Durvalumab with platinum-pemetrexed for unresectable pleural mesothelioma: survival, genomic and immunologic analyses from the phase 2 PrE0505 trial

Abstract

Mesothelioma is a rare and fatal cancer with limited therapeutic options until the recent approval of combination immune checkpoint blockade. Here we report the results of the phase 2 PrE0505 trial ( NCT02899195 ) of the anti-PD-L1 antibody durvalumab plus platinum-pemetrexed chemotherapy for 55 patients with previously untreated, unresectable pleural mesothelioma. The primary endpoint was overall survival compared to historical control with cisplatin and pemetrexed chemotherapy; secondary and exploratory endpoints included safety, progression-free survival and biomarkers of response. The combination of durvalumab with chemotherapy met the pre-specified primary endpoint, reaching a median survival of 20.4 months versus 12.1 months with historical control. Treatment-emergent adverse events were consistent with known side effects of chemotherapy, and all adverse events due to immunotherapy were grade 2 or lower. Integrated genomic and immune cell repertoire analyses revealed that a higher immunogenic mutation burden coupled with a more diverse T cell repertoire was linked to favorable clinical outcome. Structural genome-wide analyses showed a higher degree of genomic instability in responding tumors of epithelioid histology. Patients with germline alterations in cancer predisposing genes, especially those involved in DNA repair, were more likely to achieve long-term survival. Our findings indicate that concurrent durvalumab with platinum-based chemotherapy has promising clinical activity and that responses are driven by the complex genomic background of malignant pleural mesothelioma.

Conflict of interest statement

V.A. receives research funding to her institution from Bristol Myers Squibb and AstraZeneca. P.M.F. has received research funding to his institution from AstraZeneca, Bristol Myers Squibb, Novartis, Corvus and Kyowa. He has also served as a consultant for Amgen, AstraZeneca, Bristol Myers Squibb, Daiichi Sankyo, Iteos, Janssen, Mirati, Novartis and Sanofi and as a data and safety monitoring board member for Polaris and Flame Therapeutics. A.D. is Chief Medical Officer and Chief Executive Officer at TTC Oncology, Chief Medical Officer at Luminary Therapeutics, Chief Medical Officer at IGF Oncology, Chief Medical Officer at Squarex and an advisor to the Martell Diagnostic Laboratory. K.N.S. receives research funding to her institution from Bristol Myers Squibb, AstraZeneca and Enara Bio and holds founder’s equity in ManaT Bio. V.E.V. is a founder of Delfi Diagnostics and Personal Genome Diagnostics, serves on the board of directors and as a consultant for both organizations and owns Delfi Diagnostics and Personal Genome Diagnostics stock, which are subject to certain restrictions under university policy. Additionally, Johns Hopkins University owns equity in Delfi Diagnostics and Personal Genome Diagnostics. V.E.V. is an inventor of multiple licensed patents related to technologies from Johns Hopkins University. Some of these licenses and relationships are associated with equity or