RESUMEN
Chimeric antigen receptor (CAR) T-cells are an emerging therapy for refractory lymphomas. Clonal hematopoiesis (CH), the preferential outgrowth of mutated bone marrow progenitors, is enriched in lymphoma patients receiving CAR-T cells. CAR-T therapy requires conditioning chemotherapy and often induces systemic inflammatory reactions, both of which have been shown to promote expansion of CH clones. Thus, we hypothesized that pre-existing CH clones could expand during CAR-T cell treatment. We measured CH at 154 timepoints longitudinally sampled from 26 patients receiving CD30.CAR-T therapy for CD30+ lymphomas on an investigational protocol (NCT02917083). Pre-treatment CH was present in 54% of individuals and did not correlate with survival outcomes or inflammatory toxicities. Longitudinal tracking of single clones in individual patients revealed distinct clone growth dynamics. Initially small clones, defined as VAF <1%, expanded following CAR-T administration, compared with relatively muted expansions of larger clones (3.37-fold vs. 1.20-fold, P = 0.0014). Matched clones were present at low magnitude in the infused CD30.CAR-T product for all CH cases but did not affect the product's immunophenotype or transduction efficiency. As cellular immunotherapies expand to become frontline treatments for hematological malignancies, our data indicates CAR-T recipients could be enriched for CH, and further longitudinal studies centered on CH complications in this population are warranted.
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Linfoma , Receptores Quiméricos de Antígenos , Humanos , Hematopoyesis Clonal , Inmunoterapia Adoptiva/efectos adversos , Inmunoterapia Adoptiva/métodos , Linfoma/terapia , Inmunoterapia , Hematopoyesis/genéticaRESUMEN
Chimeric antigen receptor (CAR)-mediated targeting of T lineage antigens for the therapy of blood malignancies is frequently complicated by self-targeting of CAR T cells or their excessive differentiation driven by constant CAR signaling. Expression of CARs targeting CD7, a pan-T cell antigen highly expressed in T cell malignancies and some myeloid leukemias, produces robust fratricide and often requires additional mitigation strategies, such as CD7 gene editing. In this study, we show fratricide of CD7 CAR T cells can be fully prevented using ibrutinib and dasatinib, the pharmacologic inhibitors of key CAR/CD3ζ signaling kinases. Supplementation with ibrutinib and dasatinib rescued the ex vivo expansion of unedited CD7 CAR T cells and allowed regaining full CAR-mediated cytotoxicity in vitro and in vivo on withdrawal of the inhibitors. The unedited CD7 CAR T cells persisted long term and mediated sustained anti-leukemic activity in two mouse xenograft models of human T cell acute lymphoblastic leukemia (T-ALL) by self-selecting for CD7-, fratricide-resistant CD7 CAR T cells that were transcriptionally similar to control CD7-edited CD7 CAR T cells. Finally, we showed feasibility of cGMP manufacturing of unedited autologous CD7 CAR T cells for patients with CD7+ malignancies and initiated a phase I clinical trial (ClinicalTrials.gov: NCT03690011) using this approach. These results indicate pharmacologic inhibition of CAR signaling enables generating functional CD7 CAR T cells without additional engineering.
