RESUMO
Exhausted CD8 T (Tex) cells in chronic viral infection and cancer have sustained co-expression of inhibitory receptors (IRs). Tex cells can be reinvigorated by blocking IRs, such as PD-1, but synergistic reinvigoration and enhanced disease control can be achieved by co-targeting multiple IRs including PD-1 and LAG-3. To dissect the molecular changes intrinsic when these IR pathways are disrupted, we investigated the impact of loss of PD-1 and/or LAG-3 on Tex cells during chronic infection. These analyses revealed distinct roles of PD-1 and LAG-3 in regulating Tex cell proliferation and effector functions, respectively. Moreover, these studies identified an essential role for LAG-3 in sustaining TOX and Tex cell durability as well as a LAG-3-dependent circuit that generated a CD94/NKG2+ subset of Tex cells with enhanced cytotoxicity mediated by recognition of the stress ligand Qa-1b, with similar observations in humans. These analyses disentangle the non-redundant mechanisms of PD-1 and LAG-3 and their synergy in regulating Tex cells.
Assuntos
Antígenos CD , Linfócitos T CD8-Positivos , Antígenos de Histocompatibilidade Classe I , Proteína do Gene 3 de Ativação de Linfócitos , Subfamília D de Receptores Semelhantes a Lectina de Células NK , Receptor de Morte Celular Programada 1 , Animais , Antígenos CD/metabolismo , Linfócitos T CD8-Positivos/imunologia , Linfócitos T CD8-Positivos/metabolismo , Camundongos , Receptor de Morte Celular Programada 1/metabolismo , Subfamília D de Receptores Semelhantes a Lectina de Células NK/metabolismo , Antígenos de Histocompatibilidade Classe I/metabolismo , Humanos , Subfamília C de Receptores Semelhantes a Lectina de Células NK/metabolismo , Camundongos Endogâmicos C57BL , Proteínas de Grupo de Alta Mobilidade/metabolismo , Proteínas de Grupo de Alta Mobilidade/genética , Citotoxicidade Imunológica , Proliferação de Células , Células Matadoras Naturais/metabolismo , Células Matadoras Naturais/imunologiaRESUMO
Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection of vaccinated individuals is increasingly common but rarely results in severe disease, likely due to the enhanced potency and accelerated kinetics of memory immune responses. However, there have been few opportunities to rigorously study early recall responses during human viral infection. To better understand human immune memory and identify potential mediators of lasting vaccine efficacy, we used high-dimensional flow cytometry and SARS-CoV-2 antigen probes to examine immune responses in longitudinal samples from vaccinated individuals infected during the Omicron wave. These studies revealed heightened spike-specific responses during infection of vaccinated compared to unvaccinated individuals. Spike-specific cluster of differentiation (CD)4 T cells and plasmablasts expanded and CD8 T cells were robustly activated during the first week. In contrast, memory B cell activation, neutralizing antibody production and primary responses to nonspike antigens occurred during the second week. Collectively, these data demonstrate the functionality of vaccine-primed immune memory and highlight memory T cells as rapid responders during SARS-CoV-2 infection.
RESUMO
Ten years since the immune checkpoint inhibitor ipilimumab was approved for advanced melanoma, it is time to reflect on the lessons learned regarding modulation of the immune system to treat cancer and on novel approaches to further extend the efficacy of current and emerging immunotherapies. Here, we review the studies that led to our current understanding of the melanoma immune microenvironment in humans and the mechanistic work supporting these observations. We discuss how this information is guiding more precise analyses of the mechanisms of action of immune checkpoint blockade and novel immunotherapeutic approaches. Lastly, we review emerging evidence supporting the negative impact of melanoma metabolic adaptation on anti-tumor immunity and discuss how to counteract such mechanisms for more successful use of immunotherapy.
