Advertisement

Immunotherapy for brain metastases and primary brain tumors

Published:December 13, 2022DOI:https://doi.org/10.1016/j.ejca.2022.11.012

      Highlights

      • Immunotherapy of brain tumors was discussed at the V NIBIT Foundation I–O Think Tank.
      • Unlike brain metastases primary brain tumors respond poorly to ICI therapies.
      • The immune suppressive microenvironment contributes to failures of ICI therapy.
      • Neo-adjuvant studies may provide insights to boost the efficacy of ICI therapy.

      Abstract

      During the V Siena Immuno-Oncology (IO) Think Tank meeting in 2021, conditions were discussed which favor immunotherapy responses in either primary or secondary brain malignancies. Core elements of these discussions have been reinforced by important publications in 2021 and 2022. In primary brain tumors (such as glioblastoma) current immunotherapies have failed to deliver meaningful clinical benefit. By contrast, brain metastases frequently respond to current immunotherapies. The main differences between both conditions seem to be related to intrinsic factors (e.g., type of driver mutations) and more importantly extrinsic factors, such as the blood brain barrier and immune suppressive microenvironment (e.g., T cell counts, functional differences in T cells, myeloid cells). Future therapeutic interventions may therefore focus on rebalancing the immune cell population in a way which enables the host to respond to current or future immunotherapies.

      Keywords

      1. Introduction

      Immunotherapies have delivered practice-changing treatments with significant overall survival (OS) benefit [
      • Ben-Aharon O.
      • Magnezi R.
      • Leshno M.
      • Goldstein D.A.
      Association of immunotherapy with durable survival as defined by value frameworks for cancer care.
      ]. For patients with primary brain tumors, the OS benefits have been less significant as compared to patients with brain metastases [
      • Naik G.S.
      • Buchbinder E.I.
      • Cohen J.V.
      • Manos M.P.
      • Johnson A.E.W.
      • Bowling P.
      • et al.
      Long-term overall survival and predictors in anti-PD-1-naive melanoma patients with brain metastases treated with immune checkpoint inhibitors in the real-world setting: a multicohort study.
      ,
      • Reardon D.A.
      • Brandes A.A.
      • Omuro A.
      • Mulholland P.
      • Lim M.
      • Wick A.
      • et al.
      Effect of nivolumab vs bevacizumab in patients with recurrent glioblastoma: the CheckMate 143 phase 3 randomized clinical trial.
      ]. The V Siena IO Think Tank meeting held (Siena, Italy) on 7–8 October 2021, dedicated a session to immunotherapy in brain malignancies of adults. During this session, underlying mechanisms of resistance to current immunotherapies were discussed and future strategies proposed to improve treatments of brain malignancies. The following article summarizes the findings from the session and as with previous Think Tank meetings identified future clinical development initiatives [
      • Maio M.
      • Lahn M.
      • Di Giacomo A.M.
      • Covre A.
      • Calabro L.
      • Ibrahim R.
      • et al.
      A vision of immuno-oncology: the Siena think tank of the Italian network for tumor biotherapy (NIBIT) foundation.
      ,
      • Maio M.
      • Coukos G.
      • Ferrone S.
      • Fox B.A.
      • Fridman W.H.
      • Garcia P.L.
      • et al.
      Addressing current challenges and future directions in immuno-oncology: expert perspectives from the 2017 NIBIT Foundation Think Tank, Siena, Italy.
      ].
      Secondary malignancies of the brain in adults are associated with a wide range of OS, which are mostly determined by the histology of the primary tumor. Because OS in patients with brain metastases varies significantly, diagnosis-specific prognostic tools are being developed, such as the diagnosis-specific Graded Prognostic Assessment (GPA) [
      • Sperduto P.W.
      • Mesko S.
      • Li J.
      • Cagney D.
      • Aizer A.
      • Lin N.U.
      • et al.
      Survival in patients with brain metastases: summary report on the updated diagnosis-specific graded prognostic assessment and definition of the eligibility quotient.
      ]. In a patient cohort of 6984 patients with brain metastases OS ranged as follows: 7–47 months (non-small cell lung cancer), 3–36 months (breast cancer), 5–34 months (melanoma), 3–17 months (gastrointestinal cancer) and 4–35 months (renal cell carcinoma) [
      • Sperduto P.W.
      • Mesko S.
      • Li J.
      • Cagney D.
      • Aizer A.
      • Lin N.U.
      • et al.
      Survival in patients with brain metastases: summary report on the updated diagnosis-specific graded prognostic assessment and definition of the eligibility quotient.
      ] (Table 1). Recent treatments with immunotherapy and targeted therapy have improved OS in the aforementioned tumor types, including in patients who developed brain metastases.
      Table 1Main Differences between glioblastoma and brain metastases.
      Brain Malignancies
      GlioblastomaBrain metastases (NSCLC, melanoma, RCC, breast cancer and GI cancer)
      Overall SurvivalApprox. 12 months3–47 months
      Immune Cell Infiltration
       CD3+ Tumor Infiltrating

      Lymphocyte (TIL)
      LowHigh
       Myeloid CellsHighLow
      Absence of Checkpoints in Tumor Tissue
       PD-L1LowHigh
       LAG-3LowHigh
      Systemic Immune Markers Inflammation
       sPD-L1HighLow
      In contrast to patients with brain metastases the OS has not significantly improved in the past decade for patients with primary brain tumors. Depending on the type of molecular and pathological stage, the OS may range between 12 and 23 months (e.g., glioblastoma, isocitrate dehydrogenase (IDH)-wildtype) and 6–15 years (e.g., lower-grade glioma, IDH-mutated) [
      • Lapointe S.
      • Perry A.
      • Butowski N.A.
      Primary brain tumours in adults.
      ]. Based on the growing understanding of molecular pathways, a new classification for primary brain tumors was introduced [
      • Berger T.R.
      • Wen P.Y.
      • Lang-Orsini M.
      • Chukwueke U.N.
      World Health organization 2021 classification of central nervous system tumors and implications for therapy for adult-type gliomas: a review.
      ,
      • Louis D.N.
      • Perry A.
      • Wesseling P.
      • Brat D.J.
      • Cree I.A.
      • Figarella-Branger D.
      • et al.
      The 2021 WHO classification of tumors of the central nervous system: a summary.
      ]. While this updated World Health Organization (WHO) classification from 2021 introduced greater clarity in defining biologically congruous tumor entities and helped estimating prognosis more accurately, the OS has not changed for most brain tumors. While immunotherapies have not shown a convincing clinical benefit in patients with primary brain malignancies, they did have significant clinical benefit in patients with brain metastases.

