The Clinical Spectrum of Leukocyte Adhesion Deficiency (LAD) III due to Defective CalDAG-GEF1
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- Kilic, S.S. & Etzioni, A. J Clin Immunol (2009) 29: 117. doi:10.1007/s10875-008-9226-z
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Leukocyte adhesion deficiency (LAD) type III is a rare syndrome characterized by severe recurrent infections, leukocytosis, and increased bleeding tendency. All integrins are normally expressed yet a defect in their activation leads to the observed clinical manifestations.
Materials and Methods
Less than 20 patients have been reported world wide and the primary genetic defect was identified in some of them. Here we describe the clinical features of patients in whom a mutation in the calcium and diacylglycerol-regulated guanine nucleotide exchange factor 1 (CalDAG GEF1) was found and compare them to other cases of LAD III and to animal models harboring a mutation in the CalDAG GEF1 gene.
The hallmarks of the syndrome are recurrent infections accompanied by severe bleeding episodes distinguished by osteopetrosis like bone abnormalities and neurodevelopmental defects.
More than 25 years ago a defect in the crucial adhesion cascade of leukocytes was described and designated as leukocyte adhesion deficiency (LAD), characterized by severe recurrent bacterial and fungal infections, delayed separation of the umbilical cord, defective wound healing without pus formation and markedly increased leukocyte count. LAD is an autosomal recessive genetic disorder caused by mutations in the ITGB2 gene which encodes for the β subunit of the integrin (CD18), leading to the inability of leukocytes to adhere to the blood vessel endothelium . Several hundreds of patients have been described worldwide and in most of them no or markedly reduced expression of CD18 on leukocyte surface was noted. Rarely, mutations in the gene lead to a non-functional but expressed CD18 and these cases are referred to as LAD1/variant .
Ten years later a second syndrome, LAD II, has been described . This rare condition (less than ten patients reported so far) is characterized by milder infectious episodes, severe psychomotor and growth retardation, the Bombay blood phenotype and marked leukocytosis. The syndrome is caused by a general defect in fucose metabolism due to mutations in the gene FUCT1, which encodes for the specific transporter of fucose from the cytoplasma to the Golgi apparatus where the incorporation of fucose to various glycoprotein occurs . The defect leads to the absence of Sialyl Lewis X (CD15), the ligand for the selectin, essential for the first step in the adhesion cascade, the rolling phase .
We previously described three LAD III patients singled by transcriptional and translational defects in calcium and diacylglycerol-regulated guanine nucleotide exchange factor 1 (CalDAG-GEF1), an important Rap1 and integrin activator in hematopoietic cells . In the current report the clinical characteristics of our patients are highlighted and compared with those reported in other, genetically distinct, LAD III cases and in recently published animal models with CalDAG-GEF1 deficiency.
Materials and Methods
Case 1 and 2
Laboratory Features of LAD-III Patients
Pt 1 YB
Pt 2 AB
Pt 3 SA
Pt 4 MK
Age at onset
Age at diagnosis
Umbilical cord detachment (day)
Mild motor retardation
Recurrent severe infections
Increase bone density
Laboratory data (earliest recorded)
Plt (× 109/L)
Platelet aggregation response to ADP, epinephrine, and collagen
Lymphocyte subsets %, (absolute mm3)
Neutrophil gate (%)
Died at 15 months
Died at 7 months
Died at 1.5 years
A healthy appearing male infant was delivered at 38 weeks gestation by vaginal delivery. He was the second product of second degree-related parents (Fig. 2). His birth weight was 2,450 g. At delivery, mucosal and skin bleeding was observed. He received erythrocyte and platelet infusions. His clinical features are presented in Table I. Laboratory investigation showed leukocytosis and Glanzmann disease like thrombasthenia. As of 2 weeks of age, he was frequently hospitalized because of bacterial and fungal sepsis episodes and died at 7 months of age. Quantitative PCR analysis showed very low levels of CalDAG-GEF1.
Leukocyte arrest at target endothelial sites is nearly exclusively mediated by integrin receptors expressed on all circulating hematopoetic cells. These maintain their integrins (β1 and β2) in a generally non-adhesive state that can be rapidly activated subject to interactions with various agonists, predominantly chemoattractants and chemokines presented by the endothelium . Likewise, platelets maintain their major fibrinogen receptor, the integrin αIIbβ3 in an inactive conformation, which is converted by chemokines which bind to GPCR . The small GTPase Rap-1 has been implicated as the major intracellular activator of integrins both in leukocytes and platelets. A crucial molecule for Rap-1 activation is CalDAG-GEF1, which was found to be defective in our patients. It belongs to the CalDAG-GEF/Ras GRP family of intracellular signaling molecule with a guanine nucleotide exchange factor (GEP) domain that catalyzes the exchange of GTP for GDP bond to Rap1 .
