82
period between tests is not stipulated but is "a matter for discussion" by the two doctors diagnosing brainstem death.13 Donor numbers could probably be increased if the interval between the tests of brainstem function were shortened in potential donors who were cardiovascularly unstable. Sometimes the circulation can be improved by use of antidiuretic agents and hormones in addition to the more usual methods. Intensive care units should be able to obtain advice on these manoeuvres from the transplant coordinators. If the circulation collapses and asystole develops in the potential donor, it may still be possible to salvage the organs if the transplant team is on hand. "1n the English audit, five successful donations occurred under these circumstances.S Finally, there is a large group of patients who die in hospital with a possible diagnosis of brainstem death, in whom formal testing of brainstem function is never considered. In the English audit, such patients comprised 26% of those with possible brainstem death who died in intensive care units. Additionally, the Welsh audit has pointed to the existence of a large number of potentially suitable patients who die, unventilated, outside intensive care units. If such patients were to have become donors (which may well have been their wish), they would have needed to be ventilated and formally tested for brainstem death as already mentioned. In most hospitals the intensive care units are very busy and would have great difficulty in taking on this task. Ventilation, however, could take place in a high-dependency unit or even in a cubicle on a general ward. Since the potential for increasing donor numbers is greatest in this area, acute hospitals should be provided with funds to increase their facilities for ventilating patients. How then might the numbers work out? Improved measures for identifying and obtaining donors from intensive care units could raise the annual donation rate by 3 donors pmp. A rise of this order, albeit brief, is often observed when there has been nationwide publicity on the need for donor organs.14 The use of potential .donors dying on medical wards might provide another 5 donors pmp and this figure, when added to the current rate of donation, would give a total of 22 donors or 44 kidneys pmp. Such a figure is not too far removed from the perceived annual need of 48 kidneys pmp as calculated earlier. There is still a problem of finding kidneys for the large pool of patients already on the transplant waiting list. Some of this shortfall could be made up by increasing the use of
living related kidney donors. Raising the rate of kidney donation would also provide more hearts, heart-lungs, and livers for transplantation. The need for these organs is as great as for kidneys; at the end of October, 1989, 232 patients were waiting for heart transplants, 213 for heart-lungs, and 47 for liver transplants.4 The prospects for transplantation in the 1990s are not as bleak as the figures would at first suggest and there are practical ways by which donor numbers can be increased. Acute hospitals should look to how this
goal can be achieved in their own locality. Kingdom Transplant Service 1989 Annual Report. Obtainable from UKTS, Southmead Hospital, Bristol BS10 5NB. 2. Smith WGJ, Cohen DR, Asscher AW. Evaluation of renal services m Wales 1989. Report to the Welsh Office. 3. Jennett B. Brain death. Br J Anaesth 1981; 53: 1111-19. 4. Data from United Kingdom Transplant Service, Southmead Hospital, Bristol BS10 5NB. 5. Gore SM, Hinds CJ, Rutherford AJ. Organ donation from intensive care units in England. Br Med J 1989; 299: 1193-97. 6. Collins C. Organs for transplantation. Br Med J 1989; 299: 1463. 7. Combined report on regular dialysis and transplantation in Europe XIX. EDTA, 1988. 8. Vanrenterghem Y, Waer M, Roels T, et al. Shortage of kidneys, a solvable problem? The Leuven experience. Clinical transplants 1988. In: Terasaki P, ed. Los Angeles: UCLA Tissue Typing Laboratory, 1989. 9. Eurotransplant Newsletter no 69, November, 1989. 10. Road accidents. Great Britain 1988—The Casualty Report. London: HM Stationery Office, 1989. 11. Bodenham A, Berridge JC, Park GR. Brain stem death and organ donation. Br Med J 1989; 299: 1009-10. 12. Jennett B, Gentleman D. Brain stem death and organ donation. Br Med J 1989; 299: 1398-99. 13. Cadaveric organs for tranpslantation. A code of practice including the diagnosis of brain death. London: Department of Health, 1983. 14. United Kingdom Transplant Service bulletin no 68, 1989. Obtainable from UKTS, Southmead Hospital, Bristol BS10 5NR. 1. United
CHEMOTHERAPY: TOPOISOMERASES AS TARGETS An obvious goal of molecular studies in oncology is the identification of biochemical processes so characteristic of malignant cells that it will be possible to design and develop drugs with specific and predictable anti-cancer effects. This may be some way off in practice; most of the drugs in use today have evolved through empirical, even serendipitous, observation followed by chemical manipulation of a parent molecule and systematic screening until the most useful analogues have been identified. Somewhere between these two approaches comes the detailed study of compounds of proven value in cancer chemotherapy. The answers to the question "How do they work?" can be remarkably illuminating. Thus several drugs formerly regarded as
DNA-intercalating or chromosome-fragmenting agents are now known to act as topoisomerase "poisons". Two classes of DNA topoisomerase (types I and II) are recognised in mammalian cells and, in man at least, each is probably encoded by a single gene.1.2 The function of these enzyme systems appears to be to facilitate the
relaxation,