Assuntos
Inibidores de Checkpoint Imunológico , Melanoma , Humanos , Imunoterapia , Ipilimumab/uso terapêutico , Melanoma/tratamento farmacológico , Microambiente TumoralRESUMO
Naïve CD8+ T cells can differentiate into effector (Teff), memory (Tmem) or exhausted (Tex) T cells. These developmental pathways are associated with distinct transcriptional and epigenetic changes that endow cells with different functional capacities and therefore therapeutic potential. The molecular circuitry underlying these developmental trajectories and the extent of heterogeneity within Teff, Tmem and Tex populations remain poorly understood. Here, we used the lymphocytic choriomeningitis virus model of acute-resolving and chronic infection to address these gaps by applying longitudinal single-cell RNA-sequencing (scRNA-seq) and single-cell assay for transposase-accessible chromatin sequencing (scATAC-seq) analyses. These analyses uncovered new subsets, including a subpopulation of Tex cells expressing natural killer cell-associated genes that is dependent on the transcription factor Zeb2, as well as multiple distinct TCF-1+ stem/progenitor-like subsets in acute and chronic infection. These data also revealed insights into the reshaping of Tex subsets following programmed death 1 (PD-1) pathway blockade and identified a key role for the cell stress regulator, Btg1, in establishing the Tex population. Finally, these results highlighted how the same biological circuits such as cytotoxicity or stem/progenitor pathways can be used by CD8+ T cell subsets with highly divergent underlying chromatin landscapes generated during different infections.
Assuntos
Linfócitos T CD8-Positivos , Coriomeningite Linfocítica , Humanos , Linfócitos T CD8-Positivos/metabolismo , Transcriptoma , Vírus da Coriomeningite Linfocítica , Epigênese Genética , Cromatina/genética , Cromatina/metabolismo , Coriomeningite Linfocítica/metabolismoRESUMO
Anti-programmed death-1 (anti-PD-1) immunotherapy reinvigorates CD8 T cell responses in patients with cancer but PD-1 is also expressed by other immune cells, including follicular helper CD4 T cells (Tfh) which are involved in germinal centre responses. Little is known, however, about the effects of anti-PD-1 immunotherapy on noncancer immune responses in humans. To investigate this question, we examined the impact of anti-PD-1 immunotherapy on the Tfh-B cell axis responding to unrelated viral antigens. Following influenza vaccination, a subset of adults receiving anti-PD-1 had more robust circulating Tfh responses than adults not receiving immunotherapy. PD-1 pathway blockade resulted in transcriptional signatures of increased cellular proliferation in circulating Tfh and responding B cells compared with controls. These latter observations suggest an underlying change in the Tfh-B cell and germinal centre axis in a subset of immunotherapy patients. Together, these results demonstrate dynamic effects of anti-PD-1 therapy on influenza vaccine responses and highlight analytical vaccination as an approach that may reveal underlying immune predisposition to adverse events.
Assuntos
Vacinas contra Influenza , Adulto , Humanos , Imunidade Humoral , Estações do Ano , Linfócitos T Auxiliares-Indutores , VacinaçãoRESUMO
Rewiring exhausted CD8+ T (Tex) cells toward functional states remains a therapeutic challenge. Tex cells are epigenetically programmed by the transcription factor Tox. However, epigenetic remodeling occurs as Tex cells transition from progenitor (Texprog) to intermediate (Texint) and terminal (Texterm) subsets, suggesting development flexibility. We examined epigenetic transitions between Tex cell subsets and revealed a reciprocally antagonistic circuit between Stat5a and Tox. Stat5 directed Texint cell formation and re-instigated partial effector biology during this Texprog-to-Texint cell transition. Constitutive Stat5a activity antagonized Tox and rewired CD8+ T cells from exhaustion to a durable effector and/or natural killer (NK)-like state with superior anti-tumor potential. Temporal induction of Stat5 activity in Tex cells using an orthogonal IL-2:IL2Rß-pair fostered Texint cell accumulation, particularly upon PD-L1 blockade. Re-engaging Stat5 also partially reprogrammed the epigenetic landscape of exhaustion and restored polyfunctionality. These data highlight therapeutic opportunities of manipulating the IL-2-Stat5 axis to rewire Tex cells toward more durably protective states.