royalty payments directly to Johns Hopkins and V.E.V. V.E.V. is an advisor to Bristol Myers Squibb, Danaher, Genentech and Takeda Pharmaceuticals. Within the last 5 years, V.E.V. has been an advisor to Merck and Ignyta. These arrangements have been reviewed and approved by Johns Hopkins University in accordance with its conflict of interest policies. D.P. is a consultant for Aduro Biotech, Amgen, Bayer, Dynavax, Enara, FLX Bio, Immunomic Therapeutics, Janssen, Merck, Rock Springs Capitol, Tizona and Trieza; is on the Board of Directors of DNAtrix; is on the scientific advisory board of Immununica, WindMil, Dracen, Camden Partners and Astellas; receives research support from Bristol Myers Squibb and Compugen; and is a founder of ManaT Bio. C.C. is the founder of CM Cherry Consulting. J.W. is a consultant for Personal Genome Diagnostics, is the founder and owner of Resphera Biosciences and holds patents, royalties or other intellectual property from Personal Genomic Diagnostics. R.S.-D. is a consultant for Genentech/Roche, Bayer, Bristol Myers Squibb, Eli Lilly, AstraZeneca, NGM Biopharmaceuticals and Takeda Science Foundation. H.B. receives research support from Millennium, Merck, Celgene, Bristol Myers Squibb and Eli Lilly; is on the advisory board and/or a consultant for Bristol Myers Squibb, Eli Lilly, Genentech, Celgene, Pfizer, Merck, EMD-Serono, Boehringer-Ingelheim, AstraZeneca, Novartis, Genmab, Regeneron, BioNTech, Cantargia AB, Amgen, Abbvie, Axiom, PharmaMar, Takeda, Huya Bio, Mirati and Daiichi Sankyo; is on the data and safety monitoring board of the University of Pennsylvania, CAR T Program, Takeda and Incyte; is on the scientific advisory board for Sonnetbio (stock options), Rgenix (stock options) and Nucleai (stock options); receives honoraria from Amgen, Pfizer and Daiichi Sankyo; and receives travel support from Amgen, Bristol Myers Squibb, Merck, Eli Lilly, EMD-Serono and Genentech. J.B. is on the advisory board and/or a consultant for Amgen, AstraZeneca, Bristol Myers Squibb, Genentech/Roche, Eli Lilly, GlaxoSmithKline, Merck, Sanofi and Regeneron; receives grant research funding from AstraZeneca, Bristol Myers Squibb, Genentech/Roche, Merck, RAPT Therapeutics and Revolution Medicines; and is on the data and safety monitoring board/committees of GlaxoSmithKline, Sanofi and Janssen. H.L.K. reports personal fees from Aldeyra Therapeutics, AstraZeneca, Bayer, Boehringer-Ingelheim, Bristol Myers Squibb, Kyowa, Merck, Paredox Therapeutics, Deciphera, Inhibrx and Inventiva; and non-financial support from AstraZeneca, Boehringer-Ingelheim, Merck, Paredox Therapeutics and Inventiva. H.L.K. also reports funds given to support clinical trials at her institution from Aduro, AstraZeneca, Bayer, Bristol Myers Squibb, Deciphera, GlaxoSmithKline, Eli Lilly, Merck, Polaris, Verastem, Blueprint, Tesaro and Inhibrx. S.R. is a consultant for Amgen, Bristol Myers Squibb, Genentech/Roche, Merck, AstraZeneca, Takeda, Eisai, Daiichi Sankyo, Sanofi, GlaxoSmithKline and Eli Lilly and receives grants from Tesaro, Merck, AstraZeneca, Advaxis, Bristol Myers Squibb, Amgen, Takeda, Genmab and GlaxoSmithKline. All other authors declare no competing interests.

