Background
The BBB and immune microenvironment in GBM: implications for the development of new therapies
GBM immunotherapy: disappointing initial clinical results of monotherapies
Trial | Treatment | Outcome | Reference |
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CheckMate 143 phase III (NCT02017717) | anti-PD-1 (nivolumab) vs anti-VEGF (bevacizumab) | Primary endpoint not reached ↔ No improvement of mOS with anti-PD-1 (9.8 months) vs anti-VEGF (10 months) | Reardon et al., (2020) [110] |
CheckMate 498 phase III (NCT02617589) | anti-PD-1 (nivolumab) + RT vs TMZ + RT | Primary endpoint not reached ↔ No improvement of mOS with anti-PD-1 + RT (13.4 months) vs control treatment (14.9 months) | BMS press release; ClinicalTrials.gov |
CheckMate 548 phase III (NCT02667587) | anti-PD-1 (nivolumab) + SOC vs placebo + SOC | Primary endpoint not reached Data not yet released | BMS press release |
ACT IV phase III (NCT01480479) | Peptide vaccine targeting EGFRvIII (rindopepimut) + TMZ vs placebo + TMZ | Primary endpoint not reached ↔ No improvement of mOS with rindopepimut (20.1 months) vs control group (20.0 months) | Weller et al., (2017) [111] |
NCT00045968 phase III | Dendritic cell vaccine (DCVax®-L) + SOC vs placebo + SOC | Put on hold For unidentified reasons | Liau et al., (2018) [112]; ClinicalTrials.gov |
Targeting different arms of the GBM immunity cycle to improve immunotherapy efficacy: preclinical aspects
Combination of immunotherapy with chemotherapy
Combination treatment | Protocol | Cell line and model | Outcome | Reference |
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anti-PD-1 TMZ | • Tumor implantation: 2 × 105 cells • TMZ: 30 mg/kg, 5 consecutive days starting at d8, IP • anti-PD-1: 10 mg/kg 2x on d13 and d15, IV | GL261-Luc orthotopic syngeneic | • Combined therapy showed better antitumor efficacy than monotherapies with 100% tumor regression • TMZ abrogated the favorable immunological effects of anti-PD-1 (increased TIL numbers, decreased Treg and exhausted T cell frequencies, increased immunological memory) | Park J., et al. (2018) [123] |
anti-PD-1 TMZ (standard or metronomic dose) | • Tumor implantation: / • TMZ: - Standard dose (SD): 50 mg/kg for 5 consecutive days, IP - Metronomic dose (MD): 25 mg/kg for 10 consecutive days, IP • anti-PD-1: 10 mg/kg 4x every 5 days, IP | GL261 orthotopic syngeneic | • SD TMZ increased exhaustion markers on T cells, while MD TMZ did not lead to T cell exhaustion • anti-PD-1 reversed the exhaustion induced by SD TMZ in peripheral T cells but not in TILs • The survival benefit of anti-PD-1 therapy was abrogated by SD TMZ but not by MD TMZ | Karachi A., et al. (2019) [124] |
anti-PD-1 TMZ (low dose) | • Tumor implantation: 5 × 104 cells, right cerebral cortex • TMZ: 50 μg/kg, 5 consecutive days, IP • anti-PD-1: 200 μg 3x, IP | GL261 orthotopic syngeneic | • Combined therapy synergistically inhibited GBM tumor growth with a higher median survival time, a reduced tumor volume and 40% long-term survivors • Combined therapy increased CD4 and CD8 T cell infiltration in tumor lesions | Dai B., et al. (2018) [126] |
anti-PD-1 TMZ or carmustine (BCNU) (systemic or local administration) | • Tumor implantation: 1.3 × 105 cells, left striatum • Systemic chemotherapy (SC): - TMZ: 66 mg/kg, daily from d10 to d14, IP - BCNU: 5, 15 and 30 mg/kg, 3x/week for 2 weeks starting at d14, IP • Local chemotherapy (LC): - TMZ: implanted at d10 - BCNU: implanted at d14 Polymer impregnated with chemotherapy (wafer), allowing constant release in the TME for at least 2 weeks, placed directly on top of the tumor • anti-PD-1: 200 μg 3x, on d0, d12, and d14, IP | GL261-Luc orthotopic syngeneic | • Combination of LC and anti-PD-1 induced a robust immune response and survival benefit, with higher numbers of TILs and IFN-γ-secreting CD8 T cells in the brain, a higher Teff/Treg ratio and a higher tumor-infiltrating DC % • LC preserved the memory response upon rechallenge; SC abrogated it • SC abrogated the immunological benefits of anti-PD-1, did not provide survival benefit and resulted in severe lymphodepletion and severe depletion of TILs • SC alone or in combination with anti-PD-1 delayed tumor progression, but tumors recurred | Mathios D., et al. (2016) [125] |