Assuntos
Linfócitos T CD8-Positivos , Fatores de Transcrição , Fatores de Transcrição/genética , Interleucina-2 , Regulação da Expressão Gênica , Receptor de Morte Celular Programada 1/metabolismoRESUMO
The clinical benefit of T cell immunotherapies remains limited by incomplete understanding of T cell differentiation and dysfunction. We generated an epigenetic and transcriptional atlas of T cell differentiation from healthy humans that included exhausted CD8 T cells and applied this resource in three ways. First, we identified modules of gene expression and chromatin accessibility, revealing molecular coordination of differentiation after activation and between central memory and effector memory. Second, we applied this healthy molecular framework to three settings-a neoadjuvant anti-PD1 melanoma trial, a basal cell carcinoma scATAC-seq dataset, and autoimmune disease-associated SNPs-yielding insights into disease-specific biology. Third, we predicted genome-wide cis-regulatory elements and validated this approach for key effector genes using CRISPR interference, providing functional annotation and demonstrating the ability to identify targets for non-coding cellular engineering. These studies define epigenetic and transcriptional regulation of human T cells and illustrate the utility of interrogating disease in the context of a healthy T cell atlas.
Assuntos
Epigenômica , Ativação Linfocitária , Linfócitos T CD8-Positivos , Diferenciação Celular/genética , Cromatina/genética , Cromatina/metabolismo , Epigênese Genética , Humanos , Ativação Linfocitária/genéticaRESUMO
Therapeutic blocking of the PD1 pathway results in significant tumor responses, but resistance is common. We demonstrate that prolonged interferon signaling orchestrates PDL1-dependent and PDL1-independent resistance to immune checkpoint blockade (ICB) and to combinations such as radiation plus anti-CTLA4. Persistent type II interferon signaling allows tumors to acquire STAT1-related epigenomic changes and augments expression of interferon-stimulated genes and ligands for multiple T cell inhibitory receptors. Both type I and II interferons maintain this resistance program. Crippling the program genetically or pharmacologically interferes with multiple inhibitory pathways and expands distinct T cell populations with improved function despite expressing markers of severe exhaustion. Consequently, tumors resistant to multi-agent ICB are rendered responsive to ICB monotherapy. Finally, we observe that biomarkers for interferon-driven resistance associate with clinical progression after anti-PD1 therapy. Thus, the duration of tumor interferon signaling augments adaptive resistance and inhibition of the interferon response bypasses requirements for combinatorial ICB therapies.
Assuntos
Antígeno CTLA-4/antagonistas & inibidores , Melanoma/imunologia , Melanoma/terapia , Radioimunoterapia , Animais , Antígeno B7-H1/metabolismo , Linhagem Celular Tumoral , Resistencia a Medicamentos Antineoplásicos , Xenoenxertos , Humanos , Interferons/imunologia , Melanoma/tratamento farmacológico , Melanoma/radioterapia , Camundongos , Transplante de Neoplasias , Fator de Transcrição STAT1 , Linfócitos T/imunologiaRESUMO
CD8+ T cell exhaustion is a major barrier to current anti-cancer immunotherapies. Despite this, the developmental biology of exhausted CD8+ T cells (Tex) remains poorly defined, restraining improvement of strategies aimed at "re-invigorating" Tex cells. Here, we defined a four-cell-stage developmental framework for Tex cells. Two TCF1+ progenitor subsets were identified, one tissue restricted and quiescent and one more blood accessible, that gradually lost TCF1 as it divided and converted to a third intermediate Tex subset. This intermediate subset re-engaged some effector biology and increased upon PD-L1 blockade but ultimately converted into a fourth, terminally exhausted subset. By using transcriptional and epigenetic analyses, we identified the control mechanisms underlying subset transitions and defined a key interplay between TCF1, T-bet, and Tox in the process. These data reveal a four-stage developmental hierarchy for Tex cells and define the molecular, transcriptional, and epigenetic mechanisms that could provide opportunities to improve cancer immunotherapy.
