av) aie 4SSee
`OVID
`
`Williams&Wilkins Forename|
`
`SEPTEMBER 2013
`
`CrBote Kluwer Lippincott
`
`

`

`Journal
`
`of Immunotherapy
`
`
`
`Volume 36
`
`Number 7
`
`September 2013
`
`[spc Supplemental Digital Content is
`available in the article
`
`Contents
`
`Invited Commentary
`
`359
`
`362
`
`CD40 Therapy and Surgery: A Potential Immunologic Partnership
`Katelyn T. Byrne and Robert H. Vonderheide
`(See Article, page 365)
`
`Cracking the Stone: Combination Vaccination and CTLA-4 Blockade in
`Pancreatic Cancer
`David L. Bajor and Robert H. Vonderheide
`(See Article, page 382)
`
`Basic Studies
`
`365
`
` Agonistic Anti-CD40 Antibody Therapyis Effective Against Postoperative Cancer
`Recurrence and Metastasis in a Murine Tumor Model
`Andrea Khong, Matthew D. Brown, Justin B. Vivian, Bruce W. S. Robinson,
`and Andrew J. Currie
`(See Commentary, page 359)
`
`(continued next page }
`
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`Journal of Immunotherapy * Volume 36, Number 7, September 2013
`
`Co n te nts (continued)
`
`373—Irradiated and Activated Autologous PBMCs Induce Expansion of Highly
`Cytotoxic Human NK Cells In Vitro
`Yong-Oon Ahn, Saerom Kim, Tae Min Kim, Eun Young Song, Myoung Hee Park,
`and Dae Seog Heo
`
`Clinical Study
`
`382
`
`Evaluation of Ipilimumab in Combination With Allogeneic Pancreatic Tumor Cells
`Transfected With a GM-CSF Genein Previously Treated Pancreatic Cancer
`Dung T. Le, Eric Lutz, Jennifer N. Uram, Elizabeth A. Sugar, Beth Onners, Sara Solt,
`Lei Zheng, Luis A. Diaz, Jr, Ross C. Donehower, Elizabeth M. Jaffee, and
`Daniel A, Laheru
`(See Commentary, page 362)
`
`
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`

`Evaluation of Ipilimumab in Combination With Allogenei¢
`Pancreatic TumorCells Transfected With a GM-CSF Genein
`Previously Treated Pancreatic Cancer
`*
`Dung T. Le*} Eric Lutz,* Jennifer N. Uram,* Elizabeth A. Sugar,*~ Beth Onners,* Sara Solt. :
`Lei Zheng,* Luis A. Diaz, Jr, *§ Ross C. Donehower,* Elizabeth M. Jaffee,* and Daniel A. Lahertt
`
`
`
`(/ dnmunother 2013:36:382-389)
`
`vaccine,
`
`ne
`PDAis a particularly nonimmunogenic cancer, data peoh
`the concept of synergy
`reports support
`Summary: Preclinical
`gest that immuneresponses and antitumor responses ca"
`between cancer vaccines and immune checkpoint blockade in
`induced in PDAS !!
`TLA’
`honimmunogenic tumors. In particular, cytotoxic T lymphocyte-
`Cytotoxic T lymphocyte-associated antigen-4 (C eh
`associated antigen-4 (CTLA-4) antibodies have been successfully
`4) functions as a negative regulator of T-cell aei
`combined with GM-CSFcell-based vaccines (GVAX). Ipilimumab
`(anti-CTLA-4) has been tested as a single agent
`in patients with
`Ipilimumab,
`a CTLA-4 antagonist antibody, has nae
`pancreatic ductal adenocarcinoma (PDA) resulting in a delayed
`tested in patients with advanced PDA.!? Although sine 5
`response at a dose of 3mg/kg. Our study evaluated ipilimumab
`agent
`ipilimumab at 3mg/kg was minimally effective
`lOmg/kg (arm 1) andipilimumab l0me/kg + GVAX (arm 2). A
`significant delayed response in
`| patient
`suggests ! 2
`total of 30 patients with previously treated advanced PDA were
`immunotherapy could play a role in PDA. Melanen
`randomized(1:1). Induction doses were administered every 3 weeks
`studies demonstrated a dose-response relationship fee
`for a total of 4 doses followed by maintenance dosing every 12
`ipilimumab, and the dose of 10mg/kg was selected tn
`weeks. Twopatients in arm | showedevidence ofstable disease (7
`prior studies.*'* Furthermore, preclinical studies ee
`and 22 wk) but none demonstrated CA19-9 biochemical responses.