© 2021. The Author(s).

Figures

Fig. 1. Outcomes with chemo-immunotherapy in unresectable MPM.

a, Kaplan–Meier curve of OS in patients treated with durvalumab and platinum plus pemetrexed (n = 55). One-sided P value based on the Wald test for the log failure rate parameter is P = 0.0014, indicating significantly longer OS than the historical control of 12 months. b, Kaplan–Meier curve of PFS in patients treated with durvalumab and platinum plus pemetrexed (n = 55). c, Waterfall plot of best change in target lesions to treatment by histological subtype, based on maximal percentage of tumor reduction from baseline (n = 53 patients). Two patients without follow-up measurements in targeted lesions (one with best response unevaluable, the other with best response PD) were excluded. d, Spider plot of change in target lesions over time (n = 53 patients); notably, four patients had continued response or SD at the time of analysis. Two patients without follow-up measurements in targeted lesions (one with best response unevaluable, the other with best response PD) were excluded. e, Kaplan–Meier curves of OS according to histology; two-sided P value with significance level set at 0.05. f, Kaplan–Meier curves of PFS according to histology; two-sided P value with significance level set at 0.05. Epi, epithelioid. Source data

Fig. 2. Genomic landscape of chemo-immunotherapy-treated mesotheliomas.

MPMs of patients with a radiographic response harbored a higher number of non-synonymous missense sequence mutations (n = 40 MPM tumors; on average, 23 versus 18 mutations per exome for responding and non-responding tumors, respectively; Mann–Whitney P = 0.086). Epithelioid MPMs responding to therapy harbored a higher number of clonal missense mutations (n = 29 tumors; Mann–Whitney P = 0.051 and P = 0.025 for missense mutation load and clonal mutations, respectively); the numbers of subclonal mutations are shown as yellow inserts. Recurring inactivating alterations in BAP1, CDKN2A, NF2, TP53, SETD2 and PBRM1 did not differentially cluster with regard to therapeutic responses. Similarly, somatic BAP1 sequence alterations and CDKN2A homozygous deletions were detected in 32.5% (13 of 40) and 30% (12 of 40) of MPMs, without a notable enrichment with respect to therapeutic response. Specific genotypes were associated with exceptional therapeutic outcome (PFS ≥12 months and/or OS ≥24 months): patient 178 harbored tumor biallellic inactivation of NF2 and the histone methyltransferase SETD2; patient 926 harbored tumor biallellic inactivation of BAP1; and patient 361 harbored tumor homozygous deletions in BAP1 and in the SWI/SNF nucleosome remodeling gene PBRM1. An enrichment in mutations in chromatin-regulating genes was observed for patients achieving an OS of 12 or more months (Fisher’s exact P = 0.063). We identified a higher contribution of an HRD mutation signature in responsive tumors (n = 40 patients; average HRD contribution of 9.1% versus 1.6% in responding and non-responding tumors, respectively; Mann–Whitney P = 0.043). Conversely, an APOBEC mutation signature was found to be more enriched in non-responding MPM and epithelioid MPM tumors (Mann–Whitney P = 0.058 and P = 0.031, respectively). Mutations were characterized by consequence (missense, frameshift, nonsense and splice site) and recurrence (hotspots, depicted as solid circles), and loss of the wild-type allele was considered in case of truncating mutations (biallellic inactivation, marked with an ‘x’). Tumor samples from patients 329, 351, 629 and 923 were excluded from analyses of somatic alterations owing to tumor tissue quality and are not shown here; these patients were included in the germline analyses, with patient 629 harboring a deleterious mutation in BAP1. BOR, best overall response; Epi, epithelioid; Sarc, sarcomatoid. Source data

Fig. 3. Effect of germline mutations in cancer susceptibility genes on outcome from combined immuno-chemotherapy.

a, b, Patients harboring known deleterious germline mutations in mesothelioma-predisposing genes (Methods) had a longer OS (log-rank P = 0.05), especially in the epithelioid MPM group (log-rank P = 0.032). c, d, A focused analysis including deleterious germline mutations in BAP1, BRCA2, MSH6 and BLM—all genes involved in DNA damage repair—showed the same trends toward a longer OS for patients harboring germline mutations in DDR genes (log-rank P = 0.12 for all patients and log-rank P = 0.082 for patients with epithelioid tumors). All P values are two sided. DDR; DNA damage repair.

Fig. 4. Large-scale copy number analyses.

a, Genome-wide copy number analyses predominantly revealed genomic regions with copy number losses (shown in blue) and were used to determine the extent of copy number breakpoints and fraction of genome with complete allelic imbalance, reflecting genomic instability and tumor aneuploidy. The relative copy ratio (log copy ratio) values quantifying the abundance of each genomic region compared to the average genome ploidy are shown per chromosome after correction for tumor purity. Red and blue shades indicate copy gains and losses, respectively, whereas white marks indicate copy neutral regions. An HRD score was computed, taking into account telomeric allelic imbalance, LOH and large-scale state transitions. Three extreme cases of LOH were noted, with a copy number pattern that was suggestive of genome near-haploidization; these patients had an OS of 12 or more months after chemo-immunotherapy. b, c, A higher number of copy number breakpoints and a higher HRD score distinguished epithelioid MPM from patients with an OS of 12 or more months (n = 28 epithelioid MPM tumors; Mann–Whitney P = 0.0.05 and P = 0.014, respectively). d, Responding tumors harbored a higher number of mutations in single-copy regions of the genome, suggesting that these ‘difficult’ to eliminate alterations and associated neoantigens might be important drivers of the anti-tumor immune response (n = 40 MPM tumors; Mann–Whitney P = 0.027). The center line in the box plots represents the median; the upper limit of the box plots represents the third quantile (75th percentile); the lower limit of the box plots represents the first quantile (25th percentile); the upper whisker is the maximum value of the data that are within 1.5 times the interquartile range over the 75th percentile; and the lower whisker is the minimum value of the data that are within 1.5 times the interquartile range under the 25th percentile. All P values are two sided. Allelic imb. frac., fraction of genome with allelic imbalance; BOR, best overall response; CNA, copy number alteration. Source data

