`Mechanismsof Resistance to Immune Checkpoint
`Blockade: Why Does Checkpoint Inhibitor
`Immunotherapy Not Work for All Patients?
`
`Charlene M, Fares, MD"; Eezer M. Van Allen, MD?; Charles G. Drake, MD, PhD®; James P. Allison, PhD*; and
`Slwen Hu-Lieskovan, MD, PhD®
`
`AADTATOAO
`
`The emergence of immune checkpoint blockade therapies over the last decade has transformed cancer
`treatment in a wide range of tumor types. Unprecedented and durable clinical responsesin difficult-to-treat
`cancer histologies have been observed. However, despite these promising long-term responses, the majority of
`. patients fail to respond to immune checkpoint blockade, demonstrating primary resistance. Additionally,
`many of those who initially respond to treatment eventually experience relapse secondary to acquired re-
`sistance. Both primary and acquired resistance are a result of complex and constantly evolving interactions
`between cancer cells and the immune system. Many mechanismsof resistance have been characterized to
`date, and more continue to be uncovered. By elucidating and targeting mechanismsof resistance, treatments
`can be tailored to improve clinical outcomes. This review will discuss the landscape of immune checkpoint
`blockade response data, different resistance mechanisms, and potential therapeutic strategies to overcome
`resistance.
`
`INTRODUCTION
`
`Immunotherapy has recently become a viable option
`for cancer treatment; however,
`the concept of har-
`nessing the immune system to fight malignancy dates
`back over a century.
`In the 1890s, Dr. William Coley
`observed improved clinical outcomes in patients with
`cancer who experienced postsurgica? infections.
`Based on these observations, Coley experimented by
`introducing bacterial toxins to patients with sarcoma.
`Although results were inconsistent, he was able to
`demonstrate turior regression in a subsetof patients.
`However, with the advent of chemotherapy and ra-
`diotherapy, immunotherapy wentlargely overlooked, In
`the 1950s, Ehrlich formulated the concept of immu-
`nosurveiflance, which propased that emergence of
`matignant cells is a frequent event, but evolution to
`clinically relevant disease is suppressed by the im-
`mune system uniess immunity is weakened.? Although
`these early hypotheses fueled the field of cancer im-
`munotherapy, better understanding of immune acti-
`vation, regulation, and interaction with tumorcells and
`the microenvironment was needed.
`
`Now we know that the process of T-cell-mediated
`immunity is a complex sequence of events, with
`constant interplay between stimulatory and inhibitory
`signals that promote adaptive responses against for-
`eign antigens while avoiding autoimmunity. Antigen-
`specific T cells initially undergo clonal selection, with
`
`subsequent priming and activation following T-cell
`receptor recognition of corresponding antigens on
`Major histocompatibility complexes (MHCs) expressed
`by antigen-presenting ceils. For full activation, a cos-
`timulatory signal is needed between antigen-presenting
`cells and T calls, After activation and proliferation, T cells
`are trafficked to specific sites by following a chemokine
`gradient. Upon encountering cognate antigen on MHCs,
`effector T cells (Teffs) release interferon garmma (IFN-y)
`and other cytokines, promoting cytotoxicity and tumor
`cell killing. Following cancer cell eradication, memory
`¥ cells form and remain quiescent until antigen re-
`exposure.
`
`Under normal physiologic conditions, immune check-
`points function as negative feedback to regulate in-
`flammatory responses following T-cell activation, The
`CTLA-4 immune checkpoint receptor was first char-
`acterized by Brunetet al? in the 1980s. Seminal work by
`Krummel and Allison? demonstrated that CTLA-4 an
`T cells competitively binds to B7 ligands on antigen-
`presenting cells, interfering with CD28 interactions, thus
`preventing costimulation and the priming phase of T-cell
`activation (Fig. 1A). Subsequently, blockade of CTLA-4
`with antibodies demonstrated tumor
`rejection and
`emerged as proof of concept for immune checkpoint
`inhibitors.° Another immune checkpoint receptor, PD-1,
`was cloned in 1992* with subsequent characterization
`of its ligand, PD-L1,7° Interaction of PD-1 with its
`
`
`
`Authoraffiliations
`and support
`information ¢if
`applicable) appear
`al the end of this
`atticle,
`Accepted an May 17,
`2019 and published
`al ascopubs.org on
`May 17, 2018;
`DG! https/tolorg?
