Michael H. Holzscheiter1 SPSC Meeting October 26 Status Report on AD-4/ACE Antiproton Cell Experiment The Biological Effectiveness of Antiproton Annihilation

Michael H. Holzscheiter 1 SPSC Meeting October 26

Status Report on AD-4/ACEStatus Report on AD-4/ACEAntiproton Cell ExperimentAntiproton Cell Experiment

The Biological Effectiveness of Antiproton Annihilation

Nzhde Agazaryan1, Niels Bassler2, Gerd Beyer3, John J. DeMarco1, Michael Doser4,

Dragan Hajdukovic5, Michael H. Holzscheiter6, Toshiyasu Ichioka2, Keisuke S. Iwamoto1, Sandra Kovacevic5, Helge V. Knudsen2, Rolf

Landua4, Carl J. Maggiore6, William H. McBride1, Søren Pape Møller2, Jorgen

Petersen7, Vesna Popovic5, James B. Smathers1, Lloyd D. Skarsgard8, Timothy D.

Solberg1, Ulrik I. Uggerhøj2, Sanja Vranjes9, H. Rodney Withers1, Michelle Wong8,

Bradly G. Wouters10

1 UCLA Medical School, 2 University of Aarhus, 3 Geneva University Hospital4 CERN, 5 University of Montenegro, 6 PBar Labs, LLC,

7 Aarhus University Hospital, 8 British Columbia Cancer Research Centre9 Vinca Institute Belgrade 10University of Maastricht


Antiprotons deliver a higher biological dose for an Antiprotons deliver a higher biological dose for an equal effect in the entrance channel than protonsequal effect in the entrance channel than protons(and possibly heavy ions).(and possibly heavy ions).

The damage outside the beam path due to long and The damage outside the beam path due to long and medium range annihilation products is small and medium range annihilation products is small and does not significantly effect treatment planning.does not significantly effect treatment planning.

Antiprotons offer the possibility of real time imaging Antiprotons offer the possibility of real time imaging using high energy gammas and pions, even at low using high energy gammas and pions, even at low (pre-therapeutical) beam intensity.(pre-therapeutical) beam intensity.

Antiproton Therapy is based on three claims which need

proof:


•We have measured cell survival in the peak and plateau We have measured cell survival in the peak and plateau regions of an antiproton beam stopped in a biological regions of an antiproton beam stopped in a biological medium.medium.

•Extracting the relative doses which produce equivalentExtracting the relative doses which produce equivalent cell kill in the peak and the plateau region we can define cell kill in the peak and the plateau region we can define

the BEDR (Biological Effective Dose Ratio) as the ratio of the BEDR (Biological Effective Dose Ratio) as the ratio of these doses. (We only need to know the relative dose).these doses. (We only need to know the relative dose).

Results from the 2003 run period

• We can compare these results to the same experiment We can compare these results to the same experiment using a proton beam of comparable energyusing a proton beam of comparable energy..


• Irradiate sample tube with living cells suspended in gel.

• Slice sample tube in <1 mm slices and determine survival fraction for each slice.

Repeat for varying (peak) doses.

Biological Analysis Technique


Calculate “plateau” survival using slices 1 – 4.

Determine “peak” survival from slice 8 and 9.

Plot “peak” and “plateau” survival vs. relative dose (Plateau dose, particle fluence, etc.) and extract the Biological Effective Dose Ratio (BEDR).

Biological Analysis Technique

Dose (arb. units)


Cell Survival Measurements

Axial tube

Radial tubes

Location of Bragg Peak


0.0001

0.001

0.01

0.1

1

10

0 5 10 15 20 25

Tube B 1 Gy

Tube D 2.7 Gy

Tube F 4.6 Gy

Tube J 13.6 Gy

Sur

viva

l Fra

ctio

n

Depth in Sample [mm]

Data points inserted based on aminimum count of 1 surviving cell and known plating efficiency.

Cell Survival Measurements

3.8 Gy

8.9 Gy

15.5 Gy

40.0 Gy


Plateau average (slices 1,2)

Broad peak average (slices 8,8’,9,9’)

Narrow peak average (slices 8’,9)

0 2 4 6 8 10 12 14 16 18 20 22 24

0.01

0.1

1

B - 1Gy E - 1Gy C - 2Gy D - 3Gy F - 5Gy J - 25 GyS

urvi

ving

Fra

ctio

n

Depth (mm)


0 5 10 15 20 25 3010-2

10-1

100

BEDR(20%S)=9.2 - Broad peakBEDR(20%S)=9.8 - Narrow peak

Plateau Broad peak average Narrow peak average

Su

rviv

ing

Fra

ctio

n

Particle Fluence (arb. units)