Fig. 5. Baseline TCR repertoire characteristics and dynamic changes at the time of therapeutic resistance.

a, The intratumoral T cell repertoire was interrogated by TCR Vβ sequencing; clonality of the TCR repertoire was computed; and the representation of dominant clones (Methods) as a proportion of the whole TCR repertoire was determined. CD8+ T cell density and PD-L1 tumor proportion scores for each evaluable case are shown (missing cases are shown in gray). b, These analyses revealed a less clonal TCR repertoire in tumors from patients achieving an OS of 12 or more months (Mann–Whitney P = 0.018). c, A higher representation of dominant clones was also detected in tumors from patients with a shorter OS (Mann–Whitney P = 0.006). d, Differential abundance analyses of three cases with available tumor samples before therapy initiation (295, 459 and 926) and at the time of acquired resistance revealed TCR clonotypic expansions (labeled as significant positive) and regressions (labeled as significant negative) as shown for patient 926, who had a PR and an OS of 27.8 months. Fold change of intra-tumoral TCR clones is plotted on the x axis (log scale), and the adjusted corresponding Mann–Whitney P value is shown on the y axis (−log scale) of the volcano plot. All P values are two sided. Biph; biphasic; BOR, best overall response; Epi, epithelioid; IHC, immunohistochemistry; NE, non-evaluable; Neg, significantly negative (regressing TCR clones); NS, not significant; Pos, significantly positive (expanding TCR clones); Sarc, sarcomatoid. Source data

Extended Data Fig. 1. CONSORT trial diagram for the PrE0505 study.

Outline of number of patients available for analyses.

Extended Data Fig. 2. Analyses Overview.

Whole exome sequencing data were analyzed to determine somatic sequence and focal copy number changes as well as germline variants in cancer predisposing genes and HLA class I and II haplotypes. Mutational spectra and enrichment in individual genes and gene sets were assessed. Genome-wide copy number analyses were performed to estimate genome aneuploidy, number of copy number breakpoints, homologous recombination deficiency as well as utilized to compute mutation cellular fractions. Computational predictions of mutation-associated neoantigens and immunogenic mutations were coupled with analyses of the intratumoral T cell repertoire and functional analyses of neopeptides-stimulated autologous peripheral T cells. PD-L1 and CD8+T cell expression was assessed by immunohistochemistry. This figure was developed using BioRender under a full license.

Extended Data Fig. 3. Treatment emergent adverse events and outcome by platinum agent used.

(a) All grades and grade 3/4 treatment emergent adverse events; events reported in greater than 20% subjects or grade 3/4 events reported in greater than 5% of patients, worst grade reported per patient (CTCAE Version 4.03) (b) Adverse events of special interest, investigator reported assessment and attribution as possibly, probably, or related for selected events, worst grade overall as reported for each patient. (c) Kaplan-Meier curves of OS according to the platinum agent used at the start of the treatment. (d) Kaplan-Meier curves of PFS according to the platinum agent used at the start of the treatment. There was no significant difference in OS or PFS for carboplatin vs. cisplatin-based chemotherapy (log rank p = 0.18 and p = 0.4 respectively). All p values are two-sided. Source data

Extended Data Fig. 4. Distribution of mutations in genes involved in DNA damage repair.

We investigated co-occurrence of mutations in DNA damage repair (DDR) genes involved in base excision repair, DNA damage sensoring, the Fanconi anemia pathway, homologous recombination, mismatch repair, nucleotide excision repair, non-homologous end joining and translesion DNA synthesis. Mutations were characterized by consequence (missense, frameshift, nonsense, splice site, in-frame) and recurrence (hotspots) and loss of the wild type allele was considered in case of truncating mutations (biallellic inactivation, marked with an ‘x’). There were no significant differences in somatic genomic alterations in the DDR gene set between tumors from responders and non-responders. When confirmed pathogenic germline mutations in DDR genes were considered, 5 patients harboring such alterations (629-BAP1 c.1717del, 295-BRCA2 c.6730 A>T, 628-BAP1 c.581-1 G > T, 976-MSH6 c.3261dup and 506-BLM c.968 A>G) had a longer overall survival as shown in Fig. 2. Abbreviations: TMB; tumor mutation burden, BOR; best overall response, CR; complete response, PR; partial response, SD; stable disease, PD; progressive disease, LOH; loss of heterozygosity, HRD; homologous recombination defect, CNA; copy number aberration.

