Updated1.interaction of-radiation-with-biological-matter

http://dmco.ucla.edu/McBride_Labhttp://dmco.ucla.edu/McBride_Lab
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Interaction of Radiation withInteraction of Radiation with
Biological Matter:Biological Matter:
(what is biological dose?)(what is biological dose?)
Bill McBrideBill McBride
Division of Cellular and Molecular OncologyDivision of Cellular and Molecular Oncology
Dept. Radiation OncologyDept. Radiation Oncology
David Geffen School MedicineDavid Geffen School Medicine
UCLA, Los Angeles, Ca.UCLA, Los Angeles, Ca.
wmcbride@mednet.ucla.eduwmcbride@mednet.ucla.edu
Room B3-109, x47051Room B3-109, x47051

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Objectives:Objectives:
– Know the characteristics of ionizingKnow the characteristics of ionizing
radiation that make it useful for RTradiation that make it useful for RT
– Define LET and RBE and what is meant byDefine LET and RBE and what is meant by
quality of radiationquality of radiation
– Know the difference between direct andKnow the difference between direct and
indirect action of radiation and the role ofindirect action of radiation and the role of
free radicalsfree radicals
– Recognize the impact of oxygen on initialRecognize the impact of oxygen on initial
radiation damage and of hypoxia in tumorradiation damage and of hypoxia in tumor
RTRT
– Understand how biological radiation doseUnderstand how biological radiation dose
and physical radiation dose differand physical radiation dose differ

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Radiation TherapyRadiation Therapy
• Approximately 50% of cancer patientsApproximately 50% of cancer patients
receive RT with curative intentreceive RT with curative intent
• Approximately half of these are curedApproximately half of these are cured
Radiation Therapy has a long history!

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Roentgen with his wifeRoentgen with his wife’s’s
hand, 1895hand, 1895
X-rays were rapidly
adapted for use as a
clinical treatment,
initially for non-
cancerous conditions,
but soon for cancer,
as well.

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FIRST CURE OF CANCER BY X-RAYSFIRST CURE OF CANCER BY X-RAYS
1899 - BASAL CELL CARCINOMA1899 - BASAL CELL CARCINOMA
X-rays were used to cure cancer very soon after their discovery

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And rapidly became a standardAnd rapidly became a standard
treatmenttreatment
Hammersmith Hospital, London, 1905

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Although side-effects wereAlthough side-effects were
encountered!encountered!
This is a picture of a 70 year oldThis is a picture of a 70 year old
person who was irradiated byperson who was irradiated by
Freund at the of age 5 in AustriaFreund at the of age 5 in Austria
1896 for nevus pigmentosus1896 for nevus pigmentosus
piliferus.piliferus.
L. Freund, Ein mit Rontgenstrahlen behandelterL. Freund, Ein mit Rontgenstrahlen behandelter
fall von nevus pigmentosus piliferus. Wein. Med.fall von nevus pigmentosus piliferus. Wein. Med.
Wochschr. 47, 428-434 (1987).Wochschr. 47, 428-434 (1987).

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EpilepsyEpilepsy
LupusLupus
Initially more non-cancerous diseases were
treated that cancer (still popular in Europe)

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The Nobel Prize in Physiology or Medicine 1946The Nobel Prize in Physiology or Medicine 1946
"for the discovery of the production of mutations by means of X-ray irradiation
Hermann J. Muller
However, its use for benign
conditions has been limited
in most countries for fear of
radiation-induced cancer.
The carcinogenic effects of
X-rays was discovered
using fruit flies by Muller in
1946.

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Natural radioactivity was discovered by Becquerel, who was awardedNatural radioactivity was discovered by Becquerel, who was awarded
the Nobel Prize in Physics in 1903 along with Marie and Pierrethe Nobel Prize in Physics in 1903 along with Marie and Pierre
Curie "in recognition of the extraordinary services they haveCurie "in recognition of the extraordinary services they have
rendered by their joint researches on therendered by their joint researches on the radiationradiation phenomena"phenomena"
“One wraps a Lumiere photographic plate with a bromide emulsion in two sheets of very thick black paper, such that the plate does not become
clouded upon being exposed to the sun for a day. One places on the sheet of paper, on the outside, a slab of the phosphorescent substance, and
one exposes the whole to the sun for several hours. When one then develops the photographic plate, one recognizes that the silhouette of the
phosphorescent substance appears in black on the negative. If one places between the phosphorescent substance and the paper a piece of
money or a metal screen pierced with a cut-out design, one sees the image of these objects appear on the negative. One must conclude from
these experiments that the phosphorescent substance in question emits rays which pass through the opaque paper and reduces silver salts.”
Paris 1896
Maltese crossHenri BecquerelHenri Becquerel Marie CurieMarie Curie

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Natural RadioactivityNatural Radioactivity
∀ αα particlesparticles
– Positively charged, helium nucleusPositively charged, helium nucleus
∀ ββ particlesparticles
– Negatively charged, electronsNegatively charged, electrons
∀ γγ-rays-rays
– No charge, EMRNo charge, EMR

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Radioisotopes also were soon beingRadioisotopes also were soon being
used to treat and cure cancer.used to treat and cure cancer.
Radium applicators were used for
many other conditions!
Radioactive
plaques and
implants are still in
common use, for
example in prostate
implant seeds.
First cure ofFirst cure of
cancer bycancer by
radium plaque -radium plaque -
19221922

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Therapeutic Benefit and R.T.Therapeutic Benefit and R.T.
There is always a need to derive a therapeuticThere is always a need to derive a therapeutic
benefit from RT. There are 2 main ways bybenefit from RT. There are 2 main ways by
which this is achieved:which this is achieved:
1. Physical means1. Physical means
– distributing dose by treatment planningdistributing dose by treatment planning
2. Biological means2. Biological means
– dose fractionationdose fractionation

