RESUMEN
Granulocyte macrophage colony stimulating factor (GM-CSF) is a key participant in, and a clinical target for, the treatment of inflammatory diseases including rheumatoid arthritis (RA). Therapeutic inhibition of GM-CSF signalling using monoclonal antibodies to the α-subunit of the GM-CSF receptor (GMCSFRα) has shown clear benefit in patients with RA, giant cell arteritis (GCAs) and some efficacy in severe SARS-CoV-2 infection. However, GM-CSF autoantibodies are associated with the development of pulmonary alveolar proteinosis (PAP), a rare lung disease characterised by alveolar macrophage (AM) dysfunction and the accumulation of surfactant lipids. We assessed how the anti-GMCSFRα approach might impact surfactant turnover in the airway. Female C57BL/6J mice received a mouse-GMCSFRα blocking antibody (CAM-3003) twice per week for up to 24 weeks. A parallel, comparator cohort of the mouse PAP model, GM-CSF receptor ß subunit (GMCSFRß) knock-out (KO), was maintained up to 16 weeks. We assessed lung tissue histopathology alongside lung phosphatidylcholine (PC) metabolism using stable isotope lipidomics. GMCSFRß KO mice reproduced the histopathological and biochemical features of PAP, accumulating surfactant PC in both broncho-alveolar lavage fluid (BALF) and lavaged lung tissue. The incorporation pattern of methyl-D9-choline showed impaired catabolism and not enhanced synthesis. In contrast, chronic supra-pharmacological CAM-3003 exposure (100 mg/kg) over 24 weeks did not elicit a histopathological PAP phenotype despite some changes in lung PC catabolism. Lack of significant impairment of AM catabolic function supports clinical observations that therapeutic antibodies to this pathway have not been associated with PAP in clinical trials.
Asunto(s)
Artritis Reumatoide/metabolismo , COVID-19/terapia , Proteinosis Alveolar Pulmonar/inmunología , Surfactantes Pulmonares/metabolismo , Receptores de Factor Estimulante de Colonias de Granulocitos y Macrófagos/antagonistas & inhibidores , Receptores de Factor Estimulante de Colonias de Granulocitos y Macrófagos/metabolismo , Animales , Anticuerpos Monoclonales Humanizados/farmacología , Artritis Reumatoide/terapia , Autoanticuerpos/química , Líquido del Lavado Bronquioalveolar , COVID-19/inmunología , Colina/análogos & derivados , Femenino , Factor Estimulante de Colonias de Granulocitos y Macrófagos/química , Inflamación , Interleucina-6/metabolismo , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Fenotipo , Proteinosis Alveolar Pulmonar/genética , SARS-CoV-2/inmunología , TensoactivosRESUMEN
Tumor programmed cell death ligand-1 (PD-L1) expression is a key biomarker to identify patients with non-small cell lung cancer who may have an enhanced response to anti-programmed cell death-1 (PD-1)/PD-L1 treatment. Such treatments are used in conjunction with PD-L1 diagnostic immunohistochemistry assays. We developed a computer-aided automated image analysis with customized PD-L1 scoring algorithm that was evaluated via correlation with manual pathologist scores and used to determine comparability across PD-L1 immunohistochemistry assays. The image analysis scoring algorithm was developed to quantify the percentage of PD-L1 positive tumor cells on scans of whole-slide images of archival tumor samples from commercially available non-small cell lung cancer cases, stained with four immunohistochemistry PD-L1 assays (Ventana SP263 and SP142 and Dako 22C3 and 28-8). The scans were co-registered and tumor and exclusion annotations aligned to ensure that analysis of each case was restricted to comparable tissue areas. Reference pathologist scores were available from previous studies. F1, a statistical measure of precision and recall, and overall percentage agreement scores were used to assess concordance between pathologist and image analysis scores and between immunohistochemistry assays. In total, 471 PD-L1-evalulable samples were amenable to image analysis scoring. Image analysis and pathologist scores were highly concordant, with F1 scores ranging from 0.8 to 0.9 across varying matched PD-L1 cutoffs. Based on F1 and overall percentage agreement scores (both manual and image analysis scoring), the Ventana SP263 and Dako 28-8 and 22C3 assays were concordant across a broad range of cutoffs; however, the Ventana SP142 assay showed very different characteristics. In summary, a novel automated image analysis scoring algorithm was developed that was highly correlated with pathologist scores. The algorithm permitted quantitative comparison of existing PD-L1 diagnostic assays, confirming previous findings that indicate a high concordance between the Ventana SP263 and Dako 22C3 and 28-8 PD-L1 immunohistochemistry assays.