Extended Data Fig. 5. Immunogenic mutation load may be linked with functional neopeptide-specific anti-tumor immune response and therapeutic response.

(a) Patients with MPM harboring a high non synonymous mutation load were more likely to achieve a radiographic response to chemo-immunotherapy (n = 40 MPM tumors, Mann Whitney p = 0.086). (b) MPMs responsive to chemo-immunotherapy also harbored a more clonal TMB (n = 40 MPM tumors, Mann Whitney p = 0.076). (c-d) These findings were more prominent in the epithelioid MPM group (n = 29 epithelioid MPM tumors, Mann Whitney p = 0.051 for non synonymous mutation load and Mann Whitney p = 0.025 for clonal mutation burden). (e,f) Immunogenic mutations associated with HLA class I and II neoantigens (Methods) better distinguished responders from non responders (n = 40 MPM tumors, Mann Whitney p = 0.064 for class I IMMs and Mann Whitney p = 0.023 for class II IMMs). (g,h) These findings were more pronounced for patients with epithelioid MPM (n = 29 epithelioid MPM tumors, Mann Whitney p = 0.035 for class I IMMs and Mann Whitney p = 0.038 for class II IMMs). The center line in the boxplots represents the median, the upper limit of the boxplots represents the third quantile (75th percentile), the lower limit of the boxplots represents the first quantile (25th percentile), the upper whiskers is the maximum value of the data that is within 1.5 times the interquartile range over the 75th percentile, and the lower whisker is the minimum value of the data that is within 1.5 times the interquartile range under the 25th percentile. (i) Survival analyses did not reveal any significant differences in overall survival between MPM tumors in the TCGA cohort with high TMB vs. those harboring a low TMB (log rank p = 0.21). (j) Analyses of progression-free survival revealed a trend towards longer PFS for patients with MPM tumors in the TCGA cohort harboring a low TMB (log rank p = 0.05). The TMB-high and low groups were defined using the 2nd quartile of TMB in the TCGA mesothelioma cohort, which corresponds to a non-synonymous mutation burden ≥33 alterations/exome. All p values are two-sided. Abbreviations: IMM; immunogenic mutation, CR; complete response, PR; partial response, SD; stable disease, PD; progressive disease.

Extended Data Fig. 6. Germline and tumor HLA class I and II genetic variation.

(a) Tumors from responders harbored a higher load of HLA class I and II immunogenic mutations (Methods, Mann Whitney p = 0.064 and 0.023 respectively) especially in the epithelioid MPM group (Mann-Whitney p = 0.035 and p = 0.038 respectively). There were no differences in the degree of germline homozygosity found between responders and non-responders. HLA class I and II germline zygosity and somatic LOH events were combined to calculate the unique number of HLA class I and class II alleles on cancer cells, which did not differ between responders and non responders. Similarly, there was no evidence of biallellic inactivation of β2-microglobulin or differential abundance of β2-microglobulin LOH events with respect to therapeutic response. (b) When sequence divergence of the allele’s peptide-binding domains was computed for HLA class I alleles, patients with high pairwise divergence between HLA alleles and in particular in HLA-B achieved a radiographic response (Mann Whitney p = 0.06), suggesting that in high HLA divergence may be linked with an increased diversity of the neopeptide repertoire presented and reflective of a more potent-antitumor immune response. (c) While there were no differences in mean HLA class I divergence for patients with epithelioid MPM (Mann Whitney p = 0.3), (d) a higher HLA-B divergence was linked with radiographic response (Mann Whitney p = 0.003). All p values are two-sided and unadjusted for multiple comparisons. Abbreviations: IMM; immunogenic mutation, B2M; β2-microglobulin, LOH; loss of heterozygosity.

Extended Data Fig. 7. TCR clonotypic expansions to neopeptides derived from immunogenic mutations.