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Grenz Rays
Megavoltag
e
Orthovoltag
e
Superficial Therapy
Contact Therapy
20 KeV
50 KeV
150 KeV
500 KeV
1-25 MeV Major improvements in RTMajor improvements in RT
during the mid-1900s cameduring the mid-1900s came
from improved penumbrafrom improved penumbra
and decreased skin doseand decreased skin dose
associated with higherassociated with higher
energy x-rays, cobalt, andenergy x-rays, cobalt, and
high energy photons.high energy photons.
More recently conformalMore recently conformal
RT, IMRT, IGRT,RT, IMRT, IGRT,
Gammaknife, Cyberknife,Gammaknife, Cyberknife,
tomotherapy, SRS, SRT,tomotherapy, SRS, SRT,
protons, heavy ions, etc.protons, heavy ions, etc.
have added considerablehave added considerable
variety to the choices forvariety to the choices for
physical radiation deliveryphysical radiation delivery
and present radiobiologicaland present radiobiological
challenges.challenges.

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18961896 FreundFreund - treated hairy nevus with fractionated doses- treated hairy nevus with fractionated doses
19001900 Stenbeck - cured skin cancer with single dosesStenbeck - cured skin cancer with single doses
19061906 Bergonie and Trubandeau introduced theBergonie and Trubandeau introduced the “Law” that radiosensitivity is related“Law” that radiosensitivity is related
to cell proliferation (NOT TRUE!) to explain why fractionated doses sterilizedto cell proliferation (NOT TRUE!) to explain why fractionated doses sterilized
rams without skin reactionsrams without skin reactions
Regaud - treated uterine cancer with fractionated dosesRegaud - treated uterine cancer with fractionated doses
19141914 SchwartzSchwartz
- Fractionation is superior because of cell cycle redistribution- Fractionation is superior because of cell cycle redistribution
19191919 Coutard cures deep-seated H&N tumorsCoutard cures deep-seated H&N tumors
19321932 Coutard shows fractionation superior to single doseCoutard shows fractionation superior to single dose
19441944 Strandquist - empirical laws for changing dose per fractionStrandquist - empirical laws for changing dose per fraction
19671967 Ellis - Nominal Standard Dose (NSD) formulaEllis - Nominal Standard Dose (NSD) formula
1980s Linear Quadratic formula gains favor1980s Linear Quadratic formula gains favor
History of FractionationHistory of Fractionation

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The First Radiation Dosimeter!The First Radiation Dosimeter!
Early x-ray machines took a long time to deliver effective dose and
gave skin reactions that could be circumvented by dose fractionation.

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From Amaldi and Kraft, “Radiotherapy with beams of carbon ions, Reports on Progress in Physics, 68, (2005)

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““In order to save machine time, a 3-day-a-week schedule wasIn order to save machine time, a 3-day-a-week schedule was
initiated in 1962. This schedule was quickly abandoned in pre-initiated in 1962. This schedule was quickly abandoned in pre-
operative irradiation because of increased wound healing problems.operative irradiation because of increased wound healing problems.
Although acute reactions in the 3-day-a-week schedule for protractedAlthough acute reactions in the 3-day-a-week schedule for protracted
radical irradiation were not excessive,radical irradiation were not excessive, late radiation sequelaelate radiation sequelae areare
probably more pronounced as observed 2 or more years later.probably more pronounced as observed 2 or more years later.””
Fletcher, 1966.Fletcher, 1966.
3 x 3.3 Gy3 x 3.3 Gy 5 x 2 Gy5 x 2 Gy
History has repeatedly shown that dose fractionationHistory has repeatedly shown that dose fractionation
results in a therapeutic advantageresults in a therapeutic advantage

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Clinical RT is Changing, which PresentsClinical RT is Changing, which Presents
Challenges and Opportunities forChallenges and Opportunities for
RadiobiologyRadiobiology
Conventional treatment:Conventional treatment:
Tumors are irradiated to a specified dose with 2Gy fractions delivered, more or lessTumors are irradiated to a specified dose with 2Gy fractions delivered, more or less
homogeneously, in a 6 week time periodhomogeneously, in a 6 week time period
• Varying this schedule impacts outcomeVarying this schedule impacts outcome
• Radiobiological modeling attempts to provide guidelines for customization of RT usingRadiobiological modeling attempts to provide guidelines for customization of RT using
– Radiobiological principlesRadiobiological principles derived from preclinical dataderived from preclinical data
– Radiobiological parameters derived from clinical altered fractionation protocolsRadiobiological parameters derived from clinical altered fractionation protocols
Modern treatment:Modern treatment:
IMRT etc allows optimized non-homogeneous dose distributions, concomitant boosts,IMRT etc allows optimized non-homogeneous dose distributions, concomitant boosts,
dose painting -dose painting - dose heterogeneitydose heterogeneity
SRS, SRT, HDR, Protons, Heavy Ions -SRS, SRT, HDR, Protons, Heavy Ions - high dose/fx issueshigh dose/fx issues
Molecular and chemical targeting -Molecular and chemical targeting - dose adjustmentdose adjustment
Molecular prognosis and diagnosis promise individualized treatment plans andMolecular prognosis and diagnosis promise individualized treatment plans and biologicalbiological
treatment planningtreatment planning

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• Radiobiology has derived means ofRadiobiology has derived means of
understanding why dose fractionation gives aunderstanding why dose fractionation gives a
therapeutic benefit.therapeutic benefit.
• New physical delivery methods need toNew physical delivery methods need to
incorporate and/or modify these concepts.incorporate and/or modify these concepts.
• In order to understand either conventional orIn order to understand either conventional or
newer treatment effects, one needs to knownewer treatment effects, one needs to know
the differences between physical andthe differences between physical and
biological radiation dosebiological radiation dose

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What is Radiation?What is Radiation?
• Radiation is classified into two main categories:Radiation is classified into two main categories:
- Non-ionizing radiationNon-ionizing radiation
- Ionizing radiationIonizing radiation

