Aneurysmal bone cyst and osteoblastoma: an extremely rare combination in the pediatric spine.

STUDY DESIGN: Case report. OBJECTIVE: To report the clinical and imaging findings of a patient with the extremely rare association of aneurysmal bone cyst and osteoblastoma in the cervical spine. To our knowledge, only three cases have been reported in the published literature in children under 16 years of age with this condition in the cervical spine. METHODS: The patient's history, physical examination, imaging findings, and management with a complete 4-year medical history, surgical intervention and radiological follow-up are reported. RESULTS: A 4-year 11-month-old boy was diagnosed with aneurysmal bone cyst in association of osteoblastoma and was treated with CT-guided intralesional injection calcitonin and methylprednisolone. During the course of intralesional therapy, a pathological fracture of C2 was produced. Subsequently, a widened intralesional excision and instrumented fusion from occiput to cervical spine (C0-C4) was performed. CONCLUSION: The association of aneurysmal bone cyst and osteoblastoma in spine is extremely rare. Although both are benign lesions, in the cervical location, complete removal of the tumors is challenging. Wide resection with reconstruction of the segments for stability associated with adjuvant treatment with calcitonin and corticosteroids provides a good option.

Chiari 1: Is decompression always necessary previous to scoliosis surgery?

The association between Chiari 1 malformation and scoliosis is well known in the literature. Prevalence has increased after the advent of magnetic resonance imaging. In children with this association, prophylactic suboccipital decompression prior to scoliosis correction is a common surgical procedure although the rationale for this surgical management and whether not performing it may lead to spinal cord injury has not been clearly elucidated. We conducted a systematic review of the literature with the aim to obtain strong data to support the hypothesis that it is safe to proceed with scoliosis correction without prior prophylactic suboccipital decompression for Chiari 1 in an asymptomatic population. Using the Prisma methodology, we analyzed 3250 studies published between 1972 and 2018. Only four studies met the inclusion criteria. None of the studies had a level of evidence high enough to recommend prophylactic decompression previous to correction of the spinal deformity.

COVA1-18 neutralizing antibody protects against SARS-CoV-2 in three preclinical models.

One year into the Coronavirus Disease 2019 (COVID-19) pandemic caused by Severe Acute Respiratory Syndrome coronavirus 2 (SARS-CoV-2), effective treatments are still needed 1-3 . Monoclonal antibodies, given alone or as part of a therapeutic cocktail, have shown promising results in patients, raising the hope that they could play an important role in preventing clinical deterioration in severely ill or in exposed, high risk individuals 4-6 . Here, we evaluated the prophylactic and therapeutic effect of COVA1-18 in vivo , a neutralizing antibody isolated from a convalescent patient 7 and highly potent against the B.1.1.7. isolate 8,9 . In both prophylactic and therapeutic settings, SARS-CoV-2 remained undetectable in the lungs of COVA1-18 treated hACE2 mice. Therapeutic treatment also caused a dramatic reduction in viral loads in the lungs of Syrian hamsters. When administered at 10 mg kg - 1 one day prior to a high dose SARS-CoV-2 challenge in cynomolgus macaques, COVA1-18 had a very strong antiviral activity in the upper respiratory compartments with an estimated reduction in viral infectivity of more than 95%, and prevented lymphopenia and extensive lung lesions. Modelling and experimental findings demonstrate that COVA1-18 has a strong antiviral activity in three different preclinical models and could be a valuable candidate for further clinical evaluation.

Role of chain pairing for the production of functional soluble IA major histocompatibility complex class II molecules.

Structural studies of cellular receptor molecules involved in immune recognition require the production of large quantities of the extracellular domains of these glycoproteins. The murine major histocompatibility complex (MHC) class II-restricted response has been extensively studied by functional means, but the engineering and purification of the native, empty form of the most-studied murine MHC class II molecule, IA, has been difficult to achieve. IA molecules, which are the murine equivalent of human histocompatibility leukocyte antigen-DQ molecules, have a low efficiency of chain pairing, which results in poor transport to the cell surface and in the appearance of mixed isotype pairs. We have engineered soluble IA molecules whose pairing has been forced by the addition of leucine zipper peptide dimers at their COOH-terminus. The molecules are secreted "empty" into the extracellular medium and can be loaded with single peptide after purification. These IA molecules have been expressed in milligram quantity for crystallization as well as for activation of T cells and measurement of MHC class II-T cell receptor interactions.

