantation of the CDRs into the human frameworks was achieved by using Ml 3 mutagenesis vectors and synthetic oligonucleotides containing the HP 1/2 CDR-encoding sequences flanked by short sequences derived from the frameworks.
  • the VH mutagenesis vector, M13VHPCR1 contains the NEWM framework and has been described by Orlandi et al., Proc. Natl. Acad. Sci USA £6:3833-3837 (1989).
  • M13VKPCR2 contains essentially the REI framework and is identical to the M13VKPCR1 vector described by Orlandi et al.. except that there is a single amino acid change from Val to Glu in framework 4.
  • Transplanted product was recovered by PCR and cloned into M13mpl9 for sequencing.
  • the transplanted Vfj sequence [SEQ. ID NO: 3] is shown in Figure 9, panel A.
  • this product encodes the murine amino acids at positions 27-30 and an Arg to Asp change at position 94.
  • the transplanted VK sequence [SEQ. ID NO: 4] is shown in Figure 9, panel B.
  • the entire VH and VK regions of humanized HP 1/2 were cloned into appropriate expression vectors.
  • the appropriate human IgGl, IgG4 or kappa constant region was then added to the vector in appropriate reading frame with respect to the murine variable regions.
  • the vectors were then cotransfected into YB2/0 ray myeloma cells (available from ATCC), which were then selected for the presence of both vectors.
  • ELISA analysis of cell supematants demonstrated that the humanized antibody produced by these cells was at least equipotent with murine HP 1/2.
  • the cell line expressing this humanized antibody was deposited with the ATCC on November 3, 1992 and given accession number CRL 11175.
  • Example 8 Erythrocite adhesive proteins in the pathogenesis of necrotizing enterocolitis in premature infants
  • NEC Necrotizing enterocolitis
  • NEC is the most common serious acquired gastrointestinal tract disorder in the neonatal intensive care unit, responsible for an increasing number of infectious disease associated deaths in low birth weight babies. It has been estimated that NEC affects at least 25,000 newborn children in the United States annually and carries an overall mortality as high as 50%. The continued improvement in the treatment of hyaline membrane disease of the lung is likely to result in an increasing population of infants at risk for developing NEC. NEC is a disease of unknown origin. Despite extensive study, the initiating events and the pathogenesis of NEC have not been established.
  • nectrotizing enterocolitis may be indistinguishable from those of neonatal sepsis and shock, with pneumatosis intestinalis and intrahepatic venous gas the radiographic hallmarks required to confirm the diagnosis.
  • Epidemiological observations emphasize the potential roles of infection, feeding, and local vascular compromise of the GI tract in the pathogenesis of NEC, but it is likely that NEC represents a final common pathway of response of the immature intestine to injury which may be initiated by multiple factors.
  • Prematurity is recognized as the primary risk factor which predisposes infants to NEC.
  • Disturbed hemodynamics associated with hyperviscosity has been implicated as a primary factor in the development of intestinal injury.
  • erythrocytes are merely passive participants in vaso- occlusive disease.
  • Adherence of erythrocytes may initiate microvascular complications in diseases such as sickle cell anemia and diabetes.
  • the primary event may be the binding of adhesion protein expressing reticulocytes to vascular endothelial cells. Vessels partially occluded by bound reticulocytes may be completely occluded by trapping of dense, sickled cells.
  • Reticulocytes may bind to endothelial cells after opsonization with thrombospondin (TSP-1 ) or multimers of von Willebrand factor.
  • TSP-1 thrombospondin
  • reticulocytes are capable of biding directly to endothelial cell surface ligands.
  • Flow cytometric analysis of sickle erythrocytes demonstrated that a percentage of reticulocytes in peripheral circulation of individuals with sickle cell anemia abnormally express specific adhesion receptors, including CD36 and the a4bl integrin complex.
  • adhesion proteins include CD36 and the a4bl integrin complex.
  • the functional importance of these adhesion proteins in erythrocyte-endothelial cell interactions in sickle cell disease and malaria has been extensively examined utilizing a flow adhesion model.
  • Reticulocyte expression of CD36 is important in TSP- 1 mediated binding of erythrocytes to endothelial cells and erythrocyte expression of the a4bl integrin complex can mediate binding to the inducible endothelial cell adhesion molecule VCAM-1 on cytokine activated endothelial cells.
  • the expression of adhesive proteins on population of erythrocytes may result in red cell binding to endothelium and vascular occlusion.
  • the expression of CD36 and the a4bl integrin complex is not limited to reticulocytes of patients with sickle cell anemia.
  • NEC may be a clinical manifestation of this predisposition.
  • the mucosal ischemia proposed as an initiating event in NEC may not be due to an abrupt cessation of mesenteric blood flow, but rather due to vascular compromise induced by the binding of "sticky" erythrocytes to activated endothelium.
  • Adherent erythrocytes may serve as a nidus for platelet activation of the clotting cascade with resultant complete occlusion and bowel necrosis.
  • the localization of pathology to bowel is not adequately understood.
  • NEC often presents after initial feeding.
  • LPS bacterial lipopolysaccharide
  • LPS is a potent induce of VCAM-1 on endothelial cells. It is likely that feeding of premature infants may initially lead to expression of the adhesive ligands on vascular endothelial cells in the bowel, thus providing the ligand pair for the a4bl integrin complex on reticulocytes.
  • VCAM-1 vascular cell adhesion molecule- 1
  • CD36 vascular cell adhesion molecule- 1
  • IAM-1 Intercellular adhesion molecule- 1
  • ICAM-1 can be increased by a variety of cytokines constituing tumor necrosis factor (TNF). interleukin 1 (IL-1). and interferon g (IFN-g). Additionally, two additional adhesion molecules, E-selectin and vascular ell adhesion molecule 1 (VCAM-1), can be induced de novo on endothelial cells. These may serve as ligands for adhesion receptors on the surface of a variety of cells. In fact, all three adhesion receptors have been shown to play a role in red cell binding to endothelial cells under certain clinical circumstances.
  • Red cells from infants at risk for NEC and from normal term infants were examined by flow cytometry for the expressing of adhesion receptors.
  • Erythrocytes from six neonates with congenital heart disease were examined for expression of adhesive ligands. Two of these specimens were obtained from infants with NEC. Blood specimens from four infants felt to be clinically at risk for NEC demonstrated reticulocyte counts ranging from 1 - 4.2% while the two infants with clinical manifestations of NEC expressed reticuloctye counts of 5.5 and 9% respectively (Table 1).