`10, 1200/EDBK_
`240837
`
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`Copyright © 2019 American Society of Clinical Oncology. All rights reserved.
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`Fares atal
`
`PD-1/PD-L1 checkpoint blockade tend to be proportional to
`their corresponding tumor mutational burden (TMB), pre-
`sumably from the immunogenic neoantigens that are rec-
`ognized as foreign by cytotoxic T lymphocytes (CTLs).4*4
`However, tumors with similar TMB can have very different
`response to checkpoint inhibitors, indicating that response
`fo immune checkpoint blockade (ICB)
`is complex, het-
`erogeneous, and inconsistent and that additional mecha-
`nisms are at play. Increased PD-L1 expression has been
`correlated with insmune response and is currently used as
`a biomarker for ICB therapy in NSCLC and urothelial car-
`cinoma.Additionally, elevated numbers of
`tumor-
`infiltrating lymphocytes (TILs) have been noted in re-
`spansive cancers.2)*
`MECHANISMS OF RESISTANCE
`
`The biggest chaflenges for the cancer immunotherapy field
`are to understand the complex resistance mechanisms and
`to develop effective combination strategies to overcome
`resistance. According to the timing of occurrence,
`re-
`sistance can be primary, as in never-responders, or ac-
`quired, which emerges after a period of
`response.
`Resistance can also be classified as intrinsic or extrinsic to
`tumorcells. Intrinsic resistance is seen when cancercells
`alter processes that are related to immune recognition, cell
`signaling, gene expression, and DNA damage response.
`Extrinsic resistance occurs external to tumorcells throughout
`the T-cell activation process.
`
`Tumor Immunogenicity
`
`The abitily for tumors to induce adaptive immune responses
`relies on recognition of cancer cells as foreign. High TMB,
`with accompanying elevated neoantigen expression, plays
`an importantrole in antiturnor immunity.*4* With improved
`sequencing techniques, nonsynonymous mutations were
`found to generate tumor neoantigens that drive cyto-
`toxic responses against cancer cells °*5 Van Allen and
`colleaguesdemonstrated that mutational
`load was sig-
`nificantly associated with response to anti-CTLA-4 treat-
`ment in patients with metastatic melanoma. Additionally,
`Rizvi et al*®45 showed that responseto anti-PD-1 treatment
`correlated with high TMB and neoantigen load in patients
`with NSCLC. In keeping with these studies, poorly immu-
`nogenic tumors with low TMB, such as pancreatic and
`prostate cancers, are inherently more resistant to treatment
`with checkpoint inhibition.“
`Extrapolating frorn these data, mechanisms leading to loss
`of neoantigen expression by cancer cells may result in
`acquired resistance to [CB. The conceptof Immunoediting
`exemplifies the impact of necantigen loss on tumor im-
`munogenicily and explains how resistance might be formed
`against cancers with high TMB. Immuncediting suggests
`that constantinteractions between the immune system and
`cancer cells result in selection of subclones within the tumor
`
`PRACTICAL APPLICATIONS
`
`® Immune checkpoint inhibitors provide durable
`clinical responses in multiple difficult-to-treat
`tumor types.
`* The tumar microenvironment, tumor immuno-
`genicity, antigen presentation, and classic on-
`cologic pathways play roles in response and
`resistance to immune checkpoint blockade.
`« By understanding resistance mechanisms to
`immune checkpoint blockade, therapies can be
`developed to overcomeresistance and treat-
`mentfailure.
`
`» Combination treatment strategies with immune
`checkpoint inhibitors are being tested in clinical
`trials, with several alreadyin clinical use.
`« Response to immunotherapy may be better
`predicted by using a wide set of biomarkers.
