The Novel IL-2 Cytokine Immune Agonist NKTR-214 Harnesses the Adaptive and Innate Immune System for the Treatment of Solid CancersSalah Eddine Bentebibel1, Chantale Bernatchez1, Cara Haymaker1, Michael Hurwitz3, Patrick Hwu1, Mario Sznol3, Nizar Tannir1, Anthony Conley1, Harriet Kluger3, Sandra Aung2, Michael Imperiale2, Mary Tagliaferri2, Christie Fanton2, Ernesto Iacucci2, Jonathan Zalevsky2, Ute Hoch2, Adi Diab1

1The University of Texas MD Anderson Cancer Center, Houston, TX, United States of America, 2Nektar Therapeutics, San Francisco, CA, United States of America, 3Yale School of Medicine, New Haven, CT, United States of America

BACKGROUND

RESULTS

RESULTS

NKTR-214• NKTR-214 is a CD122-biased cytokine agonist

conjugated with multiple releasable chains of polyethylene glycol (PEG) designed to provide sustained signaling through the heterodimeric IL-2 receptor pathway (IL-2R) to preferentially activate and expand effector CD8+ T and NK cells over Tregs1 (Figure 1).

NKTR-214 MONOTHERAPY STUDY• A phase 1, multicenter, open-label, dose-

escalation study (EXCEL) was conductedto assess the safety, preliminary efficacy, pharmacokinetics, and pharmacodynamicsof NKTR-214 in 28 patients with locally advanced or metastatic solid tumors.2-4

• Outpatient regimen with a convenient IV dosing regimen every 2 or 3 weeks.

• NKTR-214 has a favorable safety and tolerability profile.4

– No evidence of immune-mediated AEs or organ related inflammation (eg, colitis, pneumonitis, dermatitis, hepatitis, endocrinopathies). – Grade 3 hypotension occurred in 14% of

patients and was rapidly reversible with IV fluids. – No patients experienced capillary leak syndrome. – No Grade 4 TRAEs or treatment-related deaths

• Maximum-administered dose (MAD) was 0.012 mg/kg q3w.

• Sustained exposure and robust PD changes after a single dose of NKTR-214 (Figure 3).3,5

• NKTR-214 substantially increased CD8+ T cells that were newly proliferative (Ki-67+) (Figure 4).5

CONCLUSIONS

Figure 1. NKTR-214 Mechanism of Action

CLONAL EXPANSION

Stimulates Immune Response to Kill Tumor Cells

LEGEND:NKTR-214 – Inactive2-PEG – Active Cytokine1-PEG – Active Cytokine

NKTR-214 (6-PEG)

IrreversibleRelease

2-PEGActive Cytokine

1-PEG Active Cytokine

IrreversibleRelease

IL-2Rαβγ

α

β γβ γ

IL-2Rβγ

Immunosuppressivecells limit anti-tumor

responseNKNK

CD8+

CD8+

CD8+

CD4+

Helper

CD4+

Helper

CD4+

Treg

NK NK

NK, CD4+, and CD8+ T cells

CD4+

HelperCD8+

CD4+

Helper

CD4+

HelperCD8+

NK CD4+

Helper

NK

CD8+

CD4+

HelperCD4+

HelperCD8+

CD8+

NKNK

NK

CD8+

CD4+

Helper

CD4+

Helper

NKCD8+

REFERENCES1) Charych DH, Hoch U, Langowski JL, et al. Clin Cancer Res. 2016;22(3):680–90.2) Bernatchez C, Haymaker C, Tannir NM, et al. Presented at SITC 2016, National Harbor, MD. Poster #387.3) Hurwitz ME, Diab A, Bernatchez C, et al. Presented at ASCO GU 2017, Orlando, FL. Poster #D17.4) Bernatchez C, Bentebibel SE, Hurwitz ME, et al. Presented at ASCO 2017, Chicago, IL.5) Nektar Therapeutics Analyst & Investor Event at ASCO 2017 Annual Meeting. [online]

Available at: http://ir.nektar.com/events.cfm

Figure 2. Time on Study and Best Overall ResponseSD

SDSD

SDSD(uPR)

SDSD

PDSD

PDPD

PDPDPD

PDPD

SDSD

SDSD

SDNE

PDSD

PDSDPD

NE

0.003 mg/kg q3w0.006 mg/kg q3w0.012 mg/kg q3w0.009 mg/kg q3w0.006 mg/kg q2w

Discontinued treatment due to RECIST PDDiscontinued treatment due to AEDiscontinued treatment due to other reasons

PD - Best Overall Response is Progressive DiseaseSD - Best Overall Response is Stable Disease