Assuntos
Linfócitos T CD8-Positivos/imunologia , Epigênese Genética/imunologia , Neoplasias/imunologia , Subpopulações de Linfócitos T/imunologia , Transcrição Gênica/imunologia , Animais , Antígeno B7-H1/genética , Antígeno B7-H1/imunologia , Linfócitos T CD8-Positivos/citologia , Linfócitos T CD8-Positivos/metabolismo , Células Cultivadas , Epigênese Genética/genética , Fator 1-alfa Nuclear de Hepatócito/genética , Fator 1-alfa Nuclear de Hepatócito/imunologia , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/imunologia , Humanos , Imunoterapia/métodos , Camundongos Endogâmicos C57BL , Neoplasias/genética , Neoplasias/terapia , Proteínas com Domínio T/genética , Proteínas com Domínio T/imunologia , Subpopulações de Linfócitos T/metabolismo , Transcrição Gênica/genéticaRESUMO
TCF-1 is a key transcription factor in progenitor exhausted CD8 T cells (Tex). Moreover, this Tex cell subset mediates responses to PD-1 checkpoint pathway blockade. However, the role of the transcription factor TCF-1 in early fate decisions and initial generation of Tex cells is unclear. Single-cell RNA sequencing (scRNA-seq) and lineage tracing identified a TCF-1+Ly108+PD-1+ CD8 T cell population that seeds development of mature Tex cells early during chronic infection. TCF-1 mediated the bifurcation between divergent fates, repressing development of terminal KLRG1Hi effectors while fostering KLRG1Lo Tex precursor cells, and PD-1 stabilized this TCF-1+ Tex precursor cell pool. TCF-1 mediated a T-bet-to-Eomes transcription factor transition in Tex precursors by promoting Eomes expression and drove c-Myb expression that controlled Bcl-2 and survival. These data define a role for TCF-1 in early-fate-bifurcation-driving Tex precursor cells and also identify PD-1 as a protector of this early TCF-1 subset.
Assuntos
Linfócitos T CD8-Positivos/metabolismo , Redes Reguladoras de Genes , Fator 1 de Transcrição de Linfócitos T/metabolismo , Transcrição Gênica , Animais , Linfócitos T CD8-Positivos/imunologia , Diferenciação Celular/genética , Diferenciação Celular/imunologia , Doença Crônica , Perfilação da Expressão Gênica , Interações Hospedeiro-Patógeno/genética , Interações Hospedeiro-Patógeno/imunologia , Camundongos , Receptor de Morte Celular Programada 1/metabolismo , Fator 1 de Transcrição de Linfócitos T/genética , Viroses/genética , Viroses/imunologia , Viroses/virologiaRESUMO
Exhausted CD8 T (Tex) cells are immunotherapy targets in chronic infection and cancer, but a comprehensive assessment of Tex cell diversity in human disease is lacking. Here, we developed a transcriptomic- and epigenetic-guided mass cytometry approach to define core exhaustion-specific genes and disease-induced changes in Tex cells in HIV and human cancer. Single-cell proteomic profiling identified 9 distinct Tex cell clusters using phenotypic, functional, transcription factor, and inhibitory receptor co-expression patterns. An exhaustion severity metric was developed and integrated with high-dimensional phenotypes to define Tex cell clusters that were present in healthy subjects, common across chronic infection and cancer or enriched in either disease, linked to disease severity, and changed with HIV therapy. Combinatorial patterns of immunotherapy targets on different Tex cell clusters were also defined. This approach and associated datasets present a resource for investigating human Tex cell biology, with implications for immune monitoring and immunomodulation in chronic infections, autoimmunity, and cancer.
Assuntos
Linfócitos T CD8-Positivos/imunologia , Epigenômica/métodos , Citometria de Fluxo/métodos , Perfilação da Expressão Gênica/métodos , Infecções por HIV/imunologia , Neoplasias Pulmonares/imunologia , Linfócitos T CD8-Positivos/metabolismo , Infecções por HIV/genética , Infecções por HIV/metabolismo , Humanos , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/metabolismo , Proteômica/métodos , Fatores de Transcrição/genética , Fatores de Transcrição/imunologia , Fatores de Transcrição/metabolismoRESUMO
Exhausted CD8+ T (Tex) cells in chronic infections and cancer have limited effector function, high co-expression of inhibitory receptors and extensive transcriptional changes compared with effector (Teff) or memory (Tmem) CD8+ T cells. Tex cells are important clinical targets of checkpoint blockade and other immunotherapies. Epigenetically, Tex cells are a distinct immune subset, with a unique chromatin landscape compared with Teff and Tmem cells. However, the mechanisms that govern the transcriptional and epigenetic development of Tex cells remain unknown. Here we identify the HMG-box transcription factor TOX as a central regulator of Tex cells in mice. TOX is largely dispensable for the formation of Teff and Tmem cells, but it is critical for exhaustion: in the absence of TOX, Tex cells do not form. TOX is induced by calcineurin and NFAT2, and operates in a feed-forward loop in which it becomes calcineurin-independent and sustained in Tex cells. Robust expression of TOX therefore results in commitment to Tex cells by translating persistent stimulation into a distinct Tex cell transcriptional and epigenetic developmental program.