TMZ (systemic) ICD-based DC vaccine | • Tumor implantation: 5 × 105 cells • ICD-based DC vaccine: 1 × 106 DCs, IP - On d2, d9 and d15 – vaccine alone - On d21, d28 and d35 – combination Production: cancer cells were incubated with hypericin followed by light irradiation, and then, Hyp-PDT-treated cells were mixed with DCs • TMZ: 40 mg/kg, 6x on d5, d7, d9, d12, d14, and d16 | GL261 orthotopic syngeneic | • Combined therapy provided a strong survival benefit with improved median survival and 50% long-term survivors • The ICD-based vaccine partially overcame the immune-ablating effects of chemotherapy. TMZ decreased the levels of brain-infiltrating CD8 T cells, but the combination decreased the levels of Tregs in the brain | Garg AD., et al. (2016) [127] |
TMZ (systemic or local) GL-GM (whole tumor cell vaccine) | • Tumor implantation: 5 × 103 cells, right frontal lobe • GL-GM: 2 × 106 irradiated (40 Gy) GL261-GMCSF cells, on d5, d19, and d33, IP • TMZ: - Systemic (SC): 50 mg/kg, at d7, d8 and d9, IP - Local (LC): 4.2 mg/kg/day, from d7 to d9, intratumoral | GL261 orthotopic syngeneic | • Local administration of TMZ induced a higher survival rate than systemic administration, and the effect was T cell-dependent • SC but not LC TMZ depleted blood leukocytes • Combination of TMZ IC and GL-GM increased survival and induced immune benefits with increased CD4 and CD8 TILs • Immune memory was established in long-term survivors (SC TMZ + GL-GM) | Fritzell S., et al. (2013) [128] |
TMZ (local) Whole cell vaccine | • Tumor implantation: 5 × 103 cells, right frontal lobe • Whole cell immunization: 2 × 106 irradiated (40 Gy) cells (GL261 or KR158) on d5, d19, and d33, SC • TMZ: 180 μg administered over 3 days, starting on d7, convection-enhanced delivery (CED), intratumoral | GL261 or KR158-Luc orthotopic syngeneic | • CED-TMZ and the whole cell vaccine synergized in the GL261 model resulting in 93% long-term survivors • The whole cell vaccine cured some mice of the KR158 model, and CED-TMZ prolonged median survival; however, there was no synergy between chemotherapy and immunotherapy • CED-TMZ plus the vaccine significantly decreased tumor volume and increased the intratumoral influx of T cells in both models | Enriquez Pérez JE., et al. (2020) [129] |
TMZ anti-CD47 anti-PD-1 | • Tumor implantation: 1 × 105 cells, caudate putamen • TMZ: - Concurrent: 80 mg/kg, at d11, d13 and d15, IP - Sequential: metronomic dose (20 mg/kg) at d7–9 + 80 mg/kg at d11, d13 and d15, IP • anti-CD47 (MIAP-140): 100 μg, at d11, d13 and d15, IP • anti-PD-1: 100 μg, on d16, d18 and d20, IP | GL261 or CT2-A orthotopic syngeneic | • Sequential TMZ treatment combined with anti-CD47 improved tumor growth inhibition and mice survival; monotherapies and concurrent treatment did not • Combination of sequential TMZ and anti-CD47 activated immune response in vivo, with significant increase of CD4 and CD8 T cell, IFN-γ-secreting cell and activated TAM numbers • Triple combination of TMZ, anti-CD47 and anti-PD-1 further improved the survival | von Roemeling CA., et al. (2020) [130] |
Combination of multiple immunotherapies
Combination treatment | Protocol | Cell line and model | Outcome | Reference |
---|---|---|---|---|
