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1.
Nature ; 595(7868): 572-577, 2021 07.
Artículo en Inglés | MEDLINE | ID: mdl-34044428

RESUMEN

BNT162b2, a nucleoside-modified mRNA formulated in lipid nanoparticles that encodes the SARS-CoV-2 spike glycoprotein (S) stabilized in its prefusion conformation, has demonstrated 95% efficacy in preventing COVID-191. Here we extend a previous phase-I/II trial report2 by presenting data on the immune response induced by BNT162b2 prime-boost vaccination from an additional phase-I/II trial in healthy adults (18-55 years old). BNT162b2 elicited strong antibody responses: at one week after the boost, SARS-CoV-2 serum geometric mean 50% neutralizing titres were up to 3.3-fold above those observed in samples from individuals who had recovered from COVID-19. Sera elicited by BNT162b2 neutralized 22 pseudoviruses bearing the S of different SARS-CoV-2 variants. Most participants had a strong response of IFNγ+ or IL-2+ CD8+ and CD4+ T helper type 1 cells, which was detectable throughout the full observation period of nine weeks following the boost. Using peptide-MHC multimer technology, we identified several BNT162b2-induced epitopes that were presented by frequent MHC alleles and conserved in mutant strains. One week after the boost, epitope-specific CD8+ T cells of the early-differentiated effector-memory phenotype comprised 0.02-2.92% of total circulating CD8+ T cells and were detectable (0.01-0.28%) eight weeks later. In summary, BNT162b2 elicits an adaptive humoral and poly-specific cellular immune response against epitopes that are conserved in a broad range of variants, at well-tolerated doses.


Asunto(s)
Anticuerpos Neutralizantes/inmunología , Anticuerpos Antivirales/inmunología , Vacunas contra la COVID-19/inmunología , COVID-19/inmunología , SARS-CoV-2/inmunología , Linfocitos T/inmunología , Adolescente , Adulto , Vacuna BNT162 , Linfocitos T CD8-positivos/inmunología , COVID-19/virología , Vacunas contra la COVID-19/administración & dosificación , Vacunas contra la COVID-19/efectos adversos , Epítopos de Linfocito T/inmunología , Femenino , Humanos , Inmunoglobulina G/inmunología , Memoria Inmunológica , Interferón gamma/inmunología , Interleucina-2/inmunología , Masculino , Persona de Mediana Edad , SARS-CoV-2/química , Glicoproteína de la Espiga del Coronavirus/química , Glicoproteína de la Espiga del Coronavirus/inmunología , Células TH1/inmunología , Adulto Joven
2.
Nature ; 592(7853): 283-289, 2021 04.
Artículo en Inglés | MEDLINE | ID: mdl-33524990

RESUMEN

A safe and effective vaccine against COVID-19 is urgently needed in quantities that are sufficient to immunize large populations. Here we report the preclinical development of two vaccine candidates (BNT162b1 and BNT162b2) that contain nucleoside-modified messenger RNA that encodes immunogens derived from the spike glycoprotein (S) of SARS-CoV-2, formulated in lipid nanoparticles. BNT162b1 encodes a soluble, secreted trimerized receptor-binding domain (known as the RBD-foldon). BNT162b2 encodes the full-length transmembrane S glycoprotein, locked in its prefusion conformation by the substitution of two residues with proline (S(K986P/V987P); hereafter, S(P2) (also known as P2 S)). The flexibly tethered RBDs of the RBD-foldon bind to human ACE2 with high avidity. Approximately 20% of the S(P2) trimers are in the two-RBD 'down', one-RBD 'up' state. In mice, one intramuscular dose of either candidate vaccine elicits a dose-dependent antibody response with high virus-entry inhibition titres and strong T-helper-1 CD4+ and IFNγ+CD8+ T cell responses. Prime-boost vaccination of rhesus macaques (Macaca mulatta) with the BNT162b candidates elicits SARS-CoV-2-neutralizing geometric mean titres that are 8.2-18.2× that of a panel of SARS-CoV-2-convalescent human sera. The vaccine candidates protect macaques against challenge with SARS-CoV-2; in particular, BNT162b2 protects the lower respiratory tract against the presence of viral RNA and shows no evidence of disease enhancement. Both candidates are being evaluated in phase I trials in Germany and the USA1-3, and BNT162b2 is being evaluated in an ongoing global phase II/III trial (NCT04380701 and NCT04368728).