`synergy between anti-CTLA-4 antibodies and granulocy .
`In contrast, 3 patients in arm 2 hadevidence of prolonged disease
`stabilization (31, 71, and 81 wk) and 7 patients experienced CA19-9
`macrophage-colony stimulating factor
`(GM-CSF) “i
`declines. In 2 of these patients, disease stabilization occurred after
`based vaccines.'*'® The current PDA trial builds on these
`an initial period of progression. The median overall survival (OS)
`observations by evaluating ipilimumab at 10 mg/kg aT
`(3.6 vs. 5.7 mo, hazards ratio: 0.51, P = 0.072) and | year OS (7 vs.
`or in combination with allogeneic pancreatic tumor o
`27%) favored arm 2. Similar to prior ipiimumab studies, 20% of
`transfected with a GM-CSFgene (GVAX) forthe treatm’!
`patients in each arm had grade 3/4 immune-related adverse events,
`of previously treated, locally advanced, or metastatic PD”
`Amongpatients with OS > 4.3 months, there was anincrease in the
`peak mesothelin-specific T cells (P=0.014) and enhancement of
`
`
`the T-cell repertoire (?=0.031). In conclusion. checkpoint
`
`PATIENTS AND METHODS
`blockade in combination with GVAX has the potential for clinical
`Patients
`benefit and should be evaluated in a larger study.
`Study protocol (NCT00836407) was approved by the
`Key Words: CTLA-4, GVAX,
`pancreatic
`cancer.
`Johns Hopkins institutional
`review board.
`instituuion
`ipilimumab
`biosafety committee, the FDA, and the NIH Recombinant
`DNA Advisory Committee. Participating patients signe
`informed consent.
`if they had pre-
`Patients were cligible for enrollment
`viously treated,
`locally advanced, or metastatic histologr
`cally proven PDA, were > 18 years of age, had reeeivet
`gemcitabine-based chemotherapy, had Eastern Cooperative
`Oncology Group performance status 0 or | with normal
`
`hematologic aspartate—amino-and renal funetion,
`
`
`
`Fy with the recent progress in the treatment of meta-
`transferase/alanine aminotransferase/alkaline phosphatase
`static pancreatic ductal adenocarcinoma (PDA).
`the
`<2.5x upper
`limit of normal
`(ULN) (<5 ULN fo!
`median survival
`in the best performance status patients
`patients with liver metastases), bilirubin < 1.5 x ULN, and
`remains
`11 months.'? Progress has been made with
`an expected survival of 9 weeks. Individuals were excluded
`immunotherapy for
`traditionally immunogenic cancers
`if they had infection with human immunodeficiency virus.
`such as melanoma and even in some tolerogenic cancers
`hepatitis B or C, a history of brain metastases, autoimmune
`such as lung and prostate cancer.*’ Despite the viewthat
`disease, prior CTLA-4 inhibitor or agonist use, surgery.
`radiation, chemotherapy, vaccination, orsteroids within 28
`days of studyinitiation.
`
`See Commentary by Bajor and Vonderheide
`on page 362
`
`
`Received for publication October 5, 2012; accepted March 14, 2013.
`From the *The Sidney Kimmel Cancer Center, the Skip Viragh Center
`for Pancreatic Cancer, Research and Clinical Care, and the Sol
`Goldman Pancreatic Cancer Center at Johns Hopkins: #The Swim
`Across America Laboratory at
`Johns Hopkins;
`{Bloomberg
`School of Public Health at Johns Hopkins: and §The Ludwig
`Center for Cancer Genetics and Therapeutics at Johns Hopkins,
`Baltimore, MD,
`Reprints: Dung T. Le, 1650 Orleans Street, Room 407, Baltimore, MD
`21287 (e-mail: dle2ae jhmi.edu),
`Copynght
`© 2013 by Lippincott Williams & Wilkins
`
`Study Design and Treatment
`This was a phase Ib, open-label, randomized study
`performed at Johns Hopkins University (Baltimore, MD).
`The primary objective of the study was to determine the
`safety profile of ipilimumab alone or in combination with
`GVAX inpatients with previously treated PDA. Secondary
`objectives included estimation of overall survival
`(OS).
