ebook img

SITC 2016 PDF

116 Pages·2016·8.44 MB·English
Save to my drive
Quick download
Download
Most books are stored in the elastic cloud where traffic is expensive. For this reason, we have a limit on daily download.

Preview SITC 2016

31st Annual Meeting and Associated Programs of the Society for Immunotherapy of Cancer (SITC 2016): part two: National Harbor, MD, USA. 9-13 November 2016 Citation Ager, C., M. Reilley, C. Nicholas, T. Bartkowiak, A. Jaiswal, M. Curran, T. C. Albershardt, et al. 2016. “31st Annual Meeting and Associated Programs of the Society for Immunotherapy of Cancer (SITC 2016): part two: National Harbor, MD, USA. 9-13 November 2016.” Journal for Immunotherapy of Cancer 4 (Suppl 1): 107-221. doi:10.1186/s40425-016-0173-6. http:// dx.doi.org/10.1186/s40425-016-0173-6. Published Version doi:10.1186/s40425-016-0173-6 Permanent link http://nrs.harvard.edu/urn-3:HUL.InstRepos:29739091 Terms of Use This article was downloaded from Harvard University’s DASH repository, and is made available under the terms and conditions applicable to Other Posted Material, as set forth at http:// nrs.harvard.edu/urn-3:HUL.InstRepos:dash.current.terms-of-use#LAA Share Your Story The Harvard community has made this article openly available. Please share how this access benefits you. Submit a story . Accessibility JournalforImmunoTherapyofCancer2016,4(Suppl1):73 DOI10.1186/s40425-016-0173-6 MEETING ABSTRACTS Open Access 31st Annual Meeting and Associated Programs of the Society for Immunotherapy of Cancer (SITC 2016): part two NationalHarbor,MD,USA.9-13November2016 Published:16November2016 Aboutthissupplement TheseabstractshavebeenpublishedaspartofJournalforImmunoTherapyofCancerVolume4Suppl1,2016.Thefullcontentsofthesupplement areavailableonlineathttp://jitc.biomedcentral.com/articles/supplements/volume-4-supplement-1.Pleasenotethatthisispart2of2. Combinations: Immunotherapy/ intratumoral Flt3L, although administration at both flanks is re- Immunotherapy quired for full effect. Intratumoral low-dose αCTLA-4, αPD-1, and α4-1BB at a single flank induces abscopal effects in 20 % of mice, and concurrent administration of CDG enhances systemic immun- P189 ity to cure up to 50 % of mice. We observe that the level of RationalcombinationsofintratumoralTcellandmyeloidagonists STING activation required to mediate rejection without inducing mobilizeabscopalresponsesinprostatecancer ulcerative toxicity is proportional to initial tumor size. Function- CaseyAger1,MatthewReilley2,CourtneyNicholas1,ToddBartkowiak1, ally, local STING activation complements intratumoral checkpoint AshvinJaiswal1,MichaelCurran1 modulation to reduce local MDSC infiltration, enhance CD8:Treg 1DepartmentofImmunology,UniversityofTexasMDAndersonCancer ratios, and downregulate the M2 macrophage marker CD206. In Center,Houston,TX,USA;2DepartmentofCancerMedicine,Universityof contrast, local Flt3L robustly enhances immune infiltration of TexasMDAndersonCancerCenter,Houston,TX,USA injected and distal tumors, but therapeutic benefit is attenuated Correspondence:CaseyAger([email protected]) due to concomitant induction of FoxP3+ Treg. JournalforImmunoTherapyofCancer2016,4(Suppl1):P189 Conclusions Intratumoral STING activation via CDG or DC expansion with Flt3L Background potentiatesthetherapeuticeffectsofsystemically-deliveredαCTLA-4, Despite the success of checkpoint blockade immunotherapy in αPD-1, and α4-1BB against multi-focal TRAMP-C2 prostate cancer. characteristically immunogenic cancers such as melanoma, these TheabscopalpotentialofCDGaloneisweak,incontrasttopriorob- antibodies remain ineffective against poorly T cell-infiltrated malig- servations, but combining CDG with low-dose checkpoint blockade nanciesincludingprostatecancer.Sensitizingthese“cold”tumorsto intratumorallycaninducesystemicimmunity,suggestinganalterna- immunotherapywillrequireinterventionswhichenhancetumoranti- tive approach for clinical implementation of combination immuno- genpresentationandTcellpriming,whilesuppressingmicroenviron- therapiesatreduceddoses. mentalsignalswhichconstrainTcellexpansion,survival,andeffector function independent of coinhibitory signaling. We investigated whetherintratumoraladministrationofeithertheSTINGagonistc-di- GMP (CDG) or dendritic cell (DC) growth factor Flt3-ligand can po- P190 tentiatethetherapeuticeffectsofTcellcheckpointmodulationwith Multi-genomereassortantdendriticcell-tropicvectorplatform αCTLA-4, αPD-1, and α4-1BB in a bilateral subcutaneous model of (ZVex®-Multi)allowsflexibleco-expressionofmultipleantigens prostate adenocarcinoma. Additionally, we tested whether intratu- andimmunemodulatorsforoptimalinductionofanti-tumorCD8+ moraldeliveryoflow-dosecheckpointmodulatorswithCDGatasin- Tcellresponses glelesioncanachieveabscopalcontrolofdistallesions. TinaCAlbershardt,AnshikaBajaj,JacobFArcher,RebeccaSReeves,Lisa Methods YNgo,PeterBerglund,JanterMeulen Male C57BL/6 mice were challenged subcutaneously on both flanks ImmuneDesign,Seattle,WA,USA withTRAMP-C2prostateadenocarcinoma,andtreatmentwasadmin- Correspondence:TinaCAlbershardt istered intraperitoneally and/or intratumorally for 3 doses every ([email protected]) 4 days, beginning on day 14 post-implantation for survival experi- JournalforImmunoTherapyofCancer2016,4(Suppl1):P190 mentsorday31forflowanalysisexperiments. Results Background Intratumoral delivery of STING agonist CDG alone potently rejects Inductionofimmuneresponsesagainstmultipleantigensexpressed all injected TRAMP-C2 tumors, but fails to generate systemic con- from conventional vector platforms is often ineffective for reasons trol of uninjected lesions. Systemic administration of αCTLA-4, notwellunderstood.Commonmethodsofexpressingmultipleanti- αPD-1, and α4-1BB cures 40 % of mice with bilateral TRAMP-C2, gens within a single vector construct include the use of fusion pro- and concurrent administration of CDG at one or both flanks en- teins, endoprotease cleavage sites, or internal ribosome entry sites. hances survival to 75 %. Similar effects are observed with These methods often lead to decreased expression of antigens-of- ©TheAuthor(s).2016OpenAccessThisarticleisdistributedunderthetermsoftheCreativeCommonsAttribution4.0 InternationalLicense(http://creativecommons.org/licenses/by/4.0/),whichpermitsunrestricteduse,distribution,and reproductioninanymedium,providedyougiveappropriatecredittotheoriginalauthor(s)andthesource,providealinkto theCreativeCommonslicense,andindicateifchangesweremade.TheCreativeCommonsPublicDomainDedicationwaiver (http://creativecommons.org/publicdomain/zero/1.0/)appliestothedatamadeavailableinthisarticle,unlessotherwisestated. JournalforImmunoTherapyofCancer2016,4(Suppl1):73 Page108of221 interest and/or reduced induction of T cell responses against the Methods encoded antigens. Circumventing these limitations, we have devel- To assess the effect of combined blockade of NKG2A/HLA-E and oped a novel production process for our integration-deficient, den- PD-1/PD-L1 