Data cut-off: October 4, 2017

W3/C1 W15/C5 W30/C10 W45/C15 W60/C20 W75/C25 W90/C30

Breast CaRCCRCC

ChondrosarcomaLeiomyosarcoma

RCC

MelanomaRCCRCC

MelanomaRCCRCCRCC

Triple Neg BCMelanomaMelanoma

RCCColorectal

RCCRCCRCCRCCRCCRCC

MelanomaBladder CaMelanomaMelanoma

Time on Study (Weeks/Cycles)0 W8/C4 W16/C8 W24/C12 W32/C16 W40/C20 W48/C24 W56/C28 W64/C32 W72/C36 W80/C40 W88/C44 W96/C48 W104/C52

Table 1. NKTR-214 induces gene expression changes in tumors associated with innate and adaptive immune pathways

• NanoString analysis includes 10 patients with matched baseline and wk3 samples and 5 additional patients with either baseline or wk3 sample

− RCC (8), melanoma (4), chondrosarcoma (1), leiomyosarcoma (1), and triple negative breast cancer (1)• 89 genes differentially expressed between pre- and post-treatment when all patients used in the analysis

Pathway name (database source) pSize Genes in pathway differentially expressed post-treatment

T cell receptor signaling pathway (KEGG) 37 ICOS; ITK; LCK; MAPK11; ZAP70; CD3E; CD247; CD8A; CD8B; CD40LG; CTLA-4; PDCD1

TCR signaling in naïve CD8+ T cells (NCI) 20 CD8A; CD3D; LCK; CD3E; CD3G; CD8B; CD247; ZAP70; PRF1

IL-12-mediated signaling events (NCI) 43 SOCS1; IFNG; IL-2RA; CD8A; CD3D; CD3E; CD3G; CD8B; IL-2RB; CD247; IL-2RG; STAT4; IL-12RB2; EOMES; LCK; GZMB; TBX21

IL-2-mediated signaling events (NCI) 21 SOCS1; IL-2RA; LCK; IL-2RB; IL-2RG; MAPK11; IFNG

Nature killer cell mediated cytotoxicity (KEGG) 48 GZMB; IFNG; KLRC1; KLRC2; KLRD1; LCK; SH2D1A; CD244;ZAP70; CD247; KLRK1; NCR1; BID; SH2D1B; TNFRSF10C

pSize: Total # of genes in pathway covered by NanoString assay

Figure 11. Case study: Patient with RCC stage IV, age 59, male, progressed on prior TKI

TreatmentDuration ofTreatment

Time Interval toNext Treatment

Prior Therapy Sunitinib ~ 67 mos (PD) ~ 2 mos

TherapiesAdministered

NKTR-214 ~ 5 mos (SD) ~ 1 mo

OngoingNivolumab > 9 mos

H&E CD3 CD8

Baseline

Week 3

Immunological Changes onNKTR-214 Therapy

1027 meancells/mm2

675 meancells/mm2

2017 meancells/mm2

1620 meancells/mm2

EOT NKTR-214 POST Nivolumab

~1 mo

Con�rmed PRPR on 1st scan

Nivolumab

Pre NKTR-214Left Lung Nodule

NKTR-214 (8 cycles)PD-L1 status† = neg

(Max tumor response -10% per RECIST 1.1)

†PD-L1 status was obtained using the Cell Signaling antibody (Cell Signaling #13684, PD-L1 (E1L3N)) at a 1:100 dilution on the Leica BOND RXm

Robust immune-stimulatory responseIncrease in T cell receptor

clonality at wk3

• Increased T cell clonality and induction of immune-related genes in the tumor, along with an increase in the proliferative index of immune cells in the blood, indicate the ability of NKTR-214 to induce robust immune changes in patients

1.00

0.75

0.50

0.25

0.000.00 0.25 0.50

Cumulative percentage ofunique sequences

Cum

ulat

ive

per

cent

age

of

read

s

0.75 1.00

BaselineWeek 3

Increase in proliferationand PD-1+ expressionfollowing NKTR-214

0

10

20

30

40

0

10

20

30

40

50

Day

CD

4+ K

i-67

+ (%

CD

4)

CD

8+ K

i-67+ (%

CD

8)

C1D1 C1D8

CD4+ Ki67+

CD8+ Ki67+

CD

4+ K

i-67

+ P

D-1

+ (%

CD

4)

CD

8+ K

i-67+ P

D-1

+ (%C

D8)0

2

4

6

0

5

10

15

Day

CD4+ Ki67+ PD-1+

CD8+ Ki67+ PD-1+

C1D1 C1D8

>2-fold induction ofimmune checkpoint genes and

CD8-related genes at wk3

0 2 4 6 8 10Fold change (wk3/baseline)

LAG3ICOSIDO1

CTLA-4TIGITPD-1

PD-L1PD-L2

Tim3OX40TP53CD19

0 1 2 3 4 5 6 7Fold change (wk3/baseline)