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ELECTROMAGNETIC RADIATIONSELECTROMAGNETIC RADIATIONS
Photon E = hν (energy = Planck’s const x frequency)
= hc/λ (c = speed of light, λ = wave length)
1010-9-9 1010-8-8 1010-7-7 1010-6-6 1010-5-5 1010-4-4 1010-3-3 1010-2-2 1010-1-1 11 1010 101022 101033 101044
γγ raysrays
X-raysX-rays U.V.U.V.
vv
ii
ss
ii
bb
ll
ee
Infra RedInfra Red Radio WavesRadio Waves
MicrowavesMicrowaves Short WavesShort Waves
T.V.T.V.
RadioRadio
RadarRadar
IONIZINGIONIZING
RADIATIONRADIATION NON-IONIZING RADIATIONNON-IONIZING RADIATION
λ (cms)
E (eV) 1.24x107
1.24x102
1.24x10-13

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• Non-ionizing radiationNon-ionizing radiation
– Is a particle or wave that has enough kinetic energy to raise theIs a particle or wave that has enough kinetic energy to raise the
thermal energy of an outer shell electron and causethermal energy of an outer shell electron and cause excitationexcitation
with emission ofwith emission of low energy EMR (infrared)low energy EMR (infrared)
• Ionizing radiationIonizing radiation
– Ionizing radiation has enough kinetic energy to detach at least one electron from anIonizing radiation has enough kinetic energy to detach at least one electron from an
atom or molecule, creating ionsatom or molecule, creating ions
– Charged particles such as electrons, protons, heavy ions, alpha and beta particles areCharged particles such as electrons, protons, heavy ions, alpha and beta particles are
directlydirectly ionizing because they can interact directly with atomic electrons throughionizing because they can interact directly with atomic electrons through
coulombic forces and transfer a major part of their kinetic energy directlycoulombic forces and transfer a major part of their kinetic energy directly
– In contrast, photons (x rays,In contrast, photons (x rays, γγ rays) and neutrons are chargeless and therefore morerays) and neutrons are chargeless and therefore more
penetrating. They arepenetrating. They are indirectlyindirectly ionizing. They have sufficient kinetic energy to free anionizing. They have sufficient kinetic energy to free an
orbital electron producing aorbital electron producing a ‘fast’ recoil or Compton electron that is, in turn, directly‘fast’ recoil or Compton electron that is, in turn, directly
ionizingionizing
• Energy is deposited inEnergy is deposited in “packets”,“packets”, which is why, when it is deposited in DNA, ionizing radiation iswhich is why, when it is deposited in DNA, ionizing radiation is
an efficient cytotoxic agentan efficient cytotoxic agent
• Ionizing radiation has an energy in excess of 124 eV, which corresponds to aIonizing radiation has an energy in excess of 124 eV, which corresponds to a λλ < about 10< about 10-6-6
cm.cm.
γ-ray
γ’-
ray
excitation
ionization
α particle
excitation
and ionization

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Ion formation – H2OIon formation – H2O++
and e-and e-
Excitation and H and OH radical formationExcitation and H and OH radical formation
ION RADICAL LIFETIMEION RADICAL LIFETIME
FREE RADICAL LIFETIMEFREE RADICAL LIFETIME
BREAKAGE OF BONDSBREAKAGE OF BONDS
CHEMICAL REPAIR / MISREPAIRCHEMICAL REPAIR / MISREPAIR
ENZYMIC REPAIR / MISREPAIRENZYMIC REPAIR / MISREPAIR
EARLY BIOLOGICAL EFFECTSEARLY BIOLOGICAL EFFECTS
LATE BIOLOGICAL EFFECTSLATE BIOLOGICAL EFFECTS
1010-18-18
1010-12-12
1010-6-6
101000
101066
SECSSECS
Absorption of energyAbsorption of energy
Physical effectsPhysical effects
Chemical lesionsChemical lesions
Chemical repairChemical repair
Enzyme repair/lesionEnzyme repair/lesion
Cellular effectsCellular effects
Tissue effectsTissue effects
Systemic effectsSystemic effects
Days-YearsDays-Years
Hrs-DaysHrs-Days
Mins-HrsMins-Hrs
Ionization produces ions, ion radicals, and freeIonization produces ions, ion radicals, and free
radicals concentrated along tracks and especially atradicals concentrated along tracks and especially at
Bragg peak of primary and secondary electrons. TheyBragg peak of primary and secondary electrons. They
are highly reactive and cause damage to biologicalare highly reactive and cause damage to biological
mattermatter
• IonIon - atom or molecule that has lost an electron and is charged.- atom or molecule that has lost an electron and is charged.
• Free radicalFree radical - atom or group of atoms that contains an unpaired electron and is highly reactive- atom or group of atoms that contains an unpaired electron and is highly reactive
• Aqueous electronAqueous electron - has lost kinetic energy and has been captured by water - a powerful reducing- has lost kinetic energy and has been captured by water - a powerful reducing
agent.agent.
1010-16-16
1010-14-14

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The Gray is theThe Gray is the PhysicalPhysical
Unit of RadiationUnit of Radiation
• 1 GRAY, the unit of absorbed dose (1 joule / Kg),1 GRAY, the unit of absorbed dose (1 joule / Kg),
– Causes 1-2 x 10Causes 1-2 x 1055
ionization events / cellionization events / cell
– 1% in DNA1% in DNA
– A single cobalt 60 ray will deposit about 1mGy in a cellA single cobalt 60 ray will deposit about 1mGy in a cell
• Rad (Radiation Absorbed Dose) is the old unit = cGyRad (Radiation Absorbed Dose) is the old unit = cGy

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Direct and IndirectDirect and Indirect ActionAction ofof
RadiationRadiation
• Indirectly ionizing radiation canIndirectly ionizing radiation can actact directly ordirectly or
indirectlyindirectly on biological targetson biological targets
• If the ion pairs and free radicalsIf the ion pairs and free radicals are produced inare produced in aa
biologic targetbiologic target (DNA) then this is(DNA) then this is direct actiondirect action
• If water or other atoms or molecules are ionized,If water or other atoms or molecules are ionized,
diffusible free radicals can act as intermediaries todiffusible free radicals can act as intermediaries to
cause damage - this iscause damage - this is indirect actionindirect action

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p+
e-
photon
p+
photon
INDIRECT ACTION
DIRECT ACTION
Direct and Indirect Action of
Ionizing Radiation on DNA
4 nm
2 nm
e-
R.
H2O
OH
.

