slide1 n.
Download
Skip this Video
Loading SlideShow in 5 Seconds..
Experimental Medicine Program Student Research Day 2014-06-25 PowerPoint Presentation
Download Presentation
Experimental Medicine Program Student Research Day 2014-06-25

Loading in 2 Seconds...

play fullscreen
1 / 19

Experimental Medicine Program Student Research Day 2014-06-25 - PowerPoint PPT Presentation


  • 145 Views
  • Uploaded on

Targeting Treatment Naïve CML Stem/Progenitor Cells from Imatinib-Nonresponders by Combination Treatment with JAK2 (BMS-911543) and ABL Inhibitors. Leon ( Hanyang Lin) , Min Chen, Katharina Rothe, Matthew V. Lorenzi, Adrian Woolfson and Xiaoyan Jiang

loader
I am the owner, or an agent authorized to act on behalf of the owner, of the copyrighted work described.
capcha
Download Presentation

PowerPoint Slideshow about 'Experimental Medicine Program Student Research Day 2014-06-25' - nomlanga-kirby


An Image/Link below is provided (as is) to download presentation

Download Policy: Content on the Website is provided to you AS IS for your information and personal use and may not be sold / licensed / shared on other websites without getting consent from its author.While downloading, if for some reason you are not able to download a presentation, the publisher may have deleted the file from their server.


- - - - - - - - - - - - - - - - - - - - - - - - - - E N D - - - - - - - - - - - - - - - - - - - - - - - - - -
Presentation Transcript
slide1

Targeting Treatment Naïve CML Stem/Progenitor Cells from Imatinib-Nonresponders by Combination Treatment with JAK2 (BMS-911543) and ABL Inhibitors

Leon (Hanyang Lin), Min Chen, Katharina Rothe, Matthew V. Lorenzi, Adrian Woolfson and Xiaoyan Jiang

Terry Fox Laboratory, BC Cancer Agency Department of Medicine, UBC

Vancouver, Canada

Discovery Medicine Oncology, Bristol-Myers Squibb, Princeton, United States

Experimental Medicine Program Student Research Day 2014-06-25

chronic myeloid leukemia cml
Chronic Myeloid Leukemia (CML)

Chronic Phase (CP)

Accelerated Phase (AP)

Blast Phase (BP)

3-5 years

0.1-1 years

Adapted from Ren R, Nature Reviews, 2005

slide3

Chronic Myeloid Leukemia (CML)

p210BCR-ABL

BCR

ABL

Y177

Y1294

Tyrosine Kinase

DNA BD

Actin

BD

SH2

NLS

SH3

S/T kinase

Rho-GEF

PI3K

JAK2

Ras

AKT

STAT5

MAPK

Increased proliferation

Decreased apoptosis

Massive accumulation of myeloid cells in circulation

(Major characteristic of CML)

Sattler et al, Cytokine Growth Factor Rev 8:63, 1997

slide4

Challenges in CML Treatment:

CML Stem Cells Are Insensitive to TKIs

IM

IM

IM

  • Major problems associated with TKI therapy:
  • - Early relapse, acquired drug resistance
  • Ineffectiveness in eliminating CML stem cells

Current 1st line: Imatinib

Alternatives:Dasatinib, Nilotinib

Druker BJ Blood , 2008

slide5

AHI-1-BCR-ABL-JAK2 Complex Regulates TKI Response/resistance of

CD34+ CML Cells

  • Current Challenges:
  • TKI Resistance of CML Stem Cells

BCR-ABL-dependent mechanisms:

Increased BCR-ABL expression

BCR-ABL mutations

Increased drug efflux and reduced drug influx

Genomic instability

Ctrl

AHI-1sh4

Imatinib 5 µM

80

60

% inhibition relative

to untreated cells

BCR-ABL-independent mechanisms:

Not addicted to BCR-ABL for survival

Protected by BM microenvironment

Activation of pro-survival pathways: Autophagy, Wnt//β-catenin, etc

40

20

0

Non-responders

Responders

Blast crisis

Chomel et al, Oncotarget, 2011

Jiang et al, J Natl Cancer Inst, 2007;

Zhou et al, J Exp Med, 2008

Corbin et al, JCI, 2011; Hamilton et al, Blood 2012

slide6

Hypothesis and Objectives

Relapse

Stem cells are insensitive to TKIs

TKIs

TKIs

JAK2 inhibitor

(BMS-911543)

JAK2

BCR-ABL

AHI-1

Combined suppression of BCR-ABL and JAK2 activities to destabilize this protein complex may be more effective at eliminating CML stem cells

slide7

Combination Treatment with ABL and JAK2 Inhibitors Is More Effective at Reducing pSTAT5 Levels in CML Cells

1 2 3 4 5 6 7 8

K562

P < 0.05

pSTAT5

STAT5

GAPDH

2 hours after drug treatment

Relative pSTAT5 levels

1 K562 DMSO Ctrl

2 K562 1.0 µM BMS

3 K562 0.05 µm IM

4 K562 IM + BMS

5 K562R DMSO Ctrl

6 K562R 1.0 µM BMS

7 K562R 0.05 µm IM

8 K562R IM + BMS

K562R (IM-resistant cells)

P < 0.05

P < 0.05

n=3

slide8

Combination Treatment Results in a Significant Reduction in Colony Formation of CML Cells

Total CFC

P < 0.05

P < 0.05

CML cell lines

(600 cells)