Asunto(s)
Vacunas contra la COVID-19/inmunología , COVID-19/inmunología , COVID-19/prevención & control , Modelos Animales de Enfermedad , SARS-CoV-2/inmunología , Envejecimiento/inmunología , Animales , Anticuerpos Neutralizantes/inmunología , Anticuerpos Antivirales/inmunología , Antígenos Virales/química , Antígenos Virales/genética , Antígenos Virales/inmunología , Vacuna BNT162 , COVID-19/sangre , COVID-19/terapia , COVID-19/virología , Vacunas contra la COVID-19/administración & dosificación , Vacunas contra la COVID-19/química , Vacunas contra la COVID-19/genética , Línea Celular , Ensayos Clínicos como Asunto , Femenino , Humanos , Inmunización Pasiva , Internacionalidad , Macaca mulatta/inmunología , Macaca mulatta/virología , Masculino , Ratones , Ratones Endogámicos BALB C , Modelos Moleculares , Multimerización de Proteína , ARN Viral/análisis , Sistema Respiratorio/inmunología , Sistema Respiratorio/virología , SARS-CoV-2/química , SARS-CoV-2/genética , Solubilidad , Glicoproteína de la Espiga del Coronavirus/química , Glicoproteína de la Espiga del Coronavirus/genética , Glicoproteína de la Espiga del Coronavirus/inmunología , Linfocitos T/inmunología , Vacunación , Vacunas Sintéticas/administración & dosificación , Vacunas Sintéticas/química , Vacunas Sintéticas/genética , Vacunas Sintéticas/inmunología , Sueroterapia para COVID-19 , Vacunas de ARNm
3.
Nature ; 585(7823): 107-112, 2020 09.
Artículo en Inglés | MEDLINE | ID: mdl-32728218

RESUMEN

Treating patients who have cancer with vaccines that stimulate a targeted immune response is conceptually appealing, but cancer vaccine trials have not been successful in late-stage patients with treatment-refractory tumours1,2. We are testing melanoma FixVac (BNT111)-an intravenously administered liposomal RNA (RNA-LPX) vaccine, which targets four non-mutated, tumour-associated antigens that are prevalent in melanoma-in an ongoing, first-in-human, dose-escalation phase I trial in patients with advanced melanoma (Lipo-MERIT trial, ClinicalTrials.gov identifier NCT02410733). We report here data from an exploratory interim analysis that show that melanoma FixVac, alone or in combination with blockade of the checkpoint inhibitor PD1, mediates durable objective responses in checkpoint-inhibitor (CPI)-experienced patients with unresectable melanoma. Clinical responses are accompanied by the induction of strong CD4+ and CD8+ T cell immunity against the vaccine antigens. The antigen-specific cytotoxic T-cell responses in some responders reach magnitudes typically reported for adoptive T-cell therapy, and are durable. Our findings indicate that RNA-LPX vaccination is a potent immunotherapy in patients with CPI-experienced melanoma, and suggest the general utility of non-mutant shared tumour antigens as targets for cancer vaccination.


Asunto(s)
Antineoplásicos/uso terapéutico , Vacunas contra el Cáncer/genética , Vacunas contra el Cáncer/inmunología , Melanoma/inmunología , Melanoma/terapia , Receptor de Muerte Celular Programada 1/antagonistas & inhibidores , ARN Neoplásico/genética , Linfocitos T/inmunología , Antígenos de Neoplasias/inmunología , Antineoplásicos/farmacología , Vacunas contra el Cáncer/administración & dosificación , Vacunas contra el Cáncer/efectos adversos , Terapia Combinada , Humanos , Melanoma/tratamiento farmacológico , Melanoma/patología , Estadificación de Neoplasias , Linfocitos T/citología , Linfocitos T Citotóxicos/citología , Linfocitos T Citotóxicos/inmunología , Vacunación
4.
Nature ; 586(7830): 594-599, 2020 10.
Artículo en Inglés | MEDLINE | ID: mdl-32998157

RESUMEN

An effective vaccine is needed to halt the spread of the severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) pandemic. Recently, we reported safety, tolerability and antibody response data from an ongoing placebo-controlled, observer-blinded phase I/II coronavirus disease 2019 (COVID-19) vaccine trial with BNT162b1, a lipid nanoparticle-formulated nucleoside-modified mRNA that encodes the receptor binding domain (RBD) of the SARS-CoV-2 spike protein1. Here we present antibody and T cell responses after vaccination with BNT162b1 from a second, non-randomized open-label phase I/II trial in healthy adults, 18-55 years of age. Two doses of 1-50 µg of BNT162b1 elicited robust CD4+ and CD8+ T cell responses and strong antibody responses, with RBD-binding IgG concentrations clearly above those seen in serum from a cohort of individuals who had recovered from COVID-19. Geometric mean titres of SARS-CoV-2 serum-neutralizing antibodies on day 43 were 0.7-fold (1-µg dose) to 3.5-fold (50-µg dose) those of the recovered individuals. Immune sera broadly neutralized pseudoviruses with diverse SARS-CoV-2 spike variants. Most participants had T helper type 1 (TH1)-skewed T cell immune responses with RBD-specific CD8+ and CD4+ T cell expansion. Interferon-γ was produced by a large fraction of RBD-specific CD8+ and CD4+ T cells. The robust RBD-specific antibody, T cell and favourable cytokine responses induced by the BNT162b1 mRNA vaccine suggest that it has the potential to protect against COVID-19 through multiple beneficial mechanisms.