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• Since HSince H22O is the major component in cells, the most commonO is the major component in cells, the most common
ionization event is radiolysis of water, producing reactive oxygenionization event is radiolysis of water, producing reactive oxygen
species (ROS)species (ROS)
• The most relevant water is within 2nm of the DNA and tightlyThe most relevant water is within 2nm of the DNA and tightly
boundbound
• ROS produced include: HROS produced include: H..
- reducing; OH- reducing; OH..
- oxidizing; HO- oxidizing; HO22
..
- oxidizing- oxidizing
(O(O22 + H+ H..
); H); H22OO22 - oxidizing- oxidizing
• The net effect is oxidation of cellular constituentsThe net effect is oxidation of cellular constituents
• About 60% of DNA damage caused by x-rays is due to ROSAbout 60% of DNA damage caused by x-rays is due to ROS
• About 75% of the indirect action of radiation is due to hydroxyl radicalsAbout 75% of the indirect action of radiation is due to hydroxyl radicals
(OH(OH..
))
Reactive Oxygen Species (ROS)Reactive Oxygen Species (ROS)

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Free OHFree OH..
radicals generateradicals generate organic radicalsorganic radicals by:by:
– AdditionAddition R + OHR + OH.. ..
ROHROH
– Hydrogen abstractionHydrogen abstraction RH + OHRH + OH..
RR..
+ H+ H22OO
– Electron transferElectron transfer R + OHR + OH..
RR..
+ OH+ OH --
Where R is the organic moietyWhere R is the organic moiety

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Free Radicals and theirFree Radicals and their
Scavengers MatterScavengers Matter
• Biological effects of ionizing radiation are determined in large part by freeBiological effects of ionizing radiation are determined in large part by free
radicalsradicals
• Free radicals are involved in many biological processes, including cellularFree radicals are involved in many biological processes, including cellular
respirationrespiration
• We have defenses against free radicalsWe have defenses against free radicals
– Endogenous free radical scavengers - most relevant within 2nm of the DNAEndogenous free radical scavengers - most relevant within 2nm of the DNA
– Anti-oxidantsAnti-oxidants
• eg superoxide dismutase, especially in mitochondria, and catalaseeg superoxide dismutase, especially in mitochondria, and catalase
• Free radical scavengers can protect normal tissue from radiationFree radical scavengers can protect normal tissue from radiation
– eg Amifostineeg Amifostine
• Depleting free radical scavengers will radiosensitizeDepleting free radical scavengers will radiosensitize
• What interacts with free radicals, in particular radicals in biologicalWhat interacts with free radicals, in particular radicals in biological
materials will be important in determining outcome at this levelmaterials will be important in determining outcome at this level
• Oxygen interacts with free radicalsOxygen interacts with free radicals

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Oxygen MattersOxygen Matters
• Binds H radicals forming hydrogen peroxideBinds H radicals forming hydrogen peroxide
HH..
+ O+ O22 HOHO22
..
(+HO(+HO22
..
) H) H22OO22 (+O(+O22))
• Binds electrons to give superoxideBinds electrons to give superoxide
ee--
+ O+ O22 OO22
--
+ (H+ (H22O) HOO) HO22
..
+ OH+ OH--
• Binds organic radicals to form peroxidesBinds organic radicals to form peroxides
RR..
+ O+ O22 RORO22
..
(radical peroxide)(radical peroxide)
RORO22
..
+ R+ R’ H ROOH + R’ (hydroperoxide)’ H ROOH + R’ (hydroperoxide)
RORO22
..
+ R+ R’’..
ROOR’ (peroxide)ROOR’ (peroxide)
OxygenOxygen “fixes”“fixes” the radical lesions in DNA in a form that can notthe radical lesions in DNA in a form that can not
be easily chemically repaired and therefore is abe easily chemically repaired and therefore is a very powerfulvery powerful
radiosensitizer.radiosensitizer.

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Oxygen Enhancement RatioOxygen Enhancement Ratio (OER)(OER)
Dose required to produce a specific biological effect in the absence of oxygenDose required to produce a specific biological effect in the absence of oxygen
Dose required for the same effect in its presenceDose required for the same effect in its presence==
OER varies with level of effect but can be 2.5 - 3 foldOER varies with level of effect but can be 2.5 - 3 fold
1) Culture Cells1) Culture Cells
((
3) Count cells in hemocytometer3) Count cells in hemocytometer
4) irradiate under oxic or hypoxic conditions4) irradiate under oxic or hypoxic conditions
0 Gy 2Gy 4Gy 6Gy0 Gy 2Gy 4Gy 6Gy
5) Plate cells and5) Plate cells and
grow for about 12 daysgrow for about 12 days
..
..
..
..
..
.... ..
6) Count colonies6) Count colonies
Dose (Gy)Dose (Gy)
S.F.S.F.
0 2 4 6 8 100 2 4 6 8 10
1.01.0
0.10.1
0.010.01
oxicoxic
hypoxichypoxic
Physical Dose = Biological DosePhysical Dose = Biological Dose
2) Suspend Cells2) Suspend Cells
trysinization)trysinization)

