Upload
others
View
1
Download
0
Embed Size (px)
Citation preview
Dr#Giorgio#Russo,#Ricercatore#IBFM4CNR#
New$Projects$for$in$vivo$irradia2ons$
IBFM – METHODS
Proteogenomic#Analysis##
Hadron#Therapy#
Data#CorrelaBon#
CEFALU’
CATANIA
Breast#Cancer#4#Radiotherapy#OpBmizaBon#of#Breast#Cancer#(BCa)#radiotherapy#can#be#categorized#in#three#aspects:##
1. improvement#of#dose#homogeneity#within#the#target#volume#
2. maximum#reducBon#of#cardiopulmonary#dose##
3. minimizing#risks#of#second#malignancy#inducBon##
Local#radiotherapy#limited#to#the#breast#aSer#breast4conserving#surgery#(BCS),#or#to#the#chest#wall,#aSer#modified#radical#mastectomy#(MRM),#is#rouBnely#administered#using#three4dimensional#conformal#radiotherapy#(3D4CRT)#with#relaBvely#low#risks# for# late# toxicity.# More# recently,# intensity4modulated# radiotherapy# (IMRT),# has# been# rouBnely# implemented# in# the#clinic.##
#
#
#
#
#
3D5CRT$ IMRT$ Proton5therapy$
PROS$ Low#risks#for#late#toxicity# High#conformal#dose#distribuBons##
Low#entrance#dose#to#health#Bssue;#Homogeneous#dose#distribuBon;#High#conformal#dose#distribuBon;#
steep#fall4off#to#zero#dose#distally#to#the#target.#
CONS$ Low#conformal#dose#distribuBons##PotenBal#carcinogenic#effects#of#low#dose#
to#the#lung#and#contralateral#breast#/#
Breast#Cancer#–#Proton#therapy#Proton$beam$5$Accelerated$Par2al$Breast$Irradia2on$(PB5APBI)$is#one#technical#development#being#invesBgated#to#achieve:#
• saBsfactory#local#control#of#breast#cancer#with##• reduced#toxicity#to#normal#organs#through#the#use#of#hypofracBonaBon.##
Proton#beam#RT#is#characterized#by#its#unique#dose#distribuBon#involving#a#Bragg#peak#effect.#It#delivers#a#high#dose#to#a#limited#region#followed#by#rapid#fall4off,#thereby#enhancing#normal#Bssue#sparing#while#conserving#dose#homogeneity.##
[1]#Bush#DA#et#al..#A#technique#of#parBal#breast#irradiaBon#uBlizing#proton#beam#radiotherapy:#comparison#with#conformal#X4ray#therapy.#Cancer#J#2007;13:114–8.#[2]#Kozak#KR,#et#al.#Dosimetric#comparison#of#proton#and#photon#three4dimensional,#conformal,#external#beam#accelerated#parBal#breast#irradiaBon#techniques.#Int#J#Rad.#Onc.#Biol#Phys#2006;65:1572–8.#[3]#Taghian#AG,#et#al.#Accelerated#parBal#breast#irradiaBon#using#proton#beams:#iniBal#dosimetric#experience.#Int#J#Radiat#Oncol#Biol#Phys#2006;65:1404–10.#[4]#Wang#X,#et#al.#External4beam#accelerated#parBal#breast#irradiaBon#using#mulBple#proton#beam#configuraBons#
• excellent#results#in#terms#of#planning#target#volume#(PTV)#coverage##• dose#homogeneity#• significantly#reducing#the#volume#of#irradiated#non4target#breast#Bssue#
Dosimetric$comparisons$between$PB5APBI$and$3DCRT5APBI$[154]$have$shown$that:$$
PB5APBI$achieved##
MCF7 human breast cancer cell line MDA-MB-231 human metastatic breast cancer cell line
MCF10A human mammary non-tumorigenic cell line