      1.1 The immune-pathologic landscape in primary and secondary brain tumors

      Asymptomatic patients with non-small cell lung cancer (NSCLC) and melanoma brain metastases showed intracranial responses to immune checkpoint inhibitors in phase II clinical trials [
      • Goldberg S.B.
      • Gettinger S.N.
      • Mahajan A.
      • Chiang A.C.
      • Herbst R.S.
      • Sznol M.
      • et al.
      Pembrolizumab for patients with melanoma or non-small-cell lung cancer and untreated brain metastases: early analysis of a non-randomised, open-label, phase 2 trial.
      ,
      • Tawbi H.A.
      • Forsyth P.A.
      • Algazi A.
      • Hamid O.
      • Hodi F.S.
      • Moschos S.J.
      • et al.
      Combined nivolumab and ipilimumab in melanoma metastatic to the brain.
      ]. The response was observed regardless of the underlying tumor histology and occurred within the first 2 months of treatment. For patients with metastatic melanoma, intracranial complete response occurred in patients who received the combination of nivolumab and ipilimumab (26%; 24/94) [
      • Tawbi H.A.
      • Forsyth P.A.
      • Algazi A.
      • Hamid O.
      • Hodi F.S.
      • Moschos S.J.
      • et al.
      Combined nivolumab and ipilimumab in melanoma metastatic to the brain.
      ]. By contrast, patients with recurrent glioblastoma and receiving nivolumab showed few overall or durable responses and had a comparable OS to treatment with bevacizumab [
      • Reardon D.A.
      • Brandes A.A.
      • Omuro A.
      • Mulholland P.
      • Lim M.
      • Wick A.
      • et al.
      Effect of nivolumab vs bevacizumab in patients with recurrent glioblastoma: the CheckMate 143 phase 3 randomized clinical trial.
      ]. This lack of clinical benefit was also observed in patients with glioblastoma and receiving immune checkpoint inhibitors as 1st line treatment for both O6-methylguanine methyltransferase (MGMT) promoter methylated and unmethylated glioblastoma [
      • Lim M.
      • Weller M.
      • Idbaih A.
      • Steinbach J.
      • Finocchiaro G.
      • Raval R.R.
      • et al.
      Phase 3 trial of chemoradiotherapy with temozolomide plus nivolumab or placebo for newly diagnosed glioblastoma with methylated MGMT promoter.
      ,
      • Omuro A.
      • Brandes A.A.
      • Carpentier A.F.
      • Idbaih A.
      • Reardon D.A.
      • Cloughesy T.
      • et al.
      Radiotherapy combined with nivolumab or temozolomide for newly diagnosed glioblastoma with unmethylated MGMT promoter: an international randomized phase 3 trial.
      ]. To provide a framework on how to best explain this difference of responses to current immunotherapies, the “cancer immunogram” can be used to determine whether both types of malignancies are characterized by fundamental differences in their immune landscape [
      • Blank C.U.
      • Haanen J.B.
      • Ribas A.
      • Schumacher T.N.
      CANCER IMMUNOLOGY. The "cancer immunogram.
      ] (Table 1).
      First, brain metastases and primary brain malignancies differ in their number of tumor-infiltrating lymphocytes [
      • Berghoff A.S.
      • Fuchs E.
      • Ricken G.
      • Mlecnik B.
      • Bindea G.
      • Spanberger T.
      • et al.
      Density of tumor-infiltrating lymphocytes correlates with extent of brain edema and overall survival time in patients with brain metastases.
      ,
      • Berghoff A.S.
      • Kiesel B.
      • Widhalm G.
      • Wilhelm D.
      • Rajky O.
      • Kurscheid S.
      • et al.
      Correlation of immune phenotype with IDH mutation in diffuse glioma.
      ,
      • Berghoff A.S.
      • Ricken G.
      • Widhalm G.
      • Rajky O.
      • Dieckmann K.
      • Birner P.
      • et al.
      Tumour-infiltrating lymphocytes and expression of programmed death ligand 1 (PD-L1) in melanoma brain metastases.
      ,
      • Berghoff A.S.
      • Ricken G.
      • Wilhelm D.
      • Rajky O.
      • Widhalm G.
      • Dieckmann K.
      • et al.
      Tumor infiltrating lymphocytes and PD-L1 expression in brain metastases of small cell lung cancer (SCLC).
      ,
      • Berghoff A.S.
      • Schur S.
      • Fureder L.M.
      • Gatterbauer B.
      • Dieckmann K.
      • Widhalm G.
      • et al.
      Descriptive statistical analysis of a real life cohort of 2419 patients with brain metastases of solid cancers.
      ]. Brain metastases have nearly 50% more CD3+ lymphocyte infiltrations than either glioblastoma or low-grade glioma [
      • Berghoff A.S.
      • Kiesel B.
      • Widhalm G.
      • Wilhelm D.
      • Rajky O.
      • Kurscheid S.
      • et al.
      Correlation of immune phenotype with IDH mutation in diffuse glioma.
      ]. This presence of CD3+ immune cells appeared to be associated with the IDH mutational status [
      • Berghoff A.S.
      • Kiesel B.
      • Widhalm G.
      • Wilhelm D.
      • Rajky O.
      • Kurscheid S.
      • et al.
      Correlation of immune phenotype with IDH mutation in diffuse glioma.
      ,
      • Klemm F.
      • Maas R.R.
      • Bowman R.L.
      • Kornete M.
      • Soukup K.
      • Nassiri S.
      • et al.
      Interrogation of the microenvironmental landscape in brain tumors reveals disease-specific alterations of immune cells.
      ,
      • Friebel E.
      • Kapolou K.
      • Unger S.
      • Nunez N.G.
      • Utz S.
      • Rushing E.J.
      • et al.
      Single-cell mapping of human brain cancer reveals tumor-specific instruction of tissue-invading leukocytes.
      ]. The mechanistical underpinnings of IDH mutation determining the cellular composition of the inflammatory microenvironment are still under investigation. One proposed mechanism is the presence of the oncometabolite 2-hydroxyglutarate which is produced by mutant IDH and inhibits T cell activation [
      • Bunse L.
      • Pusch S.
      • Bunse T.
      • Sahm F.
      • Sanghvi K.
      • Friedrich M.
      • et al.
      Suppression of antitumor T cell immunity by the oncometabolite (R)-2-hydroxyglutarate.
      ]. The inflammatory microenvironment of gliomas is mainly characterized by microglial cells, and IDH-mutant tumors harbour a considerable fraction of monocyte-derived macrophages. Furthermore, distinct epigenetic subgroups are known to impact lymphocyte infiltration [
      • Dejaegher J.
      • Solie L.
      • Hunin Z.
      • Sciot R.
      • Capper D.
      • Siewert C.
      • et al.
      DNA methylation based glioblastoma subclassification is related to tumoral T-cell infiltration and patient survival.
      ].
      Second, the absence of checkpoints may explain why current immunotherapies have failed in glioma. For example, programmed death-ligand 1 (PD-L1) expression is higher in brain metastases (>50% of cases) compared to glioblastoma, where membranous PD-L1 expression is seen in about 37% of samples [
      • Berghoff A.S.
      • Ricken G.
      • Widhalm G.
      • Rajky O.
      • Dieckmann K.
      • Birner P.
      • et al.
      Tumour-infiltrating lymphocytes and expression of programmed death ligand 1 (PD-L1) in melanoma brain metastases.
      ,
      • Berghoff A.S.
      • Ricken G.
      • Wilhelm D.
      • Rajky O.
      • Widhalm G.
      • Dieckmann K.
      • et al.
      Tumor infiltrating lymphocytes and PD-L1 expression in brain metastases of small cell lung cancer (SCLC).
      ]. Lower-grade gliomas have even less PD-L1 expression within the tumor microenvironment [
      • Berghoff A.S.
      • Kiesel B.
      • Widhalm G.
      • Wilhelm D.
      • Rajky O.
      • Kurscheid S.
      • et al.
      Correlation of immune phenotype with IDH mutation in diffuse glioma.
      ,
      • Berghoff A.S.
      • Kiesel B.
      • Widhalm G.
      • Rajky O.
      • Ricken G.
      • Wohrer A.
      • et al.
      Programmed death ligand 1 expression and tumor-infiltrating lymphocytes in glioblastoma.
      ].
      Alternative checkpoints such as lymphocyte-activation gene 3 (LAG-3) are nearly absent in patients with glioma except for a small subset of patients with IDH wildtype tumors [
      • Mair M.J.
      • Kiesel B.
      • Feldmann K.
      • Widhalm G.
      • Dieckmann K.
      • Wohrer A.
      • et al.
      LAG-3 expression in the inflammatory microenvironment of glioma.
      ]. In peripheral blood, soluble PD-L1 (sPD-L1) is higher in glioma than in patients with brain metastases, and sPD-L1 levels were observed to be associated with survival [
      • Mair M.J.
      • Pajenda S.
      • Ilhan-Mutlu A.
      • Steindl A.
      • Kiesel B.
      • Widhalm G.
      • et al.
      Soluble PD-L1 is associated with local and systemic inflammation markers in primary and secondary brain tumours.
      ]. Consistent with the concept of an immunosuppressive environment, a higher neutrophil-to-lymphocyte ratio (NLR) was identified in patients with glioblastoma [
      • Han S.
      • Liu Y.
      • Li Q.
      • Li Z.
      • Hou H.
      • Wu A.
      Pre-treatment neutrophil-to-lymphocyte ratio is associated with neutrophil and T-cell infiltration and predicts clinical outcome in patients with glioblastoma.
      ].
      Third, tumor mutational burden (TMB) is generally lower in patients with glioblastoma than in patients with melanoma or NSCLC [
      • Alexandrov L.B.
      • Nik-Zainal S.
      • Wedge D.C.
      • Aparicio S.A.
      • Behjati S.
      • Biankin A.V.
      • et al.
      Signatures of mutational processes in human cancer.
      ]. While there is a correlation of anti-tumor responses and TMB in patients with melanoma and NSCLC receiving immune checkpoint inhibitors, there is no such correlation of responses and TMB in glioblastoma patients [
      • Vormehr M.
      • Diken M.
      • Boegel S.
      • Kreiter S.
      • Tureci O.
      • Sahin U.
      Mutanome directed cancer immunotherapy.
      ].

