Combination Immune Checkpoint Inhibitors for the …tumor-models.com/.../2016/07/1245-Joe-Murphy-YES.pdf · Combination Immune Checkpoint Inhibitors for the Treatment of Colon Carcinoma

Combination Immune Checkpoint Inhibitors for

the Treatment of Colon Carcinoma in Humanized

Mice

Joseph Murphy, PhD

Director of Science, R&D Oncology

CTLA-4

CD28

PD1

CD8/TCR

CD8 T effector

Anti-PD1

Anti-CTLA-4

Block PD1 and CTLA-4 signaling in T effector and Tregs:

• Prevent T effector cell exhaustion and death

• Decrease Treg activity and proliferation

• Induction of Treg death by ADCC

• Create a favorable CD8 T-effector:T reg ratio

Effects of PD1 and CTLA-4 Signaling

ADCC/macrophage

Anti-CTLA-4

Tumor

Lck

ZAP70

class I

PI3K Akt

CD28

TCR

CD4

CTLA-4

Treg

PD-L1

Tumor cell

CD80/86

APC

PD-L2

SHP2

PP2a

SHP2

Two Platforms Available:

- CD34+ Embryonic Cell Engrafted (Jackson Lab)

- PBMC Engrafted

huCD34 model

- Week 3: huCD34+ HSC (~3x104)

- Umbilical cord blood

- Week ~15: >25% huCD45 in blood

- Long term studies

huPBMC model

- huPBMC injected (30x106)

- Week 3: ~30% huCD45 blood cells

- Short term studies

- T cell focused

- GVHD appears

huPBMC

Myeloablation (1 Gy)

huCD34

Cell line % CD80 + % CD86 + % CTLA-4 + % PD-L1 + % PD-L2 +

A375 0.0 0.0 0.0 66.8 0.0

HT-1080 0.0 0.0 0.0 98.2 0.4

SK-MEL-5 0.0 0.0 0.0 9.9 0.0

HCC827 0.0 0.0 0.0 94.8 0.6

Colo205 0.0 0.0 0.0 40.8 0.0

SK-MEL-24 0.0 0.0 0.0 8.4 0.0

RKO 0.0 0.0 0.1 99.9 22.0

MDA-MB-231 0.0 0.0 0.1 99.7 0.7

HCT116 0.1 0.0 0.0 60.5 0.0

Calu6 1.2 0.0 0.0 53.8 0.0

SKOV3 0.0 0.0 0.0 73.1 0.8

ME-006 0.0 0.0 0.9 86.5 5.7

ME-012 0.0 0.0 0.0 15.4 0.4

ME-016 0.0 0.0 0.2 7.6 0.0

TH-005 0.0 0.0 0.4 98.3 9.3

NS-036 0.0 0.0 0.0 80.9 1.5

A2058 0.0 0.0 1.1 55.0 0.6

% subtracted from Isotype Control

Evaluation of the Expression of PD-L1, PD-L2, CTLA-4,

CD80 and CD86 in human tumor cell lines and PDX

CD34+ Cell Engrafted Studies - RKO Colon Tumor Implant (High PD-L1 Expression)

- HLA-A*01/01 matched

- Treatment with Ipilimumab and/or Pembrolizumab

Exp. I Exp. II

Initial Assessment of human immune cell engraftment

Percentages of hCD45 and mCD45 populations were

calculated relative to the live cell gate in individual samples.

Percentages of hCD3 and mCD3 (top) were calculated relative

to hCD45 and mCD45. hCD4 and hCD8 (bottom) populations

were calculated relative to hCD3 cells in individual samples.

CD34+ NSG mice engrafted studies: Exp. I and Exp. II

Control IgG 200mg biwk x3

Ipi 100mg biwk x3

Pembro 100mg biwk x3

Ipi + Pembro

Non-engrafted - NOG

RKO-e210 blood sample for human immune cells day 15

p<0.02

p<0.004 p<0.02

p<0.03

RKO-e210 blood sample for human immune cells day 15

p<0.02

CD45RO

CC

R7

Effector

Central

Memory

Effector

Memory

Naive

Control

Gate on CD8+ cells

CD45RO

CC

R7

Effector

Central

Memory

Effector

Memory

Naive

Ipi+Pembro

Gate on CD8+ cells

p<0.002

p<0.02

p<0.02

Distribution of T cell subsets

RKO-e210 tumor response to immune checkpoint

inhibitors day 15

RKO-e210 human immune TILs at day 15

p<0.02

p<0.005

p<0.002

CD34+ NOG mice Non-HLA-A matched + Pembrolizumab

RKO-e219

HLA-A matched Non HLA-A matched

Summary and conclusions

• Pembrolizumab and ipilimumab monotherapies significantly inhibit tumor growth in

human RKO colon carcinoma xenograft in the CD34+ NSG humanized mouse model.

• Combination therapy does not enhance the efficacy demonstrated by either

monotherapy.

• Activated effector T cells appear to play a role in tumor rejection

• mMDSC, gMDSC and macrophages poorly represented in this model (data not

shown).

Questions?

Call with questions: 1-877-274-8371

[email protected]

www.criver.com