in vivo, anti-mouse NKG2A and PD-1 antibodies were dritic cell-targeting lentiviral vector platform, ZVex, enabling highly used in mice engrafted with A20 mouse B lymphoma cell line. flexible and effective multigene delivery in vivo, making it possibly For in vitro assays, anti-PD-L1 antibody durvalumab, and monali- themostversatilevectorplatformintheindustry. zumab were tested in human PBMC staphylococcal enterotoxin b Methods assays. Up to five vector genome plasmids, each encoding one full-length Results antigen or immuno-modulator, were mixed with four constant plas- Whenculturedinvitro,theA20cellsexpressligandsforPD-1butnot mids,eachencodingvectorparticleproteins,priortotransfectionof forNKG2A.ExposuretoIFN-γinvitro,orsubcutaneousinjectioninto production cells. Due to the propensity of lentiviruses forming gen- mice, induced expression of Qa-1b, resulting in a tumor model co- omic reassortants, the resulting vector preparations hypothetically expressing PD-L1 and Qa-1b. Monotherapy with PD-1 or NKG2A contain a mix of homozygous and heterozygous vector particles. blockers resulted in moderate anti-tumor efficacy while treatment qRT-PCR was used to determine total and antigen-specific titers of with combination of NKG2A and PD-1 blockers resulted in a signifi- ZVex-Multivectors,definedasvectorgenomecounts.Micewereim- cantly higher anti-tumor immunity, and an increased rate of munizedwithZVex-Multi vectorsor monozygousvectorsexpressing complete tumor regression. Depletion of either NK, or CD8+ T cells, multipleantigensfromthesamebackbonetocompareimmunogen- or IFN-γ was enough to abrogate the efficacy of PD-1 and NKG2A icityviaintracellularcytokinestaining.Twotumormodelswereused blockade, indicatingthatbothoftheseeffectorpopulationscontrib- to evaluate therapeutic efficacy: 1) a B16 melanoma model, where uteto theefficacyofthecombinationtreatment.Tofurtherexplore tumorcellswereinoculatedintheflankandmeasured2–3timesper this possibility and to assess the potential therapeutic relevance in week;and2)ametastaticCT26coloncarcinomamodel,wheretumor humans,well-validatedPBMC-basedassayswereusedwhichshowed cellswereinoculatedintravenously,andlungnoduleswereenumer- thatblockingbothaxeswithacombinationofdurvalumabandmon- ated17–19dayspost-tumorinoculation. alizumabledtoincreasedproductionofcytokinesbybothTandNK Results cells. Furthermore, the magnitude of the increase in cytokine secre- Titrations by qRT-PCR of multiple ZVex-Multi vector lots demon- tionwasdependentontheproductionofhighlevelsofIFN-γ.Since stratedthatproductionyieldsofZVex-Multiexpressinguptofourdif- IFN-γ is known to induce HLA-E this suggests that blockade of ferenttumor-associatedantigens(e.g.,NY-ESO-1,MAGE-A3)andtwo NKG2A could have a beneficial role in activation of immune cells immuno-modulators (e.g., IL-12, anti-CTLA-4 or anti-PD-L1) were throughthecombinedblockadeofPD-1/PD-L1. highly reproducible. Compared to mice immunized with vectors ex- Conclusions pressing multiple antigens from the same backbone, mice immu- Together, these data indicate that blocking NKG2A in conjunction nizedwithZVex-Multi vectorsconsistentlydevelopedT cells against withPD-1/PD-L1checkpointinhibitorsprovidesincreasedanti-tumor alltargetedTAAsandexhibitedimprovedtumorgrowthcontroland efficacy mediated by IFN-γ and support the rationale for assessing survival. thiscombinationinclinicaltrials. Conclusions ZVex-Multi is a next generation DC-tropic vector platform designed to overcome limitations of single-genome vector platforms with re- P192 specttoefficientco-expressionofanycombinationofdesiredgenes. Pharmacokineticsandimmunogenicityofpembrolizumabwhen Unlikeothervectorplatforms,ZVex-Multieliminatesmultiplecloning givenincombinationwithipilimumab:datafromKEYNOTE-029 steps modifying the vector backbone, which can often result in un- MichaelBAtkins1,MatteoSCarlino2,AntoniRibas3,JohnAThompson4, predictableexpressionpatternsofcodedgeneproducts.Itsversatility ToniKChoueiri5,FStephenHodi5,Wen-JenHwu6,DavidFMcDermott7, andagilitymakesZVex-Multipotentiallythebest-in-classvectorplat- VictoriaAtkinson8,JonathanSCebon9,BernieFitzharris10,MichaelB form for co-expression of multiple tumor antigens and immuno- Jameson11,CatrionaMcNeil12,AndrewGHill13,EricMangin14,Malidi modulators for enhanced cancer immunotherapy against a broad Ahamadi14,MariannevanVugt15,MariëllevanZutphen15,Nageatte rangeoftumors. Ibrahim14,GeorginaVLong16 1Georgetown-LombardiComprehensiveCancerCenter,Washington,DC, USA;2WestmeadandBlacktownHospitals,MelanomaInstituteAustralia, P191 andtheUniversityofSydney,Westmead,NewSouthWales,Australia; NK,TcellsandIFN-gammaarerequiredfortheanti-tumorefficacy 3UniversityofCalifornia,LosAngeles,CA,USA;4Universityof ofcombination-treatmentwithNKG2AandPD-1/PD-L1checkpoint Washington,Seattle,WA,USA;5Dana-FarberCancerInstitute/Brigham inhibitorsinpreclinicalmodels andWomen’sHospital,HarvardUniversity,Boston,MA,USA;6University CarolineDenis1,HormasGhadially2,ThomasArnoux1,FabienChanuc1, ofTexasMDAndersonCancerCenter,Houston,TX,USA;7BethIsrael NicolasFuseri1,RobertWWilkinson2,NicolaiWagtmann1,YannisMorel1, DeaconessMedicalCenter,Boston,MA,USA;8GallipoliMedicalResearch PascaleAndre1 Foundation,GreenslopesPrivateHospital,andtheUniversityof 1InnatePharma,Marseille,Provence-Alpes-Coted'Azur,France; Queensland,Greenslopes,Queensland,Australia;9OliviaNewton-John 2MedImmune,Cambridge,England,UK CancerResearchInstitute,Heidelberg,Victoria,Australia;10Canterbury Correspondence:PascaleAndre([email protected]) DistrictHealthBoard,ChristchurchHospital,Christchurch,NewZealand; JournalforImmunoTherapyofCancer2016,4(Suppl1):P191 11WaikatoHospitalRegionalCancerCentre,Hamilton,NewZealand; 12RoyalPrinceAlfredHospital,MelanomaInstituteAustralia,the Background UniversityofSydney,andChrisO’BrienLifehouse,Camperdown,New Monalizumab (IPH2201) is a first-in-class humanized IgG target- SouthWales,Australia;13TasmanOncologyResearch,SouthportGold 4 ing NKG2A, which is expressed as heterodimer with CD94 on the Coast,Queensland,Australia;14Merck&Co.,Inc.,Kenilworth,NJ,USA; surface of NK, γδT and tumor infiltrating CD8+ T cells. This inhibi- 15QuantitativeSolutions,aCertaracompany,Oss,Netherlands; tory receptor binds to HLA-E in humans and to Qa-1b in mice. 16MelanomaInstituteAustralia,theUniversityofSydney,MaterHospital, HLA-E is frequently up-regulated on cancer cells, protecting from andRoyalNorthShoreHospital,Wollstonecraft,NewSouthWales, killing by NKG2A+ cells. Monalizumab blocks binding of CD94- Australia NKG2A to HLA-E, reducing inhibitory signaling thereby enhancing Correspondence:MichaelBAtkins([email protected]) NK and T cell responses. PD-1/PD-L1 inhibitors are successfully JournalforImmunoTherapyofCancer2016,4(Suppl1):P192 being used to treat patients with a wide variety of cancers. Com- bined blockade of NKG2A/HLA-E and PD-1/PD-L1 may be a prom- Background ising strategy to better fight cancer by activating both the The pharmacokinetics of pembrolizumab