      1.2 Gene signatures in glioblastoma multiforme (GBM)

      Research has focused on identifying unique driver mutations for which specific inhibitors can be developed and consequently the development of such drugs could result in improved OS [
      • He Q.
      • Li Q.
      • Lv F.
      • Kaitin K.I.
      • Shao L.
      A survey of survival outcomes for targeted cancer drugs approved by the US food and drug administration.
      ]. Hence, discovery of molecular pathways in gliomagenesis promises the identification of new targets in either primary or secondary brain malignancies [
      • Appin C.L.
      • Brat D.J.
      Molecular pathways in gliomagenesis and their relevance to neuropathologic diagnosis.
      ]. Among glioblastoma with IDH wildtype, three different subtypes have been identified and each subtype was evaluated for their presence of specific immune cells [
      • Wang Q.
      • Hu B.
      • Hu X.
      • Kim H.
      • Squatrito M.
      • Scarpace L.
      • et al.
      Tumor evolution of glioma-intrinsic gene expression subtypes associates with immunological changes in the microenvironment.
      ]. To further explore the gene expression based subsets and their relationship to immune signature, single-cell studies have uncovered four main cellular states of glioblastoma that are influenced by the microenvironment compositions and specific genetic events: (a) neural-progenitor-like cells enriched with cyclin-dependent kinase (CDK) amplifications; (b) oligodendrocyte-progenitor-like cell enriched with platelet-derived growth factor receptor alpha (PDGFRA) amplification, (c) astrocyte-like cells showing higher frequency of epidermal growth factor receptor (EGFR) amplification and (d) mesenchymal-like cells characterized by NF1 mutations [
      • Neftel C.
      • Laffy J.
      • Filbin M.G.
      • Hara T.
      • Shore M.E.
      • Rahme G.J.
      • et al.
      An integrative model of cellular states, plasticity, and genetics for glioblastoma.
      ]. When combining all molecular information, four pathway-based types can be defined: (1) glycolytic/plurimetabolic; (2) neuronal; (3) mitochondrial; (4) proliferative/progenitor [
      • Garofano L.
      • Migliozzi S.
      • Oh Y.T.
      • D'Angelo F.
      • Najac R.D.
      • Ko A.
      • et al.
      Pathway-based classification of glioblastoma uncovers a mitochondrial subtype with therapeutic vulnerabilities.
      ]. This approach has identified new subtype-specific targets pathways, such as Mitochondrial Oxidative Phosphorylation [
      • Garofano L.
      • Migliozzi S.
      • Oh Y.T.
      • D'Angelo F.
      • Najac R.D.
      • Ko A.
      • et al.
      Pathway-based classification of glioblastoma uncovers a mitochondrial subtype with therapeutic vulnerabilities.
      ].
      A second application for molecular defined pathway analyses is the evaluation of temporal changes of glioblastoma. This evaluation can determine whether a target remains stable over time and may help to uncover mechanisms of resistance [
      • Varn F.S.
      • Johnson K.C.
      • Martinek J.
      • Huse J.T.
      • Nasrallah M.P.
      • Wesseling P.
      • et al.
      Glioma progression is shaped by genetic evolution and microenvironment interactions.
      ]. This approach shows a high diversity of change over time within the different subgroups, including metabolic changes.
      Third, gene expression evaluation can also be used to characterize the immune microenvironment [
      • Sa J.K.
      • Chang N.
      • Lee H.W.
      • Cho H.J.
      • Ceccarelli M.
      • Cerulo L.
      • et al.
      Transcriptional regulatory networks of tumor-associated macrophages that drive malignancy in mesenchymal glioblastoma.
      ]. Transcriptional regulatory network found the genes PPARG, BATF and SPI1 as promoters for attracting tissue-associated macrophages (TAM) [
      • Mall R.
      • Cerulo L.
      • Garofano L.
      • Frattini V.
      • Kunji K.
      • Bensmail H.
      • et al.
      RGBM: regularized gradient boosting machines for identification of the transcriptional regulators of discrete glioma subtypes.
      ]. Associated with these TAM is the expression of macrophage receptor with collagenous structure (MARCO) to drive mesenchymal transformation [
      • Sa J.K.
      • Chang N.
      • Lee H.W.
      • Cho H.J.
      • Ceccarelli M.
      • Cerulo L.
      • et al.
      Transcriptional regulatory networks of tumor-associated macrophages that drive malignancy in mesenchymal glioblastoma.
      ,
      • Caruso F.P.
      • Garofano L.
      • D'Angelo F.
      • Yu K.
      • Tang F.
      • Yuan J.
      • et al.
      A map of tumor-host interactions in glioma at single-cell resolution.
      ,
      • Mathewson N.D.
      • Ashenberg O.
      • Tirosh I.
      • Gritsch S.
      • Perez E.M.
      • Marx S.
      • et al.
      Inhibitory CD161 receptor identified in glioma-infiltrating T cells by single-cell analysis.
      ].

      1.3 The immune microenvironment of melanoma brain metastases: implications for immunotherapy

      Based on the early responses to immunotherapies in patients with melanoma and NSCLC [
      • Tawbi H.A.
      • Forsyth P.A.
      • Algazi A.
      • Hamid O.
      • Hodi F.S.
      • Moschos S.J.
      • et al.
      Combined nivolumab and ipilimumab in melanoma metastatic to the brain.
      ,
      • Long G.V.
      • Atkinson V.
      • Lo S.
      • Sandhu S.
      • Guminski A.D.
      • Brown M.P.
      • et al.
      Combination nivolumab and ipilimumab or nivolumab alone in melanoma brain metastases: a multicentre randomised phase 2 study.
      ,
      • Tawbi H.A.
      • Forsyth P.A.
      • Hodi F.S.
      • Algazi A.P.
      • Hamid O.
      • Lao C.D.
      • et al.
      Long-term outcomes of patients with active melanoma brain metastases treated with combination nivolumab plus ipilimumab (CheckMate 204): final results of an open-label, multicentre, phase 2 study.
      ,
      • Kim P.H.
      • Suh C.H.
      • Kim H.S.
      • Kim K.W.
      • Kim D.Y.
      • Aizer A.A.
      • et al.
      Immune checkpoint inhibitor therapy may increase the incidence of treatment-related necrosis after stereotactic radiosurgery for brain metastases: a systematic review and meta-analysis.
      ], intra-tumoral CD8, stromal PD-L1 expression and density of immune cells were associated with responses in patients with brain metastases [
      • Kluger H.M.
      • Chiang V.
      • Mahajan A.
      • Zito C.R.
      • Sznol M.
      • Tran T.
      • et al.
      Long-Term survival of patients with melanoma with active brain metastases treated with pembrolizumab on a phase II trial.
      ,
      • Tumeh P.C.
      • Harview C.L.
      • Yearley J.H.
      • Shintaku I.P.
      • Taylor E.J.
      • Robert L.
      • et al.
      PD-1 blockade induces responses by inhibiting adaptive immune resistance.
      ]. The localization of the immune cells varies between the tumor types and their brain metastases [
      • Berghoff A.S.
      • Ricken G.
      • Widhalm G.
      • Rajky O.
      • Dieckmann K.
      • Birner P.
      • et al.
      Tumour-infiltrating lymphocytes and expression of programmed death ligand 1 (PD-L1) in melanoma brain metastases.
      ,
      • Harter P.N.
      • Bernatz S.
      • Scholz A.
      • Zeiner P.S.
      • Zinke J.
      • Kiyose M.
      • et al.
      Distribution and prognostic relevance of tumor-infiltrating lymphocytes (TILs) and PD-1/PD-L1 immune checkpoints in human brain metastases.
      ]. The larger the size of the metastatic lesion, the more likely a decrease of immune cells numbers will be observed, which in turn may have an impact on response to immunotherapy [
      • Berghoff A.S.
      • Ricken G.
      • Widhalm G.
      • Rajky O.
      • Dieckmann K.
      • Birner P.
      • et al.
      Tumour-infiltrating lymphocytes and expression of programmed death ligand 1 (PD-L1) in melanoma brain metastases.
      ,
      • Harter P.N.
      • Bernatz S.
      • Scholz A.
      • Zeiner P.S.
      • Zinke J.
      • Kiyose M.
      • et al.
      Distribution and prognostic relevance of tumor-infiltrating lymphocytes (TILs) and PD-1/PD-L1 immune checkpoints in human brain metastases.
      ]. When comparing brain metastases to other metastatic sites, the composition of myeloid cells differs by anatomical location [
      • Smalley I.
      • Chen Z.
      • Phadke M.
      • Li J.
      • Yu X.
      • Wyatt C.
      • et al.
      Single-cell characterization of the immune microenvironment of melanoma brain and leptomeningeal metastases.
      ]. Prior therapies, such as radiotherapy, can influence the microenvironment creating either an immune-enriched or -depleted environment [
      • Fischer G.M.
      • Jalali A.
      • Kircher D.A.
      • Lee W.C.
      • McQuade J.L.
      • Haydu L.E.
      • et al.
      Molecular profiling reveals unique immune and metabolic features of melanoma brain metastases.
      ]. Drivers of this immune composition are genes associated with metabolic features, such as oxidative phosphorylation (OXPHOS) [
      • Egelston C.A.
      • Margolin K.
      Metabolic checkpoint of immune cells in melanoma brain metastases.
      ].