`
`inhibits the effector phase of
`ligands, PD-L1 and PD-L2,
`T-cell activation, thus dampening the immune response.'°
`Many tumors are now known to hifack this mechanism to
`avoid T-cell killing, and inhibitory antibodies directed against
`the interaction between PD-1 andits ligands have demon-
`strated antitumor responses.¥
`
`GLINIGAL RESPONSE TO IMMUNE CHECKPOINT INHIBITORS
`
`To date, seven immune checkpoint inhibitors have received
`US. Food and Drug Administration approval: one CTLA-4
`inhibitor (ipilimumab), three PD-1 inhibitors (nivolumab,
`pembrolizumab, and cemiplimab), and three PD-L1 in-
`hibitors (atezolizumab, durvalumab, and avelumab}.
`Ipili-
`mumab was the first immune checkpoint inhibitor to gain
`approval in 2011 for the treatment of melanoma."* In 2014,
`hivolumab and pembrolizumab were approved in mela-
`noma and have now gained indications for use in non—small
`cell lung cancer (NSCLC), renal cell carcinoma, head and
`neck squamous cell carcinoma, urothelial carcinoma, and
`microsatellite instability-high colorectal cancer, among
`several other tumor types.5° Atezolizumab, avelumab,
`and durvalumab are approved in many of the same his-
`tologies as the PD-1 inhibitors.3”“? Most recently, cemi-
`plimab was approved fortreatment of metastatic cutaneous
`squamous cell carcinoma.*?
`
`One of the hallmarks of immunotherapy is the durability of
`the responses that can be translated into survival benefit.
`indeed,
`in approved indications, checkpoint inhibitor im-
`munotherapy profonged survival in patients with responding
`disease,raising the tail of patient survival curves, However,
`only a subset of tumor histologies and a small percentage
`of the patients in each histology are responsive to these
`inhibitors. The response rates of different tumor types to
`
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`Mechanisms of Resistance to immune Checkpoint Blockade
`
`
`
`FAGURE 1. T-Cell Activation and
`Cosignating
`
`(A) T-cell receptor interacts with
`anligen/majer histocompatibility
`complex on APCs. Castimulatory
`signal
`is provided by B7/CD28
`interaction for T-cell activation,
`CTLA-4 competes with CD28 for
`8? binding, providing cainhibitery
`signais. {B) Costimulatory signals
`currently being targeted fo im-
`prove T-cell activation,
`(C) Ex
`pression of colnhibitory receptars
`leads to T-cell exhaustion. Colnhi-
`bitory receptors serve as thera-
`peutic targets to enhance antitumor
`immune respanse.
`Abbreviation: APC,
`presenting cell.
`
`antigen-
`
`that lack expression of neoantigens, subsequently con-
`ferring poor immunogenicity and resistance to 1CB.5%5?
`With increased intratumor heterogeneity, there is greater
`likelihood that a poorly immunogenic subclone could be
`selected,
`thus decreasing sensitivity to checkpoint
`in-
`hibition.5%5° A recent study by Anagnostou et al showed
`that relapse of NSCLC tumors after treatment with PD-1/
`PD-L1 and CTLA-4 inhibitors demonstrated loss of seven
`to 18 putative neoantigens, supporting the role of immu-
`noediting in acquired resistance (Table 1). Another study
`recently showed that expression of iFN-y paradoxically fa-
`ciliates immunoediting by CTLs, with resulting gene copy
`numberalteration contributing to immune resistance”!
`Genetic instability due to alterations in DNA repair and
`replication genes can increase immunogenicity through
`high mutational burden with subsequent neoantigen for-
`mation. Patients with melanoma were found to have better
`response to anti-PD-1 treatment
`if
`tumer cells were
`enriched for mutations in BRCAZ, an important homolagous
`recombination DNA repair gene.Similar findings were
`demonstrated in ovarian cancer,
`in which BRCAL/2-
`mutated turnors demonstrated high neoantigen joads.™
`Alterations in additional DNA damage response genes,
`including ATM, POLE, FANCA, ERCC2, and MSH6, have
`recently shown correlation with high TMB and improved
`
`clinical outcornes to ICB in urothelial cancer. Further-
`more, tumors with deficiencies in DNA mismatch repair
`genesleadingto microsatellite instability demonstrated high
`mutational burden with enhanced response to ICB across
`a wide range of histologies.°
`
`The presence of PD-Ll—expressing cancer cells within tu-
`mors is known to be an important predictor of response to
`ICB therapy and is commonly used as a biomarker.® It has
`been shown that tumors lacking PD-L1 expression generally
`show inferior clinical outcomes to 1CB compared with those
`with higherlevels of ligand.° However, tumors with absent
`PD-L1 can respond to ICB, as PD-L1 expression’ can be
`induced upon activation of the IFN response pathway.