Molecular recognition of lipid antigens by T cell receptors.

The T cell antigen receptor (TCR) mediates recognition of peptide antigens bound in the groove of major histocompatibility complex (MHC) molecules. This dual recognition is mediated by the complementarity-determining residue (CDR) loops of the alpha and beta chains of a single TCR which contact exposed residues of the peptide antigen and amino acids along the MHC alpha helices. The recent description of T cells that recognize hydrophobic microbial lipid antigens has challenged immunologists to explain, in molecular terms, the nature of this interaction. Structural studies on the murine CD1d1 molecule revealed an electrostatically neutral putative antigen-binding groove beneath the CD1 alpha helices. Here, we demonstrate that alpha/beta TCRs, when transferred into TCR-deficient recipient cells, confer specificity for both the foreign lipid antigen and CD1 isoform. Sequence analysis of a panel of CD1-restricted, lipid-specific TCRs reveals the incorporation of template-independent N nucleotides that encode diverse sequences and frequent charged basic residues at the V(D)J junctions. These sequences permit a model for recognition in which the TCR CDR3 loops containing charged residues project between the CD1 alpha helices, contacting the lipid antigen hydrophilic head moieties as well as adjacent CD1 residues in a manner that explains antigen specificity and CD1 restriction.

Anti-peptide antibodies detect steps in a protein conformational change: low-pH activation of the influenza virus hemagglutinin.

At low pH, the hemagglutinin (HA) of influenza virus undergoes an irreversible conformational change that potentiates its essential membrane fusion function. We have probed the details of this conformational change using a panel of 14 anti-HA-peptide antibodies. Whereas some antibodies reacted equally well with both the neutral and low-pH HA conformations, others reacted to a significantly greater extent with the low-pH form. The locations of the peptides recognized by the latter antibodies in the three-dimensional HA structure indicated regions of the protein that change in response to low pH. Moreover, kinetic experiments suggested steps in the conformational change. In addition to their relevance to membrane fusion, our results show that anti-peptide antibodies can be used to study some types of biologically important protein conformational changes.

Crystal structure of Sar1-GDP at 1.7 A resolution and the role of the NH2 terminus in ER export.

The Sar1 GTPase is an essential component of COPII vesicle coats involved in export of cargo from the ER. We report the 1.7-A structure of Sar1 and find that consistent with the sequence divergence of Sar1 from Arf family GTPases, Sar1 is structurally distinct. In particular, we show that the Sar1 NH2 terminus contains two regions: an NH2-terminal extension containing an evolutionary conserved hydrophobic motif that facilitates membrane recruitment and activation by the mammalian Sec12 guanine nucleotide exchange factor, and an alpha1' amphipathic helix that contributes to interaction with the Sec23/24 complex that is responsible for cargo selection during ER export. We propose that the hydrophobic Sar1 NH2-terminal activation/recruitment motif, in conjunction with the alpha1' helix, mediates the initial steps in COPII coat assembly for export from the ER.

Erythropoietin receptor activation by a ligand-induced conformation change.

Erythropoietin and other cytokine receptors are thought to be activated through hormone-induced dimerization and autophosphorylation of JAK kinases associated with the receptor intracellular domains. An in vivo protein fragment complementation assay was used to obtain evidence for an alternative mechanism in which unliganded erythropoietin receptor dimers exist in a conformation that prevents activation of JAK2 but then undergo a ligand-induced conformation change that allows JAK2 to be activated. These results are consistent with crystallographic evidence of distinct dimeric configurations for unliganded and ligand-bound forms of the erythropoietin receptor.

Global efforts in structural genomics.