BEDR AnalysisBEDR Analysis

Dose (arb. units) 0 5 10 15 20 25 3010 -2

10 -1

10 0

Plateau (CERN 2) Plateau (CERN 1) Peak (CERN 2) Peak (CERN 1)


Su

rviv

ing

Fra

ctio

n

Comparison toComparison toJune 7, 2003June 7, 2003

Dose (arb. units)


0 5 10 15 20 25 3010

-2

10 -1

100

BEDR(20%S)=9.2 - Broad peakBEDR(20%S)=9.8 - Narrow peak

Plateau Broad peak average Narrow peak average

Sur

vivi

ng F

ract

ion


CERN (50 MeV Antiprotons)CERN (50 MeV Antiprotons)

Dose (arb. units)

TRIUMF TRIUMF (reanalyzed for 50 MeV Protons)(reanalyzed for 50 MeV Protons)


The method works very well. The method works very well.

We are able to measure the survival response of V79-WNRE cells in the We are able to measure the survival response of V79-WNRE cells in the plateau and peak regions of a SOBP antiproton peak. plateau and peak regions of a SOBP antiproton peak.

In the early test experiment we obtained good data at 3 different doses in the In the early test experiment we obtained good data at 3 different doses in the plateau, and complete data at one dose in the peak. plateau, and complete data at one dose in the peak.

In the September run we obtained complete survival curves for 5 different In the September run we obtained complete survival curves for 5 different doses (in 6 measurements). The sensitivity in axial direction is high enough doses (in 6 measurements). The sensitivity in axial direction is high enough to detect the dose modulation due to the degrader used.to detect the dose modulation due to the degrader used.

An analysis of the data for the BEDR gives a result which is significantly An analysis of the data for the BEDR gives a result which is significantly higher than the value for protons (obtained at slightly higher energy and higher than the value for protons (obtained at slightly higher energy and using a different degrader) . using a different degrader) .

We observe only negligible cell kill outside of the beam in either the radial or We observe only negligible cell kill outside of the beam in either the radial or axial (beyond the peak) position at even the highest dose. This means not axial (beyond the peak) position at even the highest dose. This means not only that there is no significant spread of dose outside the beam due to the only that there is no significant spread of dose outside the beam due to the annihilation event but also no significant pion contamination in the beam.annihilation event but also no significant pion contamination in the beam.

Summary at end of 2003


2004 Run Cycle

The BEDR enhancement has been proven to be The BEDR enhancement has been proven to be significant.significant.

NEXT STEPS:NEXT STEPS:

Detailed studies of the peripheral damage due to the medium andDetailed studies of the peripheral damage due to the medium and long range products from the antiproton annihilation. long range products from the antiproton annihilation.

Clonogenic studies may not be the best approach Clonogenic studies may not be the best approach search for alternative assays. search for alternative assays.

Increased efforts on dosimetry in the periphery to the beam Increased efforts on dosimetry in the periphery to the beam

Systematic studies to find faster (and more automated) methods Systematic studies to find faster (and more automated) methods to extract biological data.to extract biological data.

Preparatory studies towards real time imaging.Preparatory studies towards real time imaging.


Evidence of LOW Peripheral Damage

0 5 10 15 20

J - 25 Gy

Depth (mm)

Distal

Peripheral Damage Study

0.001

0.01

0.1

1

10

0 2 4 6 8 10 12 14 16 18

Depth in Gel [mm]

Su

rviv

al F

ractio

n

20 Gy

5 Gy

< 2

mm

At the highest doses we can seea small effect outside the Braggpeak up to 1 – 2 mm distance

Need more sensitive assay Clonogenic may not be best DNA damage is detectable

Radial

0.0001

0.001

0.01

0.1

1

10

0 5 10 15 20 25 30

Depth [mm]

Su

rviv

al

4.6 GY

13.8 GY

21

Sam

ple

Tub

e


The COMET Assay

The comet assay is a gel electrophoresis method used to visualize and measure DNA strand breaks in individual cells using microscopy:

cell with DNA fragmentation

cell with major DNA fragmentation

cell without DNA fragmentation

Automated Analysis on individual cells

Statistical accuracy through analysisof > 100 cells per sample


The COMET Assay – Early Results

0 5 10 15 20 25 30 35 400.0

0.5

1.0

1.5

2.0

2.5

3.0

3.5

Tai

l Mom

ent

Depth in Gelatin [mm]

Cell sample irradiated with 15 Gy

Slices (0.5 mm and 1 mm) forplateaupeakperipheral (distal)

COMET Assay performed by the Inst. for Occup. Medicine, Zagreb

No detectable damage above control sample


Peripheral Damage – Neutron Dosimetry

Placement of 6Li and 7Li TLD chipsat 30 to 120 mm from annihilation volume

Total dose in Peak ~ 7 GyDose seen at 20 mm < 2 cGy

Additional dose in 6Li fromthermal neutrons only

Additional measurements toestablish neutron spectrum

in preparation



Bubble Technology Industries (BTI) Neutron Dosimeters

Superheated freon bubblets in gelatin undergo phase transitions when hit by neutrons



Detectors are re-usable Detectors are energy dependent


Monte Carlo Simulations

Original GEANT4 did not properly describe antiproton annihilation!