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Algoritmos , Antígeno B7-H1/análisis , Biomarcadores de Tumor/análisis , Carcinoma de Pulmón de Células no Pequeñas/inmunología , Interpretación de Imagen Asistida por Computador , Inmunohistoquímica , Neoplasias Pulmonares/inmunología , Automatización , Carcinoma de Pulmón de Células no Pequeñas/patología , Humanos , Neoplasias Pulmonares/patología , Variaciones Dependientes del Observador , Patólogos , Valor Predictivo de las Pruebas , Reproducibilidad de los ResultadosAsunto(s)
Antiasmáticos/farmacología , Anticuerpos Monoclonales Humanizados/farmacología , Asma/patología , Bronquios/efectos de los fármacos , Bronquios/patología , Simulación por Computador , Músculo Liso/efectos de los fármacos , Músculo Liso/patología , Antiasmáticos/uso terapéutico , Anticuerpos Monoclonales Humanizados/uso terapéutico , Asma/tratamiento farmacológico , Predicción , Humanos , Tamaño de los ÓrganosRESUMEN
OBJECTIVE: To assess the pharmacodynamic effects of sifalimumab, an investigational anti-IFN-α monoclonal antibody, in the blood and muscle of adult dermatomyositis and polymyositis patients by measuring neutralisation of a type I IFN gene signature (IFNGS) following drug exposure. METHODS: A phase 1b randomised, double-blinded, placebo controlled, dose-escalation, multicentre clinical trial was conducted to evaluate sifalimumab in dermatomyositis or polymyositis patients. Blood and muscle biopsies were procured before and after sifalimumab administration. Selected proteins were measured in patient serum with a multiplex assay, in the muscle using immunohistochemistry, and transcripts were profiled with microarray and quantitative reverse transcriptase PCR assays. A 13-gene IFNGS was used to measure the pharmacological effect of sifalimumab. RESULTS: The IFNGS was suppressed by a median of 53-66% across three time points (days 28, 56 and 98) in blood (p=0.019) and 47% at day 98 in muscle specimens post-sifalimumab administration. Both IFN-inducible transcripts and proteins were prevalently suppressed following sifalimumab administration. Patients with 15% or greater improvement from baseline manual muscle testing scores showed greater neutralisation of the IFNGS than patients with less than 15% improvement in both blood and muscle. Pathway/functional analysis of transcripts suppressed by sifalimumab showed that leucocyte infiltration, antigen presentation and immunoglobulin categories were most suppressed by sifalimumab and highly correlated with IFNGS neutralisation in muscle. CONCLUSIONS: Sifalimumab suppressed the IFNGS in blood and muscle tissue in myositis patients, consistent with this molecule's mechanism of action with a positive correlative trend between target neutralisation and clinical improvement. These observations will require confirmation in a larger trial powered to evaluate efficacy.
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Anticuerpos Monoclonales/administración & dosificación , Dermatomiositis/tratamiento farmacológico , Dermatomiositis/inmunología , Inmunosupresores/administración & dosificación , Polimiositis/tratamiento farmacológico , Polimiositis/inmunología , Adulto , Anciano , Anticuerpos Monoclonales/efectos adversos , Anticuerpos Monoclonales Humanizados , Método Doble Ciego , Femenino , Expresión Génica/efectos de los fármacos , Expresión Génica/inmunología , Humanos , Inmunosupresores/efectos adversos , Interferón Tipo I/sangre , Interferón Tipo I/genética , Interferón Tipo I/inmunología , Interferón-alfa/sangre , Interferón-alfa/genética , Interferón-alfa/inmunología , Leucocitos/efectos de los fármacos , Leucocitos/inmunología , Masculino , Persona de Mediana Edad , Músculo Esquelético/inmunología , Placebos , Resultado del Tratamiento , Adulto JovenRESUMEN
OBJECTIVE: The aim of this study was to identify serum markers that are modulated by an investigational anti-IFN-α mAb, sifalimumab, in adult DM or PM patients. METHODS: In a phase 1b clinical trial, sera were collected from a total of 48 DM or PM adult patients receiving either placebo for 3 months or sifalimumab for 6 months. Samples were tested for 128 selected proteins using a multiplex luminex immunoassay. Muscle biopsies from selected patients were stained for T cell infiltration using an anti-CD3 antibody. RESULTS: A robust overexpression of multiple serum proteins in DM or PM patients was observed, particularly in patients with an elevated baseline type I IFN gene signature in the blood or muscle. Neutralization of the type I IFN gene signature by sifalimumab resulted in coordinated suppression of T cell-related proteins such as soluble IL-2RA, TNF receptor 2 (TNFR2) and IL-18. Muscle biopsies from two patients with the highest serum protein suppression were selected and found to have a pronounced reduction of muscle T cell infiltration. Down-regulation of IL-2RA correlated with favourable manual muscle test 8 (MMT-8) alterations in sifalimumab-dosed patients. CONCLUSION: A reduced level of multiple T cell-associated proteins after sifalimumab but not placebo administration suggests a suppressive effect of blocking type I IFN signalling on T cell activation and chemoattraction that may lead to a reduction of T cell infiltration in the muscle of myositis patients. Further, soluble IL-2RA changes from baseline may serve as a responsive and/or predictive marker for type I IFN-targeted therapy in adult DM or PM patients.
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Anticuerpos Monoclonales/farmacología , Dermatomiositis/inmunología , Interferón-alfa/antagonistas & inhibidores , Polimiositis/inmunología , Linfocitos T/inmunología , Angiopoyetina 2/inmunología , Anticuerpos Monoclonales/uso terapéutico , Anticuerpos Monoclonales Humanizados , Dermatomiositis/tratamiento farmacológico , Método Doble Ciego , Regulación hacia Abajo , Femenino , Humanos , Interferón-alfa/genética , Interleucina-18/inmunología , Subunidad alfa del Receptor de Interleucina-2/efectos de los fármacos , Subunidad alfa del Receptor de Interleucina-2/inmunología , Masculino , Persona de Mediana Edad , Músculo Esquelético/efectos de los fármacos , Músculo Esquelético/inmunología , Polimiositis/tratamiento farmacológico , Receptores Tipo II del Factor de Necrosis Tumoral/efectos de los fármacos , Receptores Tipo II del Factor de Necrosis Tumoral/inmunología , Índice de Severidad de la Enfermedad , Linfocitos T/efectos de los fármacos , Resultado del TratamientoRESUMEN
Mavrilimumab (CAM-3001) is an investigational human IgG4 monoclonal antibody (MAb) targeting GM-CSF receptor alpha which is currently being developed for the treatment of RA. GM-CSF plays a central role in the pathogenesis of rheumatoid arthritis (RA) through the activation, differentiation, and survival of macrophages and neutrophils. To support clinical development, the nonclinical safety of mavrilimumab was evaluated in several studies with cynomolgus monkeys as the pharmacologically relevant species. Comprehensive toxicity parameters were assessed in each study, and treatment duration ranged from 4 to 26weeks. Mavrilimumab has an acceptable safety profile in monkeys with no changes in any parameters other than microscopic findings in lung. In several studies, minimal accumulation of foamy alveolar macrophages was observed. This finding was only seen in studies of at least 11weeks duration, was reversible following a dose-free recovery period and was considered non-adverse. At higher dose levels (≥30mg/kg/week), in a 26-week repeat-IV dose study, the presence of lung foreign material, cholesterol clefts, and granulomatous inflammation was also observed in a few animals and was considered adverse. The dose- and time-related accumulation of foamy macrophages in lung following exposure to mavrilimumab observed in several NHP studies was expected based upon the known role of GM-CSFRα signaling in the function of alveolar macrophages. Overall, a clean no-observed-adverse-effect-level (NOAEL) without any effects in lung was established and provided adequate clinical safety margins. In clinical studies in RA patients, mavrilimumab has demonstrated good clinical activity with adequate safety to support further clinical development. A Phase 2b study of mavrilimumab in subjects with RA is in progress.