Asunto(s)
Anticuerpos Antivirales/inmunología , Infecciones por Coronavirus/inmunología , Neumonía Viral/inmunología , Células TH1/inmunología , Vacunas Virales/inmunología , Adulto , Anticuerpos Neutralizantes/inmunología , Linfocitos T CD8-positivos/citología , Linfocitos T CD8-positivos/inmunología , COVID-19 , Vacunas contra la COVID-19 , Infecciones por Coronavirus/prevención & control , Citocinas/inmunología , Femenino , Alemania , Humanos , Inmunoglobulina G/inmunología , Masculino , Persona de Mediana Edad , Pandemias , Células TH1/citología , Vacunas Virales/administración & dosificación , Vacunas Virales/efectos adversos , Adulto Joven
5.
Nature ; 547(7662): 222-226, 2017 07 13.
Artículo en Inglés | MEDLINE | ID: mdl-28678784

RESUMEN

T cells directed against mutant neo-epitopes drive cancer immunity. However, spontaneous immune recognition of mutations is inefficient. We recently introduced the concept of individualized mutanome vaccines and implemented an RNA-based poly-neo-epitope approach to mobilize immunity against a spectrum of cancer mutations. Here we report the first-in-human application of this concept in melanoma. We set up a process comprising comprehensive identification of individual mutations, computational prediction of neo-epitopes, and design and manufacturing of a vaccine unique for each patient. All patients developed T cell responses against multiple vaccine neo-epitopes at up to high single-digit percentages. Vaccine-induced T cell infiltration and neo-epitope-specific killing of autologous tumour cells were shown in post-vaccination resected metastases from two patients. The cumulative rate of metastatic events was highly significantly reduced after the start of vaccination, resulting in a sustained progression-free survival. Two of the five patients with metastatic disease experienced vaccine-related objective responses. One of these patients had a late relapse owing to outgrowth of ß2-microglobulin-deficient melanoma cells as an acquired resistance mechanism. A third patient developed a complete response to vaccination in combination with PD-1 blockade therapy. Our study demonstrates that individual mutations can be exploited, thereby opening a path to personalized immunotherapy for patients with cancer.


Asunto(s)
Vacunas contra el Cáncer/genética , Vacunas contra el Cáncer/inmunología , Melanoma/inmunología , Melanoma/terapia , Mutación/genética , Medicina de Precisión/métodos , ARN/genética , Anticuerpos Monoclonales/farmacología , Anticuerpos Monoclonales/uso terapéutico , Antígeno B7-H1/inmunología , Antígenos CD8/inmunología , Vacunas contra el Cáncer/uso terapéutico , Epítopos/genética , Epítopos/inmunología , Humanos , Inmunoterapia/métodos , Melanoma/genética , Metástasis de la Neoplasia , Recurrencia Local de Neoplasia/prevención & control , Nivolumab , Receptor de Muerte Celular Programada 1/antagonistas & inhibidores , Linfocitos T/inmunología , Vacunación , Microglobulina beta-2/deficiencia
6.
Gastroenterology ; 160(1): 331-345.e6, 2021 01.
Artículo en Inglés | MEDLINE | ID: mdl-33010248

RESUMEN

BACKGROUND & AIMS: Nonalcoholic steatohepatitis causes loss of hepatic CD4+ T cells and promotes tumor growth. The liver is the most common site of distant metastases from a variety of malignancies, many of which respond to immunotherapy. We investigated the effects of steatohepatitis on the efficacy of immunotherapeutic agents against liver tumors in mice. METHODS: Steatohepatitis was induced by feeding C57BL/6NCrl or BALB/c AnNCr mice a methionine and choline-deficient diet or a choline-deficient l-amino acid-defined diet. Mice were given intrahepatic or subcutaneous injections of B16 melanoma and CT26 colon cancer cells, followed by intravenous injections of M30-RNA vaccine (M30) or intraperitoneal injections of an antibody against OX40 (aOX40) on days 3, 7, and 10 after injection of the tumor cells. We measured tumor growth and analyzed immune cells in tumor tissues by flow cytometry. Mice were given N-acetylcysteine to prevent loss of CD4+ T cells from liver. RESULTS: Administration of M30 and aOX40 inhibited growth of tumors from intrahepatic injections of B16 or CT26 cells in mice on regular diet. However, M30 and/or aOX40 did not slow growth of liver tumors from B16 or CT26 cells in mice with diet-induced steatohepatitis (methionine and choline-deficient diet or choline-deficient l-amino acid-defined diet). Steatohepatitis did not affect the ability of M30 to slow growth of subcutaneous B16 tumors. In mice with steatohepatitis given N-acetylcysteine, which prevents loss of CD4+ T cells, M30 and aOX40 were able slow growth of hepatic tumors. Flow cytometry analysis of liver tumors revealed reduced CD4+ T cells and effector memory cells in mice with vs without steatohepatitis. CONCLUSIONS: Steatohepatitis reduces the abilities of immunotherapeutic agents, such as M30 and aOX40, to inhibit tumor liver growth by reducing tumor infiltration by CD4+ T cells and effector memory cells. N-acetylcysteine restores T-cell numbers in tumors and increases the ability of M30 and aOX40 to slow tumor growth in mice.