`Regardiess of PD-L1 expression, tissues that lack TiLs are
`unlikely to respond to ICB. Tumors with larger numbers of
`TILs demonstrate greater response to iCB and may serve as
`another predictive biomarker.8*7? A study in patients with
`metastatic melanoma showed that pre-existing tumoral
`CTLs are a qualification for response to antt-PD-1 therapy.
`Tumor Microenvironment
`
`The tumor microenvironment (TME) consists of factors ex-
`iinsic to cancer cells, including various immune and stromal
`cells, vascufature, extracellular matrix, and cytokines that
`influence response to therapy. Immune-suppressive cells,
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`Fares et al
`
`TABLE 1. Mechanisms of Resistance in PD-L1—Overexpressed Tumors, Combination 1CB, Tumors With Mutated Chromatin Remodeling Complexes, and
`High TMB Cancers
`Specific Circumstances
`
`Mechanismsof Resistance
`
`PD-L1—overexpressed tumors
`
`Nonreversible and severe T-cell exhaustion
`
`
`PD-1 and CTLA-4 inhibitor combination therapy
`
`Coexpression of inhibitory receptors (LAG-3, TIM-3, TIGIT, VISTA, and
`BTLA}?2
`
`Decreased ratio of TiLs to Tregs and MDSCs
`
`Altered metaboflsm through [DO and increased adenosine production
`Mutations In PTEN, EGFR, and MYC
`
`Immunsediting with loss of neoantigens®
`
`Deletions or mutations in JAKL2, IFNGRI/2Z, and [RFP
`Decreased T-cell priming and OC dysfunction
`Aberrant WNT/p-catenin signaling
`High copy number loss of tumor suppressor genes®
`
`Association with neoantigen overexpression by genetic alterations in
`mammalian SWI/SNF chromatin remodeling complexes
`
`Less-of-function mutations in chromatin remodeler genes (PBRMI, ARIDZ
`and BRD?) sensitize tumors ta (CB and increase aceessihility to
`regulatory elements of JFN-y-Inducible genes. Loss of AR/IDZA leadsto
`increased microsateltite Instability with inability to recrult mismatch repair
`genes during DNA repair, Increasing mutational burden ard neoantigen
`load, Stabilty of chromatin remedeling complexes in tumors contributes
`to ICB resistance, ‘°1&
`High mutation overload tumors
`
`Decreased antigen presentation secondary to MHC, B2-microglobulin, and
`NLRC5 alterations’
`
`Abbreviations: TIGIT, ¥-cell immunoreceptor tyrosine-based inhibition motif domain; VISTA, V-corain immunogiobulin-containing suppressor of T-cell
`activation; BTLA, B and T lymphocyte attenuator,
`
`JAKL/2 mutations and decreased !FN-y signaling
`Upregulation of alternate Inhibitory checkpoints
`
`along with inhibitory cytokines in the TME, can undermine
`the antitumor
`immune response.”°7? Regulatory T cells
`(Tregs) are knownto facilitate self-tolerance by suppressing
`Teff function through inhibitory cytokines and direct contact,
`limiting inflammiation.”””*Infiltration of tumors by Tregs has
`been observed in many tumor types, suggesting an immu-
`nosuppressive environment in same cancers.” The ratio of
`Teffs to Tregs in murine modelsis associated with response to
`ICB, in that inability to increase Teffs or decrease Tregs may
`result in resistance to irmmunotherapy,”*7”
`Myeloid-derived suppressorcells (MDSCs) are anothertype
`of regulatory cell within the TME that can promote immune
`evasion and tumor growth.”2’”9 MDSCs have been shown to
`piay a role in facilitating tumor invasion, metastasis, and
`angiogenesis.“°Clinical studies demonstrate that increased
`presence of MDSCs within the TME correlates with poor
`response to ICB.8? Accordingly, by inhibiting trafficking of
`MDSCsto the TME, enhanced response to anti-PD-1 therapy
`was seen in a murine meadel of rhabdomyosarcoma,
`
