The relative contribution of microenvironmental interactions to the regulation of gene expression programs in multiple myeloma cells

Liat Nadav

Molecular Cell Biology Department Weizmann Institute of Science

The Hematology InstituteTel-Aviv Sourasky Medical Center

Bentzi Katz, PhD

The Hematology InstituteTel-Aviv Sourasky Medical Center

Diversity – a hallmark of tumor progression Diversity – a hallmark of tumor progression

BM growth

Dissemination

Osteoclast activation

Drug resistance I

Drug resistance II

Drug resistance III

Diversity - is it a stochastic process, Diversity - is it a stochastic process,

or is there any direction, regulation ? or is there any direction, regulation ?

monocyte

lymphocyte

neutrophil

plateletsRBC

Bonemarrow

Bonemarrow

Regulation of Megakarypoiesis by BM nichesRegulation of Megakarypoiesis by BM niches (Rafii S. et al., 1994-2007)(Rafii S. et al., 1994-2007)

Soluble factorsCellular interactionsAdhesive interactions

Can adhesive interactions generate diversity of Can adhesive interactions generate diversity of malignant plasma cells ?malignant plasma cells ?

What is the physiological significance of such a process ?What is the physiological significance of such a process ?

Isolation of adhesive variantsIsolation of adhesive variantsfrom the ARH-77 linefrom the ARH-77 line

Type-F cells(floating)

Type-A cells(adherent)

No

. of

even

ts

101 102 103 104

CD138

Type-AType-F

Flow cytometry pattern

Type-A cells

Type-F cells

Differential gene expression in adhesive variantsDifferential gene expression in adhesive variants

I

II

III

IV

V

I

II

III

IV

V

5 individual enrichments RNA purification Affymetrix array

0

10

20

30

40

50

Per

cent

of

gene

s in

eac

h fu

nctio

n ca

tego

ry

Metabolism

Signal transduction

Transport

Differentiation andDevelopment

Transcription factors

Apoptosis

Cell cycle

Adhesion

Motility

Immune response

Undefined

Color Code

Type-A cells Type-F cells

Assignment of gene expressed in adhesive variantsAssignment of gene expressed in adhesive variants

Coding region

Regulatory region

Regulatory elements

Gene AGene BGene CGene D

RNA from cell I

Pathway X is active in cell I

Gene AGene BGene CGene D

RNA from cell II

Pathway Y is active in cell II

Signaling pathways

Promotor analysis

Promotor analysis

Type-A cells Type-F cells

?NFB (CCND1, CXCL11, IL6)

EGR(c-jun, EGR1, EGR2)

NFB1

SRF

NFB2

L1

L2L3

L4

L5L6L7

L8

L9L10

H1

H2H3

H4

H5H6

H7

H8

H9

0

2

4

6

8

10

12

14

0 2 4 6 8 10 12

Ave

rage

exp

ress

ion

in T

ype

F

Average expression in Type A

B-Cell differentiation state

HighLow

H1- GADD45AL1- TIMP1

H2- IgH VDJL2- BCL6

H3- IgH variableL3- IL-6

H4- Ig rearrangedL4- EGR4

H5- IgH HML5- TNF

H6- IgH JL6- TNFRSF2

H7- IgH G1-4L7- TNFRSF4

H8- IgH ML8- TNFRSF9

H9- IgH M1-2L9- MYCN

L10- BTK

Cell adhesion regulates differentiation of plasma cellsCell adhesion regulates differentiation of plasma cells

GAPDH

F A

IgJ

GAPDH

F A

EGR1

Type-A cells

AF cells

GAPDH

EGR1

IgJ

Type-A cells

AF cells

Regulation of gene expression is reversibleRegulation of gene expression is reversible

Lapsed time )days(

Tum

or F

ree

mic

e )%

(

0

20

40

60

80

100

20 40 60 80 100 120

Type-A cells

Type-F cells

Malignancy potential of plasma cell variants Malignancy potential of plasma cell variants

1. Microenvironmental interactions can generate diversity of malignant plasma cells

2. Directional diversity – differentiation

3. Specific putative targets: NFB, EGR )Type-A(

4. Type-A cells – are they represent “stem cells” or “stem state” ?

Phenotype control by microenvironmental interactions Phenotype control by microenvironmental interactions

CD138

CD

38

Co

ntr

ol

103

102

101

101 102 103

Mec

han

ical

+en

zym

es

103

102

101

Spicules

Patient BMs also contain diverse MM cellsPatient BMs also contain diverse MM cells

Fluid

MM bone marrow

Thanks…Thanks…

Benny GeigerMolecular Cell Biology Department

Weizmann Institute of Science

Tal Shay, Eytan Domany

Physics of Complex Systems Department Weizmann Institute of Science

Ella NaparstekShoshie Baron

The Hematology InstituteTel-Aviv Sourasky Medical Center