A worldwide initiative in structural genomics aims to capitalize on the recent successes of the genome projects. Substantial new investments in structural genomics in the past 2 years indicate the high level of support for these international efforts. Already, enormous progress has been made on high-throughput methodologies and technologies that will speed up macromolecular structure determinations. Recent international meetings have resulted in the formation of an International Structural Genomics Organization to formulate policy and foster cooperation between the public and private efforts.

Structural evidence for induced fit as a mechanism for antibody-antigen recognition.

The three-dimensional structure of a specific antibody (Fab 17/9) to a peptide immunogen from influenza virus hemagglutinin [HA1(75-110)] and two independent crystal complexes of this antibody with bound peptide (TyrP100-LeuP108) have been determined by x-ray crystallographic techniques at 2.0 A, 2.9 A, and 3.1 A resolution, respectively. The nonapeptide antigen assumes a type I beta turn in the antibody combining site and interacts primarily with the Fab hypervariable loops L3, H2, and H3. Comparison of the bound and unbound Fab structures shows that a major rearrangement in the H3 loop accompanies antigen binding. This conformational change results in the creation of a binding pocket for the beta turn of the peptide, allowing TyrP105 to be accommodated. The conformation of the peptide bound to the antibody shows similarity to its cognate sequence in the HA1, suggesting a possible mechanism for the cross-reactivity of this Fab with monomeric hemagglutinin. The structures of the free and antigen bound antibodies demonstrate the flexibility of the antibody combining site and provide an example of induced fit as a mechanism for antibody-antigen recognition.

Crystal structures of two viral peptides in complex with murine MHC class I H-2Kb.

The x-ray structures of a murine MHC class I molecule (H-2Kb) were determined in complex with two different viral peptides, derived from the vesicular stomatitis virus nucleoprotein (52-59), VSV-8, and the Sendai virus nucleoprotein (324-332), SEV-9. The H-2Kb complexes were refined at 2.3 A for VSV-8 and 2.5 A for SEV-9. The structure of H-2Kb exhibits a high degree of similarity with human HLA class I, although the individual domains can have slightly altered dispositions. Both peptides bind in extended conformations with most of their surfaces buried in the H-2Kb binding groove. The nonamer peptide maintains the same amino- and carboxyl-terminal interactions as the octamer primarily by the insertion of a bulge in the center of an otherwise beta conformation. Most of the specific interactions are between side-chain atoms of H-2Kb and main-chain atoms of peptide. This binding scheme accounts in large part for the enormous diversity of peptide sequences that bind with high affinity to class I molecules. Small but significant conformational changes in H-2Kb are associated with peptide binding, and these synergistic movements may be an integral part of the T cell receptor recognition process.

Emerging principles for the recognition of peptide antigens by MHC class I molecules.

Class I major histocompatibility complex (MHC) molecules interact with self and foreign peptides of diverse amino acid sequences yet exhibit distinct allele-specific selectivity for peptide binding. The structures of the peptide-binding specificity pockets (subsites) in the groove of murine H-2Kb as well as human histocompatibility antigen class I molecules have been analyzed. Deep but highly conserved pockets at each end of the groove bind the amino and carboxyl termini of peptide through extensive hydrogen bonding and, hence, dictate the orientation of peptide binding. A deep polymorphic pocket in the middle of the groove provides the chemical and structural complementarity for one of the peptide's anchor residues, thereby playing a major role in allele-specific peptide binding. Although one or two shallow pockets in the groove may also interact with specific peptide side chains, their role in the selection of peptide is minor. Thus, usage of a limited number of both deep and shallow pockets in multiple combinations appears to allow the binding of a broad range of peptides. This binding occurs with high affinity, primarily because of extensive interactions with the peptide backbone and the conserved hydrogen bonding network at both termini of the peptide. Interactions between the anchor residue (or residues) and the corresponding allele-specific pocket provide sufficient extra binding affinity not only to enhance specificity but also to endure the presentation of the peptide at the cell surface for recognition by T cells.

Crystal structure of a gammadelta T cell receptor ligand T22: a truncated MHC-like fold.

