ATM-Deficient Colorectal Cancer Cells Are Sensitive to the PARP Inhibitor Olaparib

Elisa Orozco GómezMaría Catalina Soto López

Molecular biology2017

INTRODUCTION

DEFICIENCY OR MUTATION

p53

ATM

BRCA 1/2

SENSIBILITY TO PARP INHIBITORS (OLAPARIB)

CANCER TREATMENT

CANCER Is a disease caused by abnormal growth because of the uncontrolled increase of cells in an organism

Cells become:-Abnormal SURVIVE DIVISION OF NEW CELLS -Old -Damaged TUMOR

COLORECTAL CANCER Is the cancer from the large intestine, specifically of the colon or the rectum.

ATAXIA

Lack of muscle coordination which may affect speech, eye movements, the ability to swallow, walking, picking up objects, and other voluntary movements

CAUSE: damage, degeneration or loss of nerve cells in the cerebellum.

TELANGIECTASIA

Small dilated blood vessels, near the surface of the skin or mucous membranes.

ATM GENEGene localized in 11q22-23 which codifies for the protein: ATM serine/threonine kinase, which is called to the site where a double strand break of the DNA has occurred.

It causes the phosphorylation of proteins which activate the DNA damage checkpoint, which will cause:● CELL CYCLE ARREST● DNA REPAIR● APOPTOSIS

ATM SYNDROME (Louis-Bar syndrome)

● ATM gene is mutated or missing (in both copies of the parent in each cell)● Autosomal recessive trait, childhood disease

NEURODEGENERATION

IMMUNODEFICIENCIES

CANCER PREDISPOSITION

PARP(Poly ADP ribose polymerase)

➔ Repairs Single Stranded DNA breaks by BER.

Recruit XRCC1, DNA ligase III, DNA polymerase beta, and a kinase.

BER(base excision repair)

1. DNA transcription.

2. Cell cycle regulation.

3. DNA repair

OLAPARIB(PARP inhibitor)

1. Induces lethality in BRCA 1/2 deficient tumor cells.

2. Formation of Double Stranded DNA breaks.

3. No reparation.4. Disruption of cellular homeostasis adn

cell death.

DNA DAMAGE

Normal cell:PARPBRCA 1/2

DNA REPAIR

Cancer cell: mutated BRCA ½ NORMAL PARP

OLAPARIB (-) PARP

CELL DEATHATM SYNDROME

ATM Gene

OBJECTIVE● Figure out if ATM deficiency might sensitize different CRC cell lines to

PARP inhibitors, in this case OLAPARIB.

● Prove the absence of p53 by targetting CRC cells with PARP inhibitors and ATM inhibitors.

MATERIALES Y MÉTODOS

CULTIVO CELULAR

CULTIVO CELULAR

•MEDIO RPMI 1640

•5% SUERO BOVINO

FETAL

•2mM L GLUTAMINA

•100 U/mL PENICILINA

•100 ug/mL

ESTREPTOMICINA

TRANSFECCION

TRANSFECCION

PARA LA IRRADIACION

INMUNOTRANSFERENCIA

INMUNOTRANSFERENCIA

Técnica utilizada para identificar una proteína en una muestra que

contiene varias proteínas

EN QUÉ CASOS SE PUEDE UTILIZAR1. ¿Está la proteína de interés presente en mi muestra?2. El tratamiento con un determinado agente “X”, ¿aumenta o reduce el nivel de expresión

de mi proteína? 3. 3 ¿Cómo varía el nivel de expresión de la proteína en distintos tejidos o líneas celulares?

¿Mi muestra es sana o patológica?

ENSAYOS DE SUPERVIVENCIA CLONOGENICOS

P< 0.05

ENSAYOS DE SUPERVIVENCIA CLONOGENICOS

Se utiliza comúnmente para controlar la eficacia de los compuestos de la radiación y la modificación para determinar los efectos de los agentes citotóxicos y otras terapias contra el cáncer en la capacidad de formación de colonias, en diferentes líneas celulares.

RESULTADOS

AT Y L3

FIGURA 2

FIGURA 3

FIGURA 5

Se muestra la inmunotransferencia de control de shRNA (shGFP) y shRNA ATM.

DISCUSSION

AUTHOR WHAT DID THEY SAID? YES OR NO

Seshagiri S. et al Genome sequencing results has revealed that up to 18% (13 of 72 patients) of patients with CRC had mutation in ATM (four truncations, nine missense mutations).

Brannon AR. et al It has been reported that over 80% of CRCs have mutation of TP53.

Bang YJ. et al Indeed, results of recent phase II clinical trials indicate that patients with gastric cancer and low ATM expression showed better overall survival with paclitaxel and olaparib treatment than those with gastric cancer and normal ATM levels.

Mateo J. et al And prostate cancer patients with ATM mutation responded well to olaparib

CONCLUSIONS1. Colorectal cancer is sensitive to the PARP inhibitor, olaparib, so this

might be helpful for treatment.2. Any cancer (such as CRC, MCL, gastric cancer) which involves a

mutation of ATM gene is sensitive to PARP inhibitors, and the p53 depletion may improve the sensitivity.

3. Radiation or chemotherapy can also sensibilize ATM deficient CRC cells.

4. ATM syndrome may be root to predispose anyone to develop cancer because of its ATM dysfunctional protein.

Elisa

Catalina

BIBLIOGRAFIA

Chen Wang ,Nicholas Jette ,Daniel Moussienko ,D. Gwyn

Bebb ,Susan P. Lees-Miller (2017) ATM-Deficient Colorectal

Cancer Cells Are Sensitive to the PARP Inhibitor

Olaparib.http://www.sciencedirect.com.consultaremota.upb

.edu.co/science/article/pii/S1936523316302868