      1.4 Success, pitfalls, and failures: a clinical perspective

      Combination studies were designed to overcome resistance in patients with brain metastases [
      • Di Giacomo A.M.
      • Ascierto P.A.
      • Pilla L.
      • Santinami M.
      • Ferrucci P.F.
      • Giannarelli D.
      • et al.
      Ipilimumab and fotemustine in patients with advanced melanoma (NIBIT-M1): an open-label, single-arm phase 2 trial.
      ,
      • Di Giacomo A.M.
      • Chiarion-Sileni V.
      • Del Vecchio M.
      • Ferrucci P.F.
      • Guida M.
      • Quaglino P.
      • et al.
      Primary analysis and 4-year follow-up of the phase III NIBIT-M2 trial in melanoma patients with brain metastases.
      ]. In considering treatment outcomes, disease stabilization of intracranial metastases may be an important driver for OS [
      • Di Giacomo A.M.
      • Chiarion-Sileni V.
      • Del Vecchio M.
      • Ferrucci P.F.
      • Guida M.
      • Quaglino P.
      • et al.
      Primary analysis and 4-year follow-up of the phase III NIBIT-M2 trial in melanoma patients with brain metastases.
      ]. The drug fotemustine is a commonly used drug in glioblastoma and thus the combination with an immune checkpoint inhibitor was expected to have a disease stabilizing effect.
      Single-agent or combination immune checkpoint inhibitors have been investigated in several studies but showed no OS benefit [
      • Reardon D.A.
      • Brandes A.A.
      • Omuro A.
      • Mulholland P.
      • Lim M.
      • Wick A.
      • et al.
      Effect of nivolumab vs bevacizumab in patients with recurrent glioblastoma: the CheckMate 143 phase 3 randomized clinical trial.
      ,
      • Lim M.
      • Weller M.
      • Idbaih A.
      • Steinbach J.
      • Finocchiaro G.
      • Raval R.R.
      • et al.
      Phase 3 trial of chemoradiotherapy with temozolomide plus nivolumab or placebo for newly diagnosed glioblastoma with methylated MGMT promoter.
      ,
      • Omuro A.
      • Brandes A.A.
      • Carpentier A.F.
      • Idbaih A.
      • Reardon D.A.
      • Cloughesy T.
      • et al.
      Radiotherapy combined with nivolumab or temozolomide for newly diagnosed glioblastoma with unmethylated MGMT promoter: an international randomized phase 3 trial.
      ,
      • Reardon D.A.
      • Kim T.M.
      • Frenel J.S.
      • Simonelli M.
      • Lopez J.
      • Subramaniam D.S.
      • et al.
      Treatment with pembrolizumab in programmed death ligand 1-positive recurrent glioblastoma: results from the multicohort phase 1 KEYNOTE-028 trial.
      ]. Survival benefit was observed if the programmed death protein 1 (PD-1) inhibitor pembrolizumab was given in a neo-adjuvant setting [
      • Cloughesy T.F.
      • Mochizuki A.Y.
      • Orpilla J.R.
      • Hugo W.
      • Lee A.H.
      • Davidson T.B.
      • et al.
      Neoadjuvant anti-PD-1 immunotherapy promotes a survival benefit with intratumoral and systemic immune responses in recurrent glioblastoma.
      ]. Also, intracerebral application of either cytotoxic T-lymphocyte antigen 4 (CTLA-4) or PD-1 inhibitor could be an interesting approach, as early-phase data showed promising results [
      • Duerinck J.
      • Schwarze J.K.
      • Awada G.
      • Tijtgat J.
      • Vaeyens F.
      • Bertels C.
      • et al.
      Intracerebral administration of CTLA-4 and PD-1 immune checkpoint blocking monoclonal antibodies in patients with recurrent glioblastoma: a phase I clinical trial.
      ].

