RESUMEN
Modeling biological mechanisms is a key for disease understanding and drug-target identification. However, formulating quantitative models in the field of Alzheimer's Disease is challenged by a lack of detailed knowledge of relevant biochemical processes. Additionally, fitting differential equation systems usually requires time resolved data and the possibility to perform intervention experiments, which is difficult in neurological disorders. This work addresses these challenges by employing the recently published Variational Autoencoder Modular Bayesian Networks (VAMBN) method, which we here trained on combined clinical and patient level gene expression data while incorporating a disease focused knowledge graph. Our approach, called iVAMBN, resulted in a quantitative model that allowed us to simulate a down-expression of the putative drug target CD33, including potential impact on cognitive impairment and brain pathophysiology. Experimental validation demonstrated a high overlap of molecular mechanism predicted to be altered by CD33 perturbation with cell line data. Altogether, our modeling approach may help to select promising drug targets.
Asunto(s)
Enfermedad de Alzheimer , Disfunción Cognitiva , Humanos , Teorema de Bayes , Inteligencia Artificial , Lectina 3 Similar a Ig de Unión al Ácido Siálico/química , Lectina 3 Similar a Ig de Unión al Ácido Siálico/genética , Lectina 3 Similar a Ig de Unión al Ácido Siálico/metabolismoRESUMEN
The aim of this study was to establish the therapeutic relevance of the CD33D2 isoform by developing novel antibodies targeting the IgC domain of CD33. Two novel IgC-targeting antibodies, HL2541 and 5C11-2, were developed, and CD33 isoforms were assessed using multiple assays in cells overexpressing either CD33FL or CD33D2 isoforms, unmodified acute myeloid leukemia (AML) cell lines and primary AML specimens representing different genotypes for the CD33 splicing single nucleotide polymorphism. CD33D2 was recognized on cells overexpressing CD33D2 and unmodified AML cell lines; however, minimal/no cell surface detection of CD33D2 was observed in primary AML specimens. Both isoforms were detected intracellularly using novel antibodies. Minimal cell surface expression of CD33D2 on primary AML/progenitor cells warrants further studies on anti-CD33D2 immunotherapeutics.
Asunto(s)
Anticuerpos Monoclonales/inmunología , Lectina 3 Similar a Ig de Unión al Ácido Siálico/inmunología , Lectina 3 Similar a Ig de Unión al Ácido Siálico/metabolismo , Adolescente , Animales , Anticuerpos Monoclonales/uso terapéutico , Células de la Médula Ósea/metabolismo , Células de la Médula Ósea/patología , Línea Celular Tumoral , Niño , Preescolar , Femenino , Genotipo , Humanos , Dominios de Inmunoglobulinas/inmunología , Lactante , Leucemia Mieloide Aguda/tratamiento farmacológico , Leucemia Mieloide Aguda/genética , Leucemia Mieloide Aguda/metabolismo , Leucemia Mieloide Aguda/patología , Masculino , Ratones , Isoformas de Proteínas , Lectina 3 Similar a Ig de Unión al Ácido Siálico/química , Lectina 3 Similar a Ig de Unión al Ácido Siálico/genéticaRESUMEN
BACKGROUND: Gemtuzumab ozogamicin (GO) was approved in 2017 in the US for the treatment of adults with newly diagnosed CD33-positive (CD33+) acute myeloid leukemia (AML), and adults and pediatric patients with CD33+ relapsed/refractory (R/R) AML. OBJECTIVE: The aim of this study was to estimate the budgetary impact of introducing GO to a 1-million-member US health plan over a 5-year period. METHODS: We developed models to estimate the impact of introducing GO in combination with conventional induction chemotherapy or as monotherapy for newly diagnosed AML, and as monotherapy for R/R AML. Models were built using data on drug costs and treatment-related outcomes obtained from published clinical trials and other publicly available sources. Results were reported on a per member/per year and per member/per month (PMPM) basis. RESULTS: Base-case results of the newly diagnosed model indicated that the addition of GO in the combination setting reduced the overall budget of a 1-million-member health plan. The estimated net cost (US$) savings ranged from $72,969 ($0.006 PMPM) in year 1 to $745,426 ($0.062 PMPM) in year 5. In the monotherapy setting, GO was associated with increased net costs ranging from $4118 (0.0003 PMPM) in year 1 to $31,885 ($0.003 PMPM) in year 5. Base-case results of the R/R AML model demonstrated increased net costs that ranged from $17,326 ($0.001 PMPM) in year 1 to $46,163 ($0.004 PMPM) in year 5. Scenario analyses in all settings indicated the budget impact was not overly sensitive to the selected input assumptions, with the exception of the scenario considering only the pharmacy budget impact in the combination setting. CONCLUSIONS: The introduction of GO for newly diagnosed and R/R AML would have a minimal impact on the budget of a US health plan and could result in cost savings in the combination therapy setting for newly diagnosed AML.