No ions produced above ‘sNewest version of GEANT4 with addition of (unofficial) modules

now produces ionsBut still no annihilation on periphery includedResults are still questionable – need benchmarks to test code


Real Time Imaging Tests

Antiprotons stop in target:Disc of 1.5 cm diameter and 2 mm thicknessSet-up 1 cm slit using 10 cm thick led blocksImage seen if slit is in line of sight with sourceBackground only if slit points away from source


Future Directions

Finish Laying the Foundations (2004/2006) Finalize Clonogenic Assay Studies Intensify Peripheral Damage Studies First Demonstration of Real Time Imaging of shaped targets

Source of Pbars: AD (3 – 5 x 107/85 seconds, T = 100 – 500 ns)

BEDR Measurements on antiprotons using pristine peakhigher beam energy needed (100 MeV)will allow better comparison to existing proton and heavy ion dataperform direct comparison measurement with heavy ions

Up to now we have not seen out of beam effectstill searching for more sensitive assaycan do better with tighter beam focus (DEM line

completion)complete neutron dosimetry

Initial Demonstration only established detector capabilityhigh resolution imaging at low intensity will need small focusand would be much easier with slow extraction (detector pile-

up)


Future Directions



Moving Forward: R&D towards final certification (2006 +)Development of beam delivery and energy modulation

~ 1 mm focus, scanning possibility (Complete DEM line)Real time imaging of shaped target

Implement semi-slow extraction (106 – 107/second)?

Comparison with protons and heavy ions (2005)


Hardware needed:

Excitation sextupoles: 2 XRC available in dispersion free regions (sections 16 and 41)Electrostatic septum: not available in ADMagnetic septum: SM5306 is available

More detailed design studyBeam lifetime measurements at 300 MeV/cCommissioning of this option

(Semi-)Slow Extraction?


Future Directions



Moving Forward: R&D towards certification of method (2006 +)Development of beam delivery and energy modulation

~ 1 mm focus, scanning possibility (Complete DEM line)Real time imaging of shaped target

Implement semi-slow extraction (106 – 107/second)?Initial in vivo testing? 4 x 108 pbars deliver 1 Gy to 1 cc tumor (10 shots or 15 minutes) Possibilities to increase intensity per shot exist

Need studies on life-time and radiation protection issues

Comparison with protons and heavy ions (2005)


Mode of Operation

Biological Measurements require long beam timesIrradiation of 4-5 cell samples at biological relevant

dose levels requires 24 hours of beam time. Time window between sample preparation and

analysisis maximum 72 hours.

Logistics is difficult as several teams need to be working in concert

….and have low repetition rate Cell preparation + analysis typically takes 4+ weeks

Continue with few long run periods (4 x 24 hours)


Mode of Operation

‘Physics’ studies (dosimetry, imaging, beam delivery)

are possible with shorter shifts (8 hours)can be done by separate small sub-teamsand can be performed back-to-back

This would be best if 8 hour shifts could be taken one week (5 shifts) at a time


Mode of Operation

Date Time Scheduled Topics Comments

May 21 8 hours Beam Development Cancelled due to PS delay

June 11 8 hours Focussing tests/Dosimetry Cancelled due to AD/PS Problems

June 28 16 hours Dosimetry using TLD’s Significant time lost to AD problem

July 2 8 hours Alanin tests Found misalignment of beam line

July 19 24 hours Peripheral damage studies Cancelled to PS problem (septum)

August 6 8 hours 6Li, 7Li dosimetry First smooth run of the year

August 23 24 hours Alternative assay studies Initial studies of COMET

August 27 8 hours Dosimetry Peripheral neutron dose

September 10 8 hours Dosimetry 2nd run on neutron dose

September 20 24 hours Biological studies Cancelled due to collaboration timing

September 24 8 hours Neutron Bubble Spectrometer Fast neutron spectrum

October 15 8 hours Imaging tests First high energy gamma detection

October 25 24 hours Peripheral damage studies COMET and clonogenic assays

Documents

Michael H. Holzscheiter1 SPSC Meeting October 26 Status Report on AD-4/ACE Antiproton Cell Experiment The Biological Effectiveness of Antiproton Annihilation