Asunto(s)
Inmunoterapia , Neoplasias Hepáticas/etiología , Neoplasias Hepáticas/terapia , Melanoma/terapia , Enfermedad del Hígado Graso no Alcohólico/complicaciones , Linfocitos T/fisiología , Animales , Modelos Animales de Enfermedad , Neoplasias Hepáticas/patología , Melanoma/etiología , Melanoma/patología , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Enfermedad del Hígado Graso no Alcohólico/patología
7.
Nature ; 534(7607): 396-401, 2016 06 16.
Artículo en Inglés | MEDLINE | ID: mdl-27281205

RESUMEN

Lymphoid organs, in which antigen presenting cells (APCs) are in close proximity to T cells, are the ideal microenvironment for efficient priming and amplification of T-cell responses. However, the systemic delivery of vaccine antigens into dendritic cells (DCs) is hampered by various technical challenges. Here we show that DCs can be targeted precisely and effectively in vivo using intravenously administered RNA-lipoplexes (RNA-LPX) based on well-known lipid carriers by optimally adjusting net charge, without the need for functionalization of particles with molecular ligands. The LPX protects RNA from extracellular ribonucleases and mediates its efficient uptake and expression of the encoded antigen by DC populations and macrophages in various lymphoid compartments. RNA-LPX triggers interferon-α (IFNα) release by plasmacytoid DCs and macrophages. Consequently, DC maturation in situ and inflammatory immune mechanisms reminiscent of those in the early systemic phase of viral infection are activated. We show that RNA-LPX encoding viral or mutant neo-antigens or endogenous self-antigens induce strong effector and memory T-cell responses, and mediate potent IFNα-dependent rejection of progressive tumours. A phase I dose-escalation trial testing RNA-LPX that encode shared tumour antigens is ongoing. In the first three melanoma patients treated at a low-dose level, IFNα and strong antigen-specific T-cell responses were induced, supporting the identified mode of action and potency. As any polypeptide-based antigen can be encoded as RNA, RNA-LPX represent a universally applicable vaccine class for systemic DC targeting and synchronized induction of both highly potent adaptive as well as type-I-IFN-mediated innate immune mechanisms for cancer immunotherapy.


Asunto(s)
Antígenos de Neoplasias/inmunología , Antígenos Virales/inmunología , Células Dendríticas/inmunología , Células Dendríticas/metabolismo , Inmunoterapia/métodos , Melanoma/inmunología , Melanoma/terapia , ARN/administración & dosificación , Administración Intravenosa , Animales , Presentación de Antígeno/inmunología , Antígenos de Neoplasias/genética , Antígenos Virales/genética , Autoantígenos/genética , Autoantígenos/inmunología , Vacunas contra el Cáncer/administración & dosificación , Vacunas contra el Cáncer/genética , Ensayos Clínicos Fase I como Asunto , Células Dendríticas/citología , Modelos Animales de Enfermedad , Portadores de Fármacos/administración & dosificación , Femenino , Humanos , Interferón Tipo I/inmunología , Interferón Tipo I/metabolismo , Activación de Linfocitos/inmunología , Tejido Linfoide/citología , Tejido Linfoide/inmunología , Macrófagos/inmunología , Macrófagos/metabolismo , Masculino , Glicoproteínas de Membrana/inmunología , Ratones , Ratones Endogámicos C57BL , Nanopartículas/administración & dosificación , ARN/genética , Electricidad Estática , Linfocitos T/citología , Linfocitos T/inmunología , Receptor Toll-Like 7/inmunología
9.
Mol Cancer ; 20(1): 69, 2021 04 15.
Artículo en Inglés | MEDLINE | ID: mdl-33858437

RESUMEN

Synthetic mRNA provides a template for the synthesis of any given protein, protein fragment or peptide and lends itself to a broad range of pharmaceutical applications, including different modalities of cancer immunotherapy. With the ease of rapid, large scale Good Manufacturing Practice-grade mRNA production, mRNA is ideally poised not only for off-the shelf cancer vaccines but also for personalized neoantigen vaccination. The ability to stimulate pattern recognition receptors and thus an anti-viral type of innate immune response equips mRNA-based vaccines with inherent adjuvanticity. Nucleoside modification and elimination of double-stranded RNA can reduce the immunomodulatory activity of mRNA and increase and prolong protein production. In combination with nanoparticle-based formulations that increase transfection efficiency and facilitate lymphatic system targeting, nucleoside-modified mRNA enables efficient delivery of cytokines, costimulatory receptors, or therapeutic antibodies. Steady but transient production of the encoded bioactive molecule from the mRNA template can improve the pharmacokinetic, pharmacodynamic and safety properties as compared to the respective recombinant proteins. This may be harnessed for applications that benefit from a higher level of expression control, such as chimeric antigen receptor (CAR)-modified adoptive T-cell therapies. This review highlights the advancements in the field of mRNA-based cancer therapeutics, providing insights into key preclinical developments and the evolving clinical landscape.