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Leucemia-Linfoma Linfoblástico de Células T Precursoras , Receptores Quiméricos de Antígenos , Ratones , Animales , Humanos , Linfocitos T , Inmunoterapia Adoptiva/métodos , Dasatinib/metabolismo , Estudios de Factibilidad , Receptores Quiméricos de Antígenos/genética , Receptores Quiméricos de Antígenos/metabolismoRESUMEN
Adjuvant system 04 (AS04) is in injectable human vaccines. AS04 contains two known adjuvants, 3-O-desacyl-4'-monophosphoryl lipid A (MPL) and insoluble aluminum salts. Data from previous studies showed that both MPL and insoluble aluminum salts have nasal mucosal vaccine adjuvant activity. The present study was designed to test the feasibility of using AS04 as an adjuvant to help nasally administered antigens to induce specific mucosal and systemic immunity as well as to evaluate the deposition of antigens in the upper respiratory tract when adjuvanted with AS04. Alhydrogel, an aluminum (oxy)hydroxide suspension, was mixed with MPL to form AS04, which was then mixed with ovalbumin (OVA) or 3× M2e-HA2, a synthetic influenza virus hemagglutinin fusion protein, as an antigen to prepare OVA/AS04 and 3× M2e-HA2/AS04 vaccines, respectively. In mice, AS04 enabled antigens, when given intranasally, to induce specific IgA response in nasal and lung mucosal secretions as well as specific IgG response in the serum samples of the immunized mice, whereas subcutaneous injection of the same vaccine induced specific antibody responses only in the serum samples but not in the mucosal secretions. Splenocytes isolated from mice intranasally immunized with the OVA/AS04 also proliferated and released cytokines (i.e., IL-4 and IFN-γ) after in vitro stimulation with the antigen. In the immunogenicity test, intranasal OVA/AS04 was not more effective than intranasal OVA/MPL at the dosing regimens tested. However, when compared to OVA/MPL, OVA/AS04 showed a different atomized droplet size distribution and more importantly a more favorable OVA deposition profile when atomized into a nasal cast that was 3-D printed based on the computer tomography scan of the nose of a child. It is concluded that AS04 has mucosal adjuvant activity when given intranasally. In addition, there is a reason to be optimistic about using AS04 as an adjuvant to target an antigen of interest to the right region of the nasal cavity in humans for immune response induction.
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Hidróxido de Aluminio/inmunología , Formación de Anticuerpos/inmunología , Antígenos/inmunología , Inmunogenicidad Vacunal/inmunología , Lípido A/análogos & derivados , Sistema Respiratorio/inmunología , Vacunas/inmunología , Adyuvantes Inmunológicos/farmacología , Administración Intranasal/métodos , Animales , Citocinas/inmunología , Femenino , Humanos , Inmunidad/inmunología , Inmunidad Mucosa/inmunología , Inmunización/métodos , Lípido A/inmunología , Ratones , Ratones Endogámicos BALB C , Ovalbúmina/inmunología , Vacunación/métodosRESUMEN
We report long-term outcomes up to 18 years of a clinical trial treating children with neuroblastoma with EBV-specific T lymphocytes and CD3-activated T cells - each expressing a first-generation chimeric antigen receptor targeting GD2 with barcoded transgenes to allow tracking of each population. Of 11 patients with active disease at infusion, three patients achieved a complete response that was sustained in 2, one for 8 years until lost to follow up and one for 18+ years. Of eight patients with a history of relapse or at high risk of recurrence, five are disease-free at their last follow-up between 10-14 years post-infusion. Intermittent low levels of transgene were detected during the follow up period with significantly greater persistence in those who were long-term survivors. In conclusion, patients with relapsed/refractory neuroblastoma achieved long-term disease control after receiving GD2 CAR-T cell therapy including one patient now in remission of relapsed disease for >18 years.
RESUMEN
In this prospective, interventional phase 1 study for individuals with advanced sarcoma, we infused autologous HER2-specific chimeric antigen receptor T cells (HER2 CAR T cells) after lymphodepletion with fludarabine (Flu) ± cyclophosphamide (Cy): 1 × 108 T cells per m2 after Flu (cohort A) or Flu/Cy (cohort B) and 1 × 108 CAR+ T cells per m2 after Flu/Cy (cohort C). The primary outcome was assessment of safety of one dose of HER2 CAR T cells after lymphodepletion. Determination of antitumor responses was the secondary outcome. Thirteen individuals were treated in 14 enrollments, and seven received multiple infusions. HER2 CAR T cells expanded after 19 of 21 infusions. Nine of 12 individuals in cohorts A and B developed grade 1-2 cytokine release syndrome. Two individuals in cohort C experienced dose-limiting toxicity with grade 3-4 cytokine release syndrome. Antitumor activity was observed with clinical benefit in 50% of individuals treated. The tumor samples analyzed showed spatial heterogeneity of immune cells and clustering by sarcoma type and by treatment response. Our results affirm HER2 as a CAR T cell target and demonstrate the safety of this therapeutic approach in sarcoma. ClinicalTrials.gov registration: NCT00902044 .