Asunto(s)
Anticuerpos Monoclonales/toxicidad , Antirreumáticos/toxicidad , Receptores de Factor Estimulante de Colonias de Granulocitos y Macrófagos/antagonistas & inhibidores , Pruebas de Toxicidad/métodos , Animales , Anticuerpos Monoclonales/administración & dosificación , Anticuerpos Monoclonales/farmacocinética , Anticuerpos Monoclonales Humanizados , Antirreumáticos/administración & dosificación , Antirreumáticos/farmacocinética , Relación Dosis-Respuesta a Droga , Femenino , Células Espumosas/efectos de los fármacos , Células Espumosas/patología , Humanos , Inyecciones Intravenosas , Inyecciones Subcutáneas , Pulmón/efectos de los fármacos , Pulmón/patología , Macaca fascicularis , Macrófagos Alveolares/efectos de los fármacos , Macrófagos Alveolares/patología , Masculino , Modelos Animales , Nivel sin Efectos Adversos Observados , Receptores de Factor Estimulante de Colonias de Granulocitos y Macrófagos/inmunología , Medición de RiesgoRESUMEN
Vascular inflammation, infusion reactions, glomerulopathies, and other potentially adverse effects may be observed in laboratory animals, including monkeys, on toxicity studies of therapeutic monoclonal antibodies and recombinant human protein drugs. Histopathologic and immunohistochemical (IHC) evaluation suggests these effects may be mediated by deposition of immune complexes (ICs) containing the drug, endogenous immunoglobulin, and/or complement components in the affected tissues. ICs may be observed in glomerulus, blood vessels, synovium, lung, liver, skin, eye, choroid plexus, or other tissues or bound to neutrophils, monocytes/macrophages, or platelets. IC deposition may activate complement, kinin, and/or coagulation/fibrinolytic pathways and result in a systemic proinflammatory response. IC clearance is biphasic in humans and monkeys (first from plasma to liver and/or spleen, second from liver or spleen). IC deposition/clearance is affected by IC composition, immunomodulation, and/or complement activation. Case studies are presented from toxicity study monkeys or rats and indicate IHC-IC deposition patterns similar to those predicted by experimental studies of IC-mediated reactions to heterologous protein administration to monkeys and other species. The IHC-staining patterns are consistent with findings associated with generalized and localized IC-associated pathology in humans. However, manifestations of immunogenicity in preclinical species are generally not considered predictive to humans.
Asunto(s)
Complejo Antígeno-Anticuerpo/metabolismo , Efectos Colaterales y Reacciones Adversas Relacionados con Medicamentos , Enfermedades Vasculares/patología , Animales , Anticuerpos Monoclonales/farmacología , Plaquetas/efectos de los fármacos , Plaquetas/metabolismo , Complemento C3/metabolismo , Complejo de Ataque a Membrana del Sistema Complemento/metabolismo , Relación Dosis-Respuesta a Droga , Evaluación Preclínica de Medicamentos , Femenino , Haplorrinos , Humanos , Inmunoglobulina G/metabolismo , Inmunoglobulina M/metabolismo , Inmunohistoquímica , Masculino , Monocitos/efectos de los fármacos , Monocitos/metabolismo , Neutrófilos/efectos de los fármacos , Neutrófilos/metabolismo , Fagocitosis/efectos de los fármacos , Ratas , Enfermedades Vasculares/inducido químicamenteRESUMEN
BACKGROUND: MEDI7247 is a first-in-class antibody-drug conjugate (ADC) consisting of an anti-sodium-dependent alanine-serine-cysteine transporter 2 antibody-conjugated to a pyrrolobenzodiazepine dimer. OBJECTIVE: This first-in-human phase 1 trial evaluated MEDI7247 in patients with hematological malignancies. PATIENTS AND METHODS: Adults with acute myeloid leukemia (AML), multiple myeloma (MM), or diffuse large B-cell lymphoma (DLBCL) relapsed or refractory (R/R) to standard therapies, or for whom no standard therapy exists, were eligible. Primary endpoints were safety and determination of the maximum tolerated dose (MTD). Secondary endpoints included assessments of antitumor activity, pharmacokinetics (PK), and immunogenicity. RESULTS: As of 26 March 2020, 67 patients were treated (AML: n = 27; MM: n = 18; DLBCL: n = 22). The most common MEDI7247-related adverse events (AEs) were thrombocytopenia (41.8%), neutropenia (35.8%), and anemia (28.4%). The most common treatment-related grade 3/4 AEs were thrombocytopenia (38.8%), neutropenia (34.3%), and anemia (22.4%). Anticancer activity (number of responders/total patients evaluated) was observed in 11/67 (16.4%) patients. No correlation was observed between ASCT2 expression and clinical response. Between-patient variability of systemic exposure of MEDI7247 ADC and total antibody were high (AUCinf geometric CV%: 62.3-134.2, and 74.8-126.1, respectively). SG3199 (PBD dimer) plasma concentrations were below the limit of quantification for all patients after Study Day 8. Anti-drug antibody (ADA) prevalence was 7.7%, ADA incidence was 1.9%, and persistent-positive ADA was 5.8%. CONCLUSIONS: Thrombocytopenia and neutropenia limited repeat dosing. Although limited clinical activity was detected, the dose-escalation phase was stopped early without establishing an MTD. The study was registered with ClinicalTrials.gov (NCT03106428).