Asunto(s)
Gemtuzumab , Leucemia Mieloide Aguda , Lectina 3 Similar a Ig de Unión al Ácido Siálico/farmacología , Adulto , Presupuestos , Niño , Ahorro de Costo , Costos de los Medicamentos , Humanos , Lectina 3 Similar a Ig de Unión al Ácido Siálico/química , Lectina 3 Similar a Ig de Unión al Ácido Siálico/inmunología , Lectina 3 Similar a Ig de Unión al Ácido Siálico/metabolismoRESUMEN
A single nucleotide polymorphism (SNP) in exon 2 of the CD33 gene is associated with reduced susceptibility to late-onset Alzheimer's disease (AD) and causal for elevated mRNA lacking exon 2. In contrast to full-length CD33, transcripts lacking exon 2 result in CD33 protein unable to suppress activation responses in myeloid cells, including microglia. Currently, little is known about the regulation of CD33 exon 2 splicing. Using functional genomics and proteomic approaches, we found that SRSF1 and PTBP1 act as splicing enhancers to increase CD33 exon 2 inclusion in mRNA. Binding of PTBP1 to RNA sequences proximal to the intron 1-exon 2 splice junction is altered by the SNP and represents a potential mechanism behind the SNP-genotype dependent alternative splicing. Our studies also reveal that binding of SRSF1 to the CD33 RNA is not altered by the SNP genotype. Instead, a putative SRSF1 binding sequence at the 3' end of exon 2 directs CD33 exon 2 inclusion into the mRNA, indicating that PTBP1 and SRSF1 promote full-length isoform expression through different mechanisms. Our findings shed light on molecular interactions that regulate CD33 exon 2 splicing, ultimately impacting receptor expression on the cell surface. These data aid in the understanding of CD33's regulation of microglial signaling underpinning the AD genetic associations.
Asunto(s)
Enfermedad de Alzheimer/genética , Ribonucleoproteínas Nucleares Heterogéneas/metabolismo , Polimorfismo de Nucleótido Simple , Proteína de Unión al Tracto de Polipirimidina/metabolismo , Factores de Empalme Serina-Arginina/metabolismo , Lectina 3 Similar a Ig de Unión al Ácido Siálico/genética , Edad de Inicio , Empalme Alternativo , Enfermedad de Alzheimer/metabolismo , Sitios de Unión , Exones , Predisposición Genética a la Enfermedad , Células HeLa , Humanos , Células K562 , Proteómica/métodos , ARN Mensajero/química , ARN Mensajero/metabolismo , Lectina 3 Similar a Ig de Unión al Ácido Siálico/química , Lectina 3 Similar a Ig de Unión al Ácido Siálico/metabolismo , Células THP-1RESUMEN
In 2011, genome-wide association studies implicated a polymorphism near CD33 as a genetic risk factor for Alzheimer's disease. This finding sparked interest in this member of the sialic acid-binding immunoglobulin-type lectin family which is linked to innate immunity. Subsequent studies found that CD33 is expressed in microglia in the brain and then investigated the molecular mechanism underlying the CD33 genetic association with Alzheimer's disease. The allele that protects from Alzheimer's disease acts predominately to increase a CD33 isoform lacking exon 2 at the expense of the prototypic, full-length CD33 that contains exon 2. Since this exon encodes the sialic acid ligand-binding domain, the finding that the loss of exon 2 was associated with decreased Alzheimer's disease risk was interpreted as meaning that a decrease in functional CD33 and its associated immune suppression was protective from Alzheimer's disease. However, this interpretation may need to be reconsidered given current findings that a genetic deletion which abrogates CD33 is not associated with Alzheimer's disease risk. Therefore, integrating currently available findings leads us to propose a model wherein the CD33 isoform lacking the ligand-binding domain represents a gain of function variant that reduces Alzheimer's disease risk.