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• Hypoxic areasHypoxic areas occur almost solely in tumorsoccur almost solely in tumors and are moreand are more
radioresistant than oxic areas.radioresistant than oxic areas.
• Hypoxia contributes to treatment failureHypoxia contributes to treatment failure
• Reoxygenation occurs between radiation dose fractions giving aReoxygenation occurs between radiation dose fractions giving a
rationale for dose fractionationrationale for dose fractionation
• The oxygen effect is greater for low LET than high LET radiationThe oxygen effect is greater for low LET than high LET radiation
Giacca and Brown
Pimonizadole (oxygen mimetic)
staining colorectal carcinoma
The effects of hypoxia were first
discovered in 1909 by Schwarz who
showed that strapping a radium source on
the arm gave less of a skin reaction than
just placing it there. This was used to give
higher doses to deep seated tumors.
Clinical Relevance of HypoxiaClinical Relevance of Hypoxia

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RADIATION QUALITY ANDRADIATION QUALITY AND
BIOLOGICAL EFFECTIVENESSBIOLOGICAL EFFECTIVENESS

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gamma raysgamma rays
deep therapydeep therapy
X-raysX-rays
soft X-rayssoft X-rays
alpha-particlealpha-particle
HIGH LETHIGH LET
RadiationRadiation
LOW LETLOW LET
RadiationRadiation
Separation of ion clusters in relation toSeparation of ion clusters in relation to
size of biological targetsize of biological target
LINEAR ENERGY TRANSFERLINEAR ENERGY TRANSFER
LET is average energy (dE) imparted by excitationLET is average energy (dE) imparted by excitation
and Ionization events caused by a charged particleand Ionization events caused by a charged particle
traveling a set distance (dl) -traveling a set distance (dl) - LET = dE/dl (keV/LET = dE/dl (keV/ µµm)m)

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• A dose of 1 Gy will give 2x10A dose of 1 Gy will give 2x1033
ionization events in 10ionization events in 10-10-10
g (the sizeg (the size
of a cell nucleus). This can beof a cell nucleus). This can be
achieved by:achieved by:
– 1MeV electrons1MeV electrons
•700 electrons which give 6700 electrons which give 6
ionization events perionization events per µµm.m.
– 30 keV electrons30 keV electrons
•140 electrons which give 30140 electrons which give 30
– 4 MeV protons4 MeV protons
•14 protons which give 30014 protons which give 300
• The biological effectiveness ofThe biological effectiveness of
these different radiationsthese different radiations
vary!vary!
γ-ray
γ’-ray
excitation
ionization
α particle
excitation and ionization

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Relative BiologicalRelative Biological
EffectivenessEffectiveness (RBE) of the(RBE) of the
Radiation MattersRadiation Matters
Dose of 250 kVp x-rays required to produce an effectDose of 250 kVp x-rays required to produce an effect
Dose of test radiation required for the same effectDose of test radiation required for the same effect
==
S.F.S.F.
1.01.0
0.10.1
0.010.01
0.0010.001
DOSE GyDOSE Gy
High LETHigh LET
Low LET, HDRLow LET, HDR

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Linear Energy Transfer (LET keV/Linear Energy Transfer (LET keV/µµm)m)
RBERBE
(for cell kill)(for cell kill)
10001000100100101011
00
22
44
66
88
RBERBE
DiagnosticDiagnostic
X-raysX-rays
FastFast
NeutronsNeutrons
AlphaAlpha
ParticlesParticles
overkilloverkill
0.10.1
Co-60Co-60
gamma raysgamma rays
00
11
22
33
44
OEROER
OEROER
OER is the inverse of RBE because OER depends considerably on theOER is the inverse of RBE because OER depends considerably on the
indirect action of ionizing radiationindirect action of ionizing radiation
RBE is maximal when the average distance between ionization events =RBE is maximal when the average distance between ionization events =
distance between DNA strands = 2nmdistance between DNA strands = 2nm
RBE and OER as a function of LETRBE and OER as a function of LET

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DNA is the Primary, but not the only,DNA is the Primary, but not the only,
Cellular Target for RadiationCellular Target for Radiation
• Microbeam irradiation of cell cytoplasmMicrobeam irradiation of cell cytoplasm
does not generally cause cell death, butdoes not generally cause cell death, but
irradiation of the nucleus doesirradiation of the nucleus does
• Tritiated thymidine incorporated into cellsTritiated thymidine incorporated into cells
can kill themcan kill them
• Radiation-induced chromosomalRadiation-induced chromosomal
abnormalities correlate with cell deathabnormalities correlate with cell death
and carcinogenesisand carcinogenesis
• However, irradiation of the cytoplasm isHowever, irradiation of the cytoplasm is
not without biological consequencesnot without biological consequences

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Lesion sizeLesion size
about 15-20about 15-20
nucleotidesnucleotides BlobBlob
7 nm diam.7 nm diam.
12 ion pairs12 ion pairs
OHOH .. eeaquaqu
OHOH .. eeaquaqu
OHOH .. eeaquaqu
OHOH .. eeaquaqu
OHOH .. eeaquaquOHOH .. eeaquaqu
OHOH .. eeaquaqu
OHOH .. eeaquaqu
OHOH .. eeaquaqu
OHOH .. eeaquvaquv
OHOH .. eeaquaqu
OHOH .. eeaquaqu
OHOH .. eeaquaqu
SpurSpur
4 nm diam4 nm diam
3 ion pairs3 ion pairs
100 eV energy100 eV energy
95% of energy deposition events95% of energy deposition events
The lesions in DNA that are associated withThe lesions in DNA that are associated with
cell death and carcinogenesis aftercell death and carcinogenesis after
radiation exposure areradiation exposure are largelarge
The high cytotoxic efficiency of ionizing radiation can be ascribedThe high cytotoxic efficiency of ionizing radiation can be ascribed
to the deposition of low levels of energy in small packets withinto the deposition of low levels of energy in small packets within
the DNA that cause lesions large enough to be fatalthe DNA that cause lesions large enough to be fatal