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Neoplasias Hematológicas , Inmunoconjugados , Humanos , Masculino , Femenino , Persona de Mediana Edad , Anciano , Inmunoconjugados/uso terapéutico , Inmunoconjugados/farmacología , Inmunoconjugados/farmacocinética , Adulto , Neoplasias Hematológicas/tratamiento farmacológico , Anciano de 80 o más Años , Sistema de Transporte de Aminoácidos ASC , Antígenos de Histocompatibilidad MenorRESUMEN
Respiratory syncytial virus (RSV), a common respiratory pathogen in infants and the older population, causes pulmonary inflammation and airway occlusion that leads to impairment of lung function. Here, we have established a role for receptor for advanced glycation end products (RAGE) in RSV infection. RAGE-deficient (ager(-/-)) mice were protected from RSV-induced weight loss and inflammation. This protection correlated with an early increase in type I interferons, later decreases in proinflammatory cytokines, and a reduction in viral load. To assess the contribution of soluble RAGE (sRAGE) to RSV-induced disease, wild-type and ager(-/-) mice were given doses of sRAGE following RSV infection. Of interest, sRAGE treatment prevented RSV-induced weight loss and neutrophilic inflammation to a degree similar to that observed in ager(-/-) mice. Our work further elucidates the roles of RAGE in the pathogenesis of respiratory infections and highlights the opposing roles of membrane and sRAGE in modulating the host response to RSV infection.
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Productos Finales de Glicación Avanzada/metabolismo , Receptores Inmunológicos/metabolismo , Infecciones por Virus Sincitial Respiratorio/metabolismo , Animales , Pulmón/metabolismo , Ratones , Ratones Noqueados , Nariz , Isoformas de Proteínas , Receptor para Productos Finales de Glicación Avanzada , Receptores Inmunológicos/genética , Carga ViralRESUMEN
Mass spectrometry-based targeted proteomics allows objective protein quantitation of clinical biomarkers from a single section of formalin-fixed, paraffin-embedded (FFPE) tumor tissue biopsies. We combined high-field asymmetric waveform ion mobility spectrometry (FAIMS) and parallel reaction monitoring (PRM) to increase assay sensitivity. The modular nature of the FAIMS source allowed direct comparison of the performance of FAIMS-PRM to PRM. Limits of quantitation were determined by spiking synthetic peptides into a human spleen matrix. In addition, 20 clinical samples were analyzed using FAIMS-PRM and the quantitation of HER2 was compared with that obtained with the Ventana immunohistochemistry assay. FAIMS-PRM improved the overall signal-to-noise ratio over that from PRM and increased assay sensitivity in FFPE tissue analysis for four (HER2, EGFR, cMET, and KRAS) of five proteins of clinical interest. FAIMS-PRM enabled sensitive quantitation of basal HER2 expression in breast cancer samples classified as HER2 negative by immunohistochemistry. Furthermore, we determined the degree of FAIMS-dependent background reduction and showed that this correlated with an improved lower limit of quantitation with FAIMS. FAIMS-PRM is anticipated to benefit clinical trials in which multiple biomarker questions must be addressed and the availability of tumor biopsy samples is limited.
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Neoplasias de la Mama , Proteómica , Biopsia , Neoplasias de la Mama/metabolismo , Femenino , Humanos , Espectrometría de Movilidad Iónica/métodos , Proteínas/química , Proteómica/métodosRESUMEN
BACKGROUND: Immunotherapies have shown clinical activity in patients with advanced biliary tract cancer, for which outcomes remain poor despite standard of care treatment with gemcitabine and cisplatin. We aimed to evaluate gemcitabine and cisplatin plus durvalumab with or without tremelimumab as first-line treatment in patients with advanced biliary tract cancer. METHODS: This open-label, single-centre, phase 2 study was conducted at Seoul National University Hospital. Eligible patients were treatment-naïve adults aged 18 years or older with histologically proven unresectable or recurrent biliary tract cancer, at least one measurable lesion based on the Response Evaluation Criteria in Solid Tumors (version 1.1), an Eastern Cooperative Oncology Group performance status of 0 or 1, life expectancy of 12 weeks or longer, and adequate healthy organ and bone marrow function. Initially, all patients received one 3-week cycle of gemcitabine (1000 mg/m2) and cisplatin (25 mg/m2) on day 1 and 8 followed by gemcitabine and cisplatin plus durvalumab (1120 mg) and tremelimumab (75 mg) on day 1 of each cycle, starting with the second cycle (chemotherapy followed by chemotherapy plus durvalumab and tremelimumab group). Following protocol amendment, patients were recruited to receive gemcitabine and cisplatin plus durvalumab, starting on day 1 of the first cycle (chemotherapy plus durvalumab group) or gemcitabine and cisplatin plus durvalumab and tremelimumab also from day 1 of the first cycle (chemotherapy plus durvalumab and tremelimumab group) in parallel and allocated using a random block method. Assessors and patients were not masked to the treatment group. The primary endpoint was objective response rate, assessed in the efficacy population (ie, patients who were treated at least until the first tumour response assessment). This study is registered with ClinicalTrials.gov, NCT03046862 (active). FINDINGS: Between March 2, 2017, and Feb 13, 2020, 128 patients were enrolled (32 in the chemotherapy followed by chemotherapy plus durvalumab and tremelimumab group, 49 in the chemotherapy plus durvalumab group, and 47 in the chemotherapy plus durvalumab and tremelimumab group). Four patients (two in the chemotherapy followed by chemotherapy plus durvalumab and tremelimumab group and two in the chemotherapy plus durvalumab group) were excluded and 124 were evaluable for tumour response. The median duration of follow-up was 48·2 months (IQR 41·5-49·4) for the chemotherapy followed by chemotherapy plus durvalumab and tremelimumab group, 26·6 months (19·0-27·9) for the chemotherapy plus durvalumab group, and 24·2 months (20·7-31·7) for the chemotherapy plus durvalumab and tremelimumab group. 