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Inmunoterapia Adoptiva , Receptor ErbB-2 , Receptores Quiméricos de Antígenos , Sarcoma , Humanos , Sarcoma/terapia , Sarcoma/inmunología , Persona de Mediana Edad , Femenino , Masculino , Adulto , Inmunoterapia Adoptiva/métodos , Inmunoterapia Adoptiva/efectos adversos , Anciano , Receptores Quiméricos de Antígenos/inmunología , Linfocitos T/inmunología , Depleción Linfocítica/métodos , Estudios Prospectivos , Vidarabina/análogos & derivados , Vidarabina/administración & dosificación , Vidarabina/uso terapéutico , Ciclofosfamida/uso terapéutico , Ciclofosfamida/administración & dosificación , Resultado del TratamientoRESUMEN
Interleukin-15 (IL15) promotes the survival of T lymphocytes and enhances the antitumor properties of CAR T cells in preclinical models of solid neoplasms in which CAR T cells have limited efficacy1-4. Glypican-3 (GPC3) is expressed in a group of solid cancers5-10, and here we report the first evaluation in humans of the effects of IL15 co-expression on GPC3-CAR T cells. Cohort 1 patients (NCT02905188/NCT02932956) received GPC3-CAR T cells, which were safe but produced no objective antitumor responses and reached peak expansion at two weeks. Cohort 2 patients (NCT05103631/NCT04377932) received GPC3-CAR T cells that co-expressed IL15 (15.CAR), which mediated significantly increased cell expansion and induced a disease control rate of 66% and antitumor response rate of 33%. Infusion of 15.CAR T cells was associated with increased incidence of cytokine release syndrome, which was rapidly ameliorated by activation of the inducible caspase 9 safety switch. Compared to non-responders, tumor-infiltrating 15.CAR T cells from responders showed repression of SWI/SNF epigenetic regulators and upregulation of FOS and JUN family members as well as genes related to type I interferon signaling. Collectively, these results demonstrate that IL15 increases the expansion, intratumoral survival, and antitumor activity of GPC3-CAR T cells in patients.
RESUMEN
Vα24-invariant natural killer T cells (NKTs) have anti-tumor properties that can be enhanced by chimeric antigen receptors (CARs). Here we report updated interim results from the first-in-human phase 1 evaluation of autologous NKTs co-expressing a GD2-specific CAR with interleukin 15 (IL15) (GD2-CAR.15) in 12 children with neuroblastoma (NB). The primary objectives were safety and determination of maximum tolerated dose (MTD). The anti-tumor activity of GD2-CAR.15 NKTs was assessed as a secondary objective. Immune response evaluation was an additional objective. No dose-limiting toxicities occurred; one patient experienced grade 2 cytokine release syndrome that was resolved by tocilizumab. The MTD was not reached. The objective response rate was 25% (3/12), including two partial responses and one complete response. The frequency of CD62L+NKTs in products correlated with CAR-NKT expansion in patients and was higher in responders (n = 5; objective response or stable disease with reduction in tumor burden) than non-responders (n = 7). BTG1 (BTG anti-proliferation factor 1) expression was upregulated in peripheral GD2-CAR.15 NKTs and is a key driver of hyporesponsiveness in exhausted NKT and T cells. GD2-CAR.15 NKTs with BTG1 knockdown eliminated metastatic NB in a mouse model. We conclude that GD2-CAR.15 NKTs are safe and can mediate objective responses in patients with NB. Additionally, their anti-tumor activity may be enhanced by targeting BTG1. ClinicalTrials.gov registration: NCT03294954 .