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SINGLE STRAND BREAK
1000 / CELL / GRAY
BASE CHANGE (eg C - U)
BASE LOSS
1000 / CELL / GRAY
BASE MODIFICATION
(eg thymine/cytosine glycol)
SUGAR DAMAGE
(abstraction of hydrogen atom)
INTRASTRAND
CROSSLINK
0.5 / CELL / GRAY
INTERSTRAND
CROSSLINK
DNA-PROTEIN
CROSSLINK
1 / CELL / GRAY
*
DOUBLE STRAND BREAK
30/ CELL / GRAY

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• Not all ionization events are lethal!!Not all ionization events are lethal!!
• As a rough guide the fraction of cells surviving 2Gy (SFAs a rough guide the fraction of cells surviving 2Gy (SF2Gy2Gy))
is about 0.5is about 0.5
• If the S.F. 2Gy is 0.5, what is the S.F. after 60Gy?If the S.F. 2Gy is 0.5, what is the S.F. after 60Gy?
= 0.5= 0.53030
= 0.9x10= 0.9x10-9-9
• If the S.F. 2Gy is 0.7, what is the S.F. after 60Gy?If the S.F. 2Gy is 0.7, what is the S.F. after 60Gy?
= 0.7= 0.73030
= 2.2x10= 2.2x10-5-5

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What is the Lethal Lesion?What is the Lethal Lesion?

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Repairable Sublethal DamageRepairable Sublethal Damage
X- orX- or γγ-radiation is sparsely ionizing; most-radiation is sparsely ionizing; most
damage can be repaireddamage can be repaired
4 nm4 nm
2 nm2 nm

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SingleSingle lethal hit
Also known as α - type killing
4 nm4 nm
2 nm2 nm
Unrepairable Multiply Damaged Site
It is hypothesized that the lethal
lesions are large double strand
breaks with Multiply Damaged
Sites (MDS) that can not be
repaired. They are more likely to
occur at the end of a track

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At high dose, intertrack
repairable Sublethal Damage
may Accumulate forming
unrepairable, lethal MDS
Also known asAlso known as ββ - type killing- type killing

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S.F.S.F.
1.01.0
0.10.1
0.010.01
0.0010.001
DOSE GyDOSE Gy
Low LET, HDRLow LET, HDR
Low Dose RateLow Dose Rate
allows continuous SLDRallows continuous SLDR
Dose Rate MattersDose Rate Matters

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Chromatin Structure MattersChromatin Structure Matters
• Each cell contains about 2m of DNAEach cell contains about 2m of DNA
• The basic structure is theThe basic structure is the nucleosome,nucleosome, which is 146 base
pairs of DNA wrapped around 2 copies of histones H2A,H2A,
H2B, H3, and H4H2B, H3, and H4
• Nucleosomes are in turn wrapped around other proteins toNucleosomes are in turn wrapped around other proteins to
formform compacted chromatincompacted chromatin
• Chromatin is maximally compacted during mitosisChromatin is maximally compacted during mitosis
• Transcription requires decompaction to facilitate initiationTranscription requires decompaction to facilitate initiation
(binding of transcription factors and RNAP II) and(binding of transcription factors and RNAP II) and
elongationelongation
840nm840nm
minibandminiband
- 30nm- 30nm

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Chromatin Structure andChromatin Structure and
Radiation ResponsesRadiation Responses
• Compact chromatin is more radiosensitive than non-compactedCompact chromatin is more radiosensitive than non-compacted
–Mitotic cellsMitotic cells
• are 2.8 times more sensitive to DNA breaks than interphase cellsare 2.8 times more sensitive to DNA breaks than interphase cells
• have a lower OER (eg 2.0 compared with 2.8)have a lower OER (eg 2.0 compared with 2.8)
• do not have much of ado not have much of a “shoulder” on their survival curve“shoulder” on their survival curve
–Actively transcribing genes are less sensitive to damageActively transcribing genes are less sensitive to damage
• Decompaction and compaction require acetylation and deacetylation ofDecompaction and compaction require acetylation and deacetylation of
histones by acetyltransferases (HAT) and deacetylases (HDAC)histones by acetyltransferases (HAT) and deacetylases (HDAC)
• HDAC inhibitors are entering the clinic as anti-cancer agents and can radiosensitizeHDAC inhibitors are entering the clinic as anti-cancer agents and can radiosensitize
• Radiation Damage to DNA is not randomly distributed.Radiation Damage to DNA is not randomly distributed.
• It varies with cell cycle phase and level of gene expressionIt varies with cell cycle phase and level of gene expression
S.F.S.F.
20201616121288440000
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Dose (Gy)Dose (Gy)
LATE SLATE S
EARLY SEARLY S
G1 PHASEG1 PHASE
G2/M PHASEG2/M PHASE

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12.5Gy 14.0Gy
15.5Gy 17.0Gy
Withers, H. R. and Elkind, M. M. Radiology 91:998, 1968Withers, H. R. and Elkind, M. M. Radiology 91:998, 1968
Used the macrocolony assay in mouseUsed the macrocolony assay in mouse
jejunum to assessed the effects of 2jejunum to assessed the effects of 2
radiation doses given varying times apartradiation doses given varying times apart
to measure the time to and extent of repair,to measure the time to and extent of repair,
redistribution, and repopulationredistribution, and repopulation
(regeneration) between dose fractions.(regeneration) between dose fractions.
RedistributionRedistribution
RepairRepair
RepopulationRepopulation
700R 1500R
Colony derived from a
single surviving clonogen

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ACUTE RESPONDING TISSUESACUTE RESPONDING TISSUES
(responses seen during standard therapy)(responses seen during standard therapy)
GutGut
SkinSkin
Bone MarrowBone Marrow
MucosaMucosa
LATE RESPONDING TISSUESLATE RESPONDING TISSUES
(responses seen after end of therapy)(responses seen after end of therapy)
BrainBrain
Spinal CordSpinal Cord
KidneyKidney
LungLung
BladderBladder
Tissue Type MattersTissue Type Matters
Dose (Gy)Dose (Gy)
SurvivingSurviving
FractionFraction
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Late RespondingLate Responding
TissuesTissues
Acute RespondingAcute Responding
Tissues andTissues and
Many TumorsMany Tumors