Asunto(s)
Enfermedad de Alzheimer/genética , Lectina 3 Similar a Ig de Unión al Ácido Siálico/fisiología , Secuencias de Aminoácidos , Secuencia de Consenso , Dimerización , Exones/genética , Mutación con Ganancia de Función , Predisposición Genética a la Enfermedad , Estudio de Asociación del Genoma Completo , Humanos , Desequilibrio de Ligamiento , Metaanálisis como Asunto , Microglía/fisiología , Familia de Multigenes , Proteínas del Tejido Nervioso/genética , Proteínas del Tejido Nervioso/fisiología , Polimorfismo de Nucleótido Simple , Factores de Riesgo , Eliminación de Secuencia , Lectina 3 Similar a Ig de Unión al Ácido Siálico/química , Lectina 3 Similar a Ig de Unión al Ácido Siálico/genética , Lectinas Similares a la Inmunoglobulina de Unión a Ácido Siálico/genética , Lectinas Similares a la Inmunoglobulina de Unión a Ácido Siálico/metabolismoRESUMEN
In recent years, several entry mediators have been characterized for porcine reproductive and respiratory syndrome virus (PRRSV). Porcine sialoadhesin [pSn, also known as sialic acid-binding immunoglobulin-type lectin (Siglec-1)] and porcine CD163 (pCD163) have been identified as the most important host entry mediators that can fully coordinate PRRSV infection into macrophages. However, recent isolates have not only shown a tropism for sialoadhesin-positive cells, but also for sialoadhesin-negative cells. This observation might be partly explained by the existence of additional receptors that can support PRRSV binding and entry. In the search for new receptors, recently identified porcine Siglecs (Siglec-3, Siglec-5 and Siglec-10), members of the same family as sialoadhesin, were cloned and characterized. Only Siglec-10 was able to significantly improve PRRSV infection and production in a CD163-transfected cell line. Compared with sialoadhesin, Siglec-10 performed equally effectively as a receptor for PRRSV type 2 strain MN-184, but it was less capable of supporting infection with PRRSV type 1 strain LV (Lelystad virus). Siglec-10 was demonstrated to be involved in the endocytosis of PRRSV, confirming the important role of Siglec-10 in the entry process of PRRSV. In conclusion, it can be stated that PRRSV may use several Siglecs to enter macrophages, which may explain the strain differences in the pathogenesis.