      1.5 Immunotherapy for primary brain tumors: future therapeutic strategies

      New treatment concepts have been explored in recent years, including antibody–drug conjugates, immunocytokines, chimeric antigen receptor T (CAR-T) cells, novel immune checkpoint inhibitors and vaccines (Table 2). With unique expression of EGFR in glioblastoma, antibody-conjugates targeting EGFR domain II (for example, ABT-414) exploit the overexpression of this protein to deliver chemotherapy locally. Whereas the phase 2 study appeared to deliver the expected improved OS in recurrent glioblastoma, the phase 3 trial in newly diagnosed glioblastoma failed to show a survival benefit [
      • Weller M.
      • Butowski N.
      • Tran D.D.
      • Recht L.D.
      • Lim M.
      • Hirte H.
      • et al.
      Rindopepimut with temozolomide for patients with newly diagnosed, EGFRvIII-expressing glioblastoma (ACT IV): a randomised, double-blind, international phase 3 trial.
      ].
      Table 2Examples of novel therapeutic approaches in malignant glioma.
      General ApproachExample
      Combination with Chemotherapy and Immune checkpoint inhibitorFotemustine + immune checkpoint inhibitor
      Intracranial DeliveryImmune Checkpoint Inhibitors, oncolytic virus
      Antibody–Drug ConjugateTargeting EGFR domain II (ABT-414)
      ImmunocytokinesOnfekafusp alfa
      CAR-T cellsEGFRvIII, IL13R
      Reduction of Immune Suppression
      • TGF-β/ALK5 small molecule inhibitors
      • CTLA-4 inhibitors
      Immune Checkpoint InhibitorsNovel Inhibitors, such as LAG-3
      Vaccine
      • Peptide-based vaccines
      • Dendritic cell-based
      Another intervention is based on delivering immunocytokines which are intended to convert “immune-deserted” (=“cold”) tumors into “immune-competent” (=“hot”). Examples of such immunocytokines are interleukin (IL)-2, IL-12 and Tumor Necrosis Factor alpha (TNF-α) linked to an antibody to the extra-domain B (EDB) of fibronectin [
      • Weiss T.
      • Puca E.
      • Silginer M.
      • Hemmerle T.
      • Pazahr S.
      • Bink A.
      • et al.
      Immunocytokines are a promising immunotherapeutic approach against glioblastoma.
      ]. The “Gliomoon Trial” is such an example for evaluating novel immunocytokines in glioblastoma treatment. Patients were treated with L19-TNF (onfekafusp alfa) if they had an IDH wildtype and grade 3 or 4 gliomas. A third approach consists in delivering modified T cells, such as CAR-T cells. Utilizing the overexpression of EGFR, a first-generation CAR-T cell approach was constructed targeting the EGFR variant III [
      • O'Rourke D.M.
      • Nasrallah M.P.
      • Desai A.
      • Melenhorst J.J.
      • Mansfield K.
      • Morrissette J.J.D.
      • et al.
      A single dose of peripherally infused EGFRvIII-directed CAR T cells mediates antigen loss and induces adaptive resistance in patients with recurrent glioblastoma.
      ]. This study in recurrent glioblastoma demonstrated that such chimeric T cells can be generated and are safe. The fourth approach targets reversing immune suppression by targeting specific signalling pathways, such as transforming growth factor beta (TGF-β)/activin receptor-like kinase 5 (ALK5), CTLA-4, tryptophane, signal transducer and activator of transcription 3 (STAT3) [
      • Weller M.
      • Roth P.
      • Preusser M.
      • Wick W.
      • Reardon D.A.
      • Platten M.
      • et al.
      Vaccine-based immunotherapeutic approaches to gliomas and beyond.
      ]. The role of TGF-β/ALK5 in glioblastoma has been recognized for many years but so far trials with TGF-β/ALK5 inhibitors have not shown clinical benefit compared to existing standard therapy [
      • Bogdahn U.
      • Hau P.
      • Stockhammer G.
      • Venkataramana N.K.
      • Mahapatra A.K.
      • Suri A.
      • et al.
      Targeted therapy for high-grade glioma with the TGF-beta2 inhibitor tradersen: results of a randomized and controlled phase IIb study.
      ,
      • Brandes A.A.
      • Carpentier A.F.
      • Kesari S.
      • Sepulveda-Sanchez J.M.
      • Wheeler H.R.
      • Chinot O.
      • et al.
      A Phase II randomized study of galunisertib monotherapy or galunisertib plus lomustine compared with lomustine monotherapy in patients with recurrent glioblastoma.
      ]. Since the prior TGF-β/ALK5 inhibitors were not studied in specifically selected patients or with drugs that may achieve optimal therapeutic levels, novel inhibitors are currently being developed [
      • Papachristodoulou A.
      • Silginer M.
      • Weller M.
      • Schneider H.
      • Hasenbach K.
      • Janicot M.
      • et al.
      Therapeutic targeting of TGFbeta ligands in glioblastoma using novel antisense oligonucleotides reduces the growth of experimental gliomas.
      ,
      • Yap T.A.
      • Vieito M.
      • Baldini C.
      • Sepulveda-Sanchez J.M.
      • Kondo S.
      • Simonelli M.
      • et al.
      First-in-Human phase I study of a next-generation, oral, TGFbeta receptor 1 inhibitor, LY3200882, in patients with advanced cancer.
      ]. A fifth option is to improve the current therapy with immune therapy. While immune checkpoint inhibitors have not shown OS benefits, subgroup evaluations showed that patients with MGMT promoter methylation in their tumor and no baseline corticosteroid treatment had higher OS than patients treated with bevacizumab [
      • Reardon D.A.
      • Brandes A.A.
      • Omuro A.
      • Mulholland P.
      • Lim M.
      • Wick A.
      • et al.
      Effect of nivolumab vs bevacizumab in patients with recurrent glioblastoma: the CheckMate 143 phase 3 randomized clinical trial.
      ]. A sixth approach consists in utilizing vaccines. There are vaccines directed against the EGFR variant III (e.g., rindopepimut), dendritic cell-based vaccines (e.g., ICT-107, DCVax), tumor-derived peptides (e.g., HSPPC-96) and personalized multipeptide vaccines (e.g., GAPVAC) [
      • Medikonda R.
      • Dunn G.
      • Rahman M.
      • Fecci P.
      • Lim M.
      A review of glioblastoma immunotherapy.
      ]. However, the phase 3 study with rindopepimut added to standard of care in the newly diagnosed setting did not show a clinical benefit compared to the control arm in patients after initial resection of their first tumor [
      • Weller M.
      • Butowski N.
      • Tran D.D.
      • Recht L.D.
      • Lim M.
      • Hirte H.
      • et al.
      Rindopepimut with temozolomide for patients with newly diagnosed, EGFRvIII-expressing glioblastoma (ACT IV): a randomised, double-blind, international phase 3 trial.
      ]. While these trials failed, this remains an area where combination immunotherapy needs to be evaluated. A recent report identified the defect in immunity to brain tumors is the absence of antigen presenting cells that can prime and boost anti-cancer immunity [
      • Simonds E.F.
      • Lu E.D.
      • Badillo O.
      • Karimi S.
      • Liu E.V.
      • Tamaki W.
      • Rancan C.
      • Downey K.M.
      • Stultz J.
      • Sinha M.
      • McHenry L.K.
      • Nasholm N.M.
      • Chuntova P.
      • Sundstr√∂m A.
      • Genoud V.
      • Shahani S.A.
      • Wang L.D.
      • Brown C.E.
      • Walker P.R.
      • Swartling F.J.
      • Fong L.
      • Okada H.
      • et al.
      Deep immune profiling reveals targetable mechanisms of immune evasion in immune checkpoint inhibitor-refractory glioblastoma.
      ]. J Immunother Cancer 2021; 9:e002181. PMID 34083417).
      . This is not a problem for melanoma and other brain metastases, as priming and expansion continues in the periphery. Advanced combination immunotherapy trials that take into consideration priming, application of immune agonists and checkpoint blockers, while also addressing suppressive mechanisms represent opportunities to design aggressive therapeutic options for patients with this disease.

      2. Discussion

      The Think Tank members identified the following main areas for future research (Table 3):
      Table 3Future areas of research.
      Influence Factors Limiting Current ImmunotherapiesDiagnostics and Biomarkers
      Genetic Predispositions
      • Hypermutation of genes affecting immune responses
      • Molecular characterization of genetics-epigenetics-transcriptomics in relationship to anti-tumor responses
      Immune Conditions
      • Antigen release and presentation
      • Subset analysis of immune cell activation and recruitment
      • Rebalance of immune subsets (e.g., increase effector T cells while reducing immune suppressive macrophages)
      Therapeutic Interventions
      • Assess current therapies on their immune suppressive effect (e.g., radiation, chemotherapy)
      • Select therapies with immune enhancing effect
      • Develop detection tools to overcome intra-tumor heterogeneity to identify novel therapeutic targets
      • Use single-cell evaluation to identify therapeutic vulnerabilities
      • Assess time-dependent changes in tumor cell population to uncover unknown factors leading to resistance to immunotherapy
      First, factors that limit current immunotherapies and diagnostic tools to identify such factors. Among the factors limiting current immunotherapies, three main contributing conditions may need to be assessed further:
      • (a)
        Genetic predisposition: Immunotherapies may have a unique activity in patients with biallelic mismatch repair deficiency (bMMRD) [
        • Bouffet E.
        • Larouche V.
        • Campbell B.B.
        • Merico D.
        • de Borja R.
        • Aronson M.
        • et al.
        Immune checkpoint inhibition for hypermutant glioblastoma multiforme resulting from germline biallelic mismatch repair deficiency.
        ]. Furthermore, hypermutations appear to influence responses [
        • Touat M.
        • Li Y.Y.
        • Boynton A.N.
        • Spurr L.F.
        • Iorgulescu J.B.
        • Bohrson C.L.
        • et al.
        Mechanisms and therapeutic implications of hypermutation in gliomas.
        ]. The consequences of such mutations on influencing immune response are not known and hence a need for molecular characterization of the genetic/epigenetic/transcriptomic landscape of primary and secondary brain malignancies is desirable.
      • (b)
        Immune cell landscape: Current understanding shows a fundamental difference between the immune cell landscape of primary and secondary malignancies, which may affect anti-tumor responses [
        • Preusser M.
        • Lim M.
        • Hafler D.A.
        • Reardon D.A.
        • Sampson J.H.
        Prospects of immune checkpoint modulators in the treatment of glioblastoma.
        ]. This difference may result in differences of antigen presentation, immune cell priming, activation, recruitment and consequently, causing an imbalance of the effector T cell population. In pediatric patients, the intra-tumor injection of the oncolytic virus-based therapy DNX-2401 altered the tumor immune environment and was associated with an OS of 17.8 months [
        • Gallego Perez-Larraya J.
        • Garcia-Moure M.
        • Labiano S.
        • Patino-Garcia A.
        • Dobbs J.
        • Gonzalez-Huarriz M.
        • et al.
        Oncolytic DNX-2401 virus for pediatric diffuse intrinsic pontine glioma.
        ]. Perhaps novel approaches of therapeutic vaccines may offer new avenues for treatment concepts (Table 2).
      • (c)
        Impact of current therapies on future immunotherapies: Current standard therapies may have to be adjusted depending on their impact of affecting the immune cell landscape. This may help in designing future studies with the appropriate background therapies.
      Second, research should focus on tools and diagnostic measures which may help to uncover unappreciated mechanisms of resistance. These measures should take into consideration the intra-tumor heterogeneity as this may hinder to identification of novel therapeutic targets. Single-cell studies while cumbersome may offer a more detailed look at the complex landscape of brain malignancies. All such diagnostic and biomarker studies may need to include time-course evaluations given the highly dynamic nature of brain malignancies. As in other disease settings, neo-adjuvant study designs may provide such a platform for using broad multi-omics study designs [
      • Cloughesy T.F.
      • Mochizuki A.Y.
      • Orpilla J.R.
      • Hugo W.
      • Lee A.H.
      • Davidson T.B.
      • et al.
      Neoadjuvant anti-PD-1 immunotherapy promotes a survival benefit with intratumoral and systemic immune responses in recurrent glioblastoma.
      ,
      • Lee A.H.
      • Sun L.
      • Mochizuki A.Y.
      • Reynoso J.G.
      • Orpilla J.
      • Chow F.
      • et al.
      Neoadjuvant PD-1 blockade induces T cell and cDC1 activation but fails to overcome the immunosuppressive tumor associated macrophages in recurrent glioblastoma.
      ].

