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1.
Cell ; 178(3): 585-599.e15, 2019 07 25.
Artículo en Inglés | MEDLINE | ID: mdl-31303383

RESUMEN

New opportunities are needed to increase immune checkpoint blockade (ICB) benefit. Whereas the interferon (IFN)γ pathway harbors both ICB resistance factors and therapeutic opportunities, this has not been systematically investigated for IFNγ-independent signaling routes. A genome-wide CRISPR/Cas9 screen to sensitize IFNγ receptor-deficient tumor cells to CD8 T cell elimination uncovered several hits mapping to the tumor necrosis factor (TNF) pathway. Clinically, we show that TNF antitumor activity is only limited in tumors at baseline and in ICB non-responders, correlating with its low abundance. Taking advantage of the genetic screen, we demonstrate that ablation of the top hit, TRAF2, lowers the TNF cytotoxicity threshold in tumors by redirecting TNF signaling to favor RIPK1-dependent apoptosis. TRAF2 loss greatly enhanced the therapeutic potential of pharmacologic inhibition of its interaction partner cIAP, another screen hit, thereby cooperating with ICB. Our results suggest that selective reduction of the TNF cytotoxicity threshold increases the susceptibility of tumors to immunotherapy.


Asunto(s)
Linfocitos T CD8-positivos/inmunología , Inmunoterapia , Factor de Necrosis Tumoral alfa/metabolismo , Animales , Apoptosis/efectos de los fármacos , Linfocitos T CD8-positivos/citología , Linfocitos T CD8-positivos/metabolismo , Línea Celular Tumoral , Humanos , Proteínas Inhibidoras de la Apoptosis/metabolismo , Interferón gamma/metabolismo , Estimación de Kaplan-Meier , Masculino , Ratones , Ratones Endogámicos C57BL , Neoplasias/mortalidad , Neoplasias/terapia , ARN Guía de Kinetoplastida/metabolismo , Proteína Serina-Treonina Quinasas de Interacción con Receptores/metabolismo , Receptores de Interferón/deficiencia , Receptores de Interferón/genética , Transducción de Señal/efectos de los fármacos , Factor 2 Asociado a Receptor de TNF/deficiencia , Factor 2 Asociado a Receptor de TNF/genética , Factor de Necrosis Tumoral alfa/farmacología , Receptor de Interferón gamma
3.
Cancer Cell ; 42(4): 623-645.e10, 2024 Apr 08.
Artículo en Inglés | MEDLINE | ID: mdl-38490212

RESUMEN

Genes limiting T cell antitumor activity may serve as therapeutic targets. It has not been systematically studied whether there are regulators that uniquely or broadly contribute to T cell fitness. We perform genome-scale CRISPR-Cas9 knockout screens in primary CD8 T cells to uncover genes negatively impacting fitness upon three modes of stimulation: (1) intense, triggering activation-induced cell death (AICD); (2) acute, triggering expansion; (3) chronic, causing dysfunction. Besides established regulators, we uncover genes controlling T cell fitness either specifically or commonly upon differential stimulation. Dap5 ablation, ranking highly in all three screens, increases translation while enhancing tumor killing. Loss of Icam1-mediated homotypic T cell clustering amplifies cell expansion and effector functions after both acute and intense stimulation. Lastly, Ctbp1 inactivation induces functional T cell persistence exclusively upon chronic stimulation. Our results functionally annotate fitness regulators based on their unique or shared contribution to traits limiting T cell antitumor activity.


Asunto(s)
Sistemas CRISPR-Cas , Neoplasias , Humanos , Linfocitos T CD8-positivos , Neoplasias/genética
4.
Cell Rep Med ; 3(6): 100655, 2022 06 21.
Artículo en Inglés | MEDLINE | ID: mdl-35688159

RESUMEN

Tumor escape mechanisms for immunotherapy include deficiencies in antigen presentation, diminishing adaptive CD8+ T cell antitumor activity. Although innate natural killer (NK) cells are triggered by loss of MHC class I, their response is often inadequate. To increase tumor susceptibility to both innate and adaptive immune elimination, we performed parallel genome-wide CRISPR-Cas9 knockout screens under NK and CD8+ T cell pressure. We identify all components, RNF31, RBCK1, and SHARPIN, of the linear ubiquitination chain assembly complex (LUBAC). Genetic and pharmacologic ablation of RNF31, an E3 ubiquitin ligase, strongly sensitizes cancer cells to NK and CD8+ T cell killing. This occurs in a tumor necrosis factor (TNF)-dependent manner, causing loss of A20 and non-canonical IKK complexes from TNF receptor complex I. A small-molecule RNF31 inhibitor sensitizes colon carcinoma organoids to TNF and greatly enhances bystander killing of MHC antigen-deficient tumor cells. These results merit exploration of RNF31 inhibition as a clinical pharmacological opportunity for immunotherapy-refractory cancers.


