Abstract
Therapeutic vaccines that elicit cytotoxic T cell responses targeting tumor-specific neoantigens hold promise for providing long-term clinical benefit to patients with cancer. Here we evaluated safety and tolerability of a therapeutic vaccine encoding 20 shared neoantigens derived from selected common oncogenic driver mutations as primary endpoints in an ongoing phase 1/2 study in patients with advanced/metastatic solid tumors. Secondary endpoints included immunogenicity, overall response rate, progression-free survival and overall survival. Eligible patients were selected if their tumors expressed one of the human leukocyte antigen-matched tumor mutations included in the vaccine, with the majority of patients (18/19) harboring a mutation in KRAS. The vaccine regimen, consisting of a chimp adenovirus (ChAd68) and self-amplifying mRNA (samRNA) in combination with the immune checkpoint inhibitors ipilimumab and nivolumab, was shown to be well tolerated, with observed treatment-related adverse events consistent with acute inflammation expected with viral vector-based vaccines and immune checkpoint blockade, the majority grade 1/2. Two patients experienced grade 3/4 serious treatment-related adverse events that were also dose-limiting toxicities. The overall response rate was 0%, and median progression-free survival and overall survival were 1.9 months and 7.9 months, respectively. T cell responses were biased toward human leukocyte antigen-matched TP53 neoantigens encoded in the vaccine relative to KRAS neoantigens expressed by the patients’ tumors, indicating a previously unknown hierarchy of neoantigen immunodominance that may impact the therapeutic efficacy of multiepitope shared neoantigen vaccines. These data led to the development of an optimized vaccine exclusively targeting KRAS-derived neoantigens that is being evaluated in a subset of patients in phase 2 of the clinical study. ClinicalTrials.gov registration: NCT03953235.
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Data availability
Deidentified individual participant clinical data that underlie the results reported in this article are available for transfer. Interested investigators can obtain and certify the data transfer agreement and submit requests to the principal investigator K.J. Investigators and institutions who consent to the terms of the data transfer agreement form, including, but not limited to, the use of these data for the purpose of a specific project and only for research purposes and to protect the confidentiality of the data and limit the possibility of identification of participants in any way whatsoever for the duration of the agreement, will be granted access. Gritstone will then facilitate the transfer of the requested deidentified data within 60 days. This mechanism is expected to be via a Gritstone Secure File Transfer Service, but Gritstone reserves the right to change the specific transfer method at any time, provided appropriate levels of access authorization and control can be maintained. Epitope selection utilized a previously published model11. Source data are provided with this paper.
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Acknowledgements
The study sponsor was Gritstone bio, Inc. We thank the patients and their families; clinical staff and study coordinators; Bristol Myers Squibb for providing ipilimumab and nivolumab; and M. Marrali, K. Taquechel, J. Busby, R. Yelensky and M. Skoberne. Funding was provided by the study sponsor, Gritstone bio. Employees of Gritstone bio received salaries for study conceptualization, design, data analyses and manuscript preparation. No other authors received specific funding for this work from the sponsor.
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A.R.R., M.L., A.R.F., M.G.H., D.S. and K.J. wrote the manuscript. A.R.R., C.D.P., M.J.D., A.R.F., M.L., A.A. and K.J. contributed to study design. A.R.R., M.L., M.G.H., D.S., C.D.P., M.J.D., L.S., L.K., S.K., G.R.B., M.E., Y.J.L., M.L., L.A., J.R.J., L.D.K.T., C.P., L.S., R.Z., A. Shen, A.Y. and K.B. contributed to experimental design, execution and data analysis. A.R.F., C.K., A.I.S., C.-Y.L., A. Shergill, M.L.J., B.S.H., D.P.C., B.J., A.M., J.R.H. and J.W.G. contributed to clinical oversight, patient recruitment, enrollment and treatment. C.D.S., R.L.V. and J.A.E. contributed to vaccine design and production.