IFNGPRF1

GZMBTBX21CD8ACD8B

IL-6SMAD3

• Sustained exposure with half-life of ~20 hours• Gradual increase in active cytokine species,

reaching Cmax 1-2 days post dose• Exposure increases in proportion to dose• Free IL-2 is not detectable

• Transient decrease followed by increase in lymphocytes from Day 3 to Day 9• Transient increase in soluble IL-2 receptor alpha (sCD25) from Day 1 to Day 8,

shed from activated T cells• sCD25 levels return to baseline by Day 15• Similar PD effects observed across dose levels

Pharmacodynamics(0.006 mg/kg, n=15)

Figure 3. Sustained exposure and robust PD changes after a single dose of NKTR-214

RC: Related Cytokine, total protein detection assay measures 6-PEG NKTR-214 and all derived species; AC: Active Cytokine, detection assay for 2-PEG and lower active species

Lymphocytes

0

1

2

3

109 /

L (M

ean+

SE

)

Time (Days)15 221 8

Time (Days)

sCD25

15 221 8

0

2

4

6

ng/m

L (M

ean+

SE

)

Pharmacokinetics(0.006 mg/kg, n=15)

ng/m

L (M

ean+

SE

)

NKTR-214-AC

NKTR-214-RC

15 221 8Time (days)

0.1

1

10

100

1000

Tumor Analysis Fresh TIL analysis by �ow cytometry IHC T cell receptor gene sequencing Gene expression analysis

Blood Analysis Flow cytometry Cytokines PK PD (sCD25, lymphocytes)

Blood and Tumor Biopsy Collection and Analysis

EOT, end of treatment.

C1D1Cycle (C)/Day (D): C1D8 C2D1

Q2W or Q3W

C3D1

EOT

NKTR-214

C2D8

BaselineTumorBiopsies: Week 3

= Blood Sample

Week 8 EOT

NKTR-214NKTR-214

ACKNOWLEDGEMENTS• The authors would like to acknowledge the contribution of patients and their families in

participation of this clinical trial.

Figure 4. NKTR-214 promotes proliferation of CD4+ T cells, CD8+ T cells, and NK cells in peripheral blood

CD4+ T cells

DayC1D1

n=16

C1D80

20

40

60

10

30

50

70

CD

4+ K

i-67

+ (%

CD

4)

Melanoma

ChondrosarcomaBreast CancerBladder Cancer

Renal Cell Carcinoma

Ki-67+ set on baseline sample at ~5%

• An increase in overall T cell clonality observed in 8/12 patients after NKTR-214 treatment

• Increase in clonality associated with increased T cell infiltrate in 7/12 patients

Figure 10. Increase in T cell clonality in tumors after treatment with NKTR-214

Melanoma

ChondrosarcomaRenal Cell Carcinoma

-6 -5 -4

-4-3-2-1012345

0 21 3 4Clonality (Log2)

In�

ltra

te (L

og

2)

Figure 5. NKTR-214 induces activating markers and co-inhibitory receptors in proliferating CD4+ T cells in peripheral blood

Ki-

67+

Ki-

67−

0

20

40

60

80

Day

CD

4+ K

i67+

ICO

S+ (%

CD

4)

C1D1 C1D8

n=15

PD-1 CTLA-4 OX40 ICOS

DayDay

0

5

10

15

CD

4+ K

i-67

− P

D-1

+ (%

CD

4)

C1D1

n=10

C1D8Day

0

5

1020

30

40

C1D1 C1D8

n=16

CD

4+ K

i-67

− C

TLA

-4+ (%

CD

4)

n=8

0

5

10

15

20

C1D1 C1D8

CD

4+ K

i-67

− O

X40

+ (%

CD

4)

n=9

0

5

10

15

20

DayC1D1 C1D8

CD

4+ K

i-67

+ O

X40

+ (%

CD

4)

0

5

1020

30

40

DayC1D1 C1D8

n=15

CD

4+ K

i-67

+ C

TLA

-4+ (%

CD

4)

0

5

10

15

DayC1D1

n=11

C1D8

CD

4+ K

i-67

+ P

D-1

+ (%

CD

4)

Activating and co-inhibitory marker gates set on C1D1 sample Legend is same as for Figure 6

0

20

40

60

80

C1D1 C1D8

n=15

Day

CD

4+ K

i-67

− IC

OS

+ (%

CD

4)

ICOS

Figure 6. NKTR-214 induces activating markers and co-inhibitory receptors in proliferating CD8+ T cells in peripheral blood

0

2

48

10

C1D1 C1D80

2

4

6

8

C1D1 C1D80

5

10

15

20

25C

D8+

Ki-

67− P

D-1

+ (%

CD

8)C

D8+

Ki-

67+ P

D-1

+ (%

CD

8)

