dna damage protein expression and migration of melanoma cells irradiated with proton beam
Download
Skip this Video
Download Presentation
DNA damage, protein expression and migration of melanoma cells irradiated with proton beam

Loading in 2 Seconds...

play fullscreen
1 / 25

DNA damage, protein expression and migration of melanoma cells irradiated with proton beam - PowerPoint PPT Presentation


  • 80 Views
  • Uploaded on

DNA damage, protein expression and migration of melanoma cells irradiated with proton beam. M. Elas 1 , S. Kędracka-Krok 1 , U. Jankowska 1 , Ł. Skalniak 1 , J. Jura 1 , E.Zuba-Surma 1 , K. Jasińska 1 , A. Pawlak 1 , U. Sowa 2 , P. Olko 2 , B. Romanowska-Dixon 3 , K. Urbańska 1

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 ' DNA damage, protein expression and migration of melanoma cells irradiated with proton beam' - makani


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
dna damage protein expression and migration of melanoma cells irradiated with proton beam

DNA damage, protein expression and migration of melanoma cells irradiated with proton beam

M. Elas1, S. Kędracka-Krok1, U. Jankowska1, Ł. Skalniak1, J. Jura1, E.Zuba-Surma1, K. Jasińska1, A. Pawlak1, U. Sowa2,

P. Olko2, B. Romanowska-Dixon3, K. Urbańska1

1Faculty of Biochemistry, Biophysics and Biotechnology, JU

2Institute of Nuclear Physics, PAS

3Department of Ophthalmology and Ophthalmic Oncology,

Jagiellonian University Medical College

slide2

Methods

BLM cells irradiated with 1-7 Gy of proton beam

58 MeV proton beam

AIC-144 cyclotron at Institute of Nuclear Physics, Polish Academy of Sciences, Kraków

LET <20 keV/µm

dose rate 0.15 Gy/s

slide3

Dose-dependent slowing down of the proliferationrate

5 & 7 Gy lethal damage

1-3 Gy repair

slide5

DNA damage

dose-dependent DNA damage

2 daysafterirradiation: no difference

5 daysafter: 11% more for 3 Gy, 14% for 5 Gy, 17% for 7 Gy

=>delayedDNA damage

slide6

Caspases activity increases with time

day 5

increase in caspases 3 and 7 activity with time

slide7

S91

S91 – day 9-10

S91 – day 5

3 Gy

3 Gy

Apoptotic Cells

Healthy Cells

proteomics
Proteomics
  • 2DE and mass spectroscopy to identifyproteinsinfluenced by 3 Gy proton beamirradiation