Abstract

L'invention décrit un modèle expérimental pour l'étude du rôle du temps de transit capillaire augmenté des réticulocytes falciformes dans les complications de l'anémie falciforme, ainsi qu'un procédé pour la prévention de ces complications chez les patients atteints d'anémie falciforme. L'invention prévoit également un procédé de traitement de l'entérocolite nécrosante et d'autres maladies caractérisées par des anomalies des interactions entre les érythrocytes et l'endothélium vasculaire.
PCT/US1994/010395 1993-09-15 1994-09-15 Procede pour inhiber la fixation de reticulocytes a l'endothelium par interference avec les interactions vla-4/vcam-1 Ceased WO1995007717A1 (fr)

Priority Applications (2)

Application Number Priority Date Filing Date Title
AU78352/94A AU7835294A (en) 1993-09-15 1994-09-15 Peripheralization of hematopoietic stem cells
EP94929205A EP0720486A1 (fr) 1993-09-15 1994-09-15 Peripherisation des cellules souches hematopoietiques

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
US12222893A 1993-09-15 1993-09-15
US08/122,228 1993-09-15

Publications (2)

Publication Number Publication Date
WO1995007717A1 WO1995007717A1 (fr) 1995-03-23
WO1995007717A9 true WO1995007717A9 (fr) 1995-08-10

Family

ID=22401460

Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/US1994/010395 Ceased WO1995007717A1 (fr) 1993-09-15 1994-09-15 Procede pour inhiber la fixation de reticulocytes a l'endothelium par interference avec les interactions vla-4/vcam-1

Country Status (5)

Country Link
EP (1) EP0720486A1 (fr)
AP (1) AP9400679A0 (fr)
AU (1) AU7835294A (fr)
WO (1) WO1995007717A1 (fr)
ZA (1) ZA947006B (fr)

Families Citing this family (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US11560433B2 (en) * 2016-05-27 2023-01-24 Albert Einstein College Of Medicine Methods of treatment by targeting VCAM1 and MAEA

Family Cites Families (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
IL113261A (en) * 1989-09-01 1996-10-16 Hutchinson Fred Cancer Res Inhibition by peptides of lymphocyte adherence to vascular endothelium utilizing an extracellular matrix receptor-ligand interaction and pharmaceutical compositions containing said peptides
CA2127462C (fr) * 1992-01-13 1999-03-23 Roy R. Lobb Traitement de l'asthme
JPH07506566A (ja) * 1992-02-12 1995-07-20 バイオゲン　インコーポレイテッド 炎症性胃腸病の処置

Similar Documents

Publication Publication Date Title
US12453770B2 (en) CLEC9A binding agents
EP1278543B1 (fr) Methode d'administration d'un anticorps
AU2006301163B2 (en) Compositions and methods for treating proliferative disorders
US8246958B2 (en) Methods of inhibiting alpha-4-dependent interactions with VCAM-1 with anti-VLA-4 antibodies
EP0625912B1 (fr) Traitement de l'inflammation intestinale
CA2237808A1 (fr) Utilisations therapeutiques d'anticorps humanises contre l'alpha-4 integrine
KR20070009637A (ko) 암 세포 증식을 억제하는 항α5β1 항체의 용도
JP2006501283A (ja) ＬＦＡ−１αサブユニット抗体と利用方法
US20250333515A1 (en) Binding agents and methods of use thereof
EP0720486A1 (fr) Peripherisation des cellules souches hematopoietiques
AU2012202895A1 (en) Compositions and methods for treating proliferative disorders
HK1054320B (en) Antibody binding alpha4beta7 integrin and its use to treat inflammatory bowel disease
HK1155394B (en) Antibody binding alpha4beta7 integrin and its use to treat inflammatory bowel disease
HK1155394A (en) Antibody binding alpha4beta7 integrin and its use to treat inflammatory bowel disease
HK1158220A (en) Compositions and methods for treating proliferative disorders
HK1007683B (en) Treatment for inflammatory bowel disease