anti-PD-1 anti-LAG-3 | • Tumor implantation: 1.3 × 105 cells, striatum • anti-PD-1: 200 μg, IP - on d7, d10, d12, and d14 (early schedule) - on d10, d12, and d14 (late schedule) • anti-LAG-3: 200 μg, IP - on d7 and d10 (early schedule) - on d10 and d12 (late schedule) | GL261-Luc orthotopic syngeneic | • Early treatment with anti-LAG-3 and/or anti-PD-1 significantly improved survival • Late treatment with anti-LAG-3 did not significantly improve survival, but the combination did • The global immunological profiles were not different between the different treatment arms • Immune memory was established in long-term survivors | Harris-Bookman S., et al. (2018) [137] |
anti-PD-1 anti-TIGIT | • Tumor implantation: 1.3 × 105 cells, striatum • anti-PD-1: 200 μg, on d10, d12, and d14, IP • anti-TIGIT: 200 μg, IP, every other day for a total of 5 doses starting on d8, d10, d12 or d14 (4 ≠ schedules: A, B, C, D) | GL261-Luc orthotopic syngeneic | • Combination therapy improved long-term survival following each schedule • Combination therapy increased immune cell tumor infiltration and cytokine production • Tumor-infiltrating DCs were reduced following anti-TIGIT and anti-PD-1 combination treatment • Immune memory was established in long-term survivors | Hung AL., et al. (2018) [138] |
anti-CTLA-4 anti-PD-L1 1-MT | • Tumor implantation: 4 × 105 cells • anti-CTLA-4: 100 μg loading dose followed by 3 × 50 μg maintenance doses every 3 days, IP • anti-PD-L1: 500 μg loading dose followed by 3 × 200 μg maintenance doses every 3 days, IP • 1-MT: in the drinking water over 30 days, starting on d7 for early blockade or on d14 for late blockade | GL261 orthotopic syngeneic | • Early blockade with the triple combination cured 100% of mice, reduced Treg infiltration and increased IFN-γ-secreting CD8 T cell infiltration • Late blockade prolonged survival (78% long-term survival rate) and reduced Treg infiltration but also reduced brain-infiltrating T cells | Wainwright DA., et al. (2014) [139] |
anti-GITR agonist SRS | • Tumor implantation: 1.3 × 105 cells, striatum • SRS: 10 Gy radiation (1.9 Gy/min), d10, focal • anti-GITR: 10 mg/kg, 3x, on d10, d13, and d16, IP | GL261-Luc orthotopic syngeneic | • Combination therapy improved survival • The combination increased the CD8 effector T cell/Treg ratio • Immune memory was established in long-term survivors | Patel MA., et al. (2016) [140] |
anti-PD-1 anti-OX40 agonist GVAX (whole tumor cell vaccine) | • Tumor implantation: 7.5 × 104 cells, right striatum • anti-PD-1: 200 μg, on d3, d6, and d9, IP • anti-OX40: 250 μg, on d3, d6, and d9, IP • GVAX: 1 × 106 irradiated (35 Gy) GL261-GMCSF cells, on d3, d6, and d9, SC | GL261 orthotopic syngeneic | • The anti-PD-1 + anti-OX40 dual combination improved survival and the CD8/Treg ratio • The anti-PD-1 + GVAX dual combination improved survival and the CD8/Treg ratio and increased brain-infiltrating CD8 T cells • The triple combination led to 100% long-term survival with an increase in IFN-γ- and IL-2-secreting splenocytes and the CD4/CD8 ratio • Immune memory is established in long-term survivors | Jahan N., et al. (2019) [141] |
anti-PD-L1 Neoantigen peptide vaccine | • Tumor implantation: 5 × 104 cells • anti-PD-L1: on d7, d9, and d11, IP • Vaccine: 50 μg of each peptide + 100 μg polyIC adjuvant, on d3, d6, and d9, SC | CT2A orthotopic syngeneic | The combination therapy significantly improved mouse survival (60% long-term survivors) | Liu CJ., et al. (2020) [142] |
anti-PD-1 anti-TIM3 SRS | • Tumor implantation: 1.3 × 105 cells, left striatum • SRS: 10 Gy radiation (1.9 Gy/min), d10, focal, using the Small Animal Radiation Research Platform (SARRP) • anti-PD-1: 200 μg, on d10, d12, and d14 • anti-TIM-3: 250 μg, on d7, d11, and d-15 | GL261-Luc orthotopic syngeneic | • The anti-PD-1 and dual therapies improved survival and led to long-term survival • The triple combination led to 100% remission • The triple combination improved the immune profile of the TME and the cytokine profile of both CD4 and CD8 T cells (increased the CD8/Treg ratio, decreased the frequency of FoxP3+ Tregs, and increased the production of the inflammatory cytokines IFN-γ, TNF-α, and IL-17a) • Immune memory was established in long-term survivors | Kim JE., et al. (2017) [143] |