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A.R.R., M.L., M.G.H., C.D.S., D.S., C.D.P., M.J.D., S.K., L.K., A.Y., Y.J.L., M.L., A. Shen, G.R.B., M.E., R.L.V., J.A.E., J.R.J., L.D.K.T., L.A., C.P., L.S., R.Z., K.B., A.A., A.R.F. and K.J. are stockholders and either current or previous employees at Gritstone bio, Inc. and may be listed as co-inventors on various pending patent applications related to the vaccine platform presented in this study. C.K. received honoraria from OncLive, Total Health, consults/has consulted for Scenic Immunology BV and has received research funding from Bristol Myers Squibb, Merus, Gritstone bio and Acrivon. C.-Y.L. consults/has consulted for AstraZeneca, Genentech, Histosonics, Incyte, Ipsen, QED, Transthera and Boston Scientific and is a speaker for AstraZeneca and Incyte. A.I.S. has a leadership role at NEXT Oncology Virginia; is a stockholder of Eli Lilly; has received honoraria from CytomX Therapeutics, AstraZeneca/MedImmune, Merck, Takeda, Amgen, Janssen Oncology, Novartis, Bristol Myers Squibb and Bayer; has consultant or advisory roles at Incyte, Amgen, Novartis, Mirati Therapeutics, Gritstone bio, Jazz Pharmaceuticals, Takeda, Janssen Research & Development, Mersana, Daiichi Sankyo/AstraZeneca, Regeneron, Lilly, Black Diamond Therapeutics, Sanofi, Array BioPharma, AstraZeneca/MedImmune, Bristol Myers Squibb and Blueprint Medicines; has received research funding from LAM Therapeutics, Regeneron, Roche, AstraZeneca, Boehringer Ingelheim, Astellas Pharma, MedImmune, Novartis, Incyte, Abbvie, Ignyta, Takeda, Macrogenics, CytomX Therapeutics, Astex Pharmaceuticals, Bristol Myers Squibb, Loxo, Arch Therapeutics, Gristone bio, Plexxikon, Amgen, Daiichi Sankyo, ADCT, Janssen Oncology, Mirati Therapeutics, Rubius, Mersana, Blueprint Medicines, Alkermes, Revolution Medicines, Medikine, Synthekine, Black Diamond Therapeutics, BluPrint Oncology, Nalo Therapeutics, Scorpion Therapeutics and ArriVent Biopharma. M.L.J.: financial interests, personal, advisory board: Astellas, Otsuka; financial interests, institutional, research grant: AbbVie, Acerta, Amgen, Apexigen, Arcus, Array, AstraZeneca, Atreca, Beigene, Birdie, Boehringer Ingelheim, Checkpoint Therapeutics, Guardant Health, Genocea, Hengrui, Immunocore, Incyte, Janseen, Jounce, Gritstone bio, Lycera, Merck, Mirati, Oncomed, Regeneron, Ribon, Sanofi, Shattuck Labs, Stem CentRx, Syndax, Takeda, Tarveda, TCR2 Therapeutics, University of Michigan and WindMIL. B.J.: financial interests, personal, advisory board: Gritstone bio, Inc., Incyte, Taiho Oncology; financial interests, personal, research grant: BMS, Syntrix. A.M.: financial interests, personal, advisory board: Taiho, Incyte. J.R.H.: financial interests, personal, advisory board: Ipsen, Merck and Acrotech Biopharma; financial interests, institutional, research grant: ARMO Biosciences, Halozyme, Amgen, Merck, AbbVie, Advaxis, Astellas, Forty Seven, Immunomedics, Lilly, Gritstone bio, GSK and Arcus. The other authors declare no competing interests.
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Extended data
Extended Data Fig. 1 Tumor response waterfall plot.
Best percent change from baseline in target lesion for each patient (n = 19).
Extended Data Fig. 2 ctDNA monitoring of tumor variants in SLATE patients.
(A-E) Levels of ctDNA over time post prime vaccination for each patient for the vaccine encoded tumor mutation (orange) as well as additionally detected somatic mutations (gray). ctDNA levels reported as variant allele frequency (VAF) as a percentage of total reads. (A-D) All 4 patients with molecular response (MR) (out of 12 patients assessed), defined as > 30% decrease in vaccine encoded tumor variant compared to baseline. (E) Representative patient with no MR, showing loss of B2M start codon. The best overall response is denoted.
Extended Data Fig. 3 HLA Loss of Heterozygosity.
(A) HLA LOH was assessed in cfDNA and biopsies as described in the methods. Patient was considered LOH if LOH was detected in a biopsy or at any cfDNA timepoint. n = 18. Patient S1 was not assessed for HLA LOH due to lack of sufficient cfDNA material. (B) Longitudinal change of read allele fraction in cfDNA relative to genomic DNA (gDNA) across HLA class I alleles of S13, a molecular responder who demonstrated focal LOH that included the SLATE-relevant HLA genotype. (C) Longitudinal change of read allele fraction in cfDNA relative to gDNA of molecular non-responder S4. Patient is homozygous in HLA-B.
Extended Data Fig. 4 Tumor neoantigen specific T cell responses measured by ex vivo IFNγ ELISpot in PBMCs at baseline and at various timepoints post ChAd68 prime for each patient.
n = 16 patients (S8 and S10 not tested by ex vivo ELISpot). LOD 30. Data presented as mean SFU/106 cells ± s.d. for 2 – 3 technical replicates per sample.
Extended Data Fig. 5 Expression levels in cell lines used for mass spectrometry and analysis of HLA-A*02 expressing cell lines.