Asunto(s)
Terapia Genética , Inmunoterapia , Neoplasias/etiología , Neoplasias/terapia , ARN Mensajero/administración & dosificación , Animales , Anticuerpos/genética , Anticuerpos/inmunología , Presentación de Antígeno/inmunología , Células Presentadoras de Antígenos/inmunología , Células Presentadoras de Antígenos/metabolismo , Antígenos de Neoplasias/genética , Antígenos de Neoplasias/inmunología , Biomarcadores de Tumor , Vacunas contra el Cáncer/administración & dosificación , Vacunas contra el Cáncer/genética , Vacunas contra el Cáncer/inmunología , Citocinas/metabolismo , Células Dendríticas/inmunología , Células Dendríticas/metabolismo , Terapia Genética/métodos , Humanos , Factores Inmunológicos/genética , Inmunoterapia/métodos , Neoplasias/patología , ARN Mensajero/química , ARN Mensajero/genética , ARN Mensajero/inmunología , Receptores de Antígenos de Linfocitos T/genética , Receptores de Antígenos de Linfocitos T/inmunología , Linfocitos T/inmunología , Linfocitos T/metabolismo
10.
Annu Rev Med ; 70: 395-407, 2019 01 27.
Artículo en Inglés | MEDLINE | ID: mdl-30691374

RESUMEN

T cells are key effectors of anticancer immunity. They are capable of distinguishing tumor cells from normal ones by recognizing major histocompatibility complex-bound cancer-specific peptides. Accumulating evidence suggests that peptides associated with T cell-mediated tumor rejection arise predominantly from somatically mutated proteins and are unique to every patient's tumor. Knowledge of an individual's cancer mutanome (the entirety of cancer mutations) allows harnessing this enormous tumor cell-specific repertoire of highly immunogenic antigens for individualized cancer vaccines. This review outlines the preclinical and clinical state of individualized cancer vaccine development and the challenges ahead.


Asunto(s)
Antígenos de Neoplasias/genética , Inmunoterapia/métodos , Neoplasias/genética , Neoplasias/terapia , Medicina de Precisión/tendencias , Animales , Vacunas contra el Cáncer/administración & dosificación , Vacunas contra el Cáncer/inmunología , Predicción , Humanos , Terapia Molecular Dirigida/métodos , Mutación , Neoplasias/patología , Linfocitos T/inmunología , Resultado del Tratamiento
11.
Nature ; 520(7549): 692-6, 2015 Apr 30.
Artículo en Inglés | MEDLINE | ID: mdl-25901682

RESUMEN

Tumour-specific mutations are ideal targets for cancer immunotherapy as they lack expression in healthy tissues and can potentially be recognized as neo-antigens by the mature T-cell repertoire. Their systematic targeting by vaccine approaches, however, has been hampered by the fact that every patient's tumour possesses a unique set of mutations ('the mutanome') that must first be identified. Recently, we proposed a personalized immunotherapy approach to target the full spectrum of a patient's individual tumour-specific mutations. Here we show in three independent murine tumour models that a considerable fraction of non-synonymous cancer mutations is immunogenic and that, unexpectedly, the majority of the immunogenic mutanome is recognized by CD4(+) T cells. Vaccination with such CD4(+) immunogenic mutations confers strong antitumour activity. Encouraged by these findings, we established a process by which mutations identified by exome sequencing could be selected as vaccine targets solely through bioinformatic prioritization on the basis of their expression levels and major histocompatibility complex (MHC) class II-binding capacity for rapid production as synthetic poly-neo-epitope messenger RNA vaccines. We show that vaccination with such polytope mRNA vaccines induces potent tumour control and complete rejection of established aggressively growing tumours in mice. Moreover, we demonstrate that CD4(+) T cell neo-epitope vaccination reshapes the tumour microenvironment and induces cytotoxic T lymphocyte responses against an independent immunodominant antigen in mice, indicating orchestration of antigen spread. Finally, we demonstrate an abundance of mutations predicted to bind to MHC class II in human cancers as well by employing the same predictive algorithm on corresponding human cancer types. Thus, the tailored immunotherapy approach introduced here may be regarded as a universally applicable blueprint for comprehensive exploitation of the substantial neo-epitope target repertoire of cancers, enabling the effective targeting of every patient's tumour with vaccines produced 'just in time'.


Asunto(s)
Epítopos de Linfocito T/genética , Antígenos de Histocompatibilidad Clase II/genética , Antígenos de Histocompatibilidad Clase II/inmunología , Inmunoterapia/métodos , Melanoma Experimental/inmunología , Melanoma Experimental/terapia , Mutación/genética , Algoritmos , Animales , Linfocitos T CD4-Positivos/inmunología , Vacunas contra el Cáncer/genética , Vacunas contra el Cáncer/inmunología , Simulación por Computador , Modelos Animales de Enfermedad , Epítopos de Linfocito T/inmunología , Exoma/genética , Femenino , Antígenos de Histocompatibilidad Clase II/metabolismo , Humanos , Melanoma Experimental/genética , Ratones , Medicina de Precisión/métodos , Análisis de Secuencia de ADN , Análisis de Supervivencia
12.
Recent Results Cancer Res ; 214: 153-167, 2020.
Artículo en Inglés | MEDLINE | ID: mdl-31473852

RESUMEN

After more than a century of efforts to establish cancer immunotherapy in clinical practice, the advent of checkpoint inhibition (CPI) therapy was a critical breakthrough toward this direction (Hodi et al. in Cell Rep 13(2):412-424, 2010; Wolchok et al. in N Engl J Med 369(2):122-133, 2013; Herbst et al. in Nature 515(7528):563-567, 2014; Tumeh et al. in Nature 515(7528):568-571, 2014). Further, CPIs shifted the focus from long studied shared tumor-associated antigens to mutated ones. As cancer is caused by mutations in somatic cells, the concept to utilize these correlates of 'foreignness' to enable recognition and lysis of the cancer cell by T cell immunity seems an obvious thing to do.