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Células T Asesinas Naturales , Neuroblastoma , Receptores Quiméricos de Antígenos , Niño , Animales , Ratones , Humanos , Citotoxicidad Inmunológica , Receptores Quiméricos de Antígenos/genética , Neuroblastoma/terapia , Inmunoterapia Adoptiva/métodosRESUMEN
Previously, we synthesized 4-(N)-docosahexaenoyl 2', 2'-difluorodeoxycytidine (DHA-dFdC), a novel lipophilic compound with a potent, broad-spectrum antitumor activity. Herein, we report a solid lipid nanoparticle (SLN) formulation of DHA-dFdC with improved apparent aqueous solubility, chemical stability, as well as efficacy in a mouse model. The SLNs were prepared from lecithin/glycerol monostearate-in-water emulsions emulsified with D-α-tocopherol polyethylene glycol 1000 succinate (TPGS) and Tween 20. The resultant DHA-dFdC-SLNs were 102.2⯱â¯7.3â¯nm in diameter and increased the apparent solubility of DHA-dFdC in water to at least 5.2â¯mg/mL, more than 200-fold higher than its intrinsic water solubility. DHA-dFdC in a lyophilized powder of DHA-dFdC-SLNs was significantly more stable than the waxy solid of pure DHA-dFdC. DHA-dFdC-SLNs also showed an increased cytotoxicity against certain tumor cells than DHA-dFdC. The plasma concentration of DHA-dFdC in mice intravenously injected with DHA-dFdC-SLNs in dispersion followed a bi-exponential model, with a half-life of ~44â¯h. In mice bearing B16-F10 murine melanoma, DHA-dFdC-SLNs were significantly more effective than DHA-dFdC in controlling the tumor growth. In addition, histology evaluation revealed a high level of apoptosis and tumor encapsulation in tumors in mice treated with DHA-dFdC-SLNs. DHA-dFdC-SLNs represents a new DHA-dFdC formulation with improved antitumor activity.
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Antineoplásicos/química , Antineoplásicos/farmacología , Desoxicitidina/química , Lípidos/química , Nanopartículas/química , Solubilidad/efectos de los fármacos , Animales , Línea Celular Tumoral , Portadores de Fármacos/química , Sistemas de Liberación de Medicamentos/métodos , Emulsiones/síntesis química , Emulsiones/farmacología , Femenino , Lecitinas/química , Ratones , Ratones Endogámicos C57BL , Tamaño de la Partícula , Polietilenglicoles/química , Vitamina E/químicaRESUMEN
In an effort to improve the adjuvanticity of insoluble aluminium salts, we discovered that the adjuvant activity of aluminium salt nanoparticles is significantly stronger than aluminium salt microparticles, likely related to nanoparticle's stronger ability to directly activate NACHT, LRR and PYD domains-containing protein 3 (NLRP3) inflammasome as the nanoparticles are more efficiently taken up by phagocytic cells. Endogenous signals such as uric acid from cell damage or death caused by the cytotoxicity of aluminium salts are thought to indirectly activate inflammasome, prompting us to hypothesise that the potent adjuvant activity of aluminium salt nanoparticles is also related to their ability to stimulate uric acid production. In the present study, we prepared aluminium (oxy)hydroxide nanoparticles (â¼ 30-100 nm) and microparticles (X50, 9.43 µm) and showed that intraperitoneal injection of mice with the nanoparticles, absorbed with ovalbumin, led to a significant increase in uric acid level in the peritoneal lavage, whereas the microparticles did not. The aluminium (oxy)hydroxide nanoparticles' ability to stimulate uric acid production was also confirmed in cell culture. We concluded that the stronger adjuvant activity of insoluble aluminium (oxy)hydroxide nanoparticles, relative to microparticles, may be attributed at least in part to their stronger ability to induce endogenous danger signals such as uric acid.
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Adyuvantes Inmunológicos/administración & dosificación , Nanopartículas , Ácido Úrico/metabolismo , Vacunas/administración & dosificación , Hidróxido de Aluminio/química , Animales , Femenino , Inflamasomas/inmunología , Inyecciones Intraperitoneales , Ratones , Ratones Endogámicos BALB C , Proteína con Dominio Pirina 3 de la Familia NLR/inmunología , Ovalbúmina/inmunología , Tamaño de la Partícula , Vacunas/inmunologíaRESUMEN
Intranasal vaccination using dry powder vaccine formulation represents an attractive, non-invasive vaccination modality with better storage stability and added protection at the mucosal surfaces. Herein we report that it is feasible to induce specific mucosal and systemic antibody responses by intranasal immunization with a dry powder vaccine adjuvanted with an insoluble aluminum salt. The dry powder vaccine was prepared by thin-film freeze-drying of a model antigen, ovalbumin, adsorbed on aluminum (oxy)hydroxide as an adjuvant. Special emphasis was placed on the characterization of the dry powder vaccine formulation that can be realistically used in humans by a nasal dry powder delivery device. The vaccine powder was found to have "passable" to "good" flow properties, and the vaccine was uniformly distributed in the dry powder. An in vitro nasal deposition study using nasal casts of adult humans showed that around 90% of the powder was deposited in the nasal cavity. Intranasal immunization of rats with the dry powder vaccine elicited a specific serum antibody response as well as specific IgA responses in the nose and lung secretions of the rats. This study demonstrates the generation of systemic and mucosal immune responses by intranasal immunization using a dry powder vaccine adjuvanted with an aluminum salt.
