Download - AXL Inhibition Improves Immunotherapy by …...Agata Rybicka3, Muntequa I. Siraji1,2, Sushil Dhaka1,2, Oddbjørn Straume2,4, Michael A. Curran5, Rolf A.Brekken6, Gro Gausdal3, James

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Page 1: AXL Inhibition Improves Immunotherapy by …...Agata Rybicka3, Muntequa I. Siraji1,2, Sushil Dhaka1,2, Oddbjørn Straume2,4, Michael A. Curran5, Rolf A.Brekken6, Gro Gausdal3, James

References: 1Gubin et al., 2018. 2Chen and Mellman, 2017. 3Lemke and Rothlin, 2008. 4Ye et al., 2010. 5Chiu et al., 2015. 6Davidsen K.T et al., 2017. 7Terry et al., 2017.

ICB + bemcentinib targets tumor-immune crosstalk AXL is highly expressed in suppressive myeloid cells AXL+ M2φs are reduced by bemcentinib treatmentAXL is expressed in distinct DC populations with highly suppressive profiles

Conclusions

••••

NK cells

NK-S1

NK-S2NK-S3

NK-S5

CD11b-

CD11b+

NK-S4 Ly6G

+

MHC

II

+

-

iNOS+pDCs

DC-S9

DC-S8

CD4+

CD4-

DC

-S5

CD4+ DC-S4

CD206+ DC-S7

cDC2

DC-S1

CD80+

DC-S6

DC-S3

DC-S2

CD24+CD8+cDC1

Neutrophils

Immature cDC2

PD

L1high

DC-T3DC-T4

Ly6C+(pDCs)

CD4+

CD4-

DC-T1

Neutrophils

CD103+cDC1

CD80 low

DC-T8

DC-T9

CD80 int

CD11b+MHCII-CD24+

DC-T2

cDC2

DC-T5

DC-T6

DC-T7

Arg

-1+

Axl

+

NK cells

NK-T2

NK-T3

NK-T

1

CD11bCD

11b-

+

-

Ly6C

+

-

Spleen Tumor

Force-directed layout of X-Shift clusters visualizes rare cell populations in spleens and tumors.

Lorem ipsum

AXL is Expressed in Rare Dendritic Cell Populations

T1Sp

leen

Axl MerTK

T2Sp

leen

Axl MerTK

Tumo

rT1

T2Tu

mor

Uniform Manifold Approximation and Projection (UMAP ) of single immune cells showing AXL and MerTK expression at timepoint 1 and timepoint 2 in spleens and tumors.

Tumors mobilize AXL in Suppressive Myeloid Cells

CD4 T cells

CD8 T cells

gMDSC

NK cells

B cells

Macrophages/monocytes

Macrophages/monocytes

T1

T2

Spleen Tumor

BALB/C mice bearing established 4T1 mammary tumors were treated as indicated, n=5 per treatment per timepoint. Spleens and tumors were analyzed by CyTOF using a 34- marker myeloid focused panel. Graphs show single immune cells mapped with UMAP (left; colored by marker expression of CD11b, right;colored by density showing changes over time in immune populations in untreated animals)

Day 0 Day 12 Day14 Day 16 Day 18 Day 22

αCTLA4/αPD1

Bemcentinib

Rand

omiza

tion

T1T2

ET2ET1

High Dimensional Single Cell Immune Phenotyping

CSF1

mRN

A fol

d cha

nge

relat

ive to

ctr

0

10

20

30

CSF1

αCTLA4/ αPD 1 αCTLA4/ αPD1/Bem

Non- responder s Responder s

P = 0.0416 Fold change of CSF1 gene expression in 4T1 tumors unresponsive to ICB (n=4) or ICB and bemcentinb (n=5) and responsive tumors treated with ICB/bemcentinib (n=2), compared to vehicle

AXL Inhibition Blocks Tumor-Myeloid Crosstalk

0 50 100 1500

50

100

Days of treatment

Perce

ntsu

rviva

l

Vehicle n= 1 1Bemcentinib n=1 2

αCTLA4 /αPD1 n=17αCTLA4 /αPD1/Bemcentinib n=2 6

p = 0.0189

Durable tumor clearance in 25.6% of animalswith established orthotopic 4T1 mammary tumors treated with bemcentinib + anti-CTLA-4 + anti-PD-1 vs 5.6% of animals treated with anti-CTLA-4 + anti-PD-1

AXL Inhibition Potentiates the Effect on Immune Checkpoint Inhibitors

Tumor

Single immune cells clustered with PhenoGraph and analyzed with Marker Enrichment Modeling (MEM). Heat map of MEM-scores with changes in individual PhenoGraph clusters over time and between treatments.