`
`382 | www.immunotherapy-journal.com
`
`This materialwaseobiZgMunother * Volume 36, Number 7, September 2013
`atthe NLM and may be
`Subject US Copyright Laws
`
`

`

`othelin-specilic T-cell responses with OS, and estimation
`of overall
`response rate and immune-related response.
`Immune-related response criteria (irRC) account for the
`kinetics of both old and new lesions, given the known for
`potential delayed responses with ipilimumab.'’ Thirty
`patients with PDA were enrolled at Johns Hopkins Uni-
`versity between March 11, 2009 and December 6, 2010 with
`follow-up censored as of January 27, 2013. All patients
`were included in the safety andefficacy analyses. Patients
`were randomized ina I:] mannerto ipilimumab alone (arm
`1) or ipilimumab + GVAX (arm 2) using a randomized
`block design,
`In both arms,
`ipilimumab (10 mg/kg) was
`administered intravenously (IV) over 90 minutes. In arm 2.
`before the ipilimumab infusion, patients received GVAX,
`which consists of 2 pancreatic tumorcell lines (Pane 6.03
`and Pane 10.05), which have been modified with a plasmid
`vector encoding the cDNA for human GM-CSFand sub-
`sequently cultured and irradiated.’ The vaccine consists of
`Pane 6.03 and Pane 10.05 cells (2.5 x 10* cells each) com-
`bined into a single vaccine and administered as intradermal
`injections, 2 each in the right andleft thighs and 2 in the
`nondominant
`arm, Biosafety
`level
`2
`practices were
`employed for the containment of GVAX.
`Treatments were administered at weeks |. 4, 7, and 10.
`Computed tomography (CT) scans (magnetic resonance
`imaging if CT contraindicated) were performed for tumor
`assessments at weeks 1, 7. 14, and 22. Patients with pro-
`gressive disease (PD) without rapid clinical deterioration
`could continue on study treatment. At week 22 evaluation,
`patients with evidence ofa response orstable disease (SD)
`were oflered maintenance dosing ofthe originally assigned
`treatment every 12 weeks. Tumor assessments were “per-
`formed every 12 weeks during maintenance. Patients with
`carly progression followed by SD or better between weeks
`Id and 22 were also eligible for maintenance. Patients were
`followed by telephone conlict every 12 weeks to evaluate
`survival. disease status, and adverse events (AB),
`
`Patient Monitoring and Toxicity Criteria
`AEs were graded using the NCI CommonTerminol-
`ovy Criteria for Adverse Events (CTCAE) v3.0. For pur-
`poses of determining unacceptable toxicity during the initial
`22-week treatment phitse, patients were followed for drug
`related > grade 4 AEorgrade 3 AEsincluding immune-
`related AE (IRAE) not
`improving to « grade 2 under
`therapy within 2 weeks. In addition, 2 grade 2 eye pam or
`reduction of visual acuity that did not respond to topical
`therapy within 2 weeks was also an unacceptable toxicity, A
`+ +3 design was used to determine whether or not
`the
`plable for the first 6 patients in “ach arm.
`loxicily was acce
`If the toxicity rate was < 33%, then the remaining patients
`would be enrolled in that arm. The proportion of patients
`with unacceptable toxicities was continuously monitored. If
`the toxicity level in the combination arm was > 2/6, then
`the dose of ipilimumab could be reduced to 5 mg/kg for the
`combination armonly. Intrapatient dose deescalations were
`not permitted. AEs 70 days alter
`the last dose were
`recorded if possibly related (o the investigational agents,
`There were no protocol-defined unacceptable toxicities
`in the first 6 patients in either arm and enrollment con-
`tinued to complete the goal of TS patients per arm.
`
`Detection of Mesothelin-specific CD8 * T Cells by
`Interferon y-ELISPOT
`Synthesis of peptides, ELISA assays for identifying
`mesothelin peptides, and ELISPOTassays have previously
`been described.” '! Peripheral blood lymphocytes (PBL)
`werecollected at baseline, before each dose, 28 days after
`maintenance doses, and at
`the off study visit. PBLs from
`patients expressing HLA-A*010L and/or HLA-A*0201
`alleles were tested if pretreatment and posttreatment sam-
`ples were available. T-cell responses to mesothelin peptides
`were adjusted for background measured against irrelevant
`melanoma or
`renal
`cell
`carcinoma
`control peptides.