82 (66%) of 124 patients achieved an objective response (15 [50%] of 30 in the chemotherapy followed by chemotherapy plus durvalumab and tremelimumab group, 34 [72%] of 47 in the chemotherapy plus durvalumab group, and 33 [70%] of 47 in the chemotherapy plus durvalumab and tremelimumab group). The most common grade 3 and 4 adverse events were decreased neutrophil count (67 [53%] of 126), anaemia (50 [40%]), and decreased platelet count (24 [19%]), with no unexpected safety events. No adverse events leading to discontinuation or death occurred. INTERPRETATION: Gemcitabine and cisplatin plus immunotherapy showed promising efficacy and acceptable safety in patients with biliary tract cancer. Gemcitabine and cisplatin plus durvalumab are being evaluated in the phase 3, TOPAZ-1 study (NCT03875235) as first-line treatment in patients with advanced biliary tract cancer. FUNDING: AstraZeneca; National Research Foundation of Korea (Grant No. 2021R1A2C2007430).
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Neoplasias del Sistema Biliar , Cisplatino , Adulto , Anticuerpos Monoclonales , Anticuerpos Monoclonales Humanizados , Protocolos de Quimioterapia Combinada Antineoplásica/efectos adversos , Neoplasias del Sistema Biliar/patología , Cisplatino/efectos adversos , Desoxicitidina/análogos & derivados , Humanos , Recurrencia Local de Neoplasia/tratamiento farmacológico , GemcitabinaRESUMEN
We report the ability of two deep learning-based decision systems to stratify non-small cell lung cancer (NSCLC) patients treated with checkpoint inhibitor therapy into two distinct survival groups. Both systems analyze functional and morphological properties of epithelial regions in digital histopathology whole slide images stained with the SP263 PD-L1 antibody. The first system learns to replicate the pathologist assessment of the Tumor Cell (TC) score with a cut-point for positivity at 25% for patient stratification. The second system is free from assumptions related to TC scoring and directly learns patient stratification from the overall survival time and event information. Both systems are built on a novel unpaired domain adaptation deep learning solution for epithelial region segmentation. This approach significantly reduces the need for large pixel-precise manually annotated datasets while superseding serial sectioning or re-staining of slides to obtain ground truth by cytokeratin staining. The capacity of the first system to replicate the TC scoring by pathologists is evaluated on 703 unseen cases, with an addition of 97 cases from an independent cohort. Our results show Lin's concordance values of 0.93 and 0.96 against pathologist scoring, respectively. The ability of the first and second system to stratify anti-PD-L1 treated patients is evaluated on 151 clinical samples. Both systems show similar stratification powers (first system: HR = 0.539, p = 0.004 and second system: HR = 0.525, p = 0.003) compared to TC scoring by pathologists (HR = 0.574, p = 0.01).
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Carcinoma de Pulmón de Células no Pequeñas , Aprendizaje Profundo , Neoplasias Pulmonares , Antígeno B7-H1 , Biomarcadores de Tumor , Humanos , Inmunohistoquímica , Neoplasias Pulmonares/diagnóstico por imagen , Análisis de SupervivenciaRESUMEN
BACKGROUND: Studies have indicated that programmed death ligand 1 (PD-L1) expression may have utility as a predictive biomarker in patients with advanced/metastatic urothelial carcinoma (UC). Different immunohistochemical (IHC) assays are in development to assess PD-L1 expression on tumor cells (TCs) and tumor-infiltrating immune cells (ICs). METHODS: In this post hoc analysis of the single-arm, phase 1/2 Study 1108 (NCT01693562), PD-L1 expression was evaluated from tumor samples obtained prior to second-line treatment with durvalumab in patients with advanced/metastatic UC using the VENTANA (SP263) IHC Assay. The primary objective was to determine whether the TC ≥ 25%/IC ≥ 25% algorithm (i.e., cutoff of ≥ 25% TC or ≥ 25% IC with PD-L1 staining at any intensity above background) was optimal for predicting response to durvalumab. PD-L1 expression data were available from 188 patients. RESULTS: After a median follow-up of 15.8 and 14.6 months, higher PD-L1 expression was associated with longer overall survival (OS) and progression-free survival (PFS), respectively, with significant separation in survival curves for PD-L1-high and-low expressing patients for the TC ≥ 25%/IC ≥ 25% cutoff (median OS: 19.8 vs 4.8 months; hazard ratio: 0.46; 90% confidence interval: 0.33, 0.639). OS was also prolonged for PD-L1-high compared with-low patients when samples were categorized using TC/IC combined positive score ≥ 10 and IC≥ 5% cutoffs. In multivariate analysis, IC but not TC PD-L1 expression was significantly associated with OS, PFS, and objective response rate (P < 0.001 for each), although interaction analysis showed similar directionality of benefit for ICs and TCs. CONCLUSIONS: These findings support the utility of a combined TC/IC algorithm for predicting response to durvalumab in patients with UC, with the TC≥ 25%/IC≥ 25% cutoff optimal when used with the VENTANA (SP263) IHC Assay.
