Modifiers of Cell Survival: Repair • Sub-lethal damage repair • Half-time of repair • Potentially lethal damage repair • Effect of dose, dose rate, and cell type • Effect of dose fractionation • Effect of LET • Effect of oxygen/ hypoxia
Sub-lethal Damage • Can be repaired under normal circumstances • Two split of doses of radiation can overcome sub-lethal damage repair to cause lethal damage.
Sublethal Damage Repair • Increasing in cell survival that results from splitting RT into 2 fractions separated by a time interval • Dose divided into 2 equal fractions, separated by time, surviving fraction increases until a plateau is reached at about 2 hours
Sublethal Damage Repair Cell cycle not in progress Cell cycle in progress
SLDR • Previous experiment shows repair of SLD uncomplicated by movement of cells through the cell cycle • Longer intervals between the 2 doses results in the surviving fraction of cells decreasing again
Diagram explanation • Cells in the sensitive part of the cell cycle are killed • In first 2 hours, SLDR occurs • Resistant cells move to more sensitive part G2/M of cell cycle (REASSORTMENT) • When time interval exceeds cell cycle time, 10-12 hours, repopulation occurs and cell survival increases
Repair SLD in-vivo • The recovery factor is the ratio of SF resulting from 2-dose fractions to the SF from a single-dose fraction In neither case, there is dramatic dip in the curve at 6 hours resulting from movement of cells through the cell cycle, because the cell cycle is long The mouse tumor data show more repair in small 1-day tumors than in large hypoxic 6-tumors
EXPERIMENT • Illustrates 3 R’s of radiobiology: • Repair • Reassortment • Repopulation Additional “R”: • Re-oxygenation
Sublethal Damage Repair Half-time of repair As the time interval between the two dose fractions is increased, there is a rapid increase in the fractions of cells surviving due to the prompt repair of sublethal damage. The half-time of sublethal damage repair in mammalian cells is about 1 hour, but it may be longer in late-responding normal tissues in vivo.
SLDR • When dose delivered in 2 fractions, increase in cell survival b/c shoulder of the curve must be expressed each time • Significant for x-rays but almost nonexistent for neutrons • Repair of SLD reflects repair of DNA (DSB) breaks before they can interact to form lethal chromosomal aberration (multi-track)
Potentially Lethal Damage • Component of radiation damage that can be modified by post-irradiation environmental conditions • If left in same environment for 6-12 hours after RT before being subcultured, survival increases • Resistant tumors like melanoma? Large amount of PLDR is present
PLDR • In general, PLD is repaired and the fraction of cells surviving a given dose is enhanced if postirradiation conditions are suboptimal for growth so that cells do not have to attempt the complex process of mitosis while their chromosomes are damaged. • If mitosis is delayed by suboptimal growth conditions, DNA damage can be repaired
Effects of dose, dose rate, and cell type Effect of dose fractionation
DOSE-RATE EFFECT • Repair secondary to long radiation exposure. As dose rate decreases, survival increases b/c of more SLDR. Curve becomes shallower and shoulder disappears • More SLDR occurs during the protracted exposure
Idealized fractionated experiment Multiple small fractions approximate to a continuous exposure to a low dose rate Because continuous low-dose rate irradiation may be considered to be an infinite number of infinitely small fractions, the survival curve under these conditions also would be expected to have no shoulder and to be shallower than for single acute exposures
Examples of the dose-rate effect in-vitro and in-vivo Survival curves for HeLa cells exposed to -rays at high and low dose rates As the dose rate is reduced, the survival curve becomes shallower tends to disappear (I.e., survival curve becomes an exponential function of the dose) The dose rate effect caused by SLD is most dramatic between 1-100 rad/min. Above or below this dose-rate range, the survival curve changes little.
Dose-response curves for Chinese hamster cells (CHL-F line) exposed to cobalt 60-g-rays at various dose rates. The decrease in cell killing becomes even more dramatic as the dose rate is reduced further
Dose survival curves at high dose rate (HDR) and low dose rate (LDR) The survival curves fan out at LDR because of the presence of a range of repair times for SLD and further is associated with a range of inherent radiation sensitivities
Examples of the dose-rate effect in-vitro and in-vivo Response of mouse jejunal crypt cells irradiated with -rays. There isa dramatic dose-rate effect owing to the repair of sub-lethal radiation damage from acute exposure of 274 rad/min to a protracted exposure at 0.92 rad/min. Cell division dominates at LDR because the exposure time is longer than the cell cycle time.
INVERSE DOSE-RATE EFFECT • In some cell lines, an inverse dose-rate effect is evident • Reducing the dose rate increases the proportion of cells killed • Accumulation of cells in G2, sensitive phase of the cell cycle
Example of the inverse dose-rate effect in-vitro Decreasing the dose-rate from 154 rad to 37 rad/h increases the efficiency of cell killing, which is as effective as an acute exposure.
Dose-Rate Effect Summary • As dose rate decreases, SLDR occurs b/c of protracted exposure • In some cell lines, lowering the dose rate further allows cells to accumulate in G2, a sensitive part of the cell cycle and survival decreases • A further reduction will allow cells to pass through G2 and proliferation occurs if exposure time long compared to length of cell cycle
Very low dose rates continuous exposures • 2 rad/day continuous testis exposure does not affect reproduction • Small intestine maintain cell division and steady-state population at dose rate as high as 4 Gy/day • The blood forming tissues are intermediate between these two extremes • These depend on three important aspects (i) The cellular sensitivity of stem cells; (ii) The duration of cell cycle; and (iii) The ability of some tissues to adapt to the new trauma of continuous irradiation
Repair and radiation quality The shoulder is present for X-rays suggesting the presence of SLD repair, whereas, in response to neutrons there is no shoulder suggesting loss of SLD repair.
Cell-survival curves for Chinese hamster cells at various stages of the cell cycle From Sinclair W.K., Radiat Res. 33:620-643, 1968. The broken line is a calculated curve expected to apply to mitotic cells under hypoxia.
Survival curves for CHO cells exposed to X-rays in the presence of various oxygen concentrations. Oxygen is introduced gradually into the biologic system. The introduction of a very small quantity of oxygen, 100 ppm, is readily noticeable in a change in the slope of the survival curves. A concentration of 2,200 ppm, which is about 0.25% oxygen, moves the survival curve halfway toward the fully aerated condition.