BY Dr.HEMANTH

MODERATERDr.SWAPNA.J

ASST.PROFESSORDEPT. OF RADIOTHERAPY, SVIMS.

Introduction of fractionation

4 “ R “ s of radiobiology

5th R of radiobiology

Probably 6th R of radiobiology

THE INTRODUCTION OF FRACTIONATION

The multifraction regimens commonly used in conventional radiation therapy are a consequence of radiobiologicexperiments performed in France in the 1920s and 1930s.

It was found that a ram could not be sterilized by exposing its testes to a single dose of radiation without extensive skin damage to the scrotum

If the radiation was spread out over a period of weeks in a series of daily fractions, sterilization was possible without producing unacceptable skin damage .

It was postulated that the testes were a model of a growing tumor, whereas the skin of the scrotum represented a dose-limiting normal-tissue.

proved that fractionation of the radiation dose produces, in most cases, better tumor control for a given level of normal-tissue toxicity than a single large dose.

THE FOUR Rs OF RADIOBIOLOGYNow, more than 80 years later, we can account for the efficacy of fractionation based on more relevant radiobiologic experiments. We can appeal to the

“four Rs” of Radiobiology:

Repair of sublethal damage

Reassortment of cells within the cell cycle

Repopulation

Reoxygenation

Radiation damage to mammalian cells can operationally be divided into three categories:

(1) lethal damage,irreversible and irreparable and causes cell death.

(2) potentially lethal damage (PLD), can be modified by postirradiation environmental conditions.

(3) sublethal damage (SLD), can be repaired in hours unless additional sublethal damage is added with which it can interact to form lethal damage

Potentially Lethal Damage (PLD) Repair

causes cell death

PLD is repaired if cells are incubated in a balanced salt solution instead of full growth medium for several hours after irradiation which does not mimic a physiologic condition

Possible only in vitro

The importance of PLD repair to clinical radiotherapy is a matter of debate.

1.Sublethal Damage (SLD) Repair

operational term for the increase in cell survival that is observed if a given radiation dose is split into two fractions separated by a time interval.

Types of DNA damage

DNA REPAIR PATHWAYS

Nucleotide Excision Repair (NER)

Base Excision Repair (BER)

DNA Double-Strand Break Repair :

homologous recombination repair(HRR) requires undamaged DNA strand as a participant in the repair as a template.

occurs primarily in the late S/G2 phase of the cell cycle, when an undamaged sister chromatid is available to act as a template.

predominant pathway in lower eukaryotes like yeast.

nonhomologous end joining (NHEJ):

occurs in the G1 phase of the cell cycle, when no such template exists.

error prone and probably accounts for many of the premutagenic lesions induced in the DNA of human cells by ionizing radiation.

MECHANISM OF SLD REPAIR

the repair of sublethal damage reflects the repair and rejoining of double-strand breaks before they can interact to form lethal lesion.

If a dose is split into two parts separated by a time interval, some of the double-strand breaks produced by the first dose are rejoined and repaired before the second dose and more cells survive.

cultured mammalian cells maintained at room temperature (24°C) between the dose fractions to prevent the cells from moving through the cell cycle during this interval. This rather special experiment is described first because it illustrates repair of sublethalradiation damage uncomplicated by the movement of cells through the cell cycle.

2.Reassortment If an asynchronous population of cells present in

different phases of cell cycle is exposed to a large dose of radiation, more cells are killed in the sensitive than in the resistant phases of the cell cycle. The surviving population of cells progresses around cell cycle and said to be partly synchronized.

In Chinese hamster cells, most of the survivors from a first dose are located in the S phase of the cell cycle.

If about 6 hours are allowed to elapse before a second dose of radiation is given, this cohort of cells progresses around the cell cycle and is in G2/M, a sensitive period of the cell cycle, at the time of the second dose.

If the increase in radiosensitivity in moving from late S to the G2/M period exceeds the effect of repair of sublethal damage, the surviving fraction falls.

3.REPOPULATION Repopulation is the increase in cell division that is

seen in normal and malignant cells at some point after radiation is delivered.