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Anticuerpos Monoclonales/uso terapéutico , Antígeno B7-H1/análisis , Neoplasias Urológicas/tratamiento farmacológico , Adulto , Algoritmos , Femenino , Humanos , Estimación de Kaplan-Meier , Masculino , Persona de Mediana Edad , Supervivencia sin Progresión , Modelos de Riesgos Proporcionales , Resultado del Tratamiento , Neoplasias Urológicas/mortalidad , Neoplasias Urológicas/patologíaRESUMEN
OBJECTIVES: The interaction between the immune system and tumor cells is an important feature for the prognosis and treatment of cancer. Multiplex immunohistochemistry (mIHC) and multiplex immunofluorescence (mIF) analyses are emerging technologies that can be used to help quantify immune cell subsets, their functional state, and their spatial arrangement within the tumor microenvironment. METHODS: The Society for Immunotherapy of Cancer (SITC) convened a task force of pathologists and laboratory leaders from academic centers as well as experts from pharmaceutical and diagnostic companies to develop best practice guidelines for the optimization and validation of mIHC/mIF assays across platforms. RESULTS: Representative outputs and the advantages and disadvantages of mIHC/mIF approaches, such as multiplexed chromogenic IHC, multiplexed immunohistochemical consecutive staining on single slide, mIF (including multispectral approaches), tissue-based mass spectrometry, and digital spatial profiling are discussed. CONCLUSIONS: mIHC/mIF technologies are becoming standard tools for biomarker studies and are likely to enter routine clinical practice in the near future. Careful assay optimization and validation will help ensure outputs are robust and comparable across laboratories as well as potentially across mIHC/mIF platforms. Quantitative image analysis of mIHC/mIF output and data management considerations will be addressed in a complementary manuscript from this task force.
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Técnica del Anticuerpo Fluorescente/métodos , Inmunohistoquímica/métodos , Inmunoterapia/métodos , Coloración y Etiquetado/métodos , Microambiente Tumoral/fisiología , HumanosRESUMEN
PURPOSE: This randomized, multicenter, open-label, phase Ib/II study assessed durvalumab and tremelimumab in combination or as monotherapy for chemotherapy-refractory gastric cancer or gastroesophageal junction (GEJ) cancer. PATIENTS AND METHODS: Second-line patients were randomized 2:2:1 to receive durvalumab plus tremelimumab (arm A), or durvalumab (arm B) or tremelimumab monotherapy (arm C), and third-line patients received durvalumab plus tremelimumab (arm D). A tumor-based IFNγ gene signature was prospectively evaluated as a potential predictive biomarker in second- and third-line patients receiving the combination (arm E). The coprimary endpoints were objective response rate and progression-free survival (PFS) rate at 6 months. RESULTS: A total of 113 patients were treated: 6 in phase Ib and 107 (arm A, 27; arm B, 24; arm C, 12; arm D, 25; arm E, 19) in phase II. Overall response rates were 7.4%, 0%, 8.3%, 4.0%, and 15.8% in the five arms, respectively. PFS rates at 6 months were 6.1%, 0%, 20%, 15%, and 0%, and 12-month overall survival rates were 37.0%, 4.6%, 22.9%, 38.8%, and NA, respectively. Treatment-related grade 3/4 adverse events were reported in 17%, 4%, 42%, 16%, and 11% of patients, respectively. CONCLUSIONS: Response rates were low regardless of monotherapy or combination strategies. No new safety signals were identified. Including use of a tumor-based IFNγ signature and change in baseline and on-treatment circulating tumor DNA are clinically feasible and may be novel strategies to improve treatment response in this difficult-to-treat population.
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Adenocarcinoma/tratamiento farmacológico , Anticuerpos Monoclonales Humanizados/uso terapéutico , Anticuerpos Monoclonales/uso terapéutico , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Unión Esofagogástrica/efectos de los fármacos , Neoplasias Gástricas/tratamiento farmacológico , Adenocarcinoma/genética , Adenocarcinoma/patología , Adulto , Anciano , Anciano de 80 o más Años , Anticuerpos Monoclonales/administración & dosificación , Anticuerpos Monoclonales/efectos adversos , Anticuerpos Monoclonales Humanizados/administración & dosificación , Anticuerpos Monoclonales Humanizados/efectos adversos , Antígeno B7-H1/antagonistas & inhibidores , Antígeno CTLA-4/antagonistas & inhibidores , ADN Tumoral Circulante/genética , Unión Esofagogástrica/patología , Femenino , Humanos , Interferón gamma/metabolismo , Masculino , Persona de Mediana Edad , Estudios Prospectivos , Neoplasias Gástricas/genética , Neoplasias Gástricas/patología , Transcriptoma , Adulto JovenRESUMEN
BACKGROUND: Antibodies targeting the programmed cell death-1 (PD-1)/PD-ligand 1 (PD-1/PD-L1) checkpoint have shown promising clinical activity in patients with advanced urothelial carcinoma (UC). Expression of PD-L1 in UC tumors has been investigated using different antibody clones, staining protocols, and scoring algorithms. The aim was to establish the extent of concordance among PD-L1 immunohistochemistry (IHC) assays. METHODS: Tumor biopsy samples (N = 335) were assessed using four commercially available PD-L1 assays: VENTANA SP263, VENTANA SP142, PD-L1 IHC 28-8 pharmDx, and PD-L1 IHC 22C3 pharmDx. PD-L1 analytical staining and classification concordance, including agreement between clinically relevant scoring algorithms, were investigated using overall/positive/negative percentage agreement (OPA/PPA/NPA). RESULTS: Good analytical correlation was observed among the VENTANA SP263, PD-L1 IHC 22C3 pharmDx, and PD-L1 IHC 28-8 pharmDx assays for tumor cell (TC) and immune cell (IC) PD-L1 staining with Spearman rank coefficients of 0.92-0.93 for TCs and 0.88-0.91 for ICs. However, concordance (preset criterion: ≥85%) between patient PD-L1 status when applying the TC or ICICArea ≥ 25% (VENTANA SP263) cutoff was only achieved for PD-L1 IHC 22C3 pharmDx versus VENTANA SP263 (OPA 92.2%, PPA 86.4%, NPA 95.4%). Differences were observed between patient populations with UC tumors classified as PD-L1 high versus PD-L1 low/negative using combined positive score (CPS) ≥1, CPS ≥10, IC ≥5%, and TC/IC ≥25%. CONCLUSIONS: The VENTANA SP263 and PD-L1 IHC 22C3 pharmDx assays are analytically similar in UC. When the different PD-L1 assays were combined with their specified clinical scoring algorithms, differences were seen in patient classification driven by substantial differences in scoring approaches.
