RESUMEN
B cell receptor (BCR) signalling has emerged as a therapeutic target in B cell lymphomas, but inhibiting this pathway in diffuse large B cell lymphoma (DLBCL) has benefited only a subset of patients1. Gene expression profiling identified two major subtypes of DLBCL, known as germinal centre B cell-like and activated B cell-like (ABC)2,3, that show poor outcomes after immunochemotherapy in ABC. Autoantigens drive BCR-dependent activation of NF-κB in ABC DLBCL through a kinase signalling cascade of SYK, BTK and PKCß to promote the assembly of the CARD11-BCL10-MALT1 adaptor complex, which recruits and activates IκB kinase4-6. Genome sequencing revealed gain-of-function mutations that target the CD79A and CD79B BCR subunits and the Toll-like receptor signalling adaptor MYD885,7, with MYD88(L265P) being the most prevalent isoform. In a clinical trial, the BTK inhibitor ibrutinib produced responses in 37% of cases of ABC1. The most striking response rate (80%) was observed in tumours with both CD79B and MYD88(L265P) mutations, but how these mutations cooperate to promote dependence on BCR signalling remains unclear. Here we used genome-wide CRISPR-Cas9 screening and functional proteomics to determine the molecular basis of exceptional clinical responses to ibrutinib. We discovered a new mode of oncogenic BCR signalling in ibrutinib-responsive cell lines and biopsies, coordinated by a multiprotein supercomplex formed by MYD88, TLR9 and the BCR (hereafter termed the My-T-BCR supercomplex). The My-T-BCR supercomplex co-localizes with mTOR on endolysosomes, where it drives pro-survival NF-κB and mTOR signalling. Inhibitors of BCR and mTOR signalling cooperatively decreased the formation and function of the My-T-BCR supercomplex, providing mechanistic insight into their synergistic toxicity for My-T-BCR+ DLBCL cells. My-T-BCR supercomplexes characterized ibrutinib-responsive malignancies and distinguished ibrutinib responders from non-responders. Our data provide a framework for the rational design of oncogenic signalling inhibitors in molecularly defined subsets of DLBCL.
Asunto(s)
Carcinogénesis , Linfoma de Células B Grandes Difuso/metabolismo , Linfoma de Células B Grandes Difuso/patología , Complejos Multiproteicos/metabolismo , Transducción de Señal , Adenina/análogos & derivados , Animales , Biopsia , Sistemas CRISPR-Cas/genética , Carcinogénesis/genética , Diseño de Fármacos , Femenino , Humanos , Linfoma de Células B Grandes Difuso/genética , Ratones , Complejos Multiproteicos/química , Mutación , Factor 88 de Diferenciación Mieloide/genética , Factor 88 de Diferenciación Mieloide/metabolismo , FN-kappa B/metabolismo , Piperidinas , Proteómica , Pirazoles/farmacología , Pirazoles/uso terapéutico , Pirimidinas/farmacología , Pirimidinas/uso terapéutico , Receptores de Antígenos de Linfocitos B/antagonistas & inhibidores , Receptores de Antígenos de Linfocitos B/genética , Receptores de Antígenos de Linfocitos B/metabolismo , Transducción de Señal/efectos de los fármacos , Transducción de Señal/genética , Serina-Treonina Quinasas TOR/antagonistas & inhibidores , Serina-Treonina Quinasas TOR/metabolismo , Receptor Toll-Like 9/genética , Receptor Toll-Like 9/metabolismo , Células Tumorales Cultivadas , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Several closely related, thermophilic and cellulolytic bacterial strains, designated JKG1(T), JKG2, JKG3, JKG4 and JKG5, were isolated from a cellulolytic enrichment (corn stover) incubated in the water column of Great Boiling Spring, NV. Strain JKG1(T) had cells of diameter 0.7-0.9 µm and length ~2.0 µm that formed non-branched, multicellular filaments reaching >300 µm. Spores were not formed and dense liquid cultures were red. The temperature range for growth was 45-65 °C, with an optimum of 55 °C. The pH range for growth was pH 5.6-9.0, with an optimum of pH 7.5. JKG1(T) grew as an aerobic heterotroph, utilizing glucose, sucrose, xylose, arabinose, cellobiose, CM-cellulose, filter paper, microcrystalline cellulose, xylan, starch, Casamino acids, tryptone, peptone, yeast extract, acetate, citrate, lactate, pyruvate and glycerol as sole carbon sources, and was not observed to photosynthesize. The cells stained Gram-negative. Phylogenetic analysis using 16S rRNA gene sequences placed the new isolates in the class Chloroflexia, but distant from other cultivated members, with the highest sequence identity of 82.5â% to Roseiflexus castenholzii. The major quinone was menaquinone-9; no ubiquinones were detected. The major cellular fatty acids (>5â%) were C18â:â0, anteiso-C17â:â0, iso-C18â:â0, iso-C17â:â0, C16â:â0, iso-C16â:â0 and C17â:â0. The peptidoglycan amino acids were alanine, ornithine, glutamic acid, serine and asparagine. Whole-cell sugars included mannose, rhamnose, glucose, galactose, ribose, arabinose and xylose. Morphological, phylogenetic and chemotaxonomic results suggest that JKG1(T) is representative of a new lineage within the class Chloroflexia, which we propose to designate Kallotenue papyrolyticum gen. nov., sp. nov., Kallotenuaceae fam. nov., Kallotenuales ord. nov. The type strain of Kallotenue papyrolyticum gen. nov., sp. nov. is JKG1(T) (â=âDSM 26889(T)â=âJCM 19132(T)).