619141334

122320252

1617101

159

2411217

1858

22

CD

206

CD

11b

CD

3

CD

4

Ly6C

MH

CII

CD

80Ly

6G-C

CD

24

CD

40iN

OS

CD

11c

CD

19

CD

25A

xl

Mer

TK

CD

49b

CD

69

CD

86

CD

45P

D1

CTL

A4

CD

83

CD

8a

PD

L1

CD

335

CD

114

Arg

-1

CD

115

CD

64

F4/8

0

CD

103

CD

116

CD

206

CD

11b

CD

3

CD

4

Ly6C

MH

CII

CD

80Ly

6G-C

CD

24

CD

40iN

OS

CD

11c

CD

19

CD

25 Axl

Mer

TK

CD

49b

CD

69

CD

86

CD

45P

D1

CTL

A4

CD

83

CD

8a

PD

L1

CD

335

CD

114

Arg

-1

CD

115

CD

64

F4/8

0

CD

103

CD

116

T1 T2

Ctr

Bem

ICB

ICB

+ B

em

ICB

+ B

emCtr

Bem

ICB

gMDSC

NK cells

DCs

mMDSC

CD4 T cells

Axl+MerTK+Mφ/M2φ

gMDSC

pDCs

Lin-

B cells

B cells

Axl+MerTK+Mo/Mφ

CD8 T cellsCD4-CD8-T cells

CD

116

CD

80C

D11

cF4

/80

CD

206

CD

64M

HC

IIC

D11

bC

D24

Ly6G

-CLy

6C Axl

Mer

TKiN

OS

Arg

-1C

D11

5C

D69

PD

L1C

D10

3C

D86

CD

3C

TLA

4C

D49

bC

D40

CD

45P

D1

CD

83C

D33

5C

D25

CD

114

CD

19C

D4

CD

8

CD

116

CD

80C

D11

cF4

/80

CD

206

CD

64M

HC

IIC

D11

bC

D24

Ly6G

-CLy

6CA

xlM

erTK

iNO

SA

rg-1

CD

115

CD

69P

DL1

CD

103

CD

86C

D3

CTL

A4

CD

49b

CD

40C

D45

PD

1C

D83

CD

335

CD

25C

D11

4C

D19

CD

4C

D8

1819

136

20

1115

16251410

2422

85

12

34

97

1723

Axl+ M2φint. Mφ

iNOS+Arg-1+MφMerTK+M2φ

M2φ

Ly6C high Mo/MφCD49b+Axl+MDSC

CD24+ Mo/Mφ

gMDSC

pDCLy6C+ CD8 T cells

DCsMo/Mφ

Lin-

CD4-CD8-T cells

CD8 T cells

CD4 T cells

NK cellsLin-

T1 T2

Ctr

Bem

ICB

ICB

+ B

em

ICB

+ B

emCtr

Bem

ICB

Spleen

AXL Inhibition Reshapes the Immune LandscapeBackground

Tumor mobilization of suppressive myeloid cells remains a primary obstacle to immune checkpoint blockade (ICB) efficacy(1).

Tumor microenvironments enriched in suppressive myeloid cells are often T cell infiltrated(2) providing a rationale for ICB combination strategies with myeloid targeting agents.

The receptor tyrosine kinase AXL is a key tolerogenic regulator of the innate immune response(3).

Tumor associated macrophages (TAM) express AXL(4) and mediates M2-macrophage polarization(5).

In cancer AXL is associated with epithelial-to-mesenchymal (EMT) -related phenotypic plasticity and therapy resistance (6).

EMT is increasingly recognized as a major contributor to tumor immune evasion(7).

The small molecule AXL inhibitor bemcentinib targets tumor cell intrinsic phenotypic plasticity and myeloid immune suppression.

Bemcentinib in combination with ICB is currently in phase II trials for NSCLC (NCT03184571), melanoma (NCT02872259) and mesothelioma (NCT0365483).

AXL Inhibition Improves Immunotherapy by Targeting Local and Systemic Tumor Myeloid Cross-Talk Kjersti T.Davidsen1,2, Katarzyna Wnuk-Lipinska3, Sturla M. Grøndal1,2, Noelly Madeleine1,2, Stacey A. D’mello Peters1,2, Magnus Blø3, Linn Hodneland3, Lavina Ahmed3, Agata Rybicka3, Muntequa I. Siraji1,2, Sushil Dhaka1,2, Oddbjørn Straume2,4, Michael A. Curran5, Rolf A.Brekken6, Gro Gausdal3, James B. Lorens1,2

1 Department of Biomedicine, 2 Centre for Cancer Biomarkers, University of Bergen, 3 BerGenBio ASA, 4 Department of Oncology, Haukeland University Hospital, 5 Department of Immunology, The University of Texas MD Anderson Cancer Center, 6 Department of Surgery and Hamon Center for Therapeutic Oncology Research, UT Southwestern Medical Center