given as monotherapy adaptive and innate immune systems. are well characterized. Consistent with other monoclonal JournalforImmunoTherapyofCancer2016,4(Suppl1):73 Page109of221 antibodies, pembrolizumab has low clearance (0.202 L/day), lim- ited central (3.53 L) and peripheral (3.85 L) volume of distribu- tion, and low variability in the central volume of distribution (19 % coefficient of variation). Pembrolizumab monotherapy has low immunogenicity potential, with an observed incidence of treatment-emergent anti-drug antibodies (ADA) of<1 %. We present data on the pharmacokinetics and immunogenicity of pembrolizumab when given in combination with ipilimumab in the phase I KEYNOTE-029 study. Methods KEYNOTE-029 included 2 cohorts that assessed the safety and antitumor activity of pembrolizumab plus ipilimumab: a safety run-in that included patients with advanced melanoma or renal cell carcinoma (RCC) (N=22) and a melanoma expansion cohort (N=153). In both cohorts, patients received 4 doses of pembroli- zumab 2 mg/kg plus ipilimumab 1 mg/kg Q3W followed by pembrolizumab2mg/kgQ3Wforupto2years.Pembrolizumabserum concentrationwasquantifiedwithanelectrochemiluminescence-based immunoassay(lowerlimitofquantitation,10ng/mL).Avalidatedbridg- ing electrochemiluminescence immunoassay using a standard 3- tiered approach (drug tolerance level, 124 μg/mL) was used to detect ADA in serum. Results Across cohorts, 175 patients received pembrolizumab plus ipili- mumab: 165 with melanoma and 10 with RCC. At least 1 evalu- Fig.2(abstractP192).Observedpembrolizumabserum able sample for pharmacokinetic assessment was available for all concentrationsfrompatientswithmelanomatreatedwith 10 patients with RCC and 162 patients with melanoma. The pre- pembrolizumabplusipilimumabinrelationtothepredicted dose serum concentration versus time profiles for pembrolizumab concentrationinterval(gray)forpembrolizumab2mg/kgQ3W were similar in patients with RCC and melanoma (Fig. 1). Ob- monotherapy(logscale) served serum concentrations were within the range predicted for pembrolizumab 2 mg/kg Q3W given as monotherapy (Fig. 2). Of the 160 patients with melanoma who provided postdose ADA samples, 156 (97.5 %) were negative, 2 (1.3 %) were inconclusive, and 2 (1.3 %) were positive for treatment-emergent ADA. Best P193 overall response in the ADA-positive patients was stable disease EstablishingamodelforsuccessfulimmunotherapywithT-Vec in one and progressive disease in the other. No patient with RCC combinedwithBRAFinhibitionandanti-PD-1ingenetically had treatment-emergent ADA. engineeredmurinemelanoma Conclusions RobynGartrell1,ZoeBlake1,InesSimoes2,YichunFu1,TakuroSaito3, The addition of ipilimumab does not appear to impact pembrolizu- YingzhiQian1,YanLu1,YvonneMSaenger4 mabserumconcentration or increasethe riskof developingADAin 1ColumbiaUniversityMedicalCenter,NewYork,NY,USA;2Institut patientswithadvancedmelanomaorRCC. d'InvestigacionsBiomediquesAugustPiiSunyer,Barcelona,Catalonia, TrialRegistration Spain;3IcahnSchoolofMedicineatMountSinai,NewYork,NY,USA; ClinicalTrials.govidentifierNCT02089685. 4NewYorkPresbyterian/ColumbiaUniversityMedicalCenter,NewYork, NY,USA Correspondence:ZoeBlake([email protected]) JournalforImmunoTherapyofCancer2016,4(Suppl1):P193 Background Talimogene laherparepvec (T-Vec) is the first oncolytic virus to be U.S. Food and Drug Administration (FDA) approved for the treat- ment of cancer. T-Vec, a modified herpes simplex type I (HSV I) virus, has two proposed mechanisms of action: direct cell lysis and immune activation. Combination immunotherapy using T-Vec and checkpoint blockade has shown promise in clinical trials. In preliminary work, our laboratory has shown that T-Vec causes up- regulation of programmed cell death protein 1 (PD-1) on infiltrat- ing T cells in mice, suggesting potential synergy of T-Vec and anti-PD-1 (αPD-1). Methods In a temporally and spatially regulated murine model of BRAFCA PTEN−/− spontaneous melanoma [1], tumors are induced on right flank. When tumors reach >5 mm in diameter, mice are random- ized into 6 treatment groups comparing combinations of BRAF inhibition (BRAFi), αPD-1, and T–Vec (Table 1). Tumor growth is Fig.1(abstractP192).Arithmeticmean(SE)predoseserum measured twice a week until end of study. Flow cytometry is per- concentration-timeprofileofpembrolizumabfollowingmultiple formed on tumor, lymph node, and spleen to assess immune dosesofpembrolizumabplusipilimumab(linear-linearscale) microenvironment. JournalforImmunoTherapyofCancer2016,4(Suppl1):73 Page110of221 Results Mean tumor volume and survival was plotted to compare groups (Figs. 3 and 4). Mice treated with triple combination have decreased tumor growth. Mice treated with combination T-Vec+BRAFi with or without αPD-1 have longer survival com- pared to mice treated with control or single drug arms. Flow cytometry shows increase in percent CD3+/CD45+ cells in tu- mors of mice treated with combination αPD-1+T-Vec compared to the control and single drug arms. Percent CD8+/CD3+ cells in tumors treated with immunotherapy appears to be increased compared to the control and BRAFi only group (Fig. 5). Add- itionally, percent of FOXP3+/CD4+ cells in tumors appears to be decreased in groups receiving T-Vec (Fig. 6) while no change in FOXP3+/CD4+ populations was observed in tumors from groups receiving αPD-1 without T-Vec or in draining lymph node or spleen. Conclusions Fig.4(abstractP193).Survivalcomparisonoftreatmentgroups Initial findings show that combination therapy of BRAFi+αPD-1 +T-Vec is more effective than any single treatment. Combination immunotherapy increases infiltration of T cells into tumor. Furthermore, oncolytic virus appears to decrease regulatory T cells infiltrating tumor. This study is ongoing and further analysis will continue as we further evaluate the immune microenvironment using flow cytometry and immunohistochemistry. Acknowledgements ThestudywasfundedbytheMelanomaResearchAllianceandAmgen (Amgen-CUMC-MRAEstablishedInvestigatorAcademic-IndustryPartnership Award). Reference 1.Dankort, Curley, Cartlidge, et al.: Braf(V600E) cooperates with Pten loss to induce metastatic melanoma. Nature Genetics 2009, 41:544–552. Table1(abstractP193).Treatmentgroups Group Treatment Group1(Red) ControlChow+IP2A3+ITPBS Group2(Orange) BRAFiChow+IP2A3+ITPBS Group3(Yellow) BRAFiChow+Ipα-PD1+ITPBS Fig.5(abstractP193).FlowcytometrydataofCD8+cellsperCD3 +cellpopulations Group4(Green) BRAFiChow+IP2A3+ITT-Vec Group5(Blue) BRAFiChow+IPα-PD1+ITT-Vec Group6(Purple) ControlChow+IPα=PD1+ITT-Vec IPintraperitoneal,ITintratumoral,BRAFibriefinhibiotor,α-PD1anti programmedcelldeath1,T-VectalimogeneLeherparepvec Fig.6(abstractP193).FlowcytometrydataofCD4+/FOXP3+cells Fig.3(abstractP193).Tumorvolumecomparisonofallmice perCD4+cellpopulations JournalforImmunoTherapyofCancer2016,4(Suppl1):73 Page111of221 P194 Background Phosphatidylserinetargetingantibodyincombinationwith In the tumor microenvironment, T cell