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Dose (Gy)Dose (Gy)
Dose FractionationDose Fractionation
242420201616121288440000
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SurvivingSurviving
FractionFraction
Single doseSingle dose
Late responding tissuesLate responding tissues Single doseSingle dose
Acute responding tissuesAcute responding tissues
Fractionated doseFractionated dose
Acute responding tissuesAcute responding tissues
Fractionated doseFractionated dose
Late responding tissuesLate responding tissues
Dose fractionation spares late responding tissues more than acuteDose fractionation spares late responding tissues more than acute
responding tissues and many tumorsresponding tissues and many tumors

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The Aim is to IncreaseThe Aim is to Increase
Therapeutic Benefit!Therapeutic Benefit!
ProbabilityProbability
of tumorof tumor
control/control/
of normalof normal
tissuetissue
damagedamage
Dose (Gy)Dose (Gy)
A B CA B C
1.01.0
00
therapeutic benefittherapeutic benefit
Normal tissue complication dose-response curves are steep!Normal tissue complication dose-response curves are steep!

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Biological effectiveness of RT varies withBiological effectiveness of RT varies with
• Size of Dose (D) - (alpha and beta)Size of Dose (D) - (alpha and beta)
• Size of Dose Per Fraction (d) - (alpha and beta)Size of Dose Per Fraction (d) - (alpha and beta)
• Time over which it is delivered (T)- (alpha and beta)Time over which it is delivered (T)- (alpha and beta)
• Time between fractions (t)Time between fractions (t)
• Volume irradiated (V)Volume irradiated (V)
• Quality of Radiation (Q) - RBEQuality of Radiation (Q) - RBE
• Presence/Absence of Oxygen - OERPresence/Absence of Oxygen - OER
• DNA Repair efficiency and completenessDNA Repair efficiency and completeness
• Cell cycle phase and level of gene activationCell cycle phase and level of gene activation
• Tissue/Tumor TypeTissue/Tumor Type

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4Rs OF RADIOBIOLOGY RELEVANT TO4Rs OF RADIOBIOLOGY RELEVANT TO
CLINICAL DOSE FRACTIONATIONCLINICAL DOSE FRACTIONATION
• Repair of sublethal damageRepair of sublethal damage
- spares late responding normal tissue preferentiallyspares late responding normal tissue preferentially
• Reassortment/Redistribution of cells in the cell cycleReassortment/Redistribution of cells in the cell cycle
– increases acute effectsincreases acute effects
– no influence on late effectsno influence on late effects
– increases damage to tumorincreases damage to tumor
• Repopulation/RegenerationRepopulation/Regeneration
– spares acute responding normal tissue preferentiallyspares acute responding normal tissue preferentially
– no influence on late effects,no influence on late effects,
– danger of tumor repopulationdanger of tumor repopulation
• ReoxygenationReoxygenation
– no influence on normal tissue responsesno influence on normal tissue responses
– increases tumor damageincreases tumor damage

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Questions onQuestions on
Interaction of Radiation with Biological Matter:Interaction of Radiation with Biological Matter:
what is biological dose?what is biological dose?
Bill McBrideBill McBride
Dept. Radiation OncologyDept. Radiation Oncology
David Geffen School MedicineDavid Geffen School Medicine
UCLA, Los Angeles, Ca.UCLA, Los Angeles, Ca.
wmcbride@mednet.ucla.eduwmcbride@mednet.ucla.edu

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1.1. The lifetime of radicals in target molecules isThe lifetime of radicals in target molecules is
aboutabout
– 1010-3-3
secssecs
– 1010-6-6
secssecs
– 1010-9-9
secssecs
– 1010-12-12
secssecs
#2 – free radicals are highly unstable and reactive

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2.2. Electromagnetic radiation is considered ionizingElectromagnetic radiation is considered ionizing
if it has a photon energy greater thanif it has a photon energy greater than
– 1.24 eV1.24 eV
– 12.4 eV12.4 eV
– 124 eV124 eV
– 1.24 keV1.24 keV
#3 – this is sufficient to break bonds in biological
molecules

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3.3. The S.I. unit of absorbed dose isThe S.I. unit of absorbed dose is
– BecquerelBecquerel
– SievertSievert
– GrayGray
– RoentgenRoentgen
#3 The International System (IS) unit is the Gray, named
after the radiobiologist Louis “Hal” Gray who was based in
London

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4.4. Which of the following are not chargedWhich of the following are not charged
particles?particles?
– ElectronsElectrons
– NeutronsNeutrons
– ProtonsProtons
– Heavy ionsHeavy ions
– Alpha particlesAlpha particles
#2 – which is why they are called NEUTRons

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5.5. Which of the following is NOT a characteristic of theWhich of the following is NOT a characteristic of the
indirect action of ionizing radiationindirect action of ionizing radiation
– Production of diffusible free radicalsProduction of diffusible free radicals
– Production of reactive oxygen speciesProduction of reactive oxygen species
– Involvement of anti-oxidant defensesInvolvement of anti-oxidant defenses
– Causes a change in redox within a cell favoringCauses a change in redox within a cell favoring
reduction of constituentsreduction of constituents
#4 the free radicals produced makes ionizing
radiation an oxidative stress overall

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6.6. Which of the following is true about the oxygenWhich of the following is true about the oxygen
enhancement ratioenhancement ratio
– Is the same at all levels of cell survivalIs the same at all levels of cell survival
– Can be measured by the dog-leg in a cell survivalCan be measured by the dog-leg in a cell survival
curve after single high dose irradiation of tumorscurve after single high dose irradiation of tumors
– Is the ratio of doses needed for an isoeffect in theIs the ratio of doses needed for an isoeffect in the
absence to the presence of oxygenabsence to the presence of oxygen
– Is low for cells in S cell cycle phase compared toIs low for cells in S cell cycle phase compared to
cells in G2/M phasecells in G2/M phase
#3 responses should be compared by the doses
needed for a particular isoeffect. The OER varies with
the level of effect eg survival