Asunto(s)
Escape del Tumor , Ubiquitina-Proteína Ligasas , Células Asesinas Naturales , Factor de Necrosis Tumoral alfa/metabolismo , Ubiquitina-Proteína Ligasas/genética , Ubiquitinación
5.
Nat Commun ; 13(1): 1923, 2022 04 08.
Artículo en Inglés | MEDLINE | ID: mdl-35395848

RESUMEN

The cytokine IFNγ differentially impacts on tumors upon immune checkpoint blockade (ICB). Despite our understanding of downstream signaling events, less is known about regulation of its receptor (IFNγ-R1). With an unbiased genome-wide CRISPR/Cas9 screen for critical regulators of IFNγ-R1 cell surface abundance, we identify STUB1 as an E3 ubiquitin ligase for IFNγ-R1 in complex with its signal-relaying kinase JAK1. STUB1 mediates ubiquitination-dependent proteasomal degradation of IFNγ-R1/JAK1 complex through IFNγ-R1K285 and JAK1K249. Conversely, STUB1 inactivation amplifies IFNγ signaling, sensitizing tumor cells to cytotoxic T cells in vitro. This is corroborated by an anticorrelation between STUB1 expression and IFNγ response in ICB-treated patients. Consistent with the context-dependent effects of IFNγ in vivo, anti-PD-1 response is increased in heterogenous tumors comprising both wildtype and STUB1-deficient cells, but not full STUB1 knockout tumors. These results uncover STUB1 as a critical regulator of IFNγ-R1, and highlight the context-dependency of STUB1-regulated IFNγ signaling for ICB outcome.


Asunto(s)
Interferón gamma , Neoplasias , Receptores de Interferón , Ubiquitina-Proteína Ligasas , Humanos , Inhibidores de Puntos de Control Inmunológico , Interferón gamma/metabolismo , Neoplasias/inmunología , Receptores de Interferón/metabolismo , Transducción de Señal , Ubiquitina-Proteína Ligasas/genética , Ubiquitina-Proteína Ligasas/metabolismo , Ubiquitinación , Receptor de Interferón gamma
6.
Cancer Immunol Res ; 9(9): 999-1007, 2021 09.
Artículo en Inglés | MEDLINE | ID: mdl-34193461

RESUMEN

Adoptive transfer of genetically modified or donor-derived T cells can efficiently eradicate human tumors but is also frequently associated with major toxicity. There are several switches that can be used to kill the infused cell pool in the case of major toxicity, but the irreversible nature of these suicide switches means that the therapeutic effect is lost when they are used. To address this issue, we engineered a small-molecule responsive genetic safety switch that in the absence of drug robustly blocked cytotoxicity and cytokine expression of primary human T cells. Upon administration of drug, T-cell functions were restored in a reversible and titratable manner. We showed that this T-cell switch was universal, as it could be combined with endogenous or transduced T-cell receptors (TCR), as well as chimeric antigen receptors. The modular nature of the Chemically Regulated - SH2-delivered Inhibitory Tail (CRASH-IT) switch concept, in which inhibitory domains are brought to activating immune receptors in a controlled manner, makes it a versatile platform to regulate the activity of cell products that signal through immunoreceptor tyrosine-based activation motif (ITAM)-containing receptors.


Asunto(s)
Receptores de Antígenos de Linfocitos T/metabolismo , Linfocitos T/metabolismo , Dominios Homologos src , Citocinas/metabolismo , Humanos , Receptores Quiméricos de Antígenos/metabolismo , Transducción de Señal
7.
Cancer Res ; 81(7): 1775-1787, 2021 04 01.
Artículo en Inglés | MEDLINE | ID: mdl-33531370

RESUMEN

Although immune checkpoint blockade (ICB) has shown remarkable clinical benefit in a subset of patients with melanoma and lung cancer, most patients experience no durable benefit. The receptor tyrosine kinase AXL is commonly implicated in therapy resistance and may serve as a marker for therapy-refractory tumors, for example in melanoma, as we previously demonstrated. Here, we show that enapotamab vedotin (EnaV), an antibody-drug conjugate targeting AXL, effectively targets tumors that display insensitivity to immunotherapy or tumor-specific T cells in several melanoma and lung cancer models. In addition to its direct tumor cell killing activity, EnaV treatment induced an inflammatory response and immunogenic cell death in tumor cells and promoted the induction of a memory-like phenotype in cytotoxic T cells. Combining EnaV with tumor-specific T cells proved superior to either treatment alone in models of melanoma and lung cancer and induced ICB benefit in models otherwise insensitive to anti-PD-1 treatment. Our findings indicate that targeting AXL-expressing, immunotherapy-resistant tumors with EnaV causes an immune-stimulating tumor microenvironment and enhances sensitivity to ICB, warranting further investigation of this treatment combination. SIGNIFICANCE: These findings show that targeting AXL-positive tumor fractions with an antibody-drug conjugate enhances antitumor immunity in several humanized tumor models of melanoma and lung cancer.


Asunto(s)
Inhibidores de Puntos de Control Inmunológico/uso terapéutico , Inmunoconjugados/uso terapéutico , Neoplasias Pulmonares/terapia , Melanoma/terapia , Proteínas Proto-Oncogénicas/inmunología , Proteínas Tirosina Quinasas Receptoras/inmunología , Animales , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Línea Celular Tumoral , Terapia Combinada , Resistencia a Antineoplásicos/inmunología , Sinergismo Farmacológico , Células HEK293 , Humanos , Inhibidores de Puntos de Control Inmunológico/administración & dosificación , Inmunoconjugados/administración & dosificación , Inmunoterapia , Neoplasias Pulmonares/patología , Masculino , Melanoma/patología , Ratones , Ratones Desnudos , Ratones Transgénicos , Terapia Molecular Dirigida/métodos , Proteínas Proto-Oncogénicas/antagonistas & inhibidores , Proteínas Tirosina Quinasas Receptoras/antagonistas & inhibidores , Ensayos Antitumor por Modelo de Xenoinjerto , Tirosina Quinasa del Receptor Axl
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