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Antígeno B7-H1/análisis , Biomarcadores de Tumor/análisis , Carcinoma de Células Transicionales/clasificación , Inmunohistoquímica/normas , Neoplasias de la Vejiga Urinaria/clasificación , Anciano , Algoritmos , Femenino , Humanos , Masculino , Persona de Mediana Edad , Reproducibilidad de los ResultadosRESUMEN
Continued developments in immuno-oncology require an increased understanding of the mechanisms of cancer immunology. The immunoprofiling analysis of tissue samples from formalin-fixed, paraffin-embedded (FFPE) biopsies has become a key tool for understanding the complexity of tumor immunology and discovering novel predictive biomarkers for cancer immunotherapy. Immunoprofiling analysis of tissues requires the evaluation of combined markers, including inflammatory cell subpopulations and immune checkpoints, in the tumor microenvironment. The advent of novel multiplex immunohistochemical methods allows for a more efficient multiparametric analysis of single tissue sections than does standard monoplex immunohistochemistry (IHC). One commercially available multiplex immunofluorescence (IF) method is based on tyramide-signal amplification and, combined with multispectral microscopic analysis, allows for a better signal separation of diverse markers in tissue. This methodology is compatible with the use of unconjugated primary antibodies that have been optimized for standard IHC on FFPE tissue samples. Herein we describe in detail an automated protocol that allows multiplex IF labeling of carcinoma tissue samples with a six-marker multiplex antibody panel comprising PD-L1, PD-1, CD68, CD8, Ki-67, and AE1/AE3 cytokeratins with 4',6-diamidino-2-phenylindole as a nuclear cell counterstain. The multiplex panel protocol is optimized in an automated IHC stainer for a staining time that is shorter than that of the manual protocol and can be directly applied and adapted by any laboratory investigator for immuno-oncology studies on human FFPE tissue samples. Also described are several controls and tools, including a drop-control method for fine quality control of a new multiplex IF panel, that are useful for the optimization and validation of the technique.
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Carcinoma/patología , Técnica del Anticuerpo Fluorescente/métodos , Formaldehído/uso terapéutico , Inmunohistoquímica/métodos , Humanos , Microambiente TumoralRESUMEN
CONTEXT.: Clinical responses to anti-programmed death receptor-1 and anti-programmed death ligand-1 (PD-L1) agents are generally improved in patients with high PD-L1 expression compared with those with low/negative expression across several tumor types, including urothelial carcinoma. OBJECTIVE.: To validate a PD-L1 immunohistochemical diagnostic test in urothelial carcinoma patients treated with the anti-PD-L1 monoclonal antibody durvalumab. DESIGN.: The Ventana PD-L1 (SP263) assay was validated for intended use in urothelial carcinoma formalin-fixed, paraffin-embedded samples in studies addressing sensitivity, specificity, robustness, and precision, and implemented in study CD-ON-MEDI4736-1108 (NCT01693562). Efficacy was analyzed in patients classified according to prespecified PD-L1 expression cutoffs: PD-L1 high (if >1% of the tumor area contained tumor-associated immune cells, ≥25% of tumor cells or ≥25% of immune cells stained for PD-L1; if ≤1% of the tumor area contained immune cells, ≥25% of tumor cells or 100% of immune cells stained for PD-L1) and PD-L1 low/negative (did not meet criteria for PD-L1 high). RESULTS.: The assay met all predefined acceptance criteria for sensitivity, specificity, and precision. Interreader and intrareader precision overall agreement were 93.0% and 92.4%, respectively. For intraday reproducibility and interday precision, overall agreement was 99.2% and 100%, respectively. Interlaboratory overall agreement was 92.6%. In study CD-ON-MEDI4736-1108, durvalumab demonstrated clinical activity and durable responses in both PD-L1-high and PD-L1-low/negative subgroups, although objective response rates tended to be higher in the PD-L1-high subgroup than in the PD-L1-low/negative subgroup. CONCLUSIONS.: Determination of PD-L1 expression in urothelial carcinoma patients using the Ventana PD-L1 (SP263) assay was precise, highly reproducible, and clinically relevant.