inhibitory checkpoint recep- checkpointblockadeandtumorradiationtherapypromotesanti- torstriggersignalsthatsuppressTcelleffectorfunction,resultingin canceractivityinmousemelanoma tumorimmuneevasion.Clinicalantibodiesblockingoneofthesere- SadnaBudhu1,OlivierDeHenau1,RobertaZappasodi1,Kyle ceptors,PD-1, yield positiveresponses inmultiplecancers;however, Schlunegger2,BruceFreimark2,JeffHutchins2,ChristopherABarker1, their efficacy is limited. Simultaneously targeting more than one in- JeddDWolchok1,TahaMerghoub1 hibitorycheckpointreceptorhasemergedasapromisingtherapeutic 1MemorialSloanKetteringCancerCenter,NewYork,NY,USA;2Peregrine strategy.Insupportofthisconcept,micedeficientinPD-1andLAG- Pharmaceuticals,Inc.,Tustin,CA,USA 3,aninhibitorycheckpointreceptoroftenco-expressedwithPD-1in Correspondence:SadnaBudhu([email protected]) the tumor microenvironment, exhibit enhanced anti-tumor activity. JournalforImmunoTherapyofCancer2016,4(Suppl1):P194 Here, we demonstrate increased anti-tumor efficacy of a combined anti–human PD-1 (hPD-1) and anti–human LAG-3 (hLAG-3) therapy Background usingfullyhumanmonoclonalantibodiesindualhumanizedPD-1x Phosphatidylserine(PS)isaphospholipidthatisexposedonthesur- LAG-3mice.Thepharmacokineticsandtoxicologyof thenovelanti- faceofapoptoticcells,sometumorcellsandtumorendothelium.PS hLAG-3 antibody were assessed in non-human primates to support hasbeenshowntopromoteanti-inflammatoryandimmunosuppres- clinicaldevelopment. sive signals in the tumor microenvironment. Antibodies that target Methods PShavebeenshowntoreactivateanti-tumorimmunitybyrepolariz- REGN2810, a high affinity anti-hPD-1 monoclonal antibody that ingtumorassociatedmacrophagestoaM1-likephenotype,reducing blocks PD-1 interaction with PD-L1 and PD-L2, and a novel high thenumberofMDSCsintumorsandpromotethematurationofden- affinity monoclonal anti–hLAG-3 antibody, which blocks the LAG- driticcellsintofunctionalAPCs.InaB16melanomamodel,targeting 3/MHC II interaction were generated. Dual humanized PD-1 x PS in combination with immune checkpoint blockade has been LAG-3 mice were engineered by replacing the extracellular do- shown to have a significantly greater anti-cancer effect than either mains of mouse Pdcd1 and Lag3 with the corresponding regions agent alone. This combination was shown to enhance CD4+ and of hPD-1 and hLAG-3 and were used for testing antibody efficacy CD8+ T cell infiltration and activation in the tumors of treated ani- in a MC38.ova syngeneic tumor model. Expression of humanized mals. Radiation therapy is an effective focal treatment of primary PD-1 and LAG-3 were analyzed by flow cytometry. Binding of solidtumors, butislesseffectiveintreatingmetastatic solidtumors hLAG-3 to mouse MHC II was confirmed with a cell adhesion asamonotherapy.Thereisevidencethatradiationinducesimmuno- assay, and binding of hPD-1 to mouse PD-L1 was confirmed genictumorcelldeathandenhancestumor-specificTcellinfiltration using surface plasmon resonance. The pharmacokinetics of anti- inirradiatedtumors.Inaddition,theabscopaleffect,aphenomenon hLAG-3 antibody following a single i.v. dose, and the safety pro- in which tumor regression occurs outside the site of radiation ther- file in a 4-week weekly i.v. dose regimen of up to 50 mg/kg/ apy,hasbeenobservedinbothpreclinicalandclinicaltrialswiththe dose, were determined in cynomolgus monkeys. combinationofradiationtherapyandimmunotherapy. Results Methods TreatmentofMC38.ovatumor-bearinghumanizedmicewithacom- Weexaminedtheeffectsofcombiningtumorradiationtherapywith bination of anti-hPD-1 and anti-hLAG-3 antibodies triggered activa- an antibody that targets PS (1 N11) and an immune checkpoint tion of intratumoral and peripheral T cells. Importantly, the blockade (anti-PD-1) usingthe mouseB16 melanoma model. Tumor combination treatment exhibited an additive, dose dependent anti- surfaceareaandoverallsurvivalofmicewereusedtodetermineeffi- tumoreffectcomparedtotherespectivemonotherapies.Anti-hLAG- cacyofthecombinations. 3 antibody pharmacokinetics in cynomolgus monkeys followed a Results standardmean concentration-time profile characterized by an initial We examined the expression of PS on immune cells infiltrating B16 briefdistributionphaseandalinearbetaeliminationphase.Exposure melanomas.CD11b+myeloidcellsexpressedthehighestlevelsofPS toanti-hLAG-3increasedinadose-proportionalmanner,withelimin- ontheirsurfacewhereasTcellsandB16tumorcellsexpresslittleto ationhalf-livesrangingfrom10.8to11.5days.Anti-hLAG-3antibody noPS.ThesedatasuggestthattargetingPSinB16melanomawould was well tolerated, and no-observed-adverse-effect level (NOAEL) induce a pro-inflammatory myeloid tumor microenvironment. We couldbeestablishedupto50mg/kg. hypothesizethattherapiesthatinduceapoptoticcelldeathontumor Conclusions cells would enhance the activity of PS-targeting antibodies. We Preclinical anti-tumor efficacy of combined REGN2810 and anti- thereforeexaminedtheeffectsofcombiningaPS-targetingantibody hLAG-3antibodytreatment,togetherwithfavorablepharmacokinetic withlocaltumorradiation.WefoundthatthePS-targetingantibody and safety data for anti-hLAG-3 antibody in cynomolgus monkeys, synergizes with both anti-PD-1 and radiation therapy to improve support clinical development of this cancer combination anti-cancer activity and overall survival. In addition, the triple com- immunotherapy. binationofthePS-targetingantibody,tumorradiationandanti-PD-1 treatmentdisplayedevengreateranti-cancerandsurvivalbenefit. Conclusions P196 Thisfindinghighlightsthepotentialofcombiningthesethreeagents CombinationofPD-L1blockadewithoncolyticvaccinesre-shapes toimproveoutcomeinpatientswithadvanced-stagemelanomaand thefunctionalstateoftumorinfiltratinglymphocytes may inform the design of future clinical trials with PS targeting in CristianCapasso1,FedericaFrascaro2,SaraCarpi3,SiriTähtinen1,Sara melanomaandothercancers. Feola4,ManlioFusciello1,KaritaPeltonen1,BeatrizMartins1,Madeleine Sjöberg1,SariPesonen5,TuuliRanki5,LukaszKyruk1,ErkkoYlösmäki1, VincenzoCerullo1 P195 1UniversityofHelsinki,Helsinki,Uusimaa,Finland;2UniversityofSiena, Anovelanti-humanLAG-3antibodyincombinationwithanti- Supersano(LE),Puglia,Italy;3UniversityofPisa,Pisa,Toscana,Italy; humanPD-1(REGN2810)showsenhancedanti-tumoractivityin 4UniversityofNapoliFedericoII,Helsinki,Uusimaa,Finland;5PeptiCRAd PD-1xLAG-3dual-humanizedmiceandfavorablepharmacokinetic Oy,Helsinki,Uusimaa,Finland andsafetyprofilesincynomolgusmonkeys Correspondence:CristianCapasso([email protected]) ElenaBurova,OmairaAllbritton,PeterHong,JieDai,JerryPei,MattLiu, JournalforImmunoTherapyofCancer2016,4(Suppl1):P196 JoelKantrowitz,VenusLai,WilliamPoueymirou,DouglasMacDonald,Ella Ioffe,MarkusMohrs,WilliamOlson,GavinThurston Background Regeneron,Tarrytown,NY,USA Theimmunologicalescapeoftumorsrepresentsoneofthemainob- Correspondence:ElenaBurova([email protected]) stacles to the treatment of malignancies. The blockade of PD-1 or JournalforImmunoTherapyofCancer2016,4(Suppl1):P195 