No. of CFC per 200 input cells

Methylcellulose ± inhibitors

No drug

No drug

No drug

No drug

BMS-911543

BMS-911543

BMS-911543

BMS-911543

BMS-911543

+ IM

BMS-911543

+ IM

BMS-911543

+ IM

BMS-911543

+ IM

IM

IM

IM

IM

12 days

37ºc

P < 0.05

P < 0.05

200 cells

200 cells

No. of CFC per 200 input cells

Small colonies: <50 cells

Medium colonies: 50-300 cells

Large colonies: >300 cells

slide9

Combination Treatment Is More Effective at Reducing pSTAT5 and pCRKL Levels in

  • CD34+ CML Cells

P<0.05

P<0.05

P<0.05

P<0.01

P-STAT5 relative

to untreated cells (%)

P<0.05

P<0.01

P<0.05

No drug

P-CRKL relative

to untreated cells (%)

DA

BMS

DA + BMS

IgG Ctrl

48 hrs treatment

72 hrs treatment

n=4

slide10

Combination Treatment Results in a Significant Increase in Apoptosis of CD34+ CML Cells

P<0.05

48 hrs

Ctrl

BMS

BMS

+ DA

BMS

+ DA

BMS

BMS

BMS

+ IM

BMS

+ IM

IM

IM

DA

DA

Apoptotic cells above untreated (%)

BMS

+ NL

BMS

+ NL

NL

NL

P<0.05

PI

P<0.05

DA

BMS+DA

P<0.05

Annexin V

72 hrs

n=3

slide11

Combination Treatment Is More Effective at Reducing Colony Formation in Treatment-naïve CD34+ CML Cells from Subsequent IM-nonresponders

Total CFC

CML blood samples

P<0.001

Methylcellulose± inhibitors

P<0.001

P<0.001

% CFCs relative to

untreated cells

Select

CD34+ cells

CD34+ cells

12 days

CFU-GM

BFU-E

P<0.001

P<0.001

P<0.01

P<0.001

P<0.05

% CFCs relative to

untreated cells

% CFCs relative to

untreated cells

P<0.01

n=10

slide12

Combination Treatment Is More Effective at Eliminating More Primitive CML Cells from Subsequent IM-nonresponders

CML blood samples

Select

CD34+ cells

Total LTC-IC derived CFCs relative to

untreated cells (%)

P<0.05

CD34+ cells

P<0.05

On feeders for

6 wks  inhibitors

Methylcellulose

12 days

37ºc

n=3

slide13

BMS-911543 Has Less Toxicity on CD34+ Normal Bone Marrow Cells than CML Cells

Total CFC

P<0.001

P<0.001

% of CFCs relative to untreated cells

P<0.01

n=7

slide14

In Vivo Combination Effects

Day 55

NSG mice age 7-10 weeks

% Survival

P<0.001

Sub-lethal cesium irradiation

Days after transplantation

I.V. injection of BV173 cells

Weight (grams)

Oral gavage drug treatments

Days after transplantation

slide15

In Vivo Combination Effects

Ctrl

Vehicle

BMS

DA

DA+BMS

Ctrl

Vehicle

BMS

DA

DA+BMS

Day 55

Spleen

CD19

1.0 mm

0.03

0.19

0.21

0.03

0.02

Liver

1.9

3.15

4.53

1.62

1.47

1.0mm

Vehicle

BMS

DA

DA+BMS

Ctrl

BM

0.07

28.9

33

1.34

0.53

CD19

FSC

BCR-ABL transcript levels

BM

Spleen

Liver

P<0.01

Fold difference relative to vehicle

P<0.05

ND

ND

ND

ND

ND

ND

slide16

In Vivo Combination Effects

Day 70

NSG mice age 7-10 weeks

% Survival

P<0.001

Sub-lethal cesium irradiation

Days after transplantation

I.V. injection of BV173 cells

Weight (grams)

Oral gavage drug treatments

Days after transplantation

slide17

In Vivo Combination Effects

Vehicle

DA

DA+BMS

Vehicle

DA

DA+BMS

Day 70

Spleen

0.14

0.11

0.03

CD19

1.0 mm

Liver

5.55

4.17

1.78

1.0mm

Vehicle

DA

DA+BMS

34.7

26.3

1.17

BM

CD19

FSC

BCR-ABL transcript levels

BM

Spleen

Liver

P<0.001

P<0.001

P<0.01

Fold difference relative to vehicle

slide18

Conclusions

TKIs

Y177

P

Y177

P

BMS-911543

JAK2

BCR-ABL

AHI-1

STAT5

STAT5

Stem cell proliferation & survival

TKI response

Stem cell proliferation & survival

TKI response

JAK2

BCR-ABL

  • Combination treatment:
    • More effectively reduces pSTAT5 and pCRKL levels
    • Increases apoptotic cells and inhibits colony growth in CD34+ CML cells
    • Prolongs survival of leukemic mice

AHI-1

Targeting both BCR-ABL and JAK2 activities may be a more effective therapeutic option for CML patients

slide19

Acknowledgments

Jiang Lab Members:

Dr. Xiaoyan Jiang

Dr. Min Chen

Katharina Rothe

Dr. Kevin Lin

Sharmin Esmailzadeh

Will Liu

Rachel Huang

Damian Lai

Stem Cell Assay Lab:

Karen Lambie

Helen Nakamoto

Kyi Min Saw

Animal Resource

Centre

FACS Facility

CML Working Group:

Dr. Connie Eaves

Dr. Donna Forrest

BMS:

Dr. Matthew Lorenzi

Dr. Adrian Woolfson

Collaborators:

Dr. T. Holyoake, Scotland

Dr. A.Turhan, France

Dr. R. Arlinghaus, Houston