Asunto(s)
Clonación Molecular , Síndrome Respiratorio y de la Reproducción Porcina/metabolismo , Virus del Síndrome Respiratorio y Reproductivo Porcino/metabolismo , Receptores Virales/metabolismo , Lectina 3 Similar a Ig de Unión al Ácido Siálico/metabolismo , Secuencia de Aminoácidos , Animales , Antígenos CD/genética , Antígenos CD/metabolismo , Antígenos de Diferenciación Mielomonocítica/genética , Antígenos de Diferenciación Mielomonocítica/metabolismo , Endocitosis , Datos de Secuencia Molecular , Síndrome Respiratorio y de la Reproducción Porcina/genética , Síndrome Respiratorio y de la Reproducción Porcina/fisiopatología , Síndrome Respiratorio y de la Reproducción Porcina/virología , Virus del Síndrome Respiratorio y Reproductivo Porcino/genética , Receptores de Superficie Celular/genética , Receptores de Superficie Celular/metabolismo , Receptores Virales/química , Receptores Virales/genética , Alineación de Secuencia , Lectina 3 Similar a Ig de Unión al Ácido Siálico/química , Lectina 3 Similar a Ig de Unión al Ácido Siálico/genética , PorcinosRESUMEN
The immunomodulatory receptor Siglec-3/CD33 influences risk for late-onset Alzheimer's disease (LOAD), an apparently human-specific post-reproductive disease. CD33 generates two splice variants: a full-length CD33M transcript produced primarily by the "LOAD-risk" allele and a shorter CD33m isoform lacking the sialic acid-binding domain produced primarily from the "LOAD-protective" allele. An SNP that modulates CD33 splicing to favor CD33m is associated with enhanced microglial activity. Individuals expressing more protective isoform accumulate less brain ß-amyloid and have a lower LOAD risk. How the CD33m isoform increases ß-amyloid clearance remains unknown. We report that the protection by the CD33m isoform may not be conferred by what it does but, rather, from what it cannot do. Analysis of blood neutrophils and monocytes and a microglial cell line revealed that unlike CD33M, the CD33m isoform does not localize to cell surfaces; instead, it accumulates in peroxisomes. Cell stimulation and activation did not mobilize CD33m to the surface. Thus, the CD33m isoform may neither interact directly with amyloid plaques nor engage in cell-surface signaling. Rather, production and localization of CD33m in peroxisomes is a way of diminishing the amount of CD33M and enhancing ß-amyloid clearance. We confirmed intracellular localization by generating a CD33m-specific monoclonal antibody. Of note, CD33 is the only Siglec with a peroxisome-targeting sequence, and this motif emerged by convergent evolution in toothed whales, the only other mammals with a prolonged post-reproductive lifespan. The CD33 allele that protects post-reproductive individuals from LOAD may have evolved by adaptive loss-of-function, an example of the less-is-more hypothesis.
Asunto(s)
Enfermedad de Alzheimer/genética , Predisposición Genética a la Enfermedad , Macrófagos/metabolismo , Microglía/metabolismo , Neutrófilos/metabolismo , Polimorfismo de Nucleótido Simple , Lectina 3 Similar a Ig de Unión al Ácido Siálico/metabolismo , Alelos , Enfermedad de Alzheimer/inmunología , Enfermedad de Alzheimer/metabolismo , Enfermedad de Alzheimer/patología , Secuencias de Aminoácidos , Proteínas Bacterianas/metabolismo , Proteínas Bacterianas/toxicidad , Línea Celular , Membrana Celular/efectos de los fármacos , Membrana Celular/metabolismo , Membrana Celular/patología , Humanos , Lipopolisacáridos/toxicidad , Activación de Macrófagos/efectos de los fármacos , Macrófagos/efectos de los fármacos , Macrófagos/inmunología , Macrófagos/patología , Microglía/citología , Microglía/inmunología , Microglía/patología , N-Formilmetionina Leucil-Fenilalanina/toxicidad , Proteínas del Tejido Nervioso/química , Proteínas del Tejido Nervioso/genética , Proteínas del Tejido Nervioso/metabolismo , Neuraminidasa/metabolismo , Neuraminidasa/toxicidad , Activación Neutrófila/efectos de los fármacos , Neutrófilos/efectos de los fármacos , Neutrófilos/inmunología , Neutrófilos/patología , Peroxisomas/efectos de los fármacos , Peroxisomas/metabolismo , Peroxisomas/patología , Filogenia , Dominios y Motivos de Interacción de Proteínas , Isoformas de Proteínas/química , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Señales de Clasificación de Proteína , Transporte de Proteínas/efectos de los fármacos , Lectina 3 Similar a Ig de Unión al Ácido Siálico/química , Lectina 3 Similar a Ig de Unión al Ácido Siálico/genéticaRESUMEN