CTLA-4 receptors represented a milestone in the history of JournalforImmunoTherapyofCancer2016,4(Suppl1):73 Page112of221 immunotherapy.However,immunecheckpointinhibitorsseemtobe P197 effective in specific cohorts of patients. It has been proposed that Invitroevaluationofimmunotherapyprotocolsthroughalabel- their efficacy relies on the presence of an immunological response. freeimpedance-basedtechnologyallowsdynamicmonitoringof Thus,wehypothesizedthatdisruptionofthePD-L1/PD-1axiswould immuneresponseandreagentefficacy synergizewithouroncolyticvaccineplatformPeptiCRAd. FabioCerignoli,BiaoXi,GarretGuenther,NaichenYu,LincolnMuir, Methods LeynaZhao,YamaAbassi We used murine B16OVA in vivo tumor models and flow cytometry ACEABiosciencesInc.,SanDiego,CA,USA analysistoinvestigatetheimmunologicalbackground. Correspondence:FabioCerignoli([email protected]) Results JournalforImmunoTherapyofCancer2016,4(Suppl1):P197 First, we found that high-burdenB16OVA tumors were refractory to combination immunotherapy. However, with a more aggressive Background schedule,tumorswithalowerburdenweremoresusceptibletothe Invitrocharacterization of reagent efficacy in the context of can- combination of PeptiCRAd and PD-L1 blockade. The therapy signifi- cer immunotherapy is a necessary step before moving to more cantly increased the median survival of mice (Fig. 7). Interestingly, expensive animal models and clinical studies. However, current the reduced growth of contralaterally injected B16F10 cells sug- in vitro assays like Chromium-51, ATP-based luminescence or flow gested the presence of a long lasting immunological memory also cytometry are either difficult to implement in high throughput against non-targeted antigens. Concerning the functional state of environments or are mainly based on endpoint methodologies tumor infiltrating lymphocytes (TILs), we found that all the immune that are unable to capture the full dynamic of the immune re- therapies would enhance the percentage of activated (PD-1pos TIM- sponse. Here, we present the adaptation of an impedance-based 3neg) T lymphocytes and reduce the amount of exhausted (PD-1pos platform to monitor cytotoxic activity of immune cells activated TIM-3pos) cells compared to placebo. As expected, we found that trough different means. PeptiCRAdmonotherapycould increase the numberofantigenspe- Methods cific CD8+ T cells comparedto other treatments. However, only the Impedance technology detects cell death and proliferation of ad- combinationwithPD-L1blockadecouldsignificantlyincreasethera- herent cells by measuring changes in conductance of microelec- tio between activated and exhausted pentamer positive cells (p= trodes embedded in 96 and 384-wells cell culture plates. We 0.0058),suggestingthatbydisruptingthePD-1/PD-L1axiswecould utilized adherent and B cell leukemia/lymphoma cell lines as well decreasetheamountofdysfunctionalantigenspecificTcells.Weob- as primary tumor cells as in vitro models for immunotherapy re- served that the anatomical location deeply influenced the state of agent evaluation. We seeded the cells on electrodes coated 96- CD4+ and CD8+ T lymphocytes. In fact, TIM-3 expression was in- well plates and monitored cell adhesion and proliferation for creasedby2foldonTILscomparedtosplenicandlymphoidTcells. 24 hours. The following day effector cells were added at multiple In the CD8+ compartment, the expression of PD-1 on the surface effector:target ratios in presence of BiTEs antibodies and/or anti seemedtoberestrictedtothetumormicro-environment,whileCD4 PD-1/PD-L1 antibodies. Impedance signal was monitored for up + T cells had a high expression of PD-1 also in lymphoid organs. to seven days. Control wells were set up with effector cells only Interestingly, we found that the levels of PD-1 were significantly or with target plus effector cells but without antibodies. We higher on CD8+ T cells than on CD4+ T cells into the tumor micro- adapted such adhesion-based technology to monitor non- environment(p<0.0001). adherent B-leukemia/lymphoma cells, by developing a strategy Conclusions where the wells are coated with an anti-CD40 antibody. The coat- In conclusion, we demonstrated that the efficacy of immune check- ing allows specific adhesion and retention of B cells and meas- point inhibitors might be strongly enhanced by their combination urement of changes in impedance that are proportional to cell withcancervaccines.PeptiCRAdwasabletoincreasethenumberof number. antigen-specific T cells and PD-L1 blockade prevented their exhaus- Results tion, resulting in long-lasting immunological memory and increased Using increasing concentrations of EpCAM/CD3 BiTE, we demon- mediansurvival. strated the suitability of an impedance-based approach to quantita- tively monitor the efficacy of immune cells-mediated cancer cell killing both under different effector:target ratios and antibody con- centrations.Combinationtreatmentswithcheckpointreducedtiming andincreasedamountofkilledcancercells.Similarresultswerealso obtained with engineered CAR-T cells against CD19 or NK cell lines, demonstratingspecifickillingoftumorBcellsatveryloweffector:tar- getratios.Theresultswerealsoconfirmedbyflowcytometry. Conclusions Overall, our results demonstrate the value of an impedance-based approach in measuring the cytotoxic response across the temporal scale, an aspect that is otherwise very difficult to assess with more canonical end point assays. Furthermore, the availability of 384-well format and minimal sample handling place the technology in an ideal spot for applications in large reagent validation screening or personalized medicine, like therapeutic protocol validation directly onpatientsamples. P198 Tumornecrosisfactoralphaandinterleukin-2expressing adenovirusplusPD-1blockadeasaboostforTcelltherapyinthe contextofsolidtumortherapies VíctorCervera-Carrascón1,MikkoSiurala1,JoãoSantos1,RiikkaHavunen2, SuviParviainen1,AkseliHemminki1 Fig.7(abstractP196).SurvivalofC57micebearingB16OVA 1TILTBiotherapeutics,Helsinki,Uusimaa,Finland;2UniversityofHelsinki, tumorsandtreatedonday6post-implantationwitheitherPBS, Helsinki,Uusimaa,Finland PDL1blockade,OVA-targetingPeptiCRAdorthecombinationof Correspondence:VíctorCervera-Carrascón([email protected]) PDL1-blockadeandOVA-PeptiCRAd. JournalforImmunoTherapyofCancer2016,4(Suppl1):P198 JournalforImmunoTherapyofCancer2016,4(Suppl1):73 Page113of221 Background IMM-101, no response (initial SD). 2011 given Ipilimumab. Patient Becauseoftheimmunosuppressivetumormicroenvironment,theim- 2: 2011 50 F axillary lump removed, melanoma (no primary). munesystemisunabletodevelopeffectiveresponsesagainsttumor Concomitant mediastinal, lung, gastric and peritoneal deposits. cells. This phenomenon also acts against the effectiveness of adop- Gastric surgery, decarbazine. Commenced IMM-101 with cyber- tiveTcelltherapy.Inordertoovercomethissituationinthetumor, knife to lung lesion. 