Assuntos
Linfócitos T CD8-Positivos/imunologia , Linfócitos T CD8-Positivos/patologia , Epistasia Genética , Proteínas de Homeodomínio/metabolismo , Transcrição Gênica , Animais , Calcineurina/metabolismo , Sinalização do Cálcio , Retroalimentação Fisiológica , Feminino , Regulação da Expressão Gênica/imunologia , Genótipo , Memória Imunológica , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Fatores de Transcrição NFATC/metabolismo , Evasão TumoralRESUMO
Tumour cells evade immune surveillance by upregulating the surface expression of programmed death-ligand 1 (PD-L1), which interacts with programmed death-1 (PD-1) receptor on T cells to elicit the immune checkpoint response1,2. Anti-PD-1 antibodies have shown remarkable promise in treating tumours, including metastatic melanoma2-4. However, the patient response rate is low4,5. A better understanding of PD-L1-mediated immune evasion is needed to predict patient response and improve treatment efficacy. Here we report that metastatic melanomas release extracellular vesicles, mostly in the form of exosomes, that carry PD-L1 on their surface. Stimulation with interferon-γ (IFN-γ) increases the amount of PD-L1 on these vesicles, which suppresses the function of CD8 T cells and facilitates tumour growth. In patients with metastatic melanoma, the level of circulating exosomal PD-L1 positively correlates with that of IFN-γ, and varies during the course of anti-PD-1 therapy. The magnitudes of the increase in circulating exosomal PD-L1 during early stages of treatment, as an indicator of the adaptive response of the tumour cells to T cell reinvigoration, stratifies clinical responders from non-responders. Our study unveils a mechanism by which tumour cells systemically suppress the immune system, and provides a rationale for the application of exosomal PD-L1 as a predictor for anti-PD-1 therapy.
Assuntos
Antígeno B7-H1/imunologia , Exossomos/metabolismo , Tolerância Imunológica/imunologia , Melanoma/imunologia , Receptor de Morte Celular Programada 1/imunologia , Evasão Tumoral/imunologia , Animais , Anticorpos Monoclonais Humanizados/farmacologia , Anticorpos Monoclonais Humanizados/uso terapêutico , Antineoplásicos Imunológicos/farmacologia , Antineoplásicos Imunológicos/uso terapêutico , Antígeno B7-H1/sangue , Antígeno B7-H1/metabolismo , Linfócitos T CD8-Positivos/efeitos dos fármacos , Linfócitos T CD8-Positivos/imunologia , Estudos de Casos e Controles , Linhagem Celular Tumoral , Progressão da Doença , Feminino , Humanos , Tolerância Imunológica/efeitos dos fármacos , Interferon gama/sangue , Interferon gama/imunologia , Melanoma/tratamento farmacológico , Melanoma/patologia , Camundongos , Camundongos Nus , Metástase Neoplásica , Prognóstico , Receptor de Morte Celular Programada 1/antagonistas & inibidores , Evasão Tumoral/efeitos dos fármacos , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Despite the success of monotherapies based on blockade of programmed cell death 1 (PD-1) in human melanoma, most patients do not experience durable clinical benefit. Pre-existing T-cell infiltration and/or the presence of PD-L1 in tumours may be used as indicators of clinical response; however, blood-based profiling to understand the mechanisms of PD-1 blockade has not been widely explored. Here we use immune profiling of peripheral blood from patients with stage IV melanoma before and after treatment with the PD-1-targeting antibody pembrolizumab and identify pharmacodynamic changes in circulating exhausted-phenotype CD8 T cells (Tex cells). Most of the patients demonstrated an immunological response to pembrolizumab. Clinical failure in many patients was not solely due to an inability to induce immune reinvigoration, but rather resulted from an imbalance between T-cell reinvigoration and tumour burden. The magnitude of reinvigoration of circulating Tex cells determined in relation to pretreatment tumour burden correlated with clinical response. By focused profiling of a mechanistically relevant circulating T-cell subpopulation calibrated to pretreatment disease burden, we identify a clinically accessible potential on-treatment predictor of response to PD-1 blockade.