`Tumor-associated macrophages, particularly M2 macro-
`phages, promote tumor progression through modifications
`of the TME.™ M2 macrophages are known to stimulate
`tumor cell motitity, angiogenesis, growth, and immune
`
`evasion.Consequently, depletion of tumor-associated
`macrophages in several different murine models corre-
`lated with reduced tumor growth.*°*" Moreover, inhibition of
`myeloid growth factor signaling in macrophages circum-
`vented therapeutic resistance to ICB in a murine model of
`pancreatic cancer.24
`
`The cytokine milieu within the TME is involved in immune
`cell recruitment, activation, and proliferation, exerting both
`immune stimulatory and suppressive effects.2° Severai
`chemokines, including CCL5, CCL17, CCL22, CXCL8, and
`CXCL12, play a role in recruiting MDSCs and Tregs to the
`TME, thus promoting an immunosuppressive climate.23%
`Consequently, inhibition of the chemokine receptor CCR4
`diminished trafficking of Tregs and promoted antitumor
`effects.%*°? Alternately, CXCL9 and CXCLLO recruit CTLs to
`the TME, with subsequent destruction of cancer cells.°4°8
`Expression of CXCLS and CXCLIO can be epigenetically
`silenced, reducing TiLs and promoting resistance to ICB.
`Epigenetic modulator therapy in a modelfor ovarian cancer
`reversed suppression of these chemokines and enhanced
`response to ICB.34
`
`Transforming growth factor beta (TGF-$) signaling in-
`fluences multiple TME eiements, including cell growth and
`
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`Mechanisms of Resistance to fmmune Checkpoint Slockade
`
`differentiation, wound healing, apoptosis, and immunc-
`suppression. TGF-6 fimits
`immunosuppression through
`inhibition of CTLs while upregulating Tregs.?” In a murine
`colorectal cancer model, elevated TGF-B signaling was
`associated with poorly immunogenic tumors and limited
`response to ICB, indicating resistance.” Jn line with these
`findings, improved antitumor response to ICB was seen with
`inhibition of TGF-p in metastatic urothelial cancer.>*
`
`In addition to promoting angiogenesis, VEGF functions as an
`immunosuppressive cytokine and is associated with re-
`sistance to ICB. VEGF levels were found to be higher in
`ant-PD-i
`therapy nonresponders compared with re-
`sponders.’”° In mouse models, VEGF impeded commitment
`of lymphoid progenitors, reducing progression to the T-celt
`lineage.’Additionally, VEGF signaling reducestrafficking
`and extravasation of CTLs into the TME while it promotes
`infiltration of Tregs through a selective endothelium?
`Furthermore, VEGF increases expression of inhibitory re-
`ceptors, contributing to CTL exhaustion. Corroborating
`this evidence, inhibition of VEGF was correlated with im-
`proved response to ICB in renal cell carcinoma2™
`
`and CTLA-4 blockade through inability to upregulate MHC-I
`and PD-L1 expression.1?41#)
`
`Overexpression of alternate immune checkpoints has been
`linked to anti-PD-1 and anti-CTLA-4 therapeutic failure.
`Adaptive resistance to ICB was observed secondary to com-
`pensatory upregulation of alternative immune checkpoint re-
`ceptors,
`including T-cell
`immunoglobulin, mucin domain-3
`protein (TIM-3), and lymphocyte-aclivation gene 3 (LAG-3},
`across multiple studies.!* Alternate immune checkpoint
`receptors continue to be discovered (Fig. 1C), including B and
`T tymphocyte attenuator
`(BTLA), T-ceil
`immunoreceptor
`tyrosine-based inhibition motif domain (TIGIT), and V-domain
`immunoglobulin-containing suppressor of T-cell activation
`(VISTA}.5?? Coexpression of multiple immune checkpoints
`has been associated with a severely exhausted T-cell state.