2013 Small bowel obstruction from new dis- anattractivetherapeuticcombinationisthecombinationofoncolytic ease. Started ipilimumab. Patient 3: 2014 79 M melanoma, left viruses and immune checkpoint inhibitors. In this case, besides the cheek, BT 2.4 mm. Regional lymph node recurrence, treated with last two therapies mentioned above, combinations with T cell ther- a left neck dissection in April 2014. Developed paracardiac apy were also included. The virus used was engineered to express nodes, adrenal, lung and multiple large subcutaneous deposits. tumor necrosis factor α (TNFα) and interleukin (IL)-2, two cytokines Commenced IMM-101 with initial shrinkage. However, new large that will boost the immunogenicity of the virus and thus its antitu- subcutaneous lesions. Commenced pembrolizumab. morproperties.Ontheotherhand,theuseofanti-PD-1willavoidex- Results haustionontumorinfiltratingTcellsandhenceremovethebarriers Patient 1 - CR on Pet CT, maintained through 2016. Patient 2 - CR thatcoulddampenthedesiredimmuneresponseagainstthetumor. maintainedfor2years.Patient3-CRofsubcutaneousdepositsfour Methods daysafterfirstinjection. In the study of the antitumor effect of this three armed treat- Conclusions ment we used an in vivo model of subcutaneous B16-OVA TheCRratetoCPI’sisdisappointing,<1%forIpilimumab.PDL-1ex- melanoma-bearing mice. Two experiments were carried out; the pressionispredictiveforPD-1responsesandalthoughCPIcombina- first one (n=47) to establish the differences between the triple, tions are clearly needed, most are very toxic. IMM-101 is relatively double, and single armed combination therapies and the second free of toxicity, enhances PD-1expressionin pre-clinical models but experiment (n=84) was focused on study the differences be- mayalsoprimetumourresponsetocheckpointinhibitorsbyitsac- tween the groups that showed the best outcomes in the first tion on macrophage function. Based on these observations, we one and also optimize viral and anti-PD-1 administration regimes. speculatethatIMM-101primesfor CPI’s and propose a trialpriming Results withIMM-101,followedbyanti-PD-1antibodies. Preliminaryresultsshowastatisticallysignificantpositiveeffectcom- ing out from the combination of virus therapy and immune check- References point blockade with regard to both tumor progression and overall 1. FowlerD,etal.:MycobacteriaactivateγδT-cellanti-tumourresponses survival, with up to 43 % complete tumor regression achieved in viacytokinesfromtype1myeloiddendriticcells:amechanismof someofthegroupsafter96daysposttreatment.Ontheotherhand, actionforcancerimmunotherapy.CeII2012,61(4):535–547. the effect of adoptive cell therapy in this combination is not com- 2. Stebbing J, et al.: An intra-patient placebo-controlled phase I trial to pletelyclear.Moreresultswillbepresentedafteranalyzingbiological evaluatethesafetyandtolerabilityofintradermalIMM-101inmelan- samplescollectedduringbothexperiments. oma.AnnOncol2012,23(5):1314–1319. Conclusions 3. Dalgleish,etal.:Randomisedopen-label,phaseIIstudyofGemcitabine Preclinical studies are a key step to detect which combinations are withandwithoutIMM-101foradvancedpancreaticcancer(IMAGE-1 moresuitableforsuccessinhumantrials.Inthisstudywedeveloped Trial).BJC2016,inpress. arationaleforthecombinationrelyingontwoconcepts:tomakesi- lent tumors more visible to the immune system and to counter im- munosuppressivemechanismstounleashthefullpotentialofTcells P200 against the tumor, rendering in a modification of the tumor micro- Immunologicalimpactofcheckpointblockadeondendriticcell environmentthatmakesitmoresusceptibleforTcellmediatedkill- drivenTcellresponses:acautionarytale ing. According to the results displayed from these experiments, the MarkDeBenedette,AnaPlachco,AliciaGamble,ElizabethWGrogan, combinationofthisgeneticallymodifiedadenovirusandPD-1block- JohnKrisko,IrinaTcherepanova,CharlesNicolette ade is an efficient combination to be considered for future applica- ArgosTherapeuticsInc.,Durham,NC,USA tioninhumans. Correspondence:MarkDeBenedette ([email protected]) JournalforImmunoTherapyofCancer2016,4(Suppl1):P200 P199 IMM-101primesforincreasedcompleteresponsesfollowing Background checkpointinhibitorsinmetastaticmelanoma;3casereports AGS-003 is an individualized, autologous, tumor antigen-loaded, AngusDalgleish1,SatvinderMudan2 dendritic cell (DC) immunotherapy currently in phase III develop- 1StGeorge'sUniversityofLondon,London,UK;2TheRoyalMarsden ment for the treatment of metastatic renal cell carcinoma (mRCC) HospitalandImperialCollegeLondon,London,UK in combination with standard-of-care. Antibodies to PD-1 on acti- Correspondence:AngusDalgleish([email protected]) vated T cells or PD-L1 expressed on APCs have now been ap- JournalforImmunoTherapyofCancer2016,4(Suppl1):P199 proved for treatment of several cancer indications including RCC. While there is a strong mechanistic rationale for the potential Background synergy of these agents in combination, data supporting the im- IMM-101, a heat-killed borate-buffered whole cell product of Myco- portance of sequencing the administration of these agents are bacteriumobuensehasbeenshowntoenhancecellmediatedcyto- limited. Since the DC-based immunotherapy, AGS-003, expresses kine responses and innate immune responses involving NK and high levels of PD-L1, combinations with checkpoint blockade may gammadeltacells[1].Completeresponses(CR)inpatientswithmel- remove a critical signal protecting DCs during the early CTL acti- anomalungmetastasesdemonstrated.Followupoforiginalpublica- vation phase in vivo. Concurrent administration of checkpoint in- tion [2] has shown a 30 % 5-year survival. Combined with hibitors with AGS-003 may, therefore, impede the proposed gemcitabine in metastatic pancreatic cancer a significant survival mechanism of action of AGS-003, which is the induction of advantageovergemcitabinemonotherapyisseen[3]. tumor-specific CTL responses. Results derived from in vitro model- Methods ing of DCs inducing T cell responses can demonstrate how to We present 3 patients with metastatic melanoma, progressed better mobilize the immune system to overcome the immunosup- after initial stabilisation with IMM-101, who showed CR after pressive environment of cancer. Therefore, it was of interest to check point inhibitors (CPI) ipilimumab (n=2), pembrolizumab test anti-PD-1/anti-PD-L1 antibody therapy in vitro in combination (n=1). Patient 1: 2006 46 M melanoma left forearm, BT 3.7 mm, with DCs representative of AGS-003, to observe the effects com- 1 positive lymph node. Recurrent disease treated with surgery, bination therapy would have on antigen-specific CTL proliferation Aldara and low dose IL-2. 