Assuntos
Linfócitos T CD8-Positivos/imunologia , Melanoma/tratamento farmacológico , Melanoma/imunologia , Receptor de Morte Celular Programada 1/antagonistas & inibidores , Receptor de Morte Celular Programada 1/imunologia , Carga Tumoral/imunologia , Anticorpos Monoclonais Humanizados/administração & dosagem , Anticorpos Monoclonais Humanizados/imunologia , Anticorpos Monoclonais Humanizados/farmacocinética , Anticorpos Monoclonais Humanizados/uso terapêutico , Linfócitos T CD8-Positivos/citologia , Linfócitos T CD8-Positivos/metabolismo , Feminino , Humanos , Antígeno Ki-67/imunologia , Antígeno Ki-67/metabolismo , Masculino , Melanoma/irrigação sanguínea , Melanoma/patologia , Estadiamento de Neoplasias , Fenótipo , Resultado do TratamentoRESUMO
BACKGROUND: Immune checkpoint blockade (ICB) has transformed melanoma treatment, but optimal sequencing of ICB and surgery for clinically evident nodal metastasis remains undefined. We evaluated adjuvant-only (AT) and neoadjuvant/adjuvant (NAT) ICB with respect to survival outcomes in this patient population. METHODS: Patients who underwent lymphadenectomy (1 January 2011 to 31 July 2018) and received perioperative ICB at an academic center were identified. AT was defined as postoperative ICB, and NAT was defined as one to two cycles of ICB prior to resection with continuation of therapy following surgery. Three-year disease-free survival (DFS), locoregional recurrence-free survival (LRFS), distant disease-free survival (DDFS), and melanoma-specific survival (MSS) were estimated. RESULTS: Of 59 patients, 18 (31%) received AT and 41 (69%) received NAT. The AT and NAT groups did not differ in age (median 53 vs. 62 years, p = 0.16) or stage (IIIB 33% vs. 29%, IIIC 56% vs. 68%, IIID 11% vs. 2%, p = 0.34). Although 3-year DFS did not differ significantly by treatment sequencing (NAT vs. AT, hazard ratio [HR] 0.56, p = 0.17), NAT was associated with improved 3-year DDFS (HR 0.38, p = 0.028). Of 39 NAT patients with evaluable pathologic response, 23 (59%) and 5 (13%) had a pathologic partial response (pPR) and pathologic complete response (pCR), respectively. Patients with pPR/pCR experienced improved 3-year DFS (HR 0.16, p = 0.001), LRFS (HR 0.17, p = 0.003), and DDFS (HR 0.26, p = 0.029) compared with those with no response. Three-year MSS did not differ significantly by response (p = 0.062). CONCLUSION: NAT may be associated with improved 3-year DDFS compared with AT sequencing, and allows for early assessment of pathologic response. Further prospective evaluation of treatment sequencing is warranted.
Assuntos
Melanoma , Neoplasias Cutâneas , Intervalo Livre de Doença , Humanos , Inibidores de Checkpoint Imunológico , Melanoma/tratamento farmacológico , Terapia Neoadjuvante , Estadiamento de Neoplasias , Neoplasias Cutâneas/tratamento farmacológico , Neoplasias Cutâneas/patologiaRESUMO
BACKGROUND: We conducted a phase I trial evaluating pembrolizumab+hypofractionated radiotherapy (HFRT) for patients with metastatic cancers. METHODS: There were two strata (12 patients each): (i) NSCLC/melanoma progressing on prior anti-PD-1 therapy, (ii) other cancer types; anti-PD-1-naive. Patients received 6 cycles of pembrolizumab, starting 1 week before HFRT. Patients had ≥2 lesions; only one was irradiated (8 Gy × 3 for first half; 17 Gy × 1 for second half in each stratum) and the other(s) followed for response. RESULTS: Of the 24 patients, 20 (83%) had treatment-related adverse events (AEs) (all grade 1 or 2). There were eight grade 3 AEs, none treatment related. There were no dose-limiting toxicities or grade 4/5 AEs. Stratum 1: two patients (of 12) with progression on prior PD-1 blockade experienced prolonged responses (9.2 and 28.1 months). Stratum 2: one patient experienced a complete response and two had prolonged stable disease (7.4 and 7.0 months). Immune profiling demonstrated that anti-PD-1 therapy and radiation induced a consistent increase in the proliferation marker Ki67 in PD-1-expressing CD8 T cells. CONCLUSIONS: HFRT was well tolerated with pembrolizumab, and in some patients with metastatic NSCLC or melanoma, it reinvigorated a systemic response despite previous progression on anti-PD-1 therapy. CLINICAL TRIAL REGISTRATION: NCT02303990 ( www.clinicaltrials.gov ).