(a) We used the MANAFEST assay, which detects memory T cell responses and identifies antigen-specific T cell receptors to test neoantigen-specific TCR reactivity. Neopeptides derived from class I immunogenic mutations were tested in vitro in autologous T cell cultures from peripheral blood of patient 459 that achieved an overall survival of 32.88 months after chemo-immunotherapy. Neopeptide-specific TCR clonotypic expansions were identified post stimulation with peptides derived from the SRPK2 p.C234Y and NDUFS2 p.V412L immunogenic mutations (orange bars, y axis denotes TCR productive frequencies of CD8+ T cells after the 10-day culture). (b) Neontigen-specific TCR clonotypic expansions were noted in autologous CD8+ T cell cultures from the peripheral blood for patient 295 that achieved an overall survival of 21.85 months and remained event-free at the time of data lock. TCR clonotypic expansions for peptides derived from the immunogenic mutations SCRN1 p.V334A, PSD2 p.C307Y, ZNF469 p.P3471S and CD72 p.T71A (orange bars, y axis denotes TCR productive frequencies of CD8+ T cells after the 10-day culture). Peptide pools of known HLA class I-restricted viral antigens (Methods) were utilized as positive controls and are shown in blue bars for both cases.

Extended Data Fig. 8. Differences in CD8+T cell infiltration in BAP1 mutant tumors.

(a) Epithelioid MPMs harboring somatic BAP1 inactivating mutations had a higher intratumoral CD8+ T cell infiltration (n = 33 epithelioid MPM tumors, Mann Whitney p = 0.036). (b) RNA sequencing of tumor samples with adequate tissue (n = 6, 4 BAP1 wild type and 2 BAP1 mutant) revealed a higher expression level for GranzymeB in BAP1 mutant tumors (n = 6 MPM tumors, Mann Whitney p = 0.0094). The center line in the boxplots represents the median, the upper limit of the boxplots represents the third quantile (75th percentile), the lower limit of the boxplots represents the first quantile (25th percentile), the upper whiskers is the maximum value of the data that is within 1.5 times the interquartile range over the 75th percentile, and the lower whisker is the minimum value of the data that is within 1.5 times the interquartile range under the 25th percentile. All p values are two-sided. Source data

Extended Data Fig. 9. Genome-wide loss of heterozygosity and genome near haploidization.

(a) MPM tumors exhibited various levels of genome-wide loss of heterozygosity. LOH in chromosomal arms 4p, 4q, 6q, 13q (containing the BRCA2, ERCC5 and LATS2 loci), 14q (containing the FANCM, MLH3 and XRCC3 loci), and 22q (containing the CHECK2, NF2 and XRCC6 loci) was observed at a significantly higher rate compared to background. In three tumors (cases 506, 628, 976), LOH was widespread throughout the genome consistent with the genome near haploidization phenotype. (b) Genome-wide copy number (top) and B-allele frequency (bottom) profiles of a tumor with GNH (976), depicting a hyper-diploid state and LOH in all chromosomes observed at normal copy number state. Abbreviations: LOH; loss of heterozygosity, GNH; genome near haploidization. (c-e) Comparison of the background rate of genomic loss in 1,086 mesothelioma and non-small cell lung cancer cases from TCGA showing that haploid regions of the genome consistently harbored a lower background rate of genomic loss compared to diploid regions (Mann Whitney p = 1e-09, p < 2.2e-16 and p < 2.2e-16 for mesothelioma, lung adenocarcinoma and lung squamous cell carcinomas respectively). (f) Patients in the PrE0505 study with a radiographic response harbored a higher number of clonal mutations within the haploid regions of their genomes (n = 40 MPM tumors, Mann Whitney p = 0.043). The center line in the boxplots represents the median, the upper limit of the boxplots represents the third quantile (75th percentile), the lower limit of the boxplots represents the first quantile (25th percentile), the upper whiskers is the maximum value of the data that is within 1.5 times the interquartile range over the 75th percentile, and the lower whisker is the minimum value of the data that is within 1.5 times the interquartile range under the 25th percentile. All p values are two-sided and unadjusted for multiple comparisons. Abbreviations: MESO; mesothelioma, LUAD; lung adenocarcinoma, LUSC; lung squamous cell carcinoma, CR; complete response, PR; partial response, SD; stable disease, PD; disease progression. Source data

Extended Data Fig. 10. Correlations between genomic and molecular features.

Non parametric correlations between sequence and structural genomic and TCR features for all patients (a) and patients with epithelioid mesothelioma (b). Correlations between tumor purity and non-synonymous missense mutation burden and associated neoantigens were noted for all patients but not for the epithelioid mesothelioma group. An asterix indicates a p value less than 0.05.