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Adyuvantes Inmunológicos/administración & dosificación , Hidróxido de Aluminio/administración & dosificación , Óxido de Aluminio/administración & dosificación , Vacunas/administración & dosificación , Adyuvantes Inmunológicos/química , Adyuvantes Inmunológicos/farmacocinética , Administración Intranasal , Hidróxido de Aluminio/química , Hidróxido de Aluminio/farmacocinética , Óxido de Aluminio/química , Óxido de Aluminio/farmacocinética , Animales , Antígenos/administración & dosificación , Antígenos/química , Antígenos/inmunología , Encéfalo/metabolismo , Líquido del Lavado Bronquioalveolar/inmunología , Femenino , Inmunización , Inmunoglobulina A/inmunología , Inmunoglobulina G/sangre , Líquido del Lavado Nasal/inmunología , Ovalbúmina/administración & dosificación , Ovalbúmina/química , Ovalbúmina/inmunología , Polvos , Ratas Sprague-Dawley , Vacunas/química , Vacunas/farmacocinéticaRESUMEN
TNF-α siRNA has shown promising therapeutic benefits in animal models of rheumatoid arthritis. However, there continues to be a need for siRNA delivery systems that have high siRNA encapsulation efficiency and minimum burst release of TNF-α siRNA, and can target inflamed tissues after intravenous administration. Herein we report a novel acid-sensitive sheddable PEGylated solid-lipid nanoparticle formulation of TNF-α-siRNA, AS-TNF-α-siRNA-SLNs, prepared by incorporating lipophilized TNF-α-siRNA into solid-lipid nanoparticles composed of biocompatible lipids such as lecithin and cholesterol. The nanoparticles are approximately 120â¯nm in diameter, have a high siRNA encapsulation efficiency (>90%) and a minimum burst release of siRNA (<5%), and increase the deilvery of the siRNA in chronic inflammation sites in mouse models, including in a mouse model with collagen-induced arthritis. Importantly, in a mouse model of collagen antibody-induced arthritis that does not respond to methotrexate therapy, intravenous injection of the AS-TNF-α-siRNA-SLNs significantly reduced paw thickness, bone loss, and histopathological scores. These findings highlight the potential of using this novel siRNA nanoparticle formulation to effectively treat arthritis, potentially in patients who do not respond adequately to methotrexate.
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Antirreumáticos/administración & dosificación , Artritis Experimental/tratamiento farmacológico , Artritis Reumatoide/tratamiento farmacológico , Nanopartículas/administración & dosificación , ARN Interferente Pequeño/administración & dosificación , Factor de Necrosis Tumoral alfa/genética , Animales , Artritis Experimental/genética , Artritis Reumatoide/genética , Línea Celular , Resistencia a Medicamentos , Femenino , Lípidos/administración & dosificación , Metotrexato/administración & dosificación , Ratones Endogámicos C57BLRESUMEN
Many human vaccines contain certain insoluble aluminum salts such as aluminum oxyhydroxide and aluminum hydroxyphosphate as vaccine adjuvants to boost the immunogenicity of the vaccines. Aluminum salts have been used as vaccine adjuvants for decades and have an established, favorable safety profile. However, preparing aluminum salts and aluminum salt-adjuvanted vaccines in a consistent manner remains challenging. This chapter discusses methods to prepare aluminum salts and aluminum salt-adjuvanted vaccines, factors to consider during preparation, and methods to characterize the vaccines after preparation.