`Thommen at al? demonstrated a positive correlation between
`progressive T-cell exhaustion and increased coexpression of
`PD-1, CTLA-4, TIM-3, LAG-3, and BTLA, with subsequent
`resistance to ICB in NSCLC. Thus, these alternative immune
`checkpoint receptors may serve as potential therapeutic tar-
`gets for blockade.
`
`Antigen Presentation and Evolution of Immune Response
`
`The evolving immune response, from initial antigen expo-
`sure to cancercell cytotoxicity and memory T-cell formation,
`can be manipulated to evade antitumor immunity. The in-
`ability of T celts to proliferate and adequately diversify likely
`contributes to [CB resistance.
`Impaired priming of naive
`T cells through suppressed dendritic cell (DC) recruitment
`was associated with lack of TiLs and ICB resistance in
`melanoma.’Deficiencies in antigen presentation have
`been shown to play a role in ICB resistance. Multiple studies
`demonstrated that downregulation of MHC class | (MHC-)
`allows tumorcellsto resist immune surveillance. ©"? Loss of
`function of B2-microglobulin results in disruption of MHC-I
`folding and transport to the cell surface, thus mediating
`immune evasion of tumorcells.) An important study of
`patients with melanoma found truncating mutations in p2-
`microglobulin, leading to loss of MHC-[ expression and ac-
`quired resistance to ICB.1!* Additionally, mutations within
`the T-cell receptor binding domain of MHC-l have been
`identified in colorectal cancer, abrogating cytotoxicity and
`contributing to immune escape.)
`
`The IFN-y signaling pathway mediates immune response
`through the JAK/STAT family of receptors and transducers.
`IFN-y signaling upregulates expression of MHC-I, resulting in
`enhanced antigen presentation (Fig. 2A).2" However, IFN-y
`aiso functions within a negative-feedback loop to increase
`expression of PD-L1, conferring adaptive resistance to tumor
`cells.445118 (n the context of PD-1 blockade, amplification of
`PD-L1 in Hodgkin lymphoma correlated with improved re-
`sponse to therapy!’ Multiple studies have demonstrated
`that foss of JAK/STATsignaling results in resistance to PD-1
`
`In addition to expression of inhibitory receptors, exhausted
`T cells demonstrate impaired effector function and altered
`transcriptional state compared with Teffs. T-cell exhaustion
`presents as a spectrum, with association seen between pro-
`gressive loss of function and antigen persistence,!” Chronic
`exposure to cognate antigen also results in elevated PD-1
`expression, with subsequently impaired T-cell function.!
`Studies have shown that tumors with tow or intermediate
`expression of PD-1 can be reinvigorated with ICB. However,
`high expression of PD-1 was correlated with accumufating
`T-cell exhaustion and poor response to therapy.@"2? Re-
`cently, epigenetic changes were finked to T-cell exhaustion, in
`that exhausted cells were found to have a unique chromatin
`landscape that influenced transcriptional state and limited
`effector function.*55 Moreover, the type of distinct chromatin
`state determinedif exhausted T cells could be reprogrammed
`after therapy to avoid terminal exhaustion.*°4
`
`Following effector activity, a minority of T cells enter
`a memory phase,
`remaining quiescent until antigen
`rechaltenge."”""" Chronic antigen exposure renders pre-
`cursor memory T cells exhausted, with eventual deletion
`and lack of memory formation.!"°4" Given that success of
`ICB is highlighted by marked response durability, memory
`T-cell formation plays an important role in avoiding re-
`currence and resistance following cessation of treatment.
`Accordingly, patients who responded poorly to anti-PD-1
`therapy were shown to harbor fewer tumor-associated
`memory T cells compared with responsive patients.