CD

8+ K

i-67

+ C

TLA

-4+ (%

CD

8)C

D8+

Ki-

67− C

TLA

-4+ (%

CD

8)

CD

8+ K

i-67

− IC

OS

+ (%

CD

8)C

D8+

Ki-

67+ IC

OS

+ (%

CD

8)

C1D1 C1D8

0

2

48

10

C1D1 C1D80

2

4

6

8

C1D1 C1D80

5

10

15

20

25

Day Day Day

Day Day Day

C1D1

n=8n=12n=8

n=10n=11n=10

C1D8K

i-67

+K

i-67

−

PD-1 CTLA-4

Activating and co-inhibitory marker gates set on C1D1 sample

Melanoma

ChondrosarcomaBreast CancerBladder Cancer

Renal Cell Carcinoma

CD8+ T cells

DayC1D1 C1D8

CD

8+ K

i-67

+ (%

CD

8)

0

20

40

60

10

30

50

70n=16

CD3-CD56+ NK cells

DayC1D1 C1D8

n=13

CD3- C

D56+ K

i-67+ (

%CD

56)

0

20

40

60

80

100

Case Study

• Immunological activity in peripheral blood and tumor tissue consistent with NKTR-214’s biological mechanism of biased IL-2 pathway activation

• NKTR-214 has a robust PK-PD profile

• Robust immune-stimulatory response in the tumor and blood– NKTR-214 induced change in proliferative index of immune cells and

costimulatory/checkpoint markers only on newly proliferating immune cells– RNA expression changes consistent with increased immune cell infiltrate and

activation of effector mechanisms– Increased frequency of specific T cell clones and associated increase in T cell

infiltrate suggests efficient remodeling of the T cell repertoire

• The ability of NKTR-214 to increase TILs and increase PD-1 expression on immune cells provides a sound biological basis for combination with anti-PD1 checkpoint inhibitors

• Clinical trials currently enrolling patients with NKTR-214 in combination with anti-PD1 checkpoint inhibitors:

– PIVOT-02 clinical trial combining NKTR-214 + nivolumab in patients with metastatic melanoma, renal cell carcinoma, non-small cell lung cancer, urothelial cancer and triple-negative breast cancers (NCT02983045)

– PROPEL clinical trial combining NKTR-214 with atezolizumab or pembrolizumab in patients with metastatic melanoma, urothelial and non-small cell lung cancers (NCT03138889)

– A study of NKTR-214 + nivolumab in patients with metastatic and/or locally advanced sarcoma (NCT03282344)

Each line represents a different patient

Each line represents a different patient

Each line represents a different patient

Poster #P77: Society for Immunotherapy of Cancer 2017 Annual Meeting

Figure 9. NKTR-214 does not alter expression of Th17- or Treg-associated genes in the tumor

Gene nameGenesymbolRORC ROR gamma; transcription factor

TGFB1 TGF beta 1

TGFB2 TGF beta 2

IL-10 Interleukin-10

0.0

5.0

10.0

15.0

2.5

7.5

12.5

TGFB2

Log

−co

un

ts

RORC TGFB1 IL-10

Figure 8. NKTR-214 induces immune checkpoint and co-inhibitory genes in the tumor

Genesymbol Gene name

PDCD1 PD-1

CTLA-4 Cytotoxic T lymphocyte associated protein 4

LAG3 Lymphocyte activating 3

ICOS Inducible T cell costimulator

IDO1 Indoleamine 2,3-dioxygenase 1

TIGIT T cell immunoreceptorwith Ig and ITIM domains

MeanMedian

Figure 7. NKTR-214 induces gene expression changes in the tumor associated with T cell activation

Genesymbol Gene name

CD8A CD8 alpha chain isoform

CD8B CD8 beta chain isoform

TBX21 Th1 cell-specific transcription factor

IFNG Interferon gamma

GZMB Granzyme B

PRF1 Perforin-1

p-value ≤0.05 for all genes, comparing baseline vs wk3 means

p-value ≤0.05 for all genes, comparing baseline vs wk3 means

p-value ≥ 0.1 for all genes, comparing baseline vs wk3 means

Baseline n=14Week 3 n=11

MeanMedian

Baseline n=14Week 3 n=11

MeanMedian

Baseline n=14Week 3 n=11

0.0

2.5

5.0

7.5

10.0

12.5

CD8A CD8B TBX21 IFNG GZMB PRF1

Lo

g−

cou

nts

0.0

4.0

8.0

2.0

6.0

10.0

12.0

PDCD1 CTLA-4 LAG3 ICOS IDO1 TIGIT

Log

−co

un

ts