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Adyuvantes Inmunológicos/química , Compuestos de Aluminio/química , Aluminio/química , Fosfatos/química , Vacunas/química , Animales , HumanosRESUMEN
Insoluble aluminum salts such as aluminum oxyhydroxide have been used for decades as adjuvants in human vaccines, and many vaccines contain aluminum salts as adjuvants. Aluminum salt-adjuvanted vaccines must be managed in cold-chain (2-8° C) during transport and storage, as vaccine antigens in general are too fragile to be stable in ambient temperatures, and unintentional slowing freezing causes irreversible aggregation and permanent damage to the vaccines. Previously, we reported that thin-film freeze-drying can be used to convert vaccines adjuvanted with an aluminum salt from liquid suspension into dry powder without causing particle aggregation or decreasing in immunogenicity following reconstitution. In the present study, using ovalbumin (OVA)-adsorbed Alhydrogel® (i.e. aluminum oxyhydroxide, 2% w/v) as a model vaccine, we showed that the immunogenicity of thin-film freeze-dried OVA-adsorbed Alhydrogel® vaccine powder was not significantly changed after it was exposed for an extended period of time in temperatures as high as 40° C or subjected to repeated slow freezing-and-thawing. It is expected that immunization programs can potentially benefit by integrating thin-film freeze-drying into vaccine preparations.
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Adyuvantes Inmunológicos/efectos de la radiación , Hidróxido de Aluminio/efectos de la radiación , Liofilización , Temperatura , Potencia de la Vacuna , Vacunas/inmunología , Vacunas/efectos de la radiación , Adyuvantes Inmunológicos/administración & dosificación , Hidróxido de Aluminio/administración & dosificación , Animales , Femenino , Ratones Endogámicos BALB C , Ovalbúmina/administración & dosificación , Ovalbúmina/inmunología , Ovalbúmina/efectos de la radiación , Vacunas/administración & dosificaciónRESUMEN
Insoluble aluminum salts such as aluminum (oxy)hydroxide are commonly used as vaccine adjuvants. Recently, there is evidence suggesting that the adjuvant activity of aluminum salt-based materials is tightly related to their physicochemical properties, including nanometer-scale size, shape with long aspect ratio, and low degree of crystallinity. Herein, for the first time, the bicontinuous reverse microemulsion (RM) technique was utilized to synthesize stick-like monodisperse aluminum (oxy)hydroxide nanoparticles with a long aspect ratio of â¼10, length of â¼80 nm, and low degree of crystallinity (denoted as Al-nanosticks). Moreover, the relationship between the physicochemical properties of Al-nanosticks and the bicontinuous RM was discussed. Compared to the commercial Alhydrogel, which contains micrometer-scale aluminum oxyhydroxide particular aggregates with moderate degree of crystallinity, the Al-nanosticks are more effective in adsorbing and delivering antigens (e.g., ovalbumin, OVA) into antigen-presenting cells, activating inflammasomes, and potentiating OVA-specific antibody responses in a mouse model. It is concluded that the aluminum (oxy)hydroxide nanosticks synthesized in the bicontinuous RM are promising new aluminum salt-based vaccine adjuvants.
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Hidróxido de Aluminio/química , Adyuvantes Inmunológicos , Animales , Humanos , Ratones , Ovalbúmina , VacunasRESUMEN
Some insoluble aluminum salts are commonly used in injectable vaccines as adjuvants to accelerate, prolong, or enhance the antigen-specific immune responses. Data from previous studies testing the nasal mucosal vaccine adjuvant activity of aluminum salts are conflicting. The present study is designed to further assess the feasibility of using aluminum salts in injectable vaccines as nasal mucosal vaccine adjuvants. Using Alhydrogel®, the international scientific standard of aluminum (oxy)hydroxide gels, and ovalbumin or 3 × M2e-HA2, a synthetic influenza virus fusion protein, as antigens, we showed in a mouse model that when dosed intranasally Alhydrogel® enables antigens adsorbed on it to induce stronger antigen-specific immune responses in both serum samples (e.g., specific IgG) and nasal and lung mucosal secretions (i.e., specific IgA) in all immunized mice, as compared with nasal immunization with the antigens alone. Rerouting insoluble aluminum salts in injectable vaccines may represent a viable approach for (nasal) mucosal vaccine adjuvant discovery.