`Responses were measured to 8 HLA-A*0101 and 6 HLA-
`A*0201 mesothelin peptides. The sum ofT-cell responses to
`mesothelin peptides were reported. The size of the meso-
`thelin-specific T-cell repertoire was defined as the percent-
`age of peptides for which an induction was measured. A
`response was considered to be induced whenthe frequency
`of specific T cells was >5 in 1% 10° CD8* PBL and
`increased by > 2-fold compared with baseline.
`
`Clinical Assessments
`the specified
`Radiographic imaging was obtained at
`lime points. Response was assessed by RECIST v1.0 and
`irRC, CA19-9 serum levels were measured at
`regular
`intervals. OS was definedas the time from enrollment until
`death orloss to follow-up.
`
`Statistical Considerations
`Fifteen patients in each arm were enrolled to refine
`estimates of toxicity and initial efficacy measurements.
`Comparisons of continuous and categorical characteristics
`were made using the Wilcoxon rank-sum.
`tests and the
`Fisher exact tests, respectively. For each arm, the Kaplan-
`Meierestimates of the survival curve were calculated and
`used to estimate median OS and the proportion ofindi-
`viduals alive at
`1 year with 95%confidence intervals (C1).
`Comparisons between groups were made using log-rank
`tests. Differences between pretreatment and posttreatment
`immune responses were compared using the Wilcoxon
`signed-rank tests.
`
`RESULTS
`
`Patient Characteristics, Safety, and Tolerability
`Patient characteristics are shown in Table |. Baseline
`characteristics were similar among patients in each arm
`with the exception that arm | had fewer patients with > 2
`Prior
`therapies (60% vs. LOOM, P
`0.017). The most
`common AEs
`reported for
`ipilimumab therapy were
`IRAEs: the most common AEsreported for GVAX vac-
`cines were localized vaccine reactions and_ self-limiting
`systemic rashes. Table 2 summarizes
`IRAEs observed
`during all treatment cycles by arm and CTCAE grade. The
`rate of IRAEs attributable to ipilimumab was similar to
`what has been reported in other studies testing ipilimumab
`at
`the 10mg/kg dose. Seventy-three percent and 80% of
`patients in arm | and2, respectively, experienced anygrade
`IRAE and 20%ofpatients in both arms experienced grade
`3 and 4 IRAEs (colitis, Guillain-Barre syndrome, nephritis
`in arm 1; colitis, rash, pneumonitis in arm 2). The case of
`nephritis was considered an unacceptable toxicity because
`although
`the
`patient
`died
`from PD,
`he
`required
`
`2073 Lippincon Williams & Wilkins
`
`This material was copied
`atthe NLM and may be
`Subject US Copyright Laws
`
`www.immunotherapy-journal com | 383
`
`

`

`TABLE 1. Patient Characteristics
`
` Total (N = 30)
`Arm 1 (N = 15)
`Arm 2 (N = 15)
`Age (y), median (range)
`62 (44-77)
`63 (44-73)
`62 (49-77)
`Sex, n (0)
`Male
`Female
`ECOG, n ("%)
`()
`|
`Prior chemotherapy regimens, n ("o)
`I
`2
`3
`4
`Metastatic sites, n ("%o)
`2 (13)
`2 (13)
`4 (13)
`O*
`3 (20)
`3 (20)
`6 (20)
`|
`10 (67)
`10 (67)
`20 (67)
`2+
`
`21 (70)
`9 (30)
`
`10 (33)
`20 (67)
`
`6 (20)
`10 (33)
`12 (40)
`2(7)
`
`IL (73)
`4 (27)
`
`6 (40)
`9 (60)
`
`6 (40)
`4 (27)
`4 (27)
`1 (7)
`
`10 (67)
`§ (33)
`
`4 (27)
`11 (73)
`
`0 (0)
`6 (40)
`8 (53)
`1 (7)
`
`P
`0.77
`> 0.99
`
`0.70
`
`0.017
`
`> 0.99
`
`*Locally advanced, recurrent.