Antigen-specific immunotherapies have emerged as important components of curative treatment algorithms for many cancers. In acute myeloid leukemia (AML), success has been less obvious. Nonetheless, among the few drugs shown to improve survival in recent randomized trials is the CD33 antibody-drug conjugate gemtuzumab ozogamicin. Significant antileukemic activity is also well documented for radioimmunoconjugates targeting CD33, CD45, or CD66. These therapeutics can intensify conditioning before hematopoietic cell transplantation, but their effect on patient outcomes needs clarification. Emerging data now suggest clinical antileukemic activity of several novel antibodies and perhaps some adoptive T-cell immunotherapies and vaccines. In parallel, numerous other agents targeting a wider variety of antigens are currently being explored. However, the antigenic heterogeneity characteristic of AML is a considerable limitation for all these therapeutics, and many important questions related to the ideal target antigen(s), disease situation in which to use these therapies, most suitable patient populations, exact treatment modalities, and details of supportive care needs remain open. Addressing such questions in upcoming studies will be required to ensure that antigen-directed therapies become an effective tool in AML, a disease for which outcomes with standard "3 + 7"-based chemotherapy have remained unsatisfactory in many patients.
Asunto(s)
Antígenos/química , Antineoplásicos/uso terapéutico , Inmunoterapia/métodos , Leucemia Mieloide Aguda/terapia , Algoritmos , Aminoglicósidos/uso terapéutico , Anticuerpos/uso terapéutico , Anticuerpos Monoclonales Humanizados/uso terapéutico , Ensayos Clínicos como Asunto , Gemtuzumab , Humanos , Inmunoterapia Adoptiva/métodos , Inmunotoxinas/uso terapéutico , Leucemia Mieloide Aguda/inmunología , Radioinmunoterapia/métodos , Ensayos Clínicos Controlados Aleatorios como Asunto , Lectina 3 Similar a Ig de Unión al Ácido Siálico/química , Vacunas/uso terapéuticoRESUMEN
In this issue of Blood, Kung Sutherland et al report on the preclinical activity of SGN-CD33A, a humanized anti-CD33 antibody conjugated to a pyrrolobenzodiazepine (PBD) dimer via a protease-cleavable linker, against acute myeloid leukemia (AML) cells in vitro and in vivo.
Asunto(s)
Anticuerpos Monoclonales Humanizados/química , Benzodiazepinas/química , Resistencia a Antineoplásicos , Inmunoconjugados/química , Leucemia Mieloide Aguda/tratamiento farmacológico , Lectina 3 Similar a Ig de Unión al Ácido Siálico/química , Animales , HumanosRESUMEN
Outcomes in acute myeloid leukemia (AML) remain unsatisfactory, and novel treatments are urgently needed. One strategy explores antibodies and their drug conjugates, particularly those targeting CD33. Emerging data with gemtuzumab ozogamicin (GO) demonstrate target validity and activity in some patients with AML, but efficacy is limited by heterogeneous drug conjugation, linker instability, and a high incidence of multidrug resistance. We describe here the development of SGN-CD33A, a humanized anti-CD33 antibody with engineered cysteines conjugated to a highly potent, synthetic DNA cross-linking pyrrolobenzodiazepine dimer via a protease-cleavable linker. The use of engineered cysteine residues at the sites of drug linker attachment results in a drug loading of approximately 2 pyrrolobenzodiazepine dimers per antibody. In preclinical testing, SGN-CD33A is more potent than GO against a panel of AML cell lines and primary AML cells in vitro and in xenotransplantation studies in mice. Unlike GO, antileukemic activity is observed with SGN-CD33A in AML models with the multidrug-resistant phenotype. Mechanistic studies indicate that the cytotoxic effects of SGN-CD33A involve DNA damage with ensuing cell cycle arrest and apoptotic cell death. Together, these data suggest that SGN-CD33A has CD33-directed antitumor activity and support clinical testing of this novel therapeutic in patients with AML.