      Funding

      This meeting was supported in part by the NIBIT Foundation.

      Author contributions

      All authors critically contributed to the final manuscript draft and approved the final version. Michele Maio conceived the review together with key participants. The manuscript was developed from meeting notes and subsequently updated by each contributing author.

      Conflict of interest statement

      Anna Maria Di Giacomo has served as a consultant and/or advisor to Incyte, Pierre Fabre, Glaxo Smith Kline, Bristol-Myers Squibb, Merck Sharp Dohme, and Sanofi and has received compensated educational activities from Bristol Myers Squibb, Merck Sharp Dohme, Pierre Fabre and Sanofi.
      Maximilian J. Mair has received travel support from Pierre Fabre.
      Bernard A. Fox: has served as a consultant and/or advisor to Akoya/PerkinElmer, AstraZeneca/Definiens, Boehringer Ingelheim, Bristol-Myers Squibb, CALiDi, Hooka, Incyte, Macrogenics, NeoGenomics, Pfizer, PrimeVax, Turnstone, Ultivue. BAF is a Founder and CEO of UbiVac and has shares or options in CALiDi, PrimeVax, Turnstone and UbiVac. He has received research support from: Akoya, Bristol-Myers Squibb, Macrogenics, NanoString, OncoSec, Shimadzu, Viralytics/Merck.
      Alexander M.M. Eggermont has served as a consultant and/or advisor to: Agenus, Biocad, BioInvent, Brenus, CatalYm, Clover, Ellipses, GSK, IO Biotech, IQVIA, ISA Pharmaceuticals, Merck/MSD, Nektar, Pfizer, Sairopa, Scorpion, Sellas, SkylineDx, TigaTx, Trained Immunity Therapeutics. He has Equity in IO Biotech, Sairopa, SkylineDX. Educational Activities for BMS and Merck/MSD.
      Michele Maio has served as a consultant and/or advisor to Roche, Bristol-Myers Squibb, Merck Sharp Dohme, Incyte, AstraZeneca, Amgen, Pierre Fabre, Eli Lilly, Glaxo Smith Kline, Sciclone, Sanofi, Alfasigma, and Merck Serono; and own shares in Theravance and Epigen Therapeutics, Srl.
      Ramy Ibrahim is on the board of directors of Surface Oncology and 2Seventy. He is a consultant to Georgiammune, Harpoon and Altos labs.
      Michael Weller has received research grants from Quercis and Versameb, and honoraria for lectures or advisory board participation or consulting from Bayer, Medac, Merck (EMD), Novartis, Orbus, and Philogen.
      Michael Lahn is employed by iOnctura SA and holds company stocks.
      All remaining authors declare no conflict of interest.

      Acknowledgements

      The meeting was organized with scientific support of the NIBIT Foundation. The contents and topics of the panel discussion were not influenced by the sponsors. Development of this manuscript was funded by the NIBIT Foundation. The authors thank Michael Smith for providing scientific writing support.