In normal tissues Repopulation occurs in different speeds depending on the tissue.

early responding tissues begin repopulation at about 4 weeks.

By increasing treatment time over this amount, it is possible to reduce early toxicity in that tissue.

Late responding tissues only begin repopulation after a conventional course of radiation has been completed, and therefore repopulation has minimal effect on these tissues (the repair 'R' is more important for late tissues).

Accelerated repopulation

Treatment with any cytotoxic agent, including radiation, can trigger surviving cells (clonogens) in a tumor to divide faster than before. This is known as accelerated repopulation.

There is evidence for a similar phenomenon in human tumors.

Withers and his colleagues surveyed the literature on radiotherapy for head and neck cancer.

The analysis suggests that clonogen repopulation in this human cancer accelerates at about 28 days after the initiation of radiotherapy in a fractionated regimen.

A dose increment of about 0.6 Gy (60 rad) per day is required to compensate for this repopulation.

Such a dose increment is consistent with a 4-day clonogen doubling rate, compared with a median of about 60 days for unperturbed growth.

The conclusion to be drawn from this is that radiotherapy, at least for head and neck cancer and probably in other instances also, should be completed as soon after it has begun as is practicable.

It may be better to delay initiation of treatment than to introduce delays during treatment.

If overall treatment time is too long, the effectiveness of later dose fractions is compromised because the surviving clonogens in the tumor have been triggered into rapid repopulation.

Effect of oxygen in fixing radiation damage

representation of the dependence of radiosensitivity on oxygen concentration. If the radiosensitivity under extremely anoxic conditions is arbitrarily assigned unity the relative radiosensitivity is about 3 under well-oxygenated conditions. Most of this change of sensitivity occurs as the oxygen tension increases from 0 to 30 mm Hg. A further increase of oxygen content to that characteristic of air or even pure oxygen at high pressure has little further effect. A relative radiosensitivity halfway between anoxia and full oxygenation occurs for a pO2 of about 3 mm Hg, which corresponds to a concentration of about 0.5% oxygen.

Chronic hypoxia

(1) those that appear to be proliferating well and (2) those that are dead or dying.

Between these two extremes, region in which cells would be at an oxygen tension high enough for cells to be clonogenic but low enough to render the cells

protected from the effect of ionizing radiation.

Cells in this region would be relatively protected from a treatment with x-rays because of their low oxygen tension and could provide a focus for the subsequent regrowth of the tumor.

Acute hypoxia Regions of acute hypoxia develop in tumors as a

result of the temporary closing or blockage of a particular blood vessel. If this blockage were permanent, the cells downstream, would eventually die and be of no further consequence.

But tumor blood vessels open and close in a random fashion, so that different regions of the tumor become hypoxic intermittently.

In fact, acute hypoxia results from transient fluctuations in blood flow due to the malformed vasculature.

At the moment when a dose of radiation is delivered, a proportion of tumor cells may be hypoxic, but if the radiation is delayed until a later time, a different group of cells may be hypoxic.

4.REOXYGENATION Phenomenon by

which hypoxic cells become oxygenated after a dose of radiation is termed reoxygenation.

A modest dose of x-rays to a mixed population of aerated and hypoxic cells results in significant killing of aerated cells, but little killing of hypoxic cells.

Consequently, the viable cell population immediately after irradiation is dominated by hypoxic cells.

If sufficient time is allowed before the next radiation dose, the process of reoxygenation restores the proportion of hypoxic cells to about 15%.

If this process is repeated many times, the tumor cell population is depleted, despite the resistence of hypoxic cells to killing by x-rays.

In other words, if reoxygenation is efficient between dose fractions, the presence of hypoxic cells does not have a significant effect on the outcome of a multifractionregimen.