`ECOG indicates Eastern Cooperative Oncology Group,
`
`hemodialysis. The case of pneumonitis was considered an
`unacceptable toxicity because it took 25 days to resolve to
`«grade 2. Arm 2 did have a higher rate of dermatological
`reactions (73%vs. 60%,
`P = 0.45). Although only | rash
`was classified as a grade 3, the rashes in these patients were
`more symptomatic but often lessened in severity with sub-
`sequent doses. Grade | and 2 fatigue was also frequently
`
`reported in arm 2. Seven patients (3 in arm 1, 4 in arm2)
`with symptomatic diarrhea were diagnosed with diarrhea/
`colitis. The median time to onset to the highest grade of an
`IRAE was 53 days (range, 15 to 221d). Median time to
`resolution to grade | with steroids was 19 days with a range
`of | to 38 days. With the exception of the case of nephritis.
`the IRAEs did respond to steroids. The Guillain-Barre
`
`
`
`
`
`
`
`
`TABLE 2. Immune-related Adverse Events
`Arm 1 N= 15
`Arm 2 N = 15
`Total N = 30
`Grade 1/2 Grade 3/4 Grade 3/4 Grade 1/2
`Grade 3/4
`Grade 1/2
`Immune-related adverse events
`Dermatological
`Localized
`Systemic*
`Endocrine
`Adrenal insufliciency
`Hypophysitis
`Ophthalmologic
`Conjunctivitis
`Gastrointestinal
`Colitis
`Neurological
`Guillain-Barre syndrome
`Pulmonary
`Pneumonitis
`Renal
`Nephritis
`Other related adverse events
`Constitutional
`0
`|
`0
`l
`0)
`0
`Arthralgias
`0
`2
`0
`2
`0
`0
`Anorexia
`)
`|
`0
`0
`0
`|
`Cramps
`0
`2
`0
`2
`0
`0
`Diarrhea
`0)
`|
`0
`0
`0
`|
`Dry eye
`0
`I
`0
`0
`0
`|
`Dry skin
`0
`6
`0
`6
`0
`0
`Fatigue
`0)
`8
`0
`5
`0
`3
`Fever
`0
`6
`0
`4
`0
`2
`Flu-like symptoms
`0
`3
`0
`3
`0
`0
`Headache
`
`
`
`
`
`
`| 0 2 0 3Nausea 0
`
`0
`9
`
`0
`2
`
`|
`
`2
`
`0
`
`0
`
`()
`
`0
`0
`
`0
`()
`
`0)
`
`|
`
`I
`
`0
`
`|
`
`15
`|
`
`|
`|
`
`0
`
`3
`
`0
`
`0
`
`0)
`
`0)
`|
`
`0
`()
`
`0
`
`|
`
`0
`
`|
`
`0
`
`1s
`20
`
`|
`3
`
`|
`
`5
`
`0
`
`0
`
`0
`
`0
`|
`
`0
`0
`
`0
`
`2
`
`|
`
`|
`
`|
`
`“Although there was only one grade 3 rash observed in arm 2, rashes were more severe/symptomatic than in arm I.Neennncnee
`
`384 | www.immunotherapy-journal.com
`
`This material was copied
`at the NLMand may be
`Subject US Copyright Laws
`
`© 2013 Lippincott Williams & Wilkins
`
`

`

`noglobulin and technically resolved only to a grade 2
`because the patient continued to use a cane.
`
`Overall Survival
`Median OSforthe all patients was 4.3 months (95%
`Cl, 3.65-8.11). Median OS was 3.6 months (95%CI,
`2.5-9.2) for arm | and 5.7 months (95% CI, 4.3-14.7) for
`arm 2 (HR: 0.51; 95% CI, 0.23-1.08; P = 0.072). The
`percent alive after | year also favored the combination arm
`(7%vs. 27%) (95% CI. 1% 45%vs. 11%-62%) (Fig. 1).
`
`Mesothelin-specific T-cell Responses
`Enhanced post-vaccination mesothelin-specific T-cell
`responses were associated with increased disease-free
`survival and OS in prior GVAX studies. !' Mesothelin-
`specific T-cell
`responses were analyzed in PBL from 19
`HLA-Al
`'
`and/or HLA-A2*
`patients with at
`least
`|
`posttreatment PBL sample. Baseline, posttreatment
`|, and
`peak-induction T-cell responses are shownin Figure 2A as
`a correlate of OS for
`the combined treatment arms.