Assuntos
Anticorpos Monoclonais Humanizados/uso terapêutico , Antineoplásicos Imunológicos/uso terapêutico , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Carcinoma Pulmonar de Células não Pequenas/radioterapia , Quimiorradioterapia , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/radioterapia , Melanoma/tratamento farmacológico , Melanoma/radioterapia , Hipofracionamento da Dose de Radiação , Neoplasias Cutâneas/tratamento farmacológico , Neoplasias Cutâneas/radioterapia , Adulto , Idoso , Idoso de 80 Anos ou mais , Carcinoma Pulmonar de Células não Pequenas/patologia , Intervalo Livre de Doença , Feminino , Humanos , Neoplasias Pulmonares/patologia , Masculino , Melanoma/patologia , Pessoa de Meia-Idade , Metástase Neoplásica/tratamento farmacológico , Metástase Neoplásica/radioterapia , Neoplasias Cutâneas/patologiaRESUMO
PURPOSE: Immunologic response to anti-programmed cell death protein 1 (PD-1) therapy can occur rapidly with T-cell responses detectable in as little as one week. Given that activated immune cells are FDG avid, we hypothesized that an early FDG PET/CT obtained approximately 1 week after starting pembrolizumab could be used to visualize a metabolic flare (MF), with increased tumor FDG activity due to infiltration by activated immune cells, or a metabolic response (MR), due to tumor cell death, that would predict response. PATIENTS AND METHODS: Nineteen patients with advanced melanoma scheduled to receive pembrolizumab were prospectively enrolled. FDG PET/CT imaging was performed at baseline and approximately 1 week after starting treatment. FDG PET/CT scans were evaluated for changes in maximum standardized uptake value (SUVmax) and thresholds were identified by ROC analysis; MF was defined as >70% increase in tumor SUVmax, and MR as >30% decrease in tumor SUVmax. RESULTS: An MF or MR was identified in 6 of 11 (55%) responders and 0 of 8 (0%) nonresponders, with an objective response rate (ORR) of 100% in the MF-MR group and an ORR of 38% in the stable metabolism (SM) group. An MF or MR was associated with T-cell reinvigoration in the peripheral blood and immune infiltration in the tumor. Overall survival at 3 years was 83% in the MF-MR group and 62% in the SM group. Median progression-free survival (PFS) was >38 months (median not reached) in the MF-MR group and 2.8 months (95% confidence interval, 0.3-5.2) in the SM group (P = 0.017). CONCLUSIONS: Early FDG PET/CT can identify metabolic changes in melanoma metastases that are potentially predictive of response to pembrolizumab and significantly correlated with PFS.
Assuntos
Anticorpos Monoclonais Humanizados , Fluordesoxiglucose F18 , Melanoma , Tomografia por Emissão de Pósitrons combinada à Tomografia Computadorizada , Humanos , Melanoma/tratamento farmacológico , Melanoma/patologia , Melanoma/diagnóstico por imagem , Melanoma/mortalidade , Tomografia por Emissão de Pósitrons combinada à Tomografia Computadorizada/métodos , Anticorpos Monoclonais Humanizados/administração & dosagem , Anticorpos Monoclonais Humanizados/uso terapêutico , Masculino , Feminino , Fluordesoxiglucose F18/administração & dosagem , Pessoa de Meia-Idade , Idoso , Adulto , Resultado do Tratamento , Antineoplásicos Imunológicos/uso terapêutico , Antineoplásicos Imunológicos/administração & dosagem , Estudos Prospectivos , Prognóstico , Idoso de 80 Anos ou mais , Compostos RadiofarmacêuticosRESUMO
Persistent inflammation driven by cytokines such as type-one interferon (IFN-I) can cause immunosuppression. We show that administration of the Janus kinase 1 (JAK1) inhibitor itacitinib after anti-PD-1 (programmed cell death protein 1) immunotherapy improves immune function and antitumor responses in mice and results in high response rates (67%) in a phase 2 clinical trial for metastatic non-small cell lung cancer. Patients who failed to respond to initial anti-PD-1 immunotherapy but responded after addition of itacitinib had multiple features of poor immune function to anti-PD-1 alone that improved after JAK inhibition. Itacitinib promoted CD8 T cell plasticity and therapeutic responses of exhausted and effector memory-like T cell clonotypes. Patients with persistent inflammation refractory to itacitinib showed progressive CD8 T cell terminal differentiation and progressive disease. Thus, JAK inhibition may improve the efficacy of anti-PD-1 immunotherapy by pivoting T cell differentiation dynamics.