Murine T10 and T22 are highly related nonclassical major histocompatibility complex (MHC) class Ib proteins that bind to certain gammadelta T cell receptors (TCRs) in the absence of other components. The crystal structure of T22b at 3.1 angstroms reveals similarities to MHC class I molecules, but one side of the normal peptide-binding groove is severely truncated, which allows direct access to the beta-sheet floor. Potential gammadelta TCR-binding sites can be inferred from functional mapping of T10 and T22 point mutants and allelic variants. Thus, T22 represents an unusual variant of the MHC-like fold and indicates that gammadelta and alphabeta TCRs interact differently with their respective MHC ligands.

Crystal structures of an antibody to a peptide and its complex with peptide antigen at 2.8 A.

The three-dimensional structures of an antibody to a peptide and its complex with the peptide antigen have been determined at 2.8 A resolution. The antigen is a synthetic 19-amino acid peptide homolog of the C helix of myohemerythrin (Mhr). The unliganded Fab' crystals are orthorhombic with two molecules per asymmetric unit, whereas the complex crystals are hexagonal with one molecule per asymmetric unit. The Fab' and the Fab'-peptide complex structures have been solved independently by molecular replacement methods and have crystallographic R factors of 0.197 and 0.215, respectively, with no water molecules included. The amino-terminal portion of the peptide sequence (NH2-Glu-Val-Val-Pro-His-Lys-Lys) is clearly interpretable in the electron density map of the Fab'-peptide complex and adopts a well-defined type II beta-turn in the concave antigen binding pocket. This same peptide amino acid sequence in native Mhr is alpha-helical. The peptide conformation when bound to the Fab' is inconsistent with binding of the Fab' to native Mhr, and suggests that binding of the Fab' to conformationally altered forms of the native Mhr or to apo-Mhr. Immunological mapping previously identified this sequence as the peptide epitope, and its fine specificity correlates well with the structural analysis. The binding pocket includes a large percentage of hydrophobic residues. The buried surfaces of the peptide and the antibody are complementary in shape and cover 460 A2 and 540 A2, respectively. These two structures now enable a comparison of a specific monoclonal Fab' both in its free and antigen complexed state. While no major changes in the antibody were observed when peptide was bound, there were some small but significant side chain and main chain rearrangements.

Glycosylation and the immune system.

Almost all of the key molecules involved in the innate and adaptive immune response are glycoproteins. In the cellular immune system, specific glycoforms are involved in the folding, quality control, and assembly of peptide-loaded major histocompatibility complex (MHC) antigens and the T cell receptor complex. Although some glycopeptide antigens are presented by the MHC, the generation of peptide antigens from glycoproteins may require enzymatic removal of sugars before the protein can be cleaved. Oligosaccharides attached to glycoproteins in the junction between T cells and antigen-presenting cells help to orient binding faces, provide protease protection, and restrict nonspecific lateral protein-protein interactions. In the humoral immune system, all of the immunoglobulins and most of the complement components are glycosylated. Although a major function for sugars is to contribute to the stability of the proteins to which they are attached, specific glycoforms are involved in recognition events. For example, in rheumatoid arthritis, an autoimmune disease, agalactosylated glycoforms of aggregated immunoglobulin G may induce association with the mannose-binding lectin and contribute to the pathology.

Routes to catalysis: structure of a catalytic antibody and comparison with its natural counterpart.

The three-dimensional structure of a catalytic antibody (1F7) with chorismate mutase activity has been determined to 3.0 A resolution as a complex with a transition state analog. The structural data suggest that the antibody stabilizes the same conformationally restricted pericyclic transition state as occurs in the uncatalyzed reaction. Overall shape and charge complementarity between the combining site and the transition state analog dictate preferential binding of the correct substrate enantiomer in a conformation appropriate for reaction. Comparison with the structure of a chorismate mutase enzyme indicates an overall similarity between the catalytic mechanism employed by the two proteins. Differences in the number of specific interactions available for restricting the rotational degrees of freedom in the transition state, and the lack of multiple electrostatic interactions that might stabilize charge separation in this highly polarized metastable species, are likely to account for the observed 10(4) times lower activity of the antibody relative to that of the natural enzymes that catalyze this reaction. The structure of the 1F7 Fab'-hapten complex provides confirmation that the properties of an antibody catalyst faithfully reflect the design of the transition state analog.