`Although mesothelin-specific T-cell
`responses were not
`enhancedafterthe first treatment in either group, there was
`a significant
`induction of peak posttreatment
`responses
`among patients with OS > 4.3 months (P = 0.014). Meso-
`thelin-specific T-cell responses were measured after 2 or
`more treatments in 14 of the 19 patients and are shown
`in Figure 2B. Similar to the analysis for all
`19 patients
`evaluated, T-cell responses were not enhancedafterthefirst
`treatment
`for either group, und peak posttreatment
`responses were enhanced only in patients with OS > 4.3
`months (P = 0.020) and remained elevated throughout
`treatment, albeit not
`significantly (P = 0.13). Similarly.
`whenlevels of circulating mesothelin-specific T cells were
`compared with O§, significant correlations were only seen
`after the second treatment (P = 0.045) and at
`the time of
`
`— Ipilimumab plus GVAX
`
`1,0
`
`0.8
`
`
`
`Proportionalive
`
`0.6
`
`0.4
`
`0.2
`
`or after the first treatment (P = 0.50) (Pigs. 3A-D). Com-
`parisons after the third and fourth induction treatments
`were not performed because too few PBL samples were
`analyzed at
`these later time points. The size of the meso-
`thelin-specific T-cell repertoire measured after the first and
`final treatments in the 14 patients who received 2 or more
`treatments are shown in Figure 4A. T-cell repertoire size
`was similar after the initial treatment but significantly larger
`after multiple treatments among patients with OS > 4.3
`months (P = 0.009). Furthermore, expansion in the reper-
`toire was observed in 6 of 9 patients with OS > 4.3 months
`but not
`in any of the 5 patients with OS<4.3 months
`(Fig. 4B) and was associated with longer OS (P = 0.031).
`
`Antitumor Activity
`The best RECISTresponse was SD in 2 patients in
`arm | and 2 patients in arm 2. Using the irRC, arm 2 had
`an additional patient with SD until week 81. The quality of
`the responses in the 2 arms wasdifferent. Patients with SD
`on arm | had continuous disease progression that did not
`reach the 20% growth cutofffor 7 and 22 weeks. Arm2 had
`3 SD(1 regression starting at week 14 and maintained until
`week 31,
`| stabilization starting at week 22 and maintained
`until week 81,
`1 SD lasting until week 71). Disease stabili-
`zation occurred by the week 22 scan inall of the patients.
`Maintenance phase imaging documented disease stability
`but no further regressions. Figures SA~CshowCTfindings
`of carly tumor progression followed by regression starting
`at week
`14 and the corresponding CA19-9 responses
`(Fig. 5D). Figure SE shows interesting CA19-9 kinetics ina
`patient who hadstable local disease lasting 71 weeks, The
`baseline elevated CA 19-9 increased further during high-
`dose steroid treatment for hypophysitis and then showed a
`gradual delayed normalization. Figure SF shows the CA
`19-9 kinetics of a patient with early local progression and a
`new omental lesion at week 7 followed by disease stabili-
`zation starting at week 22 andlasting until week 81. The
`CAI9-9 rise between the
`12-week maintenance doses
`declined in response to treatments. CA19-9 declines in
`association with ipilimumab + GVAX treatment were seen
`for 7/15 patients. In contrast, 0/15 patients receiving ipili-
`mumab alone had CA19-9 declines.
`
`DISCUSSION
`The data from this phase |btrial testing ipilimumab
`alone or
`in combination with GVAX in PDA patients
`report 3 newfindings. First, the safety profile for ipilimu-
`mab alone or the combination is similar for patients with
`PDA when compared with reported studies testing ipili-
`mumab in patients with melanoma. Second,
`immune
`responses can be induced even in patients with advanced
`PDA, and these responses correlate with clinical activity.
`Third, clinical
`responses
`to immunotherapy in PDA
`patients with advanced disease require prolonged treat-
`ment, similar
`to what has been observed in melanoma
`patients treated successfully with immunotherapy. Taken
`together,
`this study provides strong support
`for further
`testing of
`ipilimumab and other
`immunotherapies
`in
`patients with PDA.
`the toxicity spectrum
`This study demonstrates that
`and rates observed in PDA patients are similar to what has
`been reported for melanoma patients treated with ipilimu-
`mab. IRAEs were evident even in advanced PDA patients,
`
` 0.0
`
`0
`
`5
`
`10
`
`T
`20
`15
`Time (months)
`
`25
`
`30
`
`Nipsavax: 15
`
`8
`
`5
`
`2
`
`2
`
`2
`
`2
`
`FIGURE 1. Survival. The Kaplan-Meieroverall survival curve asof
`January 27, 2013. Onepatient in arm 2 (ipilimumab + GVAX) is
`still alive.