anti-CTLA-4 anti-4-1BB agonist SRS | • Tumor implantation: 1.3 × 105 cells, left striatum • SRS: 10 Gy radiation (1.9 Gy/min), d10, focal, using the SARRP • anti-4-1BB: 200 μg 3x, on d11, d14, and d17, IP • anti-CTLA-4: 800 μg 3x, on d11, d17, and d23, IP | GL261-Luc orthotopic syngeneic | • The SRS + anti-CTLA-4 dual therapy prolonged survival, but only the triple combination led to long-term survival • The triple therapy and double immunotherapy led to higher TILs (CD4 and CD8 T cells) • Immune memory was established in long-term survivors and was glioma-specific | Belcaid Z., et al. (2014) [144] |
anti-PD-1 anti-CXCR4 | • Tumor implantation: 1.3 × 105 cells, left striatum • anti-PD-1: 200 μg, on d10, d12, and d14, IP • anti-CXCR4: 200 μg, on d10, d12, and d14, IP | GL261-Luc orthotopic syngeneic | • The combination improved survival • Combination therapy decreased populations of suppressive myeloid cells in the brain • Combination therapy decreased the CD4/CD8 and Treg/CD8 ratios in the brain to a higher extent than did the monotherapies • The combination and monotherapies increased the levels of circulating inflammatory antitumor cytokines • Immune memory was established in long-term survivors | Wu A., et al. (2019) [145] |
anti-PD-1 Antiangiogenic therapy (anti-VEGF + anti-Ang-2) | • Tumor implantation: 1 × 105 cells, striatum • anti-PD-1: 10 mg/kg, 2x/week starting on d10 for a total of 8 doses, IP • Antiangiogenic therapy: 25 mg/kg (anti-VEGF) and 5.6 mg/kg (anti-Ang-2), 2x/week starting on d5 until symptom apparition, SC | GL261 orthotopic syngeneic | • The triple therapy significantly improved survival compared to antiangiogenic therapy alone • The triple therapy increased CD8 TIL numbers and decreased MDSCs and Tregs in the brain The antiangiogenic therapy efficacy was improved by the addition of anti-PD-1 therapy | Di Tacchio M., et al. (2019) [146] |
anti-PD-1 anti-CTLA-4 G47Δ-mIL12 (= Oncolytic herpes simplex virus expressing IL-12) | • Tumor implantation: 2 × 104 cells (005 GSCs) or 1 × 104 cells (CT-2A), striatum • anti-PD-1: 10 mg/kg, on d8, 11 and d14 (005 GSCs) or on d10, d13 and d16 (CT-2A), IP • anti-CTLA4: 5 mg/kg, on d8, d11 and d14 (005 GSCs) or on d10, d13 and d16 (CT-2A), IP • G47Δ-mIL12: 5 × 105 PFU, on d8 (005 GSCs) or on d10 (CT-2A), intratumoral | mouse 005 GSCs or CT-2A orthotopic syngeneic | • Individual ICB only minimally extended survival in 005 GBM; combination of G47Δ-mIL12 with individual ICB modestly improved efficacy • G47Δ-mIL12 decreased the % of 005 cells and Tregs and induced M1-like polarization in TAMs; these effects are further increased with the triple combination • Dual ICBs significantly increased CD8 T cells in the brain, triple combination increased CD8 T cells and decreased Tregs • Triple combination resulted in 89% or 50% long term survival (005 GSCs or CT-2A, respectively) Immune memory was established in long-term survivors | Saha D., et al. (2017) [147] |
anti-PD-1 anti-CTLA-4 anti-IL-6 anti-CD40 agonist | • Tumor implantation: 3 × 105 genetically engineered mouse tumor cells or 2 × 105 GL261 cells • anti-PD-1: 200 μg on d9, d12, d15 and d18, IP • anti-CTLA-4: 200 μg on d9, d12, d15 and d18, IP • anti-IL-6: 200 μg on d9, d12, d15 and d18, IP • anti-CD40: 100 μg on d12, IP | RCAS-genetically engineered model or GL261 orthotopic syngeneic | • Dual targeting of IL-6 and CD40 sensitized GBM to ICBs; survival was significantly improved following triple combination • Dual targeting of IL-6 and CD40 reduced tumor growth, triple combination blocked it • All treatments induced a decrease in immunosuppressive TAM activity • All treatments, except anti-CD40 alone, decreased the expression of immunosuppressive cytokines (IL-10, TGFβ) • Only triple combination induced an increase in TILs and in IFN-γ-secreting CD8 T cells | Yang F. et al. (2021) [148] |