Intra Operative Electron RadioTherapy Exclusive ! 23 Gy 10 Mev
Boost !9 Gy 10 MeV
MCF7 MCF10A
(LuminalB, Er+/Pr+) BasalB, Er-/Pr-
MDA-MB-231
(BasalB, Er-/Pr-)
Western Blot
Microarray
Immunofluorescence
Morphologic and Clonogenic Assay
Luminex
Genotyping
MCF7
9 GY- 23 Gy
Senescence radioinduced and cell death following both
treatments
MCF10A 9 GY- 23 Gy
Radioresistent cell fraction with 9 Gy
Cell death with 23 Gy
Surviving Fraction:0 Surviving Fraction:0 Surviving Fraction:9.6%
In addition, by Microarray experiments we described the involvement of known genes related to ionizing radiation and introduced novel ones able to activate molecular networks that could contribute to guide cell fate decision and in cancer care
able to provide the opportunity to explore molecular target-directed interventions
Hadrontherapy
1.In vitro approach with cell culture models
2.In vivo approach with mouse models
Biomarkers discovery by Proteogenomic technologies
Microarray
Immunofluorescence
Morphologic and Clonogenic Assay
Luminex
Genotyping
Evaluation of morphology and cell viability in terms of reproductive capacity, by clonogenic survival assay of treated cells
Comparative differential gene expression analysis beween treated and untreated cells by Microarray
Analysis of γ-H2AX foci formation after treatment, as a marker of sensitive early cell response to the presence of DNA double-strand breaks
Evaluation# of inflammatory response induced by treatment and able to influence cell fate decision
Analysis of genetic alterations that could regulate radio-sensitivity and cell survival/death balance
Pre4Clinic#study#
To#consider#all#necessiBes#that#a#preclinical#model#requires,#Monte#Carlo#calculaBons#must#give#a#considerable#contribuBon#to#characterize#the#beam's#dosimetry#simulaBng#the#physics#characterisBcs#of#the#beam,#the#beam#line#and#the#irradiaBon#setup.#
#Preclinical$models$are#a#crucial#component#of#research#in#radiaBon#therapy#and#in#nuclear#medicine.#Small4animal#irradiaBon#systems#must#mimic#the#clinical#applicaBon#of#radiaBon#therapy#as#closely#as#possible.##
Aim$of$this$experiment$is#manly:#• to#analyze#molecular)mechanisms)induced)by)hadrons)
in#preclinical#models#in#order#to#study#the#behavior#of#healthy#Bssue#exposed#to#radiaBon##
• to# achieve# a# highly) conformal) dose) to) the) target)volume,#while#sparing#the#surrounding#healthy#Bssues.#
#
In# addiBon,# we# will# compare# in# vivo# cell# response# to#treatment#in#mouse#with#or#without#BC#cell#inoculaBon#in#order# to# analyze# the# influence# of# tumours# on# healthy#Bssue#in#radiaBon#treatment#cell#response.##
BALB/c. BALB/c nude. NU/NU Nude C57BL/6 Mouse, CD1 General multipurpose model
In vivo Mouse models
Ideal for tumor biology and xenograft research. The animal lacks a thymus, is unable to produce T-cells, and is, therefore, immunodeficient.