Transcript levels of HLA-A (A) and PADRE (Pan HLA DR epitope presented at the c-terminal of each epitope cassette) (B) measured by comparative real-time PCR in engineered K562 monoallelic HLA-A*11:01 cell lines expressing different formats of epitope cassettes. Fold-change normalized to TBP housekeeping gene. Mean fold-change ± s.d. are shown for 2 technical replicates. Mean values annotated above bars. (C) Target density of KRAS and TP53 epitopes detected by mass spectrometry in HLA-A*02:01 monoallelic cell lines. TP53-R213L measured in A*02:01 K562 lines transduced with the SLATEv1 cassette (20×1, 20 unique epitopes with 1 copy each) using cassettes with doxycycline-induced expression or zeocin selection. Data presented as mean copies/cell ± s.d. of 3 independent experiments, mean value annotated. No KRAS antigens were detected in 20×1, but very low levels of KRAS G12V and G12D were detected in 3 A*02:01 K562 lines transduced with SLATE 4×4 cassettes (4 unique KRAS epitopes with 4 copies each in cassettes engineered with either blastocidin or hygromycin selection) which have no TP53 epitopes included. *Copies/cell are reported as measured by relative response of endogenous peptide to stable isotopic labeled synthetic peptide added to isolated sample immunopeptides immediately before mass spectrometry analysis, then adjusted for an average 9% pHLA process recovery.
Extended Data Fig. 6 Immunogenicity of vaccines assessed in HLA transgenic mice.
(A) Antigen-specific T cell response assessed in splenocytes of HLA-A*11 transgenic mice (n = 6/group) 2 weeks post intramuscular immunization with the specified ChAd vaccine (5×1010 VP each) by IFNγ ELISpot following overnight stimulation with the KRAS G12C minimal epitope peptide (VVVGACGVGK). (B) Antigen-specific T cell response assessed in splenocytes of HLA-A*02 transgenic mice (n = 6/group) 2 weeks post intramuscular immunization with the specified ChAd vaccine (5×1010 VP each) by IFNγ ELISpot following overnight stimulation with the TP53-R213L peptide pool containing all possible 8 – 11mer epitopes spanning the neoantigen 25mer. (C-D) Antigen-specific T cell response assessed in splenocytes of HLA-A*11 (C) or HLA-A*01 (D) transgenic mice (n = 6/group) 2 weeks post intramuscular immunization with the specified ChAd vaccine (5×1010 VP each) by IFNγ ELISpot following overnight stimulation with the PADRE peptide. Data from same experiments shown in Fig. 4, representative data from 2 studies. (A-D) Box and whiskers represent IQR and range, line is median, log2 scale. LOD 33, values < LOD set to one-half LOD.
Extended Data Fig. 7 Expression of immunodominant AH1 epitope on the same vaccine cassette leads to decreased immune response to the E2F8 sub-dominant epitope, rescued by delivery of both epitopes in different vaccines.
(A) Design of vaccine epitope cassettes evaluated for immunogenicity in Balb/c mice. Boxes represent epitopes for which immune data is presented, lines represent other epitopes. (B) Antigen-specific T cell response assessed in splenocytes of Balb/c mice (n = 6/group) 2 weeks post intramuscular immunization with the specified ChAd vaccine (5×109 VP each) by IFNγ ELISpot following overnight stimulation with the specified peptide. Single vaccines were administered bilaterally, 2.5×109 vp per leg, for a total dose of 5×109 VP. For the blended approach vaccines were combined in a single syringe and then delivered bilaterally (5×109 VP each vaccine, 1×1010 VP total), for the bilateral approach mice received 1×1010 VP of ChAd (5×109 VP of E2F8×4 in the left tibialis anterior (TA) and 5×109 VP of AH1 in the right TA). Box and whiskers represent IQR and range, line is median, log2 scale. LOD 33, values < LOD set to one-half LOD.
Extended Data Fig. 8 Antigen repetition within vaccine expression cassette leads to increased antigen-specific T cell responses.
(A-B) Antigen-specific T cell response assessed in splenocytes of C57BL6 mice (n = 8/group) 2 weeks post immunization. IFNγ ELISpot following overnight stimulation with the specified peptide. Box and whiskers represent IQR and range, line is median, log2 scale. LOD = 33, values < LOD set to ½ LOD. (A) Mice immunized with 10 µg samRNA expressing a vaccine cassette encoding 20 mouse epitopes (1x), 10 epitopes repeated twice (2x), or 5 epitopes repeated 4 times (4x). Data from 3 H2-Kb restricted epitopes derived from B16 tumor mouse model shown. (B) Mice immunized with 1x109 VP ChAd vaccine encoding 20 epitopes with no repeats (20x1), 3 epitopes repeated 5 times (5x), or 3 epitopes repeated 6-7 times (6x or 7x). Data from 3 H2-Kb restricted epitopes derived from MC38 tumor mouse model shown.
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Supplementary Table 1, Supplementary Data and clinical study protocol (redacted).
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Rappaport, A.R., Kyi, C., Lane, M. et al. A shared neoantigen vaccine combined with immune checkpoint blockade for advanced metastatic solid tumors: phase 1 trial interim results. Nat Med 30, 1013–1022 (2024). https://doi.org/10.1038/s41591-024-02851-9
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DOI: https://doi.org/10.1038/s41591-024-02851-9