`
`« 2013 Lippincott Williams & Wilkins
`
`This material was copied
`at the NLM and may be
`Subject US Copyright Laws
`
`www.immunotherapy-journal.com | 385
`
`

`

`A
`
`A Ipilimumab alone
`@ Ipilimumab + GVAX
`
`B
`
`OS < 4.3 months
`
`OS > 4.3 months
`
`A Ipilimumab alone
`®@ Ipilimumab + GVAX
`OS < 4.3 months
`OS > 4.3 months
`=
`0.44
`pre
`P=0.13
`
`= 0.62 4pase P=0.020
`p= 0.82
`P>0.99
`-—4
`'
`
`
`
`sfc/1x10°CD8*PBL 8 gIFNy
`
`p=0.014
`p=0.36
`———— 1
`p=0.36
`p=0,92
`200 <i
`——
`.
`
`.
`
`
`
`
`
`CD8*PBL So zIFNysfc/1x10°
`
`FIGURE 2. Longer survival is associated with an induction of CD8* mesothelin-specific T-cell responses, Mesothelin peptide-specific
`CD8* T cells were quantitated in pretreatment and posttreatment peripheral blood lymphocytes (PBL) Using IFNy ELISPOT assay®-
`A, Mesothelin-specific T-cell frequencies measured at baseline, after the first treatment, and at the peak of induction in 19 HLA-Al'
`and/or HLA-A2* patients receiving at least 1 treatment. B, Mesothelin-specific T-cell frequencies measured at baseline, after the first
`treatment, at the peak of induction, andafter the final treatmentin 14 of the 19 patients who received atleast 2 treatments. Patients in
`both treatment arms (open triangles: ipilimumab alone;solid circles: ipilimumab + GVAX) were grouped together based on survival of
`greater than orless than 4.3 months. Posttreatment T-cell levels were compared with baseline levels using the Wilcoxon signed-rank
`tests. OS indicates overall survival. IFN indicates interferon.
`
`which supports that these patients do have reactive immune
`systems.
`In addition,
`the nondermatologic toxicities were
`often limited to one-organ system in patients receiving the
`combinationof ipilimumab + GVAX (7/7) when compared
`with those receiving ipilimumab alone (1/4). This raises the
`question of whether the use of a vaccine can skew the
`immune response. Toxicities are likely to differ depending
`on the ipilimumab combination, Ipilimumab in combina-
`tion with dacarbazine resulted in higher than previously
`reported immune-related hepatitis.4 However, prior studies
`have not reported on the number of organ systems affected
`in-an individual patient. Further studies are needed to
`better characterize this observation.
`in OS that was asso-
`We observed an improvement
`the combination arm
`ciated with clinical activity in
`(P = 0.07). Ipilimumab was previously tested as a single
`+
`*
`.
`12
`agent in advanced PDA patients.’ Only | patient demon-
`strated clinical activity and this was at the lower dose level
`of 3mg/kg. The higher rate ofclinical activity observed in
`our study may be due to differences in ipilimumab dose or
`more likely, due to the need of combining this T-cell acti-
`valing agent with T-cell
`inducing agents (vaccines),
`Ipili-
`mumab was previously combined with prostate GVAX ina
`phase | dose-escalation trial for the treatment ofcastrate-
`resistant prostate cancer.!* The dose selected for expansion
`in that study was 3mg/kg because of signs of clinical
`activity at that dose level. Fifty percent or greater declines
`from baseline
`in prostate-specifie antigen levels were
`observed in 25% of patients. All of these patients received
`either 3 or Smg/kg of ipilimumab. HLA-DR, a marker of
`T-cell activation, was only upregulated at
`the higher dose
`levels. Induction of PSMA-specific antibody responses was
`
`associated with improved OS. Studies of immunotherapy
`induced antibody responses are Ongoing for our PDA
`study.
`This is the first study to evaluate ipilimumab-induced
`mesothelin-specific T-cell
`responses either alone or
`it
`combination with a vaccine.