`
`Classic Oncologic Pathways
`
`Through aberrations in oncogenes and tumor suppressors,
`oncologic signaling pathways can regulate immune
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` Fares et al
`
`a Inhibiter
`
`FIGURE 2, Gall Signaling Pathways
`Influencing Immune Response
`
`Interferon gamma_ signaling
`{A)
`through JAK/STAT upregulates
`both antigen presentation and
`PD-L1 expression. Loss of JAK/
`STAT confers resistance io IGB.
`(B) Aberrant signaling through
`canonical oncologic pathways
`feads to tumor formation and can
`be targeied to overcome immune
`checkpoint blackade reststance.
`
`response (Fig. 2B). These canonical pathwaysinfluence the
`TME. by altering immune cell composition and cytokine
`profile, rendering tumors resistant to ICB.
`
`The mitogen-activated protein kinase (MAPK) pathwayis
`involved in various cellular activities,
`including pro-
`liferation, apoptosis, and motility. Thus, abnormalities in
`this pathway promote oncogenesis in multiple tumor
`types.'"? Increased MAPK signaling impairs recruitment
`and function of TiLs through expression of VEGF and
`multiple other inhibitory cytokines, resulting in immune
`evasion.Multiple studies in mouse models have demon-
`strated that MAPK inhibitors enhance TiLs, IFN-y signaling,
`MHC-I expression, and PD-L1levels, thus promoting tumor
`cell cytotoxicity, -6
`
`Loss of the tumor suppressor phosphatase and tensin ho-
`molog (PTEN)
`results in constitutive activation of
`the
`phosphaticylinositel 3-kinase {PI3K) pathway,
`leading to
`tumorigenesis.'*” Additionally, PTEN detetion was shown to
`decrease CTL activity through expression of VEGF and re-
`cruitmentof inhibitery cells to the TME,4"49 Consequently,
`joss of PTEN was associated with resistance to anti-PD-1
`therapy in uterine leiamyosarcoma.’©° in addition, a PISK-y
`inhibitor was shown to decrease MDSCs in the TME and
`improve response to ICB in animal models.*5
`
`WNT/B-catenin signaling is another canonical oncogenic
`pathway that is involved in many essential cellular pro-
`cesses.’ Abnormalities In WNT/f-catenin signaling have
`been observed in multiple tumor histologies and are
`
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`Mechanismsof Resistance to Immune Checkpoint Blackade
`
`correlated with increased ceil invasiveness and metastatic
`potential." in addition to oncogenesis, constitutive WNT
`signaling with stabilization of B-catenin might be Involved in
`CB resistance through tumer T-cell exclusion. Spranger
`et a> showed a negative correlation between level of
`B-catenin and TILs, mediated by decreased expression of
`the cytokine CCL4 and inability to recruit CD103* DCs
`needed for T-cell priming.
`
`Indoleamine 2,3-dioxygenase 1 (IDOI) is a rate-limiting
`enzyme that converts tryptophan to its metabolite kynur-
`enine and has been associated with suppression of Teff
`function and resistance to ICB.**15° Additionally, accu-
`mulation of kynurenines and depletion of the essential
`amino acid tryptophan lead to immune suppression through
`T-cell anergy and apoptosis.’®” In one study, [DO-knockout
`mice showed improved overait survival
`(OS) with ICB
`compared with wild-type mice, highlighting the therapeutic
`potentia! of (DO inhibition.’