Assuntos
Linfócitos T CD8-Positivos , Carcinoma Pulmonar de Células não Pequenas , Inibidores de Checkpoint Imunológico , Janus Quinase 1 , Inibidores de Janus Quinases , Neoplasias Pulmonares , Receptor de Morte Celular Programada 1 , Animais , Feminino , Humanos , Camundongos , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Carcinoma Pulmonar de Células não Pequenas/imunologia , Carcinoma Pulmonar de Células não Pequenas/terapia , Linfócitos T CD8-Positivos/imunologia , Inibidores de Checkpoint Imunológico/uso terapêutico , Imunoterapia/métodos , Janus Quinase 1/antagonistas & inibidores , Inibidores de Janus Quinases/uso terapêutico , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/imunologia , Neoplasias Pulmonares/terapia , Receptor de Morte Celular Programada 1/antagonistas & inibidoresRESUMO
The current abundance of immunotherapy clinical trials presents an opportunity to learn about the underlying mechanisms and pharmacodynamic effects of novel drugs on the human immune system. Here, we present a protocol to study how these immune responses impact clinical outcomes using large-scale high-throughput immune profiling of clinical cohorts. We describe the Human Immune Profiling Pipeline, which comprises an end-to-end solution from flow cytometry results to computational approaches and unsupervised patient clustering based on lymphocyte landscape. For complete details on the use and execution of this protocol, please refer to Lyudovyk et al. (2022).1.
RESUMO
BACKGROUND: Only a minority of melanoma patients experience durable responses to immunotherapies due to inter- and intra-tumoral heterogeneity in melanoma. As a result, there is a pressing need for suitable preclinical models to investigate resistance mechanisms and enhance treatment efficacy. METHODS: Here, we report two different methods for generating melanoma patient-derived organoids (MPDOs), one is embedded in collagen gel, and the other is inlaid in Matrigel. MPDOs in Matrigel are used for assessing the therapeutic effects of anti-PD-1 antibodies (αPD-1), autochthonous tumor infiltrating lymphocytes (TILs), and small molecule compounds. MPDOs in collagen gel are used for evaluating the chemotaxis and migratory capacity of TILs. FINDING: The MPDOs in collagen gel and Matrigel have similar morphology and immune cell composition to their parental melanoma tissues. MPDOs show inter- and intra-tumoral heterogeneity and contain diverse immune cells such as CD4+, CD8+ T, Treg, CD14+ monocytic, CD15+, and CD11b+ myeloid cells. The tumor microenvironment (TME) in MPDOs is highly immunosuppressive, and the lymphoid and myeloid lineages express similar levels of PD-1, PD-L1, and CTLA-4 as their parental melanoma tissues. Anti-PD-1 antibodies (αPD-1) reinvigorate CD8+ T cells and induce melanoma cell death in the MPDOs. TILs expanded by IL-2 and αPD-1 show significantly lower expression of TIM-3, better migratory capacity and infiltration of autochthonous MPDOs, and more effective killing of melanoma cells than TILs expanded by IL-2 alone or IL-2 with αCD3. A small molecule screen discovers that Navitoclax increases the cytotoxicity of TIL therapy. INTERPRETATION: MPDOs may be used to test immune checkpoint inhibitors and cellular and targeted therapies. FUNDING: This work was supported by the NIH grants CA114046, CA261608, CA258113, and the Tara Miller Melanoma Foundation.