2010 pulmonary mets, commenced and functional responses. JournalforImmunoTherapyofCancer2016,4(Suppl1):73 Page114of221 Methods anti-PD-1 treated group as compared to the untreated. Anti-PD-1 DCs derived from monocytes were co-electroporated with MART-1 treatment led to a significant increase in the number of NK cells RNAandCD40ligandRNAtorepresentAGS-003DCproducts.Invitro and macrophages in the OS lung tumors suggesting these cells co-cultures were set up with autologous CTLs and MART-1/CD40L to have a potential therapeutic benefit against OS lung metasta- DCs in the presence of anti-PD-1 or anti-PD-L1 antibodies. In some ses. In addition, anti-PD-1 therapy caused a decrease in PD-L1 ex- instances,PD-1expression washyperexpressedonCTLsbyelectro- pression in the lung tumors, possibly due to a decrease in p- poratingMART-1-specficCTLswithPD-1RNA.Subsequentexpansion ERK1/2 and p-Stat3 expression. of MART-1-specific CTLs and multi-functional responses in the pres- Conclusions ence of checkpoint blockade were mapped using multi-color flow We conclude that targeting the PD-1/PD-L1 axis could be used to cytometry. treat OS lung metastasis. Therapeutic efficacy of anti-PD-1 may be Results due to an increased activity of NK cells and/or macrophages in the Combination with anti-PD-1 antibody did not did not negatively lung tumors and that inhibition of the p-Stat3/PD-L1 pathway may affect the expansion of MART-1-specific CTL responses; however, if be the mechanismimplicatedin OS lung metastases afteranti-PD-1 PD-1 was hyper-expressed on previously stimulated MART-1-specific treatment. CTLs responses were diminished. Anti-PD-1 antibody blocking re- storedCTLfunctioninthepresenceofhighlevelsofPD-1expression. Interestingly,anti-PD-L1antibodyblockingresultedinsuppressionof P202 earlyMART-1-specificCTLexpansionandsubsequentdownstreamef- EffectoftheclassI-HDACinhibitorentinostatandthepan-HDAC fectorfunction. inhibitorvorinostatonperipheralimmunecellsubsets Conclusions ItaliaGrenga,LaurenLepone,SofiaGameiro,KarinMKnudson,Massimo Our results suggest that the sequencing of AGS-003 therapy and Fantini,KwongTsang,JamesHodge,ReneeDonahue,JeffreySchlom checkpointblockadeisimportantto allowfullCTLactivationbythe LaboratoryofTumorImmunologyandBiology,NationalCancerInstitute, DCsprior to anti-PD-1/PD-L1therapy.Moreoverthe high expression ofPD-L1onDCsmayserveasa“don’tkillthemessenger”signal,crit- NationalInstitutesofHealth,Bethesda,MD,USA Correspondence:ReneeDonahue([email protected]) icaltopreventdeletionoftheDCpriortofullsignaldeliveryduring JournalforImmunoTherapyofCancer2016,4(Suppl1):P202 earlyphasesofCTLactivation. Background Cancer immunotherapy requires effective recognition and elimin- P201 ation of tumor cells identified as non-self; however, tumors can TargetingthePD-1/PD-L1signalingpathwayforthetreatmentof evade host immune surveillance through multiple mechanisms, OSlungmetastasis including epigenetic silencing of genes involved in antigen pro- PoojaDhupkar,LingYu,EugenieSKleinerman,NancyGordon cessing and immune recognition. Epigenetic therapy with histone UniversityofTexasMDAndersonCancerCenter,Houston,TX,USA deacetylase (HDAC) inhibitors has shown limited benefit as a Correspondence:PoojaDhupkar([email protected]) monotherapy in patients with solid tumors; however, recent re- JournalforImmunoTherapyofCancer2016,4(Suppl1):P201 ports suggest the potential for synergy when combined with im- munotherapy. Entinostat is a class I-HDAC inhibitor undergoing Background trials for the treatment of various cancers, while vorinostat is a Osteosarcoma (OS) is a primary bone malignancy, commonly cul- pan-HDAC inhibitor approved in the United States for the treat- minating into aggressive pulmonary metastasis. Despite chemo- ment of cutaneous T cell lymphoma. The aim of this study was therapy advances, the 5-year survival of pulmonary metastatic OS to extensively evaluate the effects of entinostat and vorinostat on remains 25-30 %. Immunotherapy is one of the promising novel human peripheral immune cell subsets in order to examine the approaches to target minimal residual and relapsed disease. The potential for combination of HDAC inhibitors with cancer objective of this study is to determine if blocking the PD-1/PD-L1 immunotherapy. immunosuppressive signaling pathway using a PD-1 checkpoint Methods inhibitor will have an effect in OS lung metastasis. Anti-PD-1 and Peripheral blood mononuclear cells (PBMCs) from metastatic anti-PD-L1 antibodies have exhibited therapeutic benefit in mel- breast cancer patients (n=7) were exposed in vitro for 48 hours anoma, and non-small cell lung carcinoma. We hypothesize that to clinically relevant exposures of entinostat, vorinostat, or ve- disruption of the PD-1/PD-L1 signaling pathway using anti-PD-1 hicle control. PBMCs were then analyzed by multicolor flow cy- antibody has an effect against OS lung metastasis and improves tometry using 27 unique markers to identify 123 immune cell overall survival. subsets, which included 9 classic cell types [CD4+ and CD8+ T Methods cells, regulatory T cells (Treg), B cells, conventional dendritic Flow cytometry and western blotting were used to analyze PD-L1 cells (cDC), plasmacytoid dendritic cells (pDC), natural killer cells expression in 7 different OS cell lines. Immunohistochemistry (NK), natural killer T cells (NKT), and myeloid derived suppressor (IHC) analysis was used to determine PD-L1 expression in OS lung cells (MDSC)], and 114 refined subsets relating to their matur- metastases from patients and mice. LM7 human OS mouse model ation and function. was used to test the effect of blocking murine PD-1 in OS lung Results metastases. Therapeutic effect of anti-PD-1 treatment was mea- Treatmentwithentinostatandvorinostatinducedseveralnotableal- sured by the number of macro and micro-metastases. IHC was terationsinperipheralimmunecells,suggestingmainlyimmuneacti- used to measure cell proliferation (Ki-67), apoptosis (TUNEL) and vatingproperties.Exposureto entinostatincreasedthefrequencyof cleaved-caspase 3 expression in addition to NK cells and macro- activatedCD4+Tcells,activatedmatureNKcells,antigenpresenting phages infiltration. Western blotting was used to address the cells(cDCs),andhighlyimmatureMDSCs,aswellasdecreasedtotal downstream components of the signaling pathway such as p- Tregs and those with a suppressive phenotype. Exposure to vorino- Stat3 and p-Erk1/2. The Simple PCI software was used to quantify statinducedfewerchangesthanentinostat,includingincreasingthe the IHC data. frequency of activated CD4+ T cells, highly immature MDSCs, and Results NKTcells. Our studies revealed surface and total PD-L1 expression in five Conclusions out of seven human OS cell lines. Primary and metastatic OS Thesefindingsshowthatwhileentinostatandvorinostathaveoverall lung tumor samples from patients demonstrated membranous immune activating properties, entinostat induced a greater number and cytoplasmic PD-L1 expression. Using a human OS mouse changes than vorinostat. This study supports the combination of model we demonstrated therapeutic effect of anti-PD-1 therapy HDAC inhibitors with immunotherapy, including therapeutic cancer as the number of macro and micro-metastases decreased in the vaccinesand/orcheckpointinhibitors. JournalforImmunoTherapyofCancer2016,4(Suppl1):73 Page115of221 