`Other Resistance Mechanisms
`
`Greater bacterial diversity and enrichment of specific spe-
`cies have shown association with improved resporise to ICB
`across multiple studies. Relative abundance of Faecali-
`bacterium and Ruminococcaceae were observed in re-
`sponders, whereas nonresponders were more likely to
`harbor Bacteroidales.\”!*73 Moreover, germ-free mice with
`fecal
`transplants from responders to ICB demonstrated
`improved outcomes with anti-PD-L1 therapy.'* Antibiotics
`can modify response to treatment by altering relative
`amounts of certain species, yielding either increased sus-
`ceptibility or increased resistance to ICB.7*"”? This asso-
`clation between intestinal microbiota and response to ICB is
`likely due to cross-reactivity between microbial and tumor
`antigens, enhancing DC activation, antigen presentation,
`and inflammatory cytokine production. ?78179
`THERAPEUTIC STRATEGIES TO OVERCOME RESISTANCE
`
`With recent insights gained into mechanisms of ICB re-
`sistance, combination strategies using multiple treatment
`modalities are emerging (Table 2). The rationale behindthis
`multimodal approach is based upon potential synergistic
`effects of targeting different
`immune escape pathways,
`resulting if improved response to iCB and better patient
`cuicormes,
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`There is now emerging evidence that chromatin remodeling
`is involved in sensitivity and resistance to ICB (Table 1). SWI/
`SNF complexes are chromatin remodelers and are fre-
`quently mutated in a variety of cancers.4® The marnmalian
`arlalogs of the SWI/SNF compiex, BRG1-associated factor
`Increasing Tumor Immunogenicity and T-Cell Priming
`(BAF), and polybromo-associated BAF (PBAF), are im-
`Oneofthefirst strategies used to bypass resistance employs
`portant tumor suppressors. Although these complexes are
`combination anti-CTLA-4 and anti-PD-1 treatment and has
`very similar, BAF exclusively contains ARIDLA/B subunits,
`already been approved in melanoma, colorectal cancer, and
`whereas ARID2, PBRM1, and BRD? subunits are unique to
`renal celf carcinoma, with many trials ongoing.!8°182
`PBAF.*® Loss-cf-function mutations in BAF and PBAF have
`Longer-term data in patients with advanced melanoma
`recently been shown to sensitize turers to PD-1 and CTLA-
`showed that the OS rate at 3 years with combined nivolu-
`4 blockade.One study demonstrated that inactivation of
`mab/ipilimumab treatment was 58%, with nivolumab
`PBAF subunits increased chromatin accessibility to tran-
`monotherapy was 52%, and with ipilimumab monotherapy
`scription regulator elements of IFN-y—inducible genes, with
`was 34%." Although the study was not powered to
`subsequent expression of CXCLOYCXCL1O cytokines and
`compare combination therapy versus nivolumab alone, OS
`recruitment of TILs.¥®? Another study showed that loss of
`was descriptively superior with dual blockade, The clinicat
`ARIDJIA increased microsatellite instability through de-
`benefit derived from ICB combination therapyis likely due to
`ficient recruitment of mismatch repair genes, enhancing
`complimentary mechanisms, as anti-CTLA-4 playsa role in
`
`mutational burden and=sensitizing tumors to PD-L1
`priming of T cells, whereas anti-PD-1 is involved in later
`blockade}
`reactivation of effector response, Additionally, anti-CTLA-4
`has been shown to deplete Tregs from the TME and en-
`hance CTiL-mediated antitumor immunity through broader
`antigen recognition,*+
`
`Under normal physiologic conditions, tumor inflammation
`causes hypoxia and ischemia, promoting generation of
`adenosine through dephosphorylation of adenosine mono-
`phosphate by CD73. Upon interaction of adenosine with the
`A2A receptor found on lymphocytes, effector function is
`suppressed,
`limiting additional tissue injury and collateral
`inflammatory damage."*5! Overexpression of CD73 has
`been associated with poor prognosis in multipte tumor types,
`encouraging metastasis and angiogenesis.°77 Moreover,
`high expression of CD73 promotes T-cell exhaustion and Is
`associated with resistance to ICB.?6170
`
`Within recent years, a connection between gut microbiome
`composition and response to CB has been uncovered.
`
`Combination chemotherapy with PD-1 blockade is approve!
`in NSCLC, with modest improvements tn progression-free
`survival and OS.145 Chemotherapylikely sensitizes tumors
`to ICB by increasing release of antigens upon cancercell
`death, leading to increased priming of CTLs.!®” Furthermore,
`chemotherapy has been associated with depletion of MDSCs
`and Tregs.!™ Raclation therapy is thought to function simi-
`larly, with increased antigen presentation secondary to turnor
`cell death, prometing an inflarmed TME and presumably
`synergy with ICB.7"? Additionally, radiation therapy increases
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