Crystal structure of mouse CD1: An MHC-like fold with a large hydrophobic binding groove.

CD1 represents a third lineage of antigen-presenting molecules that are distantly related to major histocompatibility complex (MHC) molecules in the immune system. The crystal structure of mouse CD1d1, corresponding to human CD1d, at 2.8 resolution shows that CD1 adopts an MHC fold that is more closely related to that of MHC class I than to that of MHC class II. The binding groove, although significantly narrower, is substantially larger because of increased depth and it has only two major pockets that are almost completely hydrophobic. The extreme hydrophobicity and shape of the binding site are consistent with observations that human CD1b and CD1c can present mycobacterial cell wall antigens, such as mycolic acid and lipoarabinomannans. However, mouse CD1d1 can present very hydrophobic peptides, but must do so in a very different way from MHC class Ia and class II molecules.

Crystal structure of the principal neutralization site of HIV-1.

The crystal structure of a complex between a 24-amino acid peptide from the third variable (V3) loop of human immunodeficiency virus-type 1 (HIV-1) gp 120 and the Fab fragment of a broadly neutralizing antibody (59.1) was determined to 3 angstrom resolution. The tip of the V3 loop containing the Gly-Pro-Gly-Arg-Ala-Phe sequence adopts a double-turn conformation, which may be the basis of its conservation in many HIV-1 isolates. A complete map of the HIV-1 principal neutralizing determinant was constructed by stitching together structures of V3 loop peptides bound to 59.1 and to an isolate-specific (MN) neutralizing antibody (50.1). Structural conservation of the overlapping epitopes suggests that this biologically relevant conformation could be of use in the design of synthetic vaccines and drugs to inhibit HIV-1 entry and virus-related cellular fusion.

Structural basis of plasticity in T cell receptor recognition of a self peptide-MHC antigen.

The T cell receptor (TCR) inherently has dual specificity. T cells must recognize self-antigens in the thymus during maturation and then discriminate between foreign pathogens in the periphery. A molecular basis for this cross-reactivity is elucidated by the crystal structure of the alloreactive 2C TCR bound to self peptide-major histocompatibility complex (pMHC) antigen H-2Kb-dEV8 refined against anisotropic 3.0 angstrom resolution x-ray data. The interface between peptide and TCR exhibits extremely poor shape complementarity, and the TCR beta chain complementarity-determining region 3 (CDR3) has minimal interaction with the dEV8 peptide. Large conformational changes in three of the TCR CDR loops are induced upon binding, providing a mechanism of structural plasticity to accommodate a variety of different peptide antigens. Extensive TCR interaction with the pMHC alpha helices suggests a generalized orientation that is mediated by the Valpha domain of the TCR and rationalizes how TCRs can effectively "scan" different peptides bound within a large, low-affinity MHC structural framework for those that provide the slight additional kinetic stabilization required for signaling.

Crystallographic evidence for preformed dimers of erythropoietin receptor before ligand activation.

Erythropoietin receptor (EPOR) is thought to be activated by ligand-induced homodimerization. However, structures of agonist and antagonist peptide complexes of EPOR, as well as an EPO-EPOR complex, have shown that the actual dimer configuration is critical for the biological response and signal efficiency. The crystal structure of the extracellular domain of EPOR in its unliganded form at 2.4 angstrom resolution has revealed a dimer in which the individual membrane-spanning and intracellular domains would be too far apart to permit phosphorylation by JAK2. This unliganded EPOR dimer is formed from self-association of the same key binding site residues that interact with EPO-mimetic peptide and EPO ligands. This model for a preformed dimer on the cell surface provides insights into the organization, activation, and plasticity of recognition of hematopoietic cell surface receptors.