Asunto(s)
Vacunas contra el Cáncer , Epítopos/inmunología , Inmunoterapia , Neoplasias/terapia , Antígenos de Neoplasias/inmunología , Humanos
13.
BMC Cancer ; 19(1): 914, 2019 Sep 13.
Artículo en Inglés | MEDLINE | ID: mdl-31519152

RESUMEN

BACKGROUND: NY-BR-1 has been described as a breast cancer associated differentiation antigen with intrinsic immunogenicity giving rise to endogenous T and B cell responses. The current study presents the first murine tumor model allowing functional investigation of NY-BR-1-specific immune responses in vivo. METHODS: A NY-BR-1 expressing tumor model was established in DR4tg mice based on heterotopic transplantation of stable transfectant clones derived from the murine H2 compatible breast cancer cell line EO771. Composition and phenotype of tumor infiltrating immune cells were analyzed by qPCR and FACS. MHC I binding affinity of candidate CTL epitopes predicted in silico was determined by FACS using the mutant cell line RMA-S. Frequencies of NY-BR-1 specific CTLs among splenocytes of immunized mice were quantified by FACS with an epitope loaded Db-dextramer. Functional CTL activity was determined by IFNγ catch or IFNγ ELISpot assays and statistical analysis was done applying the Mann Whitney test. Tumor protection experiments were performed by immunization of DR4tg mice with replication deficient recombinant adenovirus followed by s.c. challenge with NY-BR-1 expressing breast cancer cells. RESULTS: Our results show spontaneous accumulation of CD8+ T cells and F4/80+ myeloid cells preferentially in NY-BR-1 expressing tumors. Upon NY-BR-1-specific immunization experiments combined with in silico prediction and in vitro binding assays, the first NY-BR-1-specific H2-Db-restricted T cell epitope could be identified. Consequently, flow cytometric analysis with fluorochrome conjugated multimers showed enhanced frequencies of CD8+ T cells specific for the newly identified epitope in spleens of immunized mice. Moreover, immunization with Ad.NY-BR-1 resulted in partial protection against outgrowth of NY-BR-1 expressing tumors and promoted intratumoral accumulation of macrophages. CONCLUSION: This study introduces the first H2-Db-resctricted CD8+ T cell epitope-specific for the human breast cancer associated tumor antigen NY-BR-1. Our novel, partially humanized tumor model enables investigation of the interplay between HLA-DR4-restricted T cell responses and CTLs within their joint attack of NY-BR-1 expressing tumors.


Asunto(s)
Antígenos de Neoplasias/inmunología , Epítopos de Linfocito T/inmunología , Cadenas HLA-DRB1/genética , Neoplasias/etiología , Neoplasias/patología , Linfocitos T/inmunología , Linfocitos T/metabolismo , Animales , Antígenos de Neoplasias/genética , Biomarcadores , Línea Celular Tumoral , Modelos Animales de Enfermedad , Cadenas HLA-DRB1/inmunología , Xenoinjertos , Humanos , Inmunización , Inmunofenotipificación , Leucocitos/inmunología , Leucocitos/metabolismo , Ratones , Ratones Transgénicos , Subgrupos de Linfocitos T/inmunología , Subgrupos de Linfocitos T/metabolismo
15.
Cancer Immunol Res ; 12(10): 1409-1420, 2024 Oct 01.
Artículo en Inglés | MEDLINE | ID: mdl-38885358

RESUMEN

Interleukin 2 (IL-2) is a crucial cytokine in T-cell immunity, with a promising potential in cancer vaccines. However, therapeutic application of IL-2 is hampered by its short half-life and substantial toxicity. This study reports preclinical characterization of a mouse serum albumin-IL-2 fusion protein (Alb-IL2) encoded on nucleoside-modified RNA that is delivered via a nanoparticle formulation (Alb-IL2 RNA-NP) mediating prolonged cytokine availability. Alb-IL2 RNA-NP was combined with RNA-lipoplex (RNA-LPX) vaccines to evaluate its effect on the expansion of vaccine-induced antigen specific T-cell immunity. In mice dosed with Alb-IL2 RNA-NP, translated protein was shown to be systemically available up to 2 days, with an albumin-dependent preferred presence in the tumor and tumor-draining lymph node. Alb-IL2 RNA-NP administration prolonged serum availability of the cytokine compared with murine recombinant IL-2. In combination with RNA-LPX vaccines, Alb-IL2 RNA-NP administration highly increased the expansion of RNA-LPX vaccine-induced CD8+ T cells in the spleen and blood. The combination enhanced and sustained the fraction of IL-2 receptor (IL-2R) α-positive antigen-specific CD8+ T cells and ameliorated the functional capacity of the CD8+ T-cell population. Alb-IL2 RNA-NP strongly improved the antitumor activity and survival of concomitant RNA-LPX vaccination and PD-L1 blockade in a subcutaneous mouse tumor model. The favorable pharmacokinetic properties of Alb-IL2 RNA-NP render it an attractive modality for rationally designed combination immunotherapy. RNA vaccines that induce tumor-specific T-cell immunity for Alb-IL2 RNA-NP to further amplify are particularly attractive combination partners.