      References

        • Ben-Aharon O.
        • Magnezi R.
        • Leshno M.
        • Goldstein D.A.
        Association of immunotherapy with durable survival as defined by value frameworks for cancer care.
        JAMA Oncol. 2018; 4: 326-332
        • Naik G.S.
        • Buchbinder E.I.
        • Cohen J.V.
        • Manos M.P.
        • Johnson A.E.W.
        • Bowling P.
        • et al.
        Long-term overall survival and predictors in anti-PD-1-naive melanoma patients with brain metastases treated with immune checkpoint inhibitors in the real-world setting: a multicohort study.
        J Immunother. 2021; 44: 307-318
        • Reardon D.A.
        • Brandes A.A.
        • Omuro A.
        • Mulholland P.
        • Lim M.
        • Wick A.
        • et al.
        Effect of nivolumab vs bevacizumab in patients with recurrent glioblastoma: the CheckMate 143 phase 3 randomized clinical trial.
        JAMA Oncol. 2020; 6: 1003-1010
        • Maio M.
        • Lahn M.
        • Di Giacomo A.M.
        • Covre A.
        • Calabro L.
        • Ibrahim R.
        • et al.
        A vision of immuno-oncology: the Siena think tank of the Italian network for tumor biotherapy (NIBIT) foundation.
        J Exp Clin Cancer Res. 2021; 40: 240
        • Maio M.
        • Coukos G.
        • Ferrone S.
        • Fox B.A.
        • Fridman W.H.
        • Garcia P.L.
        • et al.
        Addressing current challenges and future directions in immuno-oncology: expert perspectives from the 2017 NIBIT Foundation Think Tank, Siena, Italy.
        Cancer Immunol Immunother. 2019; 68: 1-9
        • Sperduto P.W.
        • Mesko S.
        • Li J.
        • Cagney D.
        • Aizer A.
        • Lin N.U.
        • et al.
        Survival in patients with brain metastases: summary report on the updated diagnosis-specific graded prognostic assessment and definition of the eligibility quotient.
        J Clin Oncol. 2020; 38: 3773-3784
        • Lapointe S.
        • Perry A.
        • Butowski N.A.
        Primary brain tumours in adults.
        Lancet. 2018; 392: 432-446
        • Berger T.R.
        • Wen P.Y.
        • Lang-Orsini M.
        • Chukwueke U.N.
        World Health organization 2021 classification of central nervous system tumors and implications for therapy for adult-type gliomas: a review.
        JAMA Oncol. 2022; 8: 1493-1501
        • Louis D.N.
        • Perry A.
        • Wesseling P.
        • Brat D.J.
        • Cree I.A.
        • Figarella-Branger D.
        • et al.
        The 2021 WHO classification of tumors of the central nervous system: a summary.
        Neuro Oncol. 2021; 23: 1231-1251
        • Goldberg S.B.
        • Gettinger S.N.
        • Mahajan A.
        • Chiang A.C.
        • Herbst R.S.
        • Sznol M.
        • et al.
        Pembrolizumab for patients with melanoma or non-small-cell lung cancer and untreated brain metastases: early analysis of a non-randomised, open-label, phase 2 trial.
        Lancet Oncol. 2016; 17: 976-983
        • Tawbi H.A.
        • Forsyth P.A.
        • Algazi A.
        • Hamid O.
        • Hodi F.S.
        • Moschos S.J.
        • et al.
        Combined nivolumab and ipilimumab in melanoma metastatic to the brain.
        N Engl J Med. 2018; 379: 722-730
        • Lim M.
        • Weller M.
        • Idbaih A.
        • Steinbach J.
        • Finocchiaro G.
        • Raval R.R.
        • et al.
        Phase 3 trial of chemoradiotherapy with temozolomide plus nivolumab or placebo for newly diagnosed glioblastoma with methylated MGMT promoter.
        Neuro Oncol. 2022; : noac116
        • Omuro A.
        • Brandes A.A.
        • Carpentier A.F.
        • Idbaih A.
        • Reardon D.A.
        • Cloughesy T.
        • et al.
        Radiotherapy combined with nivolumab or temozolomide for newly diagnosed glioblastoma with unmethylated MGMT promoter: an international randomized phase 3 trial.
        Neuro Oncol. 2022; : noac099
        • Blank C.U.
        • Haanen J.B.
        • Ribas A.
        • Schumacher T.N.
        CANCER IMMUNOLOGY. The "cancer immunogram.
        Science. 2016; 352: 658-660
        • Berghoff A.S.
        • Fuchs E.
        • Ricken G.
        • Mlecnik B.
        • Bindea G.
        • Spanberger T.
        • et al.
        Density of tumor-infiltrating lymphocytes correlates with extent of brain edema and overall survival time in patients with brain metastases.
        OncoImmunology. 2016; 5e1057388
        • Berghoff A.S.
        • Kiesel B.
        • Widhalm G.
        • Wilhelm D.
        • Rajky O.
        • Kurscheid S.
        • et al.
        Correlation of immune phenotype with IDH mutation in diffuse glioma.
        Neuro Oncol. 2017; 19: 1460-1468
        • Berghoff A.S.
        • Ricken G.
        • Widhalm G.
        • Rajky O.
        • Dieckmann K.
        • Birner P.
        • et al.
        Tumour-infiltrating lymphocytes and expression of programmed death ligand 1 (PD-L1) in melanoma brain metastases.
        Histopathology. 2015; 66: 289-299
        • Berghoff A.S.
        • Ricken G.
        • Wilhelm D.
        • Rajky O.
        • Widhalm G.
        • Dieckmann K.
        • et al.
        Tumor infiltrating lymphocytes and PD-L1 expression in brain metastases of small cell lung cancer (SCLC).
        J Neuro Oncol. 2016; 130: 19-29
        • Berghoff A.S.
        • Schur S.
        • Fureder L.M.
        • Gatterbauer B.
        • Dieckmann K.
        • Widhalm G.
        • et al.
        Descriptive statistical analysis of a real life cohort of 2419 patients with brain metastases of solid cancers.
        ESMO Open. 2016; 1e000024
        • Klemm F.
        • Maas R.R.
        • Bowman R.L.
        • Kornete M.
        • Soukup K.
        • Nassiri S.
        • et al.
        Interrogation of the microenvironmental landscape in brain tumors reveals disease-specific alterations of immune cells.
        Cell. 2020; 181: 1643-1660 e17
        • Friebel E.
        • Kapolou K.
        • Unger S.
        • Nunez N.G.
        • Utz S.
        • Rushing E.J.
        • et al.
        Single-cell mapping of human brain cancer reveals tumor-specific instruction of tissue-invading leukocytes.
        Cell. 2020; 181: 1626-1642 e20
        • Bunse L.
        • Pusch S.
        • Bunse T.
        • Sahm F.
        • Sanghvi K.
        • Friedrich M.
        • et al.
        Suppression of antitumor T cell immunity by the oncometabolite (R)-2-hydroxyglutarate.
        Nat Med. 2018; 24: 1192-1203
        • Dejaegher J.
        • Solie L.
        • Hunin Z.
        • Sciot R.
        • Capper D.
        • Siewert C.
        • et al.
        DNA methylation based glioblastoma subclassification is related to tumoral T-cell infiltration and patient survival.
        Neuro Oncol. 2021; 23: 240-250
        • Berghoff A.S.
        • Kiesel B.
        • Widhalm G.
        • Rajky O.
        • Ricken G.
        • Wohrer A.
        • et al.
        Programmed death ligand 1 expression and tumor-infiltrating lymphocytes in glioblastoma.
        Neuro Oncol. 2015; 17: 1064-1075
        • Mair M.J.
        • Kiesel B.
        • Feldmann K.
        • Widhalm G.
        • Dieckmann K.
        • Wohrer A.
        • et al.
        LAG-3 expression in the inflammatory microenvironment of glioma.
        J Neuro Oncol. 2021; 152: 533-539
        • Mair M.J.
        • Pajenda S.
        • Ilhan-Mutlu A.
        • Steindl A.
        • Kiesel B.
        • Widhalm G.
        • et al.
        Soluble PD-L1 is associated with local and systemic inflammation markers in primary and secondary brain tumours.
        ESMO Open. 2020; 5e000863
        • Han S.
        • Liu Y.
        • Li Q.
        • Li Z.
        • Hou H.
        • Wu A.
        Pre-treatment neutrophil-to-lymphocyte ratio is associated with neutrophil and T-cell infiltration and predicts clinical outcome in patients with glioblastoma.
        BMC Cancer. 2015; 15: 617
        • Alexandrov L.B.
        • Nik-Zainal S.
        • Wedge D.C.
        • Aparicio S.A.
        • Behjati S.
        • Biankin A.V.
        • et al.
        Signatures of mutational processes in human cancer.
        Nature. 2013; 500: 415-421
        • Vormehr M.
        • Diken M.
        • Boegel S.
        • Kreiter S.
        • Tureci O.
        • Sahin U.
        Mutanome directed cancer immunotherapy.
        Curr Opin Immunol. 2016; 39: 14-22
        • He Q.
        • Li Q.
        • Lv F.
        • Kaitin K.I.
        • Shao L.
        A survey of survival outcomes for targeted cancer drugs approved by the US food and drug administration.
        Ther Innov Regul Sci. 2021; 55: 676-684
        • Appin C.L.
        • Brat D.J.
        Molecular pathways in gliomagenesis and their relevance to neuropathologic diagnosis.
        Adv Anat Pathol. 2015; 22: 50-58
        • Wang Q.
        • Hu B.
        • Hu X.
        • Kim H.
        • Squatrito M.
        • Scarpace L.
        • et al.
        Tumor evolution of glioma-intrinsic gene expression subtypes associates with immunological changes in the microenvironment.
        Cancer Cell. 2017; 32: 42-56 e6
        • Neftel C.
        • Laffy J.
        • Filbin M.G.
        • Hara T.
        • Shore M.E.
        • Rahme G.J.
        • et al.
        An integrative model of cellular states, plasticity, and genetics for glioblastoma.
        Cell. 2019; 178: 835-849 e21
        • Garofano L.
        • Migliozzi S.
        • Oh Y.T.
        • D'Angelo F.
        • Najac R.D.
        • Ko A.
        • et al.
        Pathway-based classification of glioblastoma uncovers a mitochondrial subtype with therapeutic vulnerabilities.
        Nat Can (Que). 2021; 2: 141-156
        • Varn F.S.
        • Johnson K.C.
        • Martinek J.
        • Huse J.T.
        • Nasrallah M.P.
        • Wesseling P.
        • et al.
        Glioma progression is shaped by genetic evolution and microenvironment interactions.
        Cell. 2022; 185: 2184-2199 e16
        • Sa J.K.
        • Chang N.
        • Lee H.W.
        • Cho H.J.
        • Ceccarelli M.
        • Cerulo L.
        • et al.
        Transcriptional regulatory networks of tumor-associated macrophages that drive malignancy in mesenchymal glioblastoma.
        Genome Biol. 2020; 21: 216