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7.7. Which of the following is true about Linear EnergyWhich of the following is true about Linear Energy
TransferTransfer
– It is a measure of the biological effectiveness ofIt is a measure of the biological effectiveness of
ionizing radiationionizing radiation
– Shows an inverse correlation with the oxygenShows an inverse correlation with the oxygen
enhancement ratioenhancement ratio
– Is maximal at a relative biological effectiveness ofIs maximal at a relative biological effectiveness of
150 keV/micrometer150 keV/micrometer
– Is measured in keV/micrometerIs measured in keV/micrometer
#4 LET is an average value imparted per unit path length.
Because the radiations vary in energy, the LET is not
biologically very useful

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8.8. The Relative Biological Effectiveness of aThe Relative Biological Effectiveness of a
radiation isradiation is
– Assessed by the dose required for toAssessed by the dose required for to
produce the same effect as 250kVp X-raysproduce the same effect as 250kVp X-rays
– Is the ratio of the dose required of 250 kVpIs the ratio of the dose required of 250 kVp
X-rays to that of the test radiation for a givenX-rays to that of the test radiation for a given
isoeffectisoeffect
– Is directly related to Linear Energy TransferIs directly related to Linear Energy Transfer
– Is about 3 for alpha particle radiationIs about 3 for alpha particle radiation
#2 - again, measured by isoeffective doses – classically
relative to 250kVp x-rays, but often more recently 60Co
has been used

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9.9. The lethal lesion caused in DNA by low LETThe lethal lesion caused in DNA by low LET
ionizing radiation isionizing radiation is
– 15-20 nucleotides in size15-20 nucleotides in size
– Caused by alpha-type eventsCaused by alpha-type events
– Does not correlate with chromosomalDoes not correlate with chromosomal
aberrationsaberrations
– Due to oxygen fixationDue to oxygen fixation
#1 The lesions are large i.e they are not point mutations.

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10. Approximately how many DNA double10. Approximately how many DNA double
strand breaks are caused per cell per Gray?strand breaks are caused per cell per Gray?
– 1-101-10
– 15-2515-25
– 30-4030-40
– 45-6045-60
– 60-7560-75
#3 This probably varies considerably depending on
numerous factors, but this is a reasonable approximation

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11.11. If the fraction of cells surviving 2Gy irradiation isIf the fraction of cells surviving 2Gy irradiation is
0.5, what is a reasonable estimate of the percent of0.5, what is a reasonable estimate of the percent of
DNA double strand breaks that are effectivelyDNA double strand breaks that are effectively
repaired?repaired?
– 99%99%
– 95%95%
– 75%75%
– 50%50%
#1 If 60-80 DSB/2Gy kills half the cells, then >99% must
be repaired

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12. If the fraction of cells surviving 2Gy is 0.4,12. If the fraction of cells surviving 2Gy is 0.4,
what is the surviving fraction after 50 Gywhat is the surviving fraction after 50 Gy
given in 2Gy fractions?given in 2Gy fractions?
– 1010-8-8
– 1010-9-9
– 1010-10-10
– 1010-11-11
#3 - 0.425
= 1.12x10-10

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13. Sublethal DNA damage is most likely to13. Sublethal DNA damage is most likely to
accumulateaccumulate
– At high total doses given at high dose rateAt high total doses given at high dose rate
– At high total doses under hypoxiaAt high total doses under hypoxia
– After high LET radiationAfter high LET radiation
– After low fractionated doses of radiationAfter low fractionated doses of radiation
#1 Intertrack interactions between ionization events are
more likely at high dose and high dose rate

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15. Sublethal DNA damage is most likely to be repaired15. Sublethal DNA damage is most likely to be repaired
– After high total doses given at high dose rateAfter high total doses given at high dose rate
– If cells are held in a non-proliferative stateIf cells are held in a non-proliferative state
– After high LET radiationAfter high LET radiation
– Between low fractionated doses of radiationBetween low fractionated doses of radiation
#4 The lower the number of ionization events in time and
space, the more likely they are to be repaired

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15.15. Which of the following is true about chromatinWhich of the following is true about chromatin
structure in cellsstructure in cells
– Compacted chromatin is more radiosensitive thanCompacted chromatin is more radiosensitive than
non-compacted chromationnon-compacted chromation
– During mitosis cells decompact their chromatinDuring mitosis cells decompact their chromatin
and become radiosensitiveand become radiosensitive
– Compact chromatin in S phase mediatesCompact chromatin in S phase mediates
radioresistancyradioresistancy
– Compaction facilitates gene transcriptionCompaction facilitates gene transcription
#1 –compaction occurs in mitosis, which is why
chromosomes can be seen under the microscope during
this phase

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16.16. Which of the following is correct aboutWhich of the following is correct about
alpha-type cell killing following radiationalpha-type cell killing following radiation
exposureexposure
– It represents single lethal hitsIt represents single lethal hits
– It is due to accumulated damageIt is due to accumulated damage
– It requires intertrack interactionsIt requires intertrack interactions
– It is not oxygen dependentIt is not oxygen dependent
#1 – Intratrack lesions dominate and accumulated
damage plays only a small role

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17.17. Which of the following radiobiologicalWhich of the following radiobiological
phenomena occurring between dosephenomena occurring between dose
fractions has little or no effect on normalfractions has little or no effect on normal
tissue radiation responses?tissue radiation responses?
– RepairRepair
– Redistribution of cells in the cell cycleRedistribution of cells in the cell cycle
– RepopulationRepopulation
– ReoxygenationReoxygenation
#4 – Normal tissues are generally considered to be well
oxygenated

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