MECHANISM OF REOXYGENATION

Some tumors take days to reoxygenate ; and in

others process completes with in 1 hr

Contain two components fast and slow reflecting two types of hypoxia acute versus chronic

Some tumors show both fast and slow components

SLOW COMPONENTTAKES PLACE OVER A PERIOD OF DAYS IN CHRONICALLY HYPOXIC

CELLS

After a dose of radiation

Tumor cells killed and removed from population

tumor shrinks in size and restructuring or a revascularization of the tumor occurs

surviving cells previously beyond the range of oxygen diffusion become closer to a blood supply and so reoxygenate.

FAST COMPONENT First component of reoxygenation

complete within hours

caused by the reoxygenation of acutely hypoxic cells.

Those cells that were hypoxic at the time of irradiation because they were in regions in which a blood vessel was temporarily closed quickly reoxygenate when that vessel is reopened.

Importance of Reoxygenation in RT

If all the human tumors reoxygenate rapidly , use of a multifraction course of radiotherapy, extending over a period of time, can deal effectively with any hypoxic cells in human tumors.

- mouse mammary carcinoma that reoxygenates rapidly and well .

- rat sarcoma that shows two waves of reoxygenation .

- mouse osteosarcoma that does not reoxygenate at all for several days and then only slowly .

- mouse fibrosarcoma that reoxygenates quickly but not as completely as the mammary carcinoma .

- mouse fibrosarcoma that reoxygenates quickly and well . The proportion of hypoxic cells as a function

of time after irradiation with a large dose of x-rays for five transplanted tumors in experimental animals.

Making optimal choice of fractionation, demands a detailed knowledge of the time course of reoxygenation in the particular tumor to be irradiated.

Unfortunately, this information is available for only a few animal tumors and no information at present for human tumors. Indeed, in humans it is not known with certainty whether any or all tumors reoxygenated

Evidence from radiotherapy clinics that many tumorsare eradicated with doses of 60 Gy in 30 fractions argues strongly in favor of reoxygenation.

SUMMARY

Fractionation

allows the repair and regeneration of normal tissues.

sensitizes the tumor cells to radiation.

Repair in between the fractions benefits the normal tissues but makes the tumor cells more resistant.

Reassortment makes the tumor sensitive to RT

Repopulation or regeneration benefits the normal tissues in between fractions but makes the tumor cells resistant to RT

Reoxygenation makes the tumor sensitive to RT

Recovery in normal tissues after RT = repair + regeneration

REASSORTMENT & REOXYGENATION - FRIENDS OF TUMOR RADIOTHERAPY

REPAIR & REPOPULATION – FOES OF TUMOR RADIOTHERAPY (but friends of normal tissues)

5th R of RADIOBIOLOGYRADIOSENSITIVITY

Law of Bergonie & Tribondeau

Radiosensitivty of living tissues varies with

maturation & metabolism;

1. Stem cells are radiosensitive. More mature cells are

more resistant

2. Younger tissues are more radiosensitive

3. Tissues with high metabolic activity are highly

radiosensitive

4. High proliferation and growth rate, high radiosensitivty

Response of tissue determined by amount ofenergydeposited per unit mass (dose in Gy)

Two identical doses may not produce identical responses due to other modifying factors

Physical Factors Biological Factors

- linear energy thansfer – Oxygen Effect

- relative biological effectiveness – Age

- fractionation – Recovery

– Chemical Agents

– Hormesis

Radiosensitivityof different cells in humans in order from least sensitive to most sensitive

6th R of radiobiology ? REMOTE CELL KILL – BYSTANDER EFFECT

Phenomenon in which unirradiated cells exhibit irradiated effects as a result of signals received from nearby irradiated cells.

In November 1992, Hatsumi Nagasawa and John B. Little first reported this radiobiological phenomenon

Might be due to activation cells of innate immune system by ionizing radiation to produce pro-inflammatory mediators of genomic instability and cause suppression of cytokine production in the surrounding non-irradiated cells via the bystander effect.

ABSCOPAL effect(ab - away from ; scopal -target)

Defined as a reaction of cells within an organism that had not been directly exposed to irradiation, but cause tumor regression of the non-irradiated tumors (Postow et al., 2012).

Thought to be mediated by activation of the immune system via cytokines.

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