P203 andinhibittumorprogression.PhaseIb/IIatrialsofcombinationther- Shifting the balance of tumor-mediated immune suppression apywithimmunecheckpointinhibitionareplanned. and augmenting immunotherapy with antibody blockade of semaphorin 4D to facilitate immune-mediated tumor rejection P204 ElizabethEvans1,HolmBussler1,CrystalMallow1,ChristineReilly1, CombinationofaglycomimeticantagonisttoE-selectinand SeboldTorno1,MariaScrivens1,CathieFoster1,AlanHowell1,Leslie CXCR4,GMI-1359,withananti-PD-L1antibodyattenuates Balch1,AlyssaKnapp1,JohnELeonard1,MarkParis1,TerryFisher1, regulatoryTcellinfiltrationandacceleratestimetocomplete SiwenHu-Lieskovan2,AntoniRibas2,ErnestSmith1,MauriceZauderer1 responseinthemurineCT26tumormodel 1Vaccinex,Rochester,NY,USA;2UniversityofCalifornia,LosAngeles, WilliamFogler1,MarilynFranklin2,MattThayer2,DanSaims2,JohnL. LosAngeles,CA,USA Magnani1 Correspondence:ElizabethEvans([email protected]) 1GlycoMimetics,Inc.,Rockville,MD,USA;2MIBioresearch,AnnArbor,MI, JournalforImmunoTherapyofCancer2016,4(Suppl1):P203 USA Correspondence:WilliamFogler([email protected]) Background JournalforImmunoTherapyofCancer2016,4(Suppl1):P204 We report a novel role for semaphorin 4D (SEMA4D, CD100) in modulatingthetumormicroenvironment(TME)toexcludeactivated Background antigen presenting cells and cytotoxic T lymphocytes so as to pro- RegulatoryT cells(T ) modulate anti-tumorimmunitybysuppress- reg motetumorgrowth.AntibodyblockadereducesexpansionofMDSC, ingTcellactivation.T areinducedandmaintainedbyimmunoreg- reg shiftsthebalanceofM1/M2,Teffector/Tregulatorycellsandassoci- ulatory receptors, such as PD-L1, and respond to homing signals atedcytokinesandchemokines,andaugmentstumorrejectionwith withintheinflamedtumormicroenvironmentthatincludetheendo- immunecheckpointinhibition. thelialcellprotein,E-selectin,andtheCXCR4ligand,SDF-1.GMI-1359 Methods is a small molecule glycomimetic beginning clinical evaluation with Anti-SEMA4D antibodies were evaluated, alone and in combination dual inhibitory activity against E-selectin and SDF-1. The aim of the with immune checkpoint antibodies. Immune response was charac- currentstudywastodetermineifGMI-1359aloneorincombination terized by immunohistochemistry, flow cytometry, functional assays, withanti-mPD-L1antibodyaffectedtheinvivogrowthofCT26colon and cytokine, chemokine and gene expression analysis. Anti-tumor carcinomaandtoassesspercentagesofinfiltrativeintratumoralcells activitywasevaluatedinvariouspreclinicalmodels.AphaseItrialfor expressingimmunemarkers. singleagentVX15/2503wascompleted. Methods Results Female Balb/c mice were implanted subcutaneously with 5x105 SEMA4D restricts migration of macrophages and promotes expan- CT26.WT tumor cells. Three days post tumor injection, mice (n= sion of suppressive myeloid cells in vitro. Strong expression of 15/group) were treated with saline, GMI-1359 (40 mg/kg for 12 SEMA4D at the invasive margins of actively growing tumors consecutive days), isotype control antibody (anti-KLH) or anti- in vivo modulates the infiltration and polarization of leukocytes in mPD-L1 antibody (10 F.9G2, 10 mg/kg on days 3, 6, 10, 13, and the TME. Antibody neutralization facilitated recruitment of acti- 17), or the combination of GMI-1359 and anti-mPD-L1 or anti- vated APCs and T lymphocytes into the TME in preclinical KLH. On day 15, tumors and spleens (n=5/group) were excised models. M-MDSCs were significantly reduced in both tumor and and T cells (total CD4+ and CD8+, and CCR7+/CD62L+ subsets of blood following treatment. This was accompanied by a significant each), regulatory T cells (T ; CD4/CD25/FoxP3), and myeloid de- reg shift towards increased Th1 cytokines and CTL-recruiting chemo- rived suppressor cells (MDSC; CD11b+/Gr1+) were determined by kines, with concurrent reduction in Treg-, MDSC-, and M2- flow cytometry. The remaining mice were followed for tumor macrophage promoting chemokines (CCL2, CXCL1, CXCL5). response. Accordingly, an increase in Teff:Treg ratio (3x, p<0.005) and CTL Results activity (4x, p<0.0001) was observed. NanoString gene expression Treatments were well tolerated. Mice in control groups and single analysis of on-treatment tumors confirms an increase in the agent GMI-1359 were all identified with progressive disease. In gamma-inflammatory gene signature (Ribas, ASCO 2015), includ- contrast, treatment with anti-mPD-L1 alone or in combination with ing significant increases in CXCL9, Gzmb, CCR5, Stat1, Lag3, Ptprc, GMI-1359produceda40%completeresponse(CR)rate.Themedian Ciita, Pdcd1 (PD-1), and Itga1. These coordinated changes in the time to CR was shorter when anti-mPD-L1 was combined with GMI- tumoral immune context are associated with durable tumor rejec- 1359comparedtoanti-mPD-L1alone(14vs.23days,respectively,p< tion and immunologic memory in preclinical colon, breast, and 0.0471).Evaluationoftumorinfiltratingcellsshowedthatcombination melanoma models. Importantly, anti-SEMA4D antibody can further therapy with GMI-1359 and anti-mPD-L1 reduced the percentage of enhance activity of immune checkpoint inhibitors and chemother- T compared to treatment with saline, GMI-1359 or anti-mPD-L1 as reg apy. Strikingly, the combination of anti-SEMA4D with anti-CTLA-4 singletreatments(0.9%vs.3.3%,2.9%and1.9%,respectively).No acts synergistically, with maximal increase in survival (p<0.01) otherTcellsubsetswereaffected.Inspleens,themedianpercentage and complete tumor regression in 100 % of mice, as compared ofT wereunaffectedbyanyofthetreatmentsandsuggestthatthe reg to 22 % with monotherapy (p<0.01). SEMA4D antibody treat- reductioninintratumoralT bycombinedtreatmentwithanti-PD-L1 reg ment was well tolerated in nonclinical and clinical studies; includ- andGMI-1359wasanattenuatedresponsetomaintenanceandhom- ing a phase I multiple ascending dose trial in patients with ingsignalsinthetumormicroenvironment. advanced refractory solid tumors. Patients with the longest dur- Conclusions ation of treatment, 48–55 weeks, included colorectal, breast, and In conclusion, these studies demonstrate that the dual E-selectin/ a papillary thyroid patient, who had a partial response by RECIST. CXCR4 antagonist, GMI-1359, in combination with anti-mPD-L1 anti- Conclusions body attenuates the induction and distribution of intratumoral T reg InhibitionofSEMA4Drepresentsanovelmechanismandtherapeutic and this reduction in T is associated with a more rapid immuno- reg strategy to promote functional immune infiltration into the tumor therapeuticanti-tumorresponse.

Description:
Despite the success of checkpoint blockade immunotherapy in . 2MedImmune, Cambridge, England, UK . able sample for pharmacokinetic assessment was available for all . Radiation therapy is an effective focal treatment of primary Correspondence: George Fromm ([email protected]).
See more

The list of books you might like

Most books are stored in the elastic cloud where traffic is expensive. For this reason, we have a limit on daily download.