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Anticuerpos Monoclonales/uso terapéutico , Antineoplásicos Inmunológicos/uso terapéutico , Antígeno B7-H1/análisis , Carcinoma de Células Transicionales/tratamiento farmacológico , Inmunohistoquímica/métodos , Neoplasias de la Vejiga Urinaria/tratamiento farmacológico , Adulto , Algoritmos , Carcinoma de Células Transicionales/metabolismo , Femenino , Humanos , Masculino , Persona de Mediana Edad , Reproducibilidad de los Resultados , Neoplasias de la Vejiga Urinaria/metabolismoRESUMEN
BACKGROUND: Immune checkpoint therapies (ICTs) targeting the programmed cell death-1 (PD1)/programmed cell death ligand-1 (PD-L1) pathway have improved outcomes for patients with non-small cell lung cancer (NSCLC), particularly those with high PD-L1 expression. However, the predictive value of manual PD-L1 scoring is imperfect and alternative measures are needed. We report an automated image analysis solution to determine the predictive and prognostic values of the product of PD-L1+ cell and CD8+ tumor infiltrating lymphocyte (TIL) densities (CD8xPD-L1 signature) in baseline tumor biopsies. METHODS: Archival or fresh tumor biopsies were analyzed for PD-L1 and CD8 expression by immunohistochemistry. Samples were collected from 163 patients in Study 1108/NCT01693562, a Phase 1/2 trial to evaluate durvalumab across multiple tumor types, including NSCLC, and a separate cohort of 199 non-ICT- patients. Digital images were automatically scored for PD-L1+ and CD8+ cell densities using customized algorithms applied with Developer XD™ 2.7 software. RESULTS: For patients who received durvalumab, median overall survival (OS) was 21.0 months for CD8xPD-L1 signature-positive patients and 7.8 months for signature-negative patients (p = 0.00002). The CD8xPD-L1 signature provided greater stratification of OS than high densities of CD8+ cells, high densities of PD-L1+ cells, or manually assessed tumor cell PD-L1 expression ≥25%. The CD8xPD-L1 signature did not stratify OS in non-ICT patients, although a high density of CD8+ cells was associated with higher median OS (high: 67 months; low: 39.5 months, p = 0.0009) in this group. CONCLUSIONS: An automated CD8xPD-L1 signature may help to identify NSCLC patients with improved response to durvalumab therapy. Our data also support the prognostic value of CD8+ TILS in NSCLC patients who do not receive ICT. TRIAL REGISTRATION: ClinicalTrials.gov identifier: NCT01693562 . Study code: CD-ON-MEDI4736-1108. Interventional study (ongoing but not currently recruiting). Actual study start date: August 29, 2012. Primary completion date: June 23, 2017 (final data collection date for primary outcome measure).
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Anticuerpos Monoclonales/uso terapéutico , Biomarcadores de Tumor/análisis , Carcinoma de Pulmón de Células no Pequeñas/tratamiento farmacológico , Procesamiento de Imagen Asistido por Computador , Neoplasias Pulmonares/tratamiento farmacológico , Pulmón/patología , Adulto , Anciano , Anciano de 80 o más Años , Anticuerpos Monoclonales/farmacología , Antígeno B7-H1/análisis , Antígeno B7-H1/antagonistas & inhibidores , Antígeno B7-H1/inmunología , Antígeno B7-H1/metabolismo , Biomarcadores de Tumor/inmunología , Biomarcadores de Tumor/metabolismo , Biopsia , Antígenos CD8/análisis , Antígenos CD8/inmunología , Antígenos CD8/metabolismo , Linfocitos T CD8-positivos , Carcinoma de Pulmón de Células no Pequeñas/inmunología , Carcinoma de Pulmón de Células no Pequeñas/mortalidad , Carcinoma de Pulmón de Células no Pequeñas/patología , Femenino , Humanos , Inmunohistoquímica , Pulmón/efectos de los fármacos , Pulmón/inmunología , Neoplasias Pulmonares/inmunología , Neoplasias Pulmonares/mortalidad , Neoplasias Pulmonares/patología , Linfocitos Infiltrantes de Tumor , Masculino , Persona de Mediana Edad , Valor Predictivo de las Pruebas , Pronóstico , Análisis de Supervivencia , Resultado del Tratamiento , Adulto JovenRESUMEN
INTRODUCTION: Durvalumab selectively blocks programmed cell death ligand-1 (PD-L1) binding to programmed cell death-1. Encouraging clinical activity and manageable safety were reported in urothelial carcinoma, non-small-cell lung cancer (NSCLC), hepatocellular carcinoma (HC) and small-cell lung cancer (SCLC) in a multicenter phase I/II study. Safety and clinical activity in recurrent/metastatic head and neck squamous cell carcinoma (HNSCC) were evaluated in the expansion phase. METHODS: Patients received 10 mg/kg of durvalumab intravenously every 2 weeks for 12 months or until confirmed progressive disease or unacceptable toxicity. The primary objective was safety; clinical activity was a secondary objective. RESULTS: Sixty-two patients were enrolled and evaluable (received first dose ≥24 weeks before data cutoff). Median age was 57 years; 40.3% were human papillomavirus (HPV)-positive; 32.3% had tumour cell PD-L1 expression ≥25%, and 62.9% were current/former smokers. They had a median of 2 prior systemic treatments (range, 1-13). All-causality adverse events (AEs) occurred in 98.4%; drug-related AEs occurred in 59.7% and were grade III-IV in 9.7%. There were no drug-related discontinuations or deaths. Objective response rate (blinded independent central review) was 6.5% (15.0% for PD-L1 ≥25%, 2.6% for <25%). Median time to response was 2.7 months (range, 1.2-5.5); median duration was 12.4 months (range, 3.5-20.5+). Median progression-free survival was 1.4 months; median overall survival (OS) was 8.4 months. OS rate was 62% at 6 months and 38% at 12 months (42% for PD-L1 ≥25%, 36% for <25%). CONCLUSIONS: Durvalumab safety in HNSCC was manageable and consistent with other cohorts of the study. Early, durable responses in these heavily pretreated patients warrant further investigation; phase III monotherapy and combination therapy studies are ongoing. CLINICAL TRIAL REGISTRY: clinicaltrials.gov NCT01693562; MedImmune study 1108.