ctrl

3 Gy

Kędracka-Krok et al., Plos One, 2014

slide9

upregulated

downregulated

13 up, 4 down

> 1.5 x

Nucleus

VCP

MVP

STRAP

FAB-2

Lamine A/C

GAPDH

Moesin

Actinin 4

FAB-2

Vimentin

Annexin 7

Lamine A/C

Lamine B

Cytoplasm

STRAP

MCM7

Annexin 7

MVP

Caprin-1

PDCD6

VCP

HSP70

P-body

PDCD6

Caprin-1

TIM, GAPDH

VCP

Stress

granule

Mitochodrium

Kędracka-Krok et al., Plos One, 2014

slide10

DNA repair, RNA regulation

VCP

MVP

STRAP

FAB-2

Lamine A/C

GAPDH

Nucleus

Moesin

Actinin 4

FAB-2

Vimentin

Annexin 7

Lamine A/C

Lamine B

Cytoplasm

STRAP

MCM7

Annexin 7

MVP

Caprin-1

PDCD6

VCP

MVP (1.9 ), LamineA/C (1.8 ), GAPDH(2.4 ) – DNA repair

VCP (1.9 ) – chromatinassociateddegradation

MVP – PTEN translocationregulation

MVP, VCP – transcriptionregulation

STRAP (4.1 ) , FAB-2(1.9 ) – mRNAregulation

Caprin-1 (1.9), PDCD6 (1.5 )– mRNAregulation

HSP70 & G3BP1 (1.8 ) – stress & mRNA stability and stress granule formation

PDCD6

P-body

Caprin-1

TIM, GAPDH

VCP

Stress

granule

Mitochodrium

Kędracka-Krok et al., Plos One, 2014

slide11

Cell survival

VCP

MVP

STRAP

FAB-2

Lamine A/C

GAPDH

Nucleus

Moesin

Actinin 4

FAB-2

Vimentin

Annexin 7

Lamine A/C

Lamine B

Cytoplasm

STRAP

MCM7

Annexin 7

MVP

Caprin-1

PDCD6

VCP

STRAP (4.1 ) – survival - apoptosisbalance

MCM7 (1.6 )– proliferationbalance

Annexin 7(2.5 ) – proliferationinhibition

MVP – survivalenhancement

Caprin-1 (1.9) – proliferation

PDCD6 (1.5 )– apoptosis, ubiquitinationregulation

VCP (1.9 ) – aggregatesmanagement, lysosomaldegradation

PDCD6

P-body

Caprin-1

TIM, GAPDH

VCP

Stress

granule

Mitochodrium

Kędracka-Krok et al., Plos One, 2014

slide12

Cell metabolism

VCP

MVP

STRAP

FAB-2

Lamine A/C

GAPDH

Nucleus

Moesin

Actinin 4

FAB-2

Vimentin

Annexin 7

Lamine A/C

Lamine B

Cytoplasm

STRAP

MCM7

Annexin 7

MVP

Caprin-1

PDCD6

VCP

TIM (2.4 ), GAPDH(2.4 ) – glycolysis

VCP(1.9 ) – mitochondria associateddegradation

PDCD6

P-body

Caprin-1

TIM, GAPDH

VCP

Stress

granule

Mitochodrium

Kędracka-Krok et al., Plos One, 2014

slide13

Cytoskeleton and motility

VCP

MVP

STRAP

FAB-2

Lamine A/C

GAPDH

Nucleus

Moesin

Actinin 4

Fab-2

Vimentin

Annexin 7

Lamine A/C

Lamine B

Cytoplasm

STRAP

MCM7

Annexin 7

MVP

Caprin-1

PDCD6

VCP

Moesin(2.4 ) – actinremodelling, motility

Actinin 4 (1.9 ) – migration and metastasis

FAB-2 (1.9 ) – migration, microtubuledestabilizer

Vimentin (2.0, 2.1, 3.4, 1.6 ) –metastasis, EMT marker

Annexin 7(2.5 ) – motility

LamineA/C (1.6, 2,4, 1,5 ) – PI3K/AKT/PTEN, adhesion, motility

Lamine B (1.4 ) - motility

PDCD6

P-body

Caprin-1

TIM, GAPDH

VCP

Stress

granule

Mitochodrium

Kędracka-Krok et al., Plos One, 2014

proteomics1
Proteomics
  • 4 groups:
          • DNA repair & mRNAregulation
          • Survival and apoptosis
          • Glycolyticmetabolism
          • Cytoskeleton and migration
  • Signalingpathways:
      • p53
      • TGFβ
      • PTEN
      • AKT
      • BAX, Bcl-2

Kędracka-Krok et al., Plos One, 2014

slide15

Interaction with tumor microenvironment

  • Heavy ionsshown to inhibitmetastasis (Takahashi, 2003; Tsuboi, 2005)
  • After proton beamirradiation:
  • strongly inhibited matrix metaloproteinase-2 activity in highly aggressive HT1080 human fibrosarcomacells in vitro, and significantly decreased the number of pulmonary metastasis in mouse osteosarcoma in vivo (Ogata et al. 2005).
  • the expression level or activity of molecules related to metastasis such as αVβ3, β1 integrin, and MMP-2 (Takahashi, 2003)
slide16

Angiogenesisgeneactivation

Real-time PCR analysis usinghuman angiogenesis TaqMan® Array Plates

Blm cells, 3 Gy, 48 hrculture

Genes with 1.5 change vs controlshown

slide17

Migration properties of melanoma cellsafterprotonotherapy

Blm, speed

Omm1.3, speed

Omm 1.3, distance

Blm, distance

slide20

WYNIKI

Inhibition of metastases of eye-implanted BHM melanoma

x 4.4

p=0.0052

β-irradiation

Romanowska et al., ABP, 2013

slide21

Conclusions

  • 1-3Gysublethal, 5-7 Gylethaldamage of BLM cells
  • Delayed DNA damage, leading to apoptosis, resulting from endogenous ROS generationdue to cellsignalling
  • Upregulation of proteinsinvolved in DNA repair and stress, apoptosis and survival, glycolyticmetabolism and migration and cytoskeleton
  • In the lattergroup, vimentin was heavilysuppressed, together with Annexin 7, whereasexpression of Moesin 7, Lamins A/C and B, Actinin 4 and FAB-2 wereincreased
  • Interaction with tumor microenvironment: Upregulation of angiogenicgenes, in vivo inhibition of metastases in BHM eye model
slide22

Obrazowanie mysich guzów: mechanizm odpowiedzi na radio- i fototerapię

MRI, angiografia ToF

MRI, perfuzja ASL

slide23

Dept of General Biochemistry

Jolanta Jura

Łukasz Skalniak

Agnieszka Cierniak

Institute of Nuclear Physics, PAS

Paweł Olko

Jan Swakoń

Urszula Sowa

Marta Ptaszkiewicz

Dept PhysicalBiochemistry

Sylwia Kędracka-Krok

Urszula Jankowska

Dept Cell Biology

Ewa Zuba-Surma

Marta Michalik

Dept of Ophthalmology and Ophthalmic Oncology, Jagiellonian University Medical College

Bożena Romanowska-Dixon

Dept Biophysics

Krystyna Urbańska

Katarzyna Jasińska

Małgorzata Szczygieł

Martyna Krzykawska-Serda

Michał Gonet

Agnieszka Drzał

slide24

ODLEGŁE PRZERZUTY CZERNIAKA

Przerzuty do płucsą inicjowane u części zwierząt (37%)gdy guz pierwotny zajmuje zaledwie 0,10 powierzchni PK(już po 2 dniach wzrostu guza)

wczesne i p ne efekty radioterapii
WCZESNE I PÓŹNE EFEKTY RADIOTERAPII

Całkowitej regresji guza po 10 Gy 125I (4,2,2,2) towarzyszy brak przerzutów w płucach u 50% leczonych chomików nawet po 70 dniach od enukleacji

29 –34 dni po ENU

Średnica guza (mm)

100% zwierząt

70 dni po ENU

Czas (dni)

50 % zwierząt

(Sawow Aneta, 2001)

ad