        • Mall R.
        • Cerulo L.
        • Garofano L.
        • Frattini V.
        • Kunji K.
        • Bensmail H.
        • et al.
        RGBM: regularized gradient boosting machines for identification of the transcriptional regulators of discrete glioma subtypes.
        Nucleic Acids Res. 2018; 46: e39
        • Caruso F.P.
        • Garofano L.
        • D'Angelo F.
        • Yu K.
        • Tang F.
        • Yuan J.
        • et al.
        A map of tumor-host interactions in glioma at single-cell resolution.
        GigaScience. 2020; 9 (giaa109)
        • Mathewson N.D.
        • Ashenberg O.
        • Tirosh I.
        • Gritsch S.
        • Perez E.M.
        • Marx S.
        • et al.
        Inhibitory CD161 receptor identified in glioma-infiltrating T cells by single-cell analysis.
        Cell. 2021; 184: 1281-1298 e26
        • Long G.V.
        • Atkinson V.
        • Lo S.
        • Sandhu S.
        • Guminski A.D.
        • Brown M.P.
        • et al.
        Combination nivolumab and ipilimumab or nivolumab alone in melanoma brain metastases: a multicentre randomised phase 2 study.
        Lancet Oncol. 2018; 19: 672-681
        • Tawbi H.A.
        • Forsyth P.A.
        • Hodi F.S.
        • Algazi A.P.
        • Hamid O.
        • Lao C.D.
        • et al.
        Long-term outcomes of patients with active melanoma brain metastases treated with combination nivolumab plus ipilimumab (CheckMate 204): final results of an open-label, multicentre, phase 2 study.
        Lancet Oncol. 2021; 22: 1692-1704
        • Kim P.H.
        • Suh C.H.
        • Kim H.S.
        • Kim K.W.
        • Kim D.Y.
        • Aizer A.A.
        • et al.
        Immune checkpoint inhibitor therapy may increase the incidence of treatment-related necrosis after stereotactic radiosurgery for brain metastases: a systematic review and meta-analysis.
        Eur Radiol. 2021; 31: 4114-4129
        • Kluger H.M.
        • Chiang V.
        • Mahajan A.
        • Zito C.R.
        • Sznol M.
        • Tran T.
        • et al.
        Long-Term survival of patients with melanoma with active brain metastases treated with pembrolizumab on a phase II trial.
        J Clin Oncol. 2019; 37: 52-60
        • Tumeh P.C.
        • Harview C.L.
        • Yearley J.H.
        • Shintaku I.P.
        • Taylor E.J.
        • Robert L.
        • et al.
        PD-1 blockade induces responses by inhibiting adaptive immune resistance.
        Nature. 2014; 515: 568-571
        • Harter P.N.
        • Bernatz S.
        • Scholz A.
        • Zeiner P.S.
        • Zinke J.
        • Kiyose M.
        • et al.
        Distribution and prognostic relevance of tumor-infiltrating lymphocytes (TILs) and PD-1/PD-L1 immune checkpoints in human brain metastases.
        Oncotarget. 2015; 6: 40836-40849
        • Smalley I.
        • Chen Z.
        • Phadke M.
        • Li J.
        • Yu X.
        • Wyatt C.
        • et al.
        Single-cell characterization of the immune microenvironment of melanoma brain and leptomeningeal metastases.
        Clin Cancer Res. 2021; 27: 4109-4125
        • Fischer G.M.
        • Jalali A.
        • Kircher D.A.
        • Lee W.C.
        • McQuade J.L.
        • Haydu L.E.
        • et al.
        Molecular profiling reveals unique immune and metabolic features of melanoma brain metastases.
        Cancer Discov. 2019; 9: 628-645
        • Egelston C.A.
        • Margolin K.
        Metabolic checkpoint of immune cells in melanoma brain metastases.
        Cancer Discov. 2019; 9: 581-583
        • Di Giacomo A.M.
        • Ascierto P.A.
        • Pilla L.
        • Santinami M.
        • Ferrucci P.F.
        • Giannarelli D.
        • et al.
        Ipilimumab and fotemustine in patients with advanced melanoma (NIBIT-M1): an open-label, single-arm phase 2 trial.
        Lancet Oncol. 2012; 13: 879-886
        • Di Giacomo A.M.
        • Chiarion-Sileni V.
        • Del Vecchio M.
        • Ferrucci P.F.
        • Guida M.
        • Quaglino P.
        • et al.
        Primary analysis and 4-year follow-up of the phase III NIBIT-M2 trial in melanoma patients with brain metastases.
        Clin Cancer Res. 2021; 27: 4737-4745
        • Reardon D.A.
        • Kim T.M.
        • Frenel J.S.
        • Simonelli M.
        • Lopez J.
        • Subramaniam D.S.
        • et al.
        Treatment with pembrolizumab in programmed death ligand 1-positive recurrent glioblastoma: results from the multicohort phase 1 KEYNOTE-028 trial.
        Cancer. 2021; 127: 1620-1629
        • Cloughesy T.F.
        • Mochizuki A.Y.
        • Orpilla J.R.
        • Hugo W.
        • Lee A.H.
        • Davidson T.B.
        • et al.
        Neoadjuvant anti-PD-1 immunotherapy promotes a survival benefit with intratumoral and systemic immune responses in recurrent glioblastoma.
        Nat Med. 2019; 25: 477-486
        • Duerinck J.
        • Schwarze J.K.
        • Awada G.
        • Tijtgat J.
        • Vaeyens F.
        • Bertels C.
        • et al.
        Intracerebral administration of CTLA-4 and PD-1 immune checkpoint blocking monoclonal antibodies in patients with recurrent glioblastoma: a phase I clinical trial.
        J Immunother Cancer. 2021; 9e002296
        • Weller M.
        • Butowski N.
        • Tran D.D.
        • Recht L.D.
        • Lim M.
        • Hirte H.
        • et al.
        Rindopepimut with temozolomide for patients with newly diagnosed, EGFRvIII-expressing glioblastoma (ACT IV): a randomised, double-blind, international phase 3 trial.
        Lancet Oncol. 2017; 18: 1373-1385
        • Weiss T.
        • Puca E.
        • Silginer M.
        • Hemmerle T.
        • Pazahr S.
        • Bink A.
        • et al.
        Immunocytokines are a promising immunotherapeutic approach against glioblastoma.
        Sci Transl Med. 2020; 12eabb2311
        • O'Rourke D.M.
        • Nasrallah M.P.
        • Desai A.
        • Melenhorst J.J.
        • Mansfield K.
        • Morrissette J.J.D.
        • et al.
        A single dose of peripherally infused EGFRvIII-directed CAR T cells mediates antigen loss and induces adaptive resistance in patients with recurrent glioblastoma.
        Sci Transl Med. 2017; 9eaaa0984
        • Weller M.
        • Roth P.
        • Preusser M.
        • Wick W.
        • Reardon D.A.
        • Platten M.
        • et al.
        Vaccine-based immunotherapeutic approaches to gliomas and beyond.
        Nat Rev Neurol. 2017; 13: 363-374
        • Bogdahn U.
        • Hau P.
        • Stockhammer G.
        • Venkataramana N.K.
        • Mahapatra A.K.
        • Suri A.
        • et al.
        Targeted therapy for high-grade glioma with the TGF-beta2 inhibitor tradersen: results of a randomized and controlled phase IIb study.
        Neuro Oncol. 2011; 13: 132-142
        • Brandes A.A.
        • Carpentier A.F.
        • Kesari S.
        • Sepulveda-Sanchez J.M.
        • Wheeler H.R.
        • Chinot O.
        • et al.
        A Phase II randomized study of galunisertib monotherapy or galunisertib plus lomustine compared with lomustine monotherapy in patients with recurrent glioblastoma.
        Neuro Oncol. 2016; 18: 1146-1156
        • Papachristodoulou A.
        • Silginer M.
        • Weller M.
        • Schneider H.
        • Hasenbach K.
        • Janicot M.
        • et al.
        Therapeutic targeting of TGFbeta ligands in glioblastoma using novel antisense oligonucleotides reduces the growth of experimental gliomas.
        Clin Cancer Res. 2019; 25: 7189-7201
        • Yap T.A.
        • Vieito M.
        • Baldini C.
        • Sepulveda-Sanchez J.M.
        • Kondo S.
        • Simonelli M.
        • et al.
        First-in-Human phase I study of a next-generation, oral, TGFbeta receptor 1 inhibitor, LY3200882, in patients with advanced cancer.
        Clin Cancer Res. 2021; 27: 6666-6676
        • Medikonda R.
        • Dunn G.
        • Rahman M.
        • Fecci P.
        • Lim M.
        A review of glioblastoma immunotherapy.
        J Neuro Oncol. 2021; 151: 41-53
        • Simonds E.F.
        • Lu E.D.
        • Badillo O.
        • Karimi S.
        • Liu E.V.
        • Tamaki W.
        • Rancan C.
        • Downey K.M.
        • Stultz J.
        • Sinha M.
        • McHenry L.K.
        • Nasholm N.M.
        • Chuntova P.
        • Sundstr√∂m A.
        • Genoud V.
        • Shahani S.A.
        • Wang L.D.
        • Brown C.E.
        • Walker P.R.
        • Swartling F.J.
        • Fong L.
        • Okada H.
        • et al.
        Deep immune profiling reveals targetable mechanisms of immune evasion in immune checkpoint inhibitor-refractory glioblastoma.
        J Immunother Cancer. 2021 Jun; 9 (PubMed [citation] PMID: 34083417, PMCID: PMC8183210)e002181https://doi.org/10.1136/jitc-2020-002181
        • Bouffet E.
        • Larouche V.
        • Campbell B.B.
        • Merico D.
        • de Borja R.
        • Aronson M.
        • et al.
        Immune checkpoint inhibition for hypermutant glioblastoma multiforme resulting from germline biallelic mismatch repair deficiency.
        J Clin Oncol. 2016; 34: 2206-2211
        • Touat M.
        • Li Y.Y.
        • Boynton A.N.
        • Spurr L.F.
        • Iorgulescu J.B.
        • Bohrson C.L.
        • et al.
        Mechanisms and therapeutic implications of hypermutation in gliomas.
        Nature. 2020; 580: 517-523
        • Preusser M.
        • Lim M.
        • Hafler D.A.
        • Reardon D.A.
        • Sampson J.H.
        Prospects of immune checkpoint modulators in the treatment of glioblastoma.
        Nat Rev Neurol. 2015; 11: 504-514
        • Gallego Perez-Larraya J.
        • Garcia-Moure M.
        • Labiano S.
        • Patino-Garcia A.
        • Dobbs J.
        • Gonzalez-Huarriz M.
        • et al.
        Oncolytic DNX-2401 virus for pediatric diffuse intrinsic pontine glioma.
        N Engl J Med. 2022; 386: 2471-2481
        • Lee A.H.
        • Sun L.
        • Mochizuki A.Y.
        • Reynoso J.G.
        • Orpilla J.
        • Chow F.
        • et al.
        Neoadjuvant PD-1 blockade induces T cell and cDC1 activation but fails to overcome the immunosuppressive tumor associated macrophages in recurrent glioblastoma.
        Nat Commun. 2021; 12: 6938