MCF7
MCF10A
MDA-MB-231 Injection of tumour cells$
Leg injection to test the device $
Breast injection
Experimental Design part 1
Cellular models MCF-7 MCF-10
MDA-MB231
Injection of tumor cells
Housing at Animal House UniCT#
Transfer to LNS #
Cells growth
MCF7
MCF10A
MDA-MB-231
Leg injection for testing the device $
Breast injection for Experimental protocol
When tumour reach 10mm about 15 days#
Caliber #
Monitoring of tumor growth for 72H (Acute response)
15 days (tardive response)
Day 2 Hadrontherapy Treatment#
Day 3 Come back to Animal House#
Day 1 Transfer to LNS #
Experimental Design part 2
1) Dissection and removal of tumour a n d h e a l t h y t i s s u e f o r anatomopathology and proteogenomic analisys
2) Removal of hearth, lung, liver, kidney to analise toxicity#
Geant4#applicaBon:#Hadrontherapy#&#Dicom#
Hadrontherapy$applica2on$can#be#used#to#provide#a#good#source#of#informaBon#about#how#to#build#and#opBmize#the#experimental#setup#for#preclinical#applicaBons.##For#example,#the#geometrical#dimensions#of#the#system,#posiBon#of#small#animal,#source#surface#distance#SSD#etc.,#are#variables#that#can#be#managed#with#Geant4#simulaBons.#
DICOM$images$will#be#given#as#input#to#hadrontherapy#applicaBon#to#study#in#more#detail#the#damage#to#healthy#Bssue#surrounding#the#tumoral#area.##Thus,#Monte#Carlo#simulaBons#will#allow#us#to#understand#how#well#we#can#deliver#dose#to#targets#in#a#small#animal#subject,#and#to#esBmate#accurately#the#dose#released#to#the#Organ#at#Risk#(OAR),#main#goal#of#this#project.#
It#will#be#developed#a#new#Geant4#applicaBon#that#will#be#based#on#the#following#advanced#Geant4#example:#
PETherapy$
Geant4:#PETherapy#The#new#Geant4#PETherapy$applicaBon$will#be#developed#to#perform:#• preliminary#dosimetric#studies##• preliminary#RBE#analysis#and#that#will#compared#with#pathologist#biological#results.##
And#to#achieve:#• the#best#irradiaBon#condiBon#(e.g.#holder,#size#of#the#housing,#etc.)#• more#knowledge#about#the#dose#distribuBon#inside#the#target#(that#will#upload#in#GEANT4#geometry#using#the#real#DICOM#TC#images#of#animals)#and#in#the#surrounding#healthy#Bssue.#
##In#parBcular,#since#not#all#quanBBes#involved#in#radiobiological#and#medical#physics#studies#are#measurable#experimentally#
(e.g.#LET),#hence#tools#such#as#Geant4#and#biophysical#modelizaBon#of#ion#acBon#at#the#subcellular#level#are#necessary#to#model# experimental# beam# lines# and# predict# parBcle# behaviour# in# the# cellular# environment# compensaBng# for# the#disconBnuous#dose#delivery#pamern#inherent#to#radiaBon,#whether#administered#externally#or#internally.#
Experimental#setup#
The#animal#housing:#
• will#be#made#of#Polymethyl#methacrylate#(PMMA)#
• will#be#design#to#allow#an#easy#placement#of#tumor#region#respect#proton#beam.#
The#irradiaBon#will#be#performed#inside#CATANA#facility#at#LNS#4#Catania.##So,#it#will#be#possible#to#use#a#modulated#clinical#beam#to#deposit#an#uniform#dose#to#enBre#tumour#volume.#
Since#breast#cancer#cells#will#be#implanted#subcutaneous,#a#set#of#tantalum#clips,#that#will#be#implanted#around#breast#cancer#cells,#will#be#used#to#perform#a#precise#posiBoning#of#animals.#
cancer#
The#hadrontherapy#treatment#was#made#of#only#one#fracBon#and#the#prescribed#dose#will#be#of#about#24#Gy*.#
*The#prescribed#dose#esBmated#by#means#of#a#bibliographic#revision.#
Dosimetry#
Usually,#the#absolute#dosimetry#will#be#performed#by#using#a#Markus#chamber#but#due#the#limle#dimension#of#the#PTV,#it#will#needed#to#use#specific#dosimetric#tools#for#the#measuring#of#the#dose#released#by#this#limle#beam#field.#
Radiochromic#film#are#present#the#following#features:#
• SpaBal#resoluBon#of#µm#
• Self#developing#• 24D#detector#• Tissue#equivalent#• Dose#rate#indipendent#(about#5%)#• Easy#to#use##
The#previous#features#permit#their#use#as#in)vivo)dosimetry.##
FUTURE PERSPECTIVE Validation of identified biomarkers
TARGETED THERAPY Hadrontherapic treatments combined with drug molecules targeted in vivo.
0 3 6 9 12 15 18 21 24 27 30 33 36
Data Integration
Pathways analysis
Comparison between in vivo and in vitro data
Irradiating in vivo mouse models
Irradiating in vitro cell models
Preliminary irradiament on murine model
Setup of the device
Protocol approval by the Ethics Committee
Simulations of irradiation region
2015 | 2016 | 2017