Asunto(s)
Vacunas contra el Cáncer , Interleucina-2 , Animales , Vacunas contra el Cáncer/inmunología , Ratones , Femenino , Ratones Endogámicos C57BL , Linfocitos T CD8-positivos/inmunología , Vacunas de ARNm , Línea Celular Tumoral , Disponibilidad Biológica , Linfocitos T/inmunología , Humanos , ARN/genética
16.
Cancer Cell ; 42(4): 568-582.e11, 2024 Apr 08.
Artículo en Inglés | MEDLINE | ID: mdl-38490213

RESUMEN

Major histocompatibility complex (MHC) class I antigen presentation deficiency is a common cancer immune escape mechanism, but the mechanistic implications and potential strategies to address this challenge remain poorly understood. Studying ß2-microglobulin (B2M) deficient mouse tumor models, we find that MHC class I loss leads to a substantial immune desertification of the tumor microenvironment (TME) and broad resistance to immune-, chemo-, and radiotherapy. We show that treatment with long-lasting mRNA-encoded interleukin-2 (IL-2) restores an immune cell infiltrated, IFNγ-promoted, highly proinflammatory TME signature, and when combined with a tumor-targeting monoclonal antibody (mAB), can overcome therapeutic resistance. Unexpectedly, the effectiveness of this treatment is driven by IFNγ-releasing CD8+ T cells that recognize neoantigens cross-presented by TME-resident activated macrophages. These macrophages acquire augmented antigen presentation proficiency and other M1-phenotype-associated features under IL-2 treatment. Our findings highlight the importance of restoring neoantigen-specific immune responses in the treatment of cancers with MHC class I deficiencies.


Asunto(s)
Linfocitos T CD8-positivos , Neoplasias , Animales , Ratones , Antígenos de Histocompatibilidad Clase I/genética , Interleucina-2/genética , Interleucina-2/inmunología , Neoplasias/genética , ARN Mensajero , Microambiente Tumoral
17.
Front Immunol ; 14: 1102282, 2023.
Artículo en Inglés | MEDLINE | ID: mdl-36969213

RESUMEN

Introduction: The cell line MC38 is a commonly used murine model for colorectal carcinoma. It has a high mutational burden, is sensitive to immune checkpoint immunotherapy and endogenous CD8+ T cell responses against neoantigens have been reported. Methods: Here, we re-sequenced exomes and transcriptomes of MC38 cells from two different sources, namely Kerafast (originating from NCI/NIH, MC38-K) and the Leiden University Medical Center cell line collection (MC38-L), comparing the cell lines on the genomic and transcriptomic level and analyzing their recognition by CD8+ T cells with known neo-epitope specificity. Results: The data reveals a distinct structural composition of MC38-K and MC38-L cell line genomes and different ploidies. Further, the MC38-L cell line harbored about 1.3-fold more single nucleotide variations and small insertions and deletions than the MC38-K cell line. In addition, the observed mutational signatures differed; only 35.3% of the non-synonymous variants and 5.4% of the fusion gene events were shared. Transcript expression values of both cell lines correlated strongly (p = 0.919), but we found different pathways enriched in the genes that were differentially upregulated in the MC38-L or MC38-K cells, respectively. Our data show that previously described neoantigens in the MC38 model such as Rpl18mut and Adpgkmut were absent in the MC38-K cell line resulting that such neoantigen-specific CD8+ T cells recognizing and killing MC38-L cells did not recognize or kill MC38-K cells. Conclusion: This strongly indicates that at least two sub-cell lines of MC38 exist in the field and underlines the importance of meticulous tracking of investigated cell lines to obtain reproducible results, and for correct interpretation of the immunological data without artifacts. We present our analyses as a reference for researchers to select the appropriate sub-cell line for their own studies.


Asunto(s)
Neoplasias Colorrectales , Transcriptoma , Humanos , Animales , Ratones , Linfocitos T CD8-positivos , Línea Celular Tumoral , Mutación
18.
Oncoimmunology ; 11(1): 2030135, 2022.
Artículo en Inglés | MEDLINE | ID: mdl-35186440

RESUMEN

Immune checkpoint inhibitors (ICI) targeting the PD-1/PD-L1 axis have changed the treatment paradigm for advanced solid tumors; however, many patients experience treatment resistance. In preclinical models 4-1BB co-stimulation synergizes with ICI by activating cytotoxic T- and NK-cell-mediated anti-tumor immunity. Here we characterize the mechanism of action of a mouse-reactive Fc-inert PD-L1×4-1BB bispecific antibody (mbsAb-PD-L1×4-1BB) and provide proof-of-concept for enhanced anti-tumor activity. In reporter assays mbsAb-PD-L1×4-1BB exhibited conditional 4-1BB agonist activity that was dependent on simultaneous binding to PD-L1. mbsAb-PD-L1×4-1BB further blocked the PD-L1/PD-1 interaction independently of 4-1BB binding. By combining both mechanisms, mbsAb-PD-L1×4-1BB strongly enhanced T-cell proliferation, cytokine production and antigen-specific cytotoxicity using primary mouse cells in vitro. Furthermore, mbsAb-PD-L1×4-1BB exhibited potent anti-tumor activity in the CT26 and MC38 models in vivo, leading to the rejection of CT26 tumors that were unresponsive to PD-L1 blockade alone. Anti-tumor activity was associated with increased tumor-specific CD8+ T cells and reduced regulatory T cells within the tumor microenvironment and tumor-draining lymph nodes. In immunocompetent tumor-free mice, mbsAb-PD-L1×4-1BB treatment neither induced T-cell infiltration into the liver nor elevated liver enzymes in the blood. Dual targeting of PD-L1 and 4-1BB with a bispecific antibody may therefore address key limitations of first generation 4-1BB-agonistic antibodies, and may provide a novel approach to improve PD-1/PD-L1 checkpoint blockade.