`Indeed. mesothelin-specilic
`T-cell responses were measured in patients after treatment
`with ipilimumab alone and with the combination, T-cell
`responses measured in both arms were analyzed together
`because the pattern of induction and association with sul-
`vival were similar between the 2 arms and also because of
`the small sample size. The induction of mesothelin-specilic
`responses in the ipilimumab alone arm support the concept
`that non-antigen-specific agents such as ipilimumab act by
`enhancing preexisting endogenous tumor-specific Tcells. A
`correlation between the magnitude of the mesothelin-spe-
`cific T-cell response and OS was only seen for postinduction
`2 and peak responses. This suggests that T-cell
`levels at
`baseline and after an initial treatment do not predict who
`will respondto therapyandthat multiple doses are required
`to induce T-cell responses in most patients, Similar to prior
`studies,’* these data also Suggest that the maintenance of
`an enhanced T-cell
`response may better predict which
`patients are morelikely to benefit from treatment. Inter-
`estingly, diversification of the mesothelin-specific T-cell
`repertoire with additional
`treatments was seen in both
`arms. However, a greater numberof patients in the com-
`bination arm exhibited these responses (4/7) compared
`with the ipilimumab alone arm (2/7) suggesting that
`the
`frequency ofpreexisting mesothelin-specific Tcells are low
`and require a vaccine to induce larger pools of precursor
`T cells. Larger clinical
`trials comparing the induction of
`
`386 | www.immunotherapy-journal.com
`
`This material was copied
`at the NLM and may be
`Subject US Copyright Laws
`
`© 2013 Lippincott Williams & Wilkins
`
`

`

`S48
`
`p=0.39
`
`
`
`
`
`OverallSurvival(Days)
`
`100
`
`150
`
`200
`
`Mesothelin-specific T cells per
`1x105 CD8* PBL
`
`Mesothelin-specific T cells per
`1x10° CD8* PBL
`
`p=0.50
`
`= 800
`%
`
`8 £gaw
`
`o oso
`
`
`
`0
`
`50
`
`100
`
`150
`
`200
`
`D
`
`a a ea ao$o
`
`e up
`
`
`
`Peak Induction
`p=0.047
`
`100
`
`150
`
`200
`
`oO
`
`“4
`
`PostInduction Dose 2
`
`
`
`
`
` p=0.045 OverallSurvival(Days)
`
`0
`
`20
`
`40
`
`60
`
`80
`
`Mesothelin-specific T cells per
`Mesothelin-specific T cells per
`1x105 CD8* PBL
`1x10° cD8* PBL
`FIGURE3. Increasedlevels of peak and postinduction dose 2 mesothelin-specific T cells are associated with longer survival. Frequencies
`of mesothelin-specific T cells measured by IFNy ELISPOTin peripheral blood lymphocytes (PBL)isolated (A) before treatment, (B) after
`theinitial treatment, (C) after the second treatment, and (D) at the time of peak induction, are plotted against overall survival. Linear
`regressions were performed and respective P values are shown for each timepoint. IFN indicates interferon.
`
`T-cell responses between these 2 arms shouldfurther clarify
`this issue. Consistent with a prior study,'” posttreatment
`expansion of the mesothelin-specific T-cell repertoire was
`associated with longer OS.
`In this study, median survival
`for
`the 6 patients demonstrating an expansion in.
`their
`
`15.7 months
`repertoires was
`mesothelin-specific T-cell
`compared with 4.1 months for the & patients whose reper-
`toires were unchangedafter treatment. When expansionsin
`the T-cell repertoire were seen, they were detected by the
`endofinduction suggesting the possibility that this measure
`
`A
`
`A |pilimumab alone
`® Ipilimumab + GVAX
`
`B
`
`OS < 4.3 months
`
`OS > 4.3 months
`
`400
`
`p=0.009
`
`0
`
`60
`
`40
`
`,
`
`A Ipilimumab alone
`@ Ipilimumab + GVAX
`
`OS > 4.3 months
`
`100
`
`60
`
`40
`
`3a$
`
`OS <4.3 months
`
`85
`
`=o
`
`£ :
`
`o#
`
`@
`
`g
`
`eg
`
`@ 100
`5
`3
`80
`7—_——4ua
`:
`Ew
`3
`3
`c
`gs
`i
`i
`£
`3
`2
`33
`i
`i.a
`3 20
`8
`3x
`0
`
`—Ser
`NK
`+
`at
`ar
`at
`e
`-
`&
`Oy
`ss
`o
`=
`C)
`<
`+
`eS
`od
`
`FIGURE 4. Longersur