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Hidróxido de Aluminio/inmunología , Formación de Anticuerpos/inmunología , Antígenos Virales/inmunología , Inmunidad Mucosa , Proteínas Virales de Fusión/inmunología , Adyuvantes Inmunológicos/administración & dosificación , Adyuvantes Inmunológicos/química , Administración Intranasal , Adsorción , Hidróxido de Aluminio/administración & dosificación , Hidróxido de Aluminio/química , Animales , Antígenos Virales/química , Antígenos Virales/genética , Femenino , Inmunoglobulina A/análisis , Inmunoglobulina A/biosíntesis , Inmunoglobulina A/inmunología , Inmunoglobulina G/biosíntesis , Inmunoglobulina G/sangre , Inmunoglobulina G/inmunología , Ratones , Ratones Endogámicos BALB C , Mucosa Nasal/inmunología , Vacunación , Proteínas Virales de Fusión/administración & dosificaciónRESUMEN
This review is intended to provide a critical account of the current goals and technologies of particle engineering regarding the production of crystalline and amorphous particles. The technologies discussed here cover traditional crystallization technologies, supercritical fluid technologies, spray drying, controlled solvent crystallization, and sonocrystallization. Also recent advancements in particle engineering including spray freezing into liquid, thin-film freeze-drying, PRINT technology are presented. The paper also examines the merits and limitations of these technologies with respect to their methods of characterization. Additionally a section discussing the utility of creating amorphous and crystalline formulation approaches in regards to bioavailability and utility in formulation is presented.
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Preparaciones Farmacéuticas/administración & dosificación , Tecnología Farmacéutica/métodos , Disponibilidad Biológica , Química Farmacéutica , Cristalización , Liofilización , Humanos , Tamaño de la Partícula , SolubilidadRESUMEN
Many currently licensed and commercially available human vaccines contain aluminum salts as vaccine adjuvants. A major limitation with these vaccines is that they must not be exposed to freezing temperatures during transport or storage such that the liquid vaccine freezes, because freezing causes irreversible coagulation that damages the vaccines (e.g., loss of efficacy). Therefore, vaccines that contain aluminum salts as adjuvants are formulated as liquid suspensions and are required to be kept in cold chain (2-8°C) during transport and storage. Formulating vaccines adjuvanted with aluminum salts into dry powder that can be readily reconstituted before injection may address this limitation. Spray freeze-drying of vaccines with low concentrations of aluminum salts and high concentrations of trehalose alone, or a mixture of sugars and amino acids, as excipients can convert vaccines containing aluminum salts into dry powder, but fails to preserve the particle size and/or immunogenicity of the vaccines. In the present study, using ovalbumin as a model antigen adsorbed onto aluminum hydroxide or aluminum phosphate, a commercially available tetanus toxoid vaccine adjuvanted with potassium alum, a human hepatitis B vaccine adjuvanted with aluminum hydroxide, and a human papillomavirus vaccine adjuvanted with aluminum hydroxyphosphate sulfate, it was shown that vaccines containing a relatively high concentration of aluminum salts (i.e., up to ~1%, w/v, of aluminum hydroxide) can be converted into a dry powder by thin-film freezing followed by removal of the frozen solvent by lyophilization while using low levels of trehalose (i.e., as low as 2% w/v) as an excipient. Importantly, the thin-film freeze-drying process did not cause particle aggregation, nor decreased the immunogenicity of the vaccines. Moreover, repeated freezing-and-thawing of the dry vaccine powder did not cause aggregation. Thin-film freeze-drying is a viable platform technology to produce dry powders of vaccines that contain aluminum salts.
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Adyuvantes Farmacéuticos/química , Compuestos de Alumbre/química , Vacunas contra Hepatitis B , Tecnología Farmacéutica/métodos , Toxoide Tetánico , Hidróxido de Aluminio/química , Animales , Rastreo Diferencial de Calorimetría , Composición de Medicamentos , Estabilidad de Medicamentos , Femenino , Liofilización , Vacunas contra Hepatitis B/química , Vacunas contra Hepatitis B/inmunología , Inmunoglobulina G/sangre , Ratones Endogámicos BALB C , Microscopía Electrónica de Rastreo , Ovalbúmina/inmunología , Tamaño de la Partícula , Fosfatos/química , Polvos , Toxoide Tetánico/química , Toxoide Tetánico/inmunologíaRESUMEN
Pneumocystis spp. are significant pathogens in a variety of mammals. We tried to establish a Drosophila model of pneumocystosis using either P. murina or P. carinii. Whereas the pathogens were competent in susceptible mice, no infection could be established even in corticosteroid-treated Toll-deficient flies. This further substantiates the tropism described for mammalian hosts.