The significance of vascular integrins has been demonstrated by numerous studies in murine knock out models as well as in rare human LAD syndromes.
In LAD I severe, life-threatening bacterial infections occur subject to mutations in the gene encoding β2 integrin. Another genetic disorder, called Glanzmann’s thrombasthenia is associated with mutations in the gene encoding the β3 integrin, resulting in dysfunctional platelet aggregation with severe bleeding tendency .
Leukocyte Adhesion Deficiency Syndromes
Number of cases reported
Less than 10
Recurrent severe infections
Delayed separation of the umbilical cord
↓↓↓ or absent
Integrin defect in
Primary genetic defect
Ca1DAGGEF1 4/13 of cases
We have previously shown that Rap1 activation is defective in some cases of LAD III  leading to integrin dysfunction. The high incidence of infections in this syndrome is mainly due to the leukocyte adhesion dysfunction. It has been also shown that defects in NADPH oxidase, observed in LAD III , may also contribute to susceptibility to infections.
Recently Kuijpers et al.  described nine patients with LAD III (referred to them as LAD I/variant). All of them originated from the Anatolia area in Turkey, while our patients are from Eastern Turkey. While recurrent infections, bleeding tendency, leukocytosis, and defective integrins activation were reported for these patients, their clinical course seems to be milder and the majority of patients remained alive even without bone marrow transplantation and no mutations in CalDAG-GEF1 were found . Thus, as previously proposed  different defects in the intracellular activation of both platelets and leukocytes integrins can lead to LAD III syndromes with differences in the clinical presentation .
CalDAG-GEF1 appears to be specifically expressed within the hematopoietic system as well as in neurons, especially in the stratum of the basal ganglia . In addition to the hematopoietic dysfunctions, other non-hematological features mark our patients.
Although the development milestones were found to be retarded in all patients they all showed normal cognitive and social skills. Two of them had epileptic activity and required antiepileptic treatment. The epileptic activity was not correlated to brain compression due to bleeding or subdural effusion. CalDAG-GEF1 protein levels in patients’ nerves were never tested and we cannot thus establish whether the expression level of the GEF in the nervous system is equally reduced as in hematopoietic cells or to a lesser degree subject to differences in mRNA processing and translation between nerve and blood cells. Whether reduced CalDAG-GEF1 levels in patient neutrons could affect neuron function remains to be determined. Future studies are needed to address multiple potential roles of CalDAG-GEF1 as a Rap-1 effector and integrator of integrin activation events in neurons.
Notably, all four patients had increased bone density on X-ray similar to that seen in patients with osteopetrosis. Mutations in several genes, including the gene encoding receptor activator of nuclear factor-kb ligand (RANKL), a crucial osteoclast growth factor involved in osteoclast differentiation and function, were identified in patients with osteopetrosis. Osteoclasts are hematopeitic derived cells of myeloid origin . It has been shown that once RANKL starts osteoclast differentiation the αvβ3 integrin on osteoclasts mediates key matrix interactions essential for osteoclast maturation . The large similarity between the platelet integrin GpIIb3 and the osteoclast αvβ3 suggests that activation of the alfavbeta3 integrin is both Rap-1 and CalDAG-GEF1 dependent and is therefore defective in the LAD III cases identified by us. Loss of osteoclast-matrix adhesion may impair osteoclast bone resorbing activities and explain the increased bone density measured in our LAD III cases.
Recently, several animal models of LAD III have been reported. Knock out CalDAG-GEFI deficient mice exhibit the same platelet and neutrophil adhesion defect . Still no spontaneous bleeding or apparent infections in these mice were observed . It should be noted that the mice were kept in a restricted pathogen-free lining space. Furthermore no abnormalities in bone density on X-ray were observed in the CalDAG-GEF1 knock out mice (Graybiel AM., personal communication).
Mutations in CalDAG-GEF1 gene were also reported in calf and dogs [20, 21]. Interestingly, in these two animal models, only platelet aggregation defect was found while leukocyte count was normal and no neutrophil adhesion defect was observed (Boudreaux MK., personal communication). As in our patients, the main cause of death was due to bleeding episodes. It appears that CalDAG GEF1 is indispensable for platelets aggregation in all species, while several other Rap-1 pathways may substitute for loss of CalDAG-GEF1 in leukocytes in different species. Interestingly, murine lymphocytes lack CalDAG-GEF1 in contrast to human lymphocytes  and thus, the dependence on this GEF for normal leukocyte integrin activation may be particularly high in humans.
We would like to thank Prof. Alon for critically reviewing the paper and to Prof. Rechavi’s group for helping in the genetic analysis of the families.
Recently we found that these patients have also mutation in Kindlin 3 (Blood Accepted).