Asunto(s)
Anticuerpos Biespecíficos , Neoplasias , Animales , Anticuerpos Biespecíficos/farmacología , Anticuerpos Biespecíficos/uso terapéutico , Antígeno B7-H1 , Linfocitos T CD8-positivos , Humanos , Inhibidores de Puntos de Control Inmunológico/farmacología , Inhibidores de Puntos de Control Inmunológico/uso terapéutico , Ratones , Neoplasias/tratamiento farmacológico , Receptor de Muerte Celular Programada 1/uso terapéutico , Microambiente Tumoral
19.
Front Oncol ; 10: 1195, 2020.
Artículo en Inglés | MEDLINE | ID: mdl-32793490

RESUMEN

Background: Tumor models are critical for our understanding of cancer and the development of cancer therapeutics. The 4T1 murine mammary cancer cell line is one of the most widely used breast cancer models. Here, we present an integrated map of the genome, transcriptome, and immunome of 4T1. Results: We found Trp53 (Tp53) and Pik3g to be mutated. Other frequently mutated genes in breast cancer, including Brca1 and Brca2, are not mutated. For cancer related genes, Nav3, Cenpf, Muc5Ac, Mpp7, Gas1, MageD2, Dusp1, Ros, Polr2a, Rragd, Ros1, and Hoxa9 are mutated. Markers for cell proliferation like Top2a, Birc5, and Mki67 are highly expressed, so are markers for metastasis like Msln, Ect2, and Plk1, which are known to be overexpressed in triple-negative breast cancer (TNBC). TNBC markers are, compared to a mammary gland control sample, lower (Esr1), comparably low (Erbb2), or not expressed at all (Pgr). We also found testis cancer antigen Pbk as well as colon/gastrointestinal cancer antigens Gpa33 and Epcam to be highly expressed. Major histocompatibility complex (MHC) class I is expressed, while MHC class II is not. We identified 505 single nucleotide variations (SNVs) and 20 insertions and deletions (indels). Neoantigens derived from 22 SNVs and one deletion elicited CD8+ or CD4+ T cell responses in IFNγ-ELISpot assays. Twelve high-confidence fusion genes were observed. We did not observe significant downregulation of mismatch repair (MMR) genes or SNVs/indels impairing their function, providing evidence for 6-thioguanine resistance. Effects of the integration of the murine mammary tumor virus were observed at the genome and transcriptome level. Conclusions: 4T1 cells share substantial molecular features with human TNBC. As 4T1 is a common model for metastatic tumors, our data supports the rational design of mode-of-action studies for pre-clinical evaluation of targeted immunotherapies.

20.
Oncoimmunology ; 9(1): 1758004, 2020 05 13.
Artículo en Inglés | MEDLINE | ID: mdl-32923115

RESUMEN

Glucocorticosteroids (GCS) have an established role in oncology and are administered to cancer patients in routine clinical care and in drug development trials as co-medication. Given their strong immune-suppressive activity, GCS may interfere with immune-oncology drugs. We are developing a therapeutic cancer vaccine, which is based on a liposomal formulation of tumor-antigen encoding RNA (RNA-LPX) and induces a strong T-cell response both in mice as well as in humans. In this study, we investigated in vivo in mice and in human PBMCs the effect of the commonly used long-acting GCS Dexamethasone (Dexa) on the efficacy of this vaccine format, with a particular focus on antigen-specific T-cell immune responses. We show that Dexa, when used as premedication, substantially blunts RNA-LPX vaccine-mediated immune effects. Premedication with Dexa inhibits vaccine-dependent induction of serum cytokines and chemokines and reduces both the number and activation of splenic conventional dendritic cells (cDC) expressing vaccine-encoded antigens. Consequently, priming of functional effector T cells and therapeutic activity is significantly impaired. Interestingly, responses are less impacted when Dexa is administered post-vaccination. Consistent with this observation, although many inflammatory cytokines are reduced, IFNα, a key cytokine in T-cell priming, is less impacted and antigen expression by cDCs is intact. These findings warrant special caution when combining GCS with immune therapies relying on priming and activation of antigen-specific T cells and suggest that careful sequencing of these treatments may preserve T-cell induction.


Asunto(s)
Neoplasias , Animales , Dexametasona , Femenino , Humanos , Inmunidad , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Neoplasias/tratamiento farmacológico , Premedicación
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