Cristel V Camacho, PhD

Sandeep Burma, PhDDepartment of Radiation Oncology

June 27, 2013

Genetic Events Underlying Radiation-Induced Gliomagenesis

The Space Radiation Problem

Galactic Cosmic Rays (GCR) • Protons (85%)• Alpha particles (14%) • High atomic number and high energy (HZE) particles (1%)

science1.nasa.gov

Solar Energetic Particles

Galactic Cosmic Rays

Solar Energetic Particles • Solar particle events• Coronal mass ejections

NASA needs to close the knowledge gap on a broad range of biological questions

X-rays 1 GeV/n Fe

γ

science1.nasa.gov

Ionizing Radiation and Carcinogenesis

NASA Space Radiation Laboratory (NSRL)

I. DNA damage responses to gamma- versus particle- irradiation

II. Cellular transformation by gamma- versus particle- irradiation

III. Mouse models of radiation-induced gliomagenesis

science1.nasa.gov

Mukherjee B, Camacho CV, et al., DNA Repair 2008

I. DNA damage responses to HZE particles versus gamma-raysG

amm

a1

GeV

/n F

e

• Fe ions produce more complex DNA damage• Fe ion-induced damage is incompletely repaired

- persistent DNA damage signaling- damage-induced senescence- reduced cell survival

0 1 2 3 40.001

0.01

0.1

1

Gamma rays

1 GeV/n Fe

Dose (Gy)

Su

rviv

al

• What are the long term carcinogenic consequences of persistent DSBs such as those induced by Fe ions?

Sur

viva

l

Dose (Gy)

1 GeV/n FeGammaMock

~3% ~84%~24%

1 GeV/n FeGamma

p53

actin

p21

p16/Ink4a

p53 (ser15)

0.5

h24

h

1 GeV/n Fe

γH2AX

53BP1

γH2AX

1 GeV/n Fe

Gamma

% f

oci r

em

aini

ngTime (hrs)

NSCs

Astrocytes

II. Ink4a/Arf-null astrocytes: A sensitive paradigm for rapidly evaluating the tumorigenic potential of HZE particles

IR

Tumor suppressor lossInk4a/Arf-/-

Oncogene activation Tumorigenesis

McEllin B, Camacho CV, et al., Cancer Research 2010Mukherjee B, McEllin B, Camacho CV, et al., Cancer Research 2009

p19/Arf

Exon 1β Exon 1α Exon 2 Exon 3

p16/Ink4a Cyclin DCDK 4/6

pRB

MDM2 p53IR Ink-/-, Pten-/-,EGFR; NSC

Gam

ma

4Gy

Fe

1 G

yMock IR

0 20 40 600

50

100

150

200

250FeGamma

Mock (0/8)g 1Gy (0/8)g 4Gy (2/8)Fe 1 Gy (12/12)

DAY

Ink4a/Arf-/- astrocytes irradiated with Fe ions are potently tumorigenic

Vo

l m

m3

Exon 1β Exon 1α Exon 2 Exon 3

p16/Ink4a

p19/Arf

Exon 2Exon 1

p15/Ink4b

#2#1

Tumor

Ch 4

0 10 20 30 400

100

200

300

400

500

p 1 5C o n tro l C o n tro l (12 /12 )

p 15 (12 /12 )

D AY

Vo

lmm

3

Ctrl p15

Fe 1 Gy (acute)3 months post-IR

Mock Fe

Days Post-IR

0 100 200 3000

20

40

60

80

100

Fe (n=40) (10)

Mock-IR (n=40) (0)

NesCre; Ink4ab-/-/Arf f/f

X-Rays (n=40) (10)Tum

or f

ree

surv

ival

III. Transgenic mouse models of radiation-induced gliomagenesis

Ink4a/Arf f/f Ink4a-/-;Ink4b-/-;Arf f/f Ink4a-/-;Ink4b-/-;Arf f/fPTEN f/+

Ink4a+/+;Arf+/+

Rapid induction of high-grade gliomas by DNA double-strand breaks

Genomic amplification of Met is a frequent event in radiation-induced gliomas

Met

Amp

Non-Amp

Met FISHAmp

Non-Amp

Met

pMet

pMet

Sox2

Actin

Par βGal Met

V5-tagged

Met

V5

βGal MetMet

Sox2

Merge

βGal Met

NS

CS

oft-

agar

βGal Met

Project Overview

Nestin-Cre Ink4a/Arf f/f

Nestin-Cre Ink4ab-/-;Arf f/f

Nestin-Cre Ink4ab-/-;Arf f/f

PTEN f/+Nestin-Cre

Ink4a+/+;Arf +/+

Mock-IR

1Gy Fe (600 MeV/n)

4Gy Gamma-rays

40

40

40

40

40

40

40

40

40

40

40

40

HZE particles vs Gamma-rays

Molecular Profiling and Validation

RNA Expression ProfilingWhole Genome aCGH Copy

Number Analysis

Radiation Protection/Mitigation

Acknowledgements

Sandeep Burma, PhDBipasha Mukherjee, PhDNozomi Tomimatsu, PhDXiaohuan Gao, PhDBrian McEllin, PhDMariya IlchevaCarlos Gil del AlcazarPavlina TodorovaMolly Gillam

UTSWMichael D Story, PhD Liang-Hao Ding, PhDDebabrata Saha, PhD Amyn A Habib, MDXian-Jin Xie, PhDKimmo Hatanpaa, MD, PhD

Adayabalam S Balajee, PhD (Columbia U.)

Funding

CBP Training Grant (PI: Jerry Shay, PhD)

NASA

BNL/ NSRLPeter Guida, PhDAdam Rusek, PhD

Thesis CommitteeRobert M Bachoo, MD, PhDDavid A Boothman, PhD Chaitanya S Nirodi, PhD

Past MembersBrandon Hahm, MDNandini PichamoorthyKaushik AmancherlaKatherine Deland