unwinding, controlled cleavage, and rejoining of the DNA helix during replication and transcription. DNA is such a large and unwieldy molecule that without topoisomerases it would be unable to participate in many biochemical reactions and would degenerate into an irretrievable tangle. Not surprisingly, topoisomerase activity tends to be high in cells that are metabolically very active, especially in those from rapidly dividing tissues. While much remains to be learned about the details of topoisomerase activity and the differences between types I and II, it seems clear that both are required for long-term cell survival. Topoisomerase I can cleave only a single DNA strand whereas type II cleaves both strands of the helix. The enzyme forms a protein bridge across the ends of the divided DNA molecule until continuity is restored, but topoisomerase poisons stabilise the DNA/protein complex so that the normally process of strand division, disentangling, and rejoining is
rapid
83
arrested at mid-stage. The mid-stage arrest appears to activate endogenous nucleases so that the cell does not merely stop growing-its DNA is degraded and it dies. The semisynthetic podophyllotoxin derivative etoposide (VP16) and its close analogue, teniposide, bind to the DNA/topoisomerase II complexes, as do the intercalating agents doxorubicin, actinomycin-D, mitoxantrone, bisantrene, and amsacrine (M-AMSA). Much, if not all, of their cytotoxic activity is attributable to their specific interactions with topoisomerase 11.3 None appears to bind to topoisomerase I, but this enzyme is the specific target for a plant alkaloid, 20(S)-camptothecin, which was investigated some twenty years ago but did not progress beyond phase I clinical trials as an anti-cancer drug."’’’ With the discovery of its mode of action, there has been renewed interest in this agent and its analogues6 and Giovanella and colleagues7 now suggest that these compounds may have a place in the chemotherapy of gastrointestinal tumours. Three human colon cancer cell lines, which could be propagated as subcutaneous tumour xenografts in nude mice, were shown to be much more sensitive to 9-amino camptothecin (9-AC) than to 5-FU, doxorubicin, or several other cytotoxic drugs. When each was tested as a single agent, only the
camptothecin-derivative was able to inhibit tumour growth completely and to induce total regression of an established xenograft at drug doses that were well tolerated by the animals. Whilst a single experiment of this type does not necessarily herald a new dawn for the clinical management of gastrointestinal tract cancer, efficacy in xenograft systems has generally proved to be a sound guide to drug sensitivity of tumours8 and there were some indications of response in patients with advanced gastrointestinal cancer in the early trials of carnptothecin.4,S That topoisomerases I and II can be "poisoned" independently of each other will undoubtedly lead to a much improved understanding of the distinct functions of the two enzyme systems in normal cell physiology. Selective resistance to inhibitors of both topoisomerases has been recorded and will obviously limit the value, for example, of 9-AC as a single agent. However, the scope for combinations of this drug with one or more of the topoisomerase II inhibitors remains to be explored. There are indications that topoisomerase levels are not only high but also that their regulation is qualitatively abnormal in tumour cells.3 If this finding can be confirmed, it may be possible to exploit the differences to increase the therapeutic index of
topoisomerase poisons. Doxorubicin and, to a far lesser extent, etoposide, have been of some value in the chemotherapy of gastrointestinal tract cancer;9.10 however, in general, results in this common group of diseases leave much to be desired. Recent advances, especially in the development of drug delivery systems, offer some grounds for optimism, but fresh studies on the camptothecins are to be welcomed not only for their potential as additions to the therapeutic arsenal but also for the insights they can provide into the molecular pathology of cancer.
1 Wang JC. DNA topoisomerases. Ann Rev Biochem 1985; 54: 665-97. 2. Futcher B. Supercoiling and transcription or vice versa. Trends Genet
1988; 4: 271-72. 3 Liu LF. DNA
topoisomerase poisons as anti-tumor drugs. Ann Rev Biochem 1989; 58: 351-75. 4 Gottlieb JA, Guarino AM, Call JB, Oliviero VT, Block JB. Preliminary pharmacological and clinical evaluation of camptothecin sodium (NSC-100880). Cancer Chemother Rep 1970; 54: 470.
Muggia FM, Creaven PJ, Hansen HH, Cohen MH, Selaway OS. Phase I clinical trials of weekly and daily treatment with camptothecin (NSC-100880): correlation with preclincial studies. Cancer Chemother Rep 1972; 56: 515-21. 6. Hsiang Y-H, Liu LF, Wall ME, et al. DNA topoisomerase I- mediated DNA cleavage and cytotoxicity of camptothecin analogues. Cancer Res 5.
1989; 49: 4385-89. 7. Giovanella BC, Stehlin
JS, Wall ME, et al. DNA topoisomerase-1targeted chemotherapy of human colon cancer in xenografts. Science 1989; 246: 1046-48. 8. Steel GG, Peckham MJ. Human tumour xenografts: a critical appraisal. Br J Cancer 1980; 41 (suppl): 133-41. 9. Cunningham D. Cytotoxic drugs for gastric and colorectal cancer. 10.
Br Med J 1989; 299: 1479-80. Fleming RA, Miller AA, Stewart CF. Etoposide:
an
update.
Clin
Pharmacol 1989; 8: 274-93.
HAEMODYNAMICS OF HYPERTENSION In established hypertension the high blood pressure is maintained by an increase in total peripheral resistance.But what initiates essential hypertension? Increased cardiac output has been reported in young hypertensive patients with normal calculated peripheral resistance.2.3 Although such subjects may well be a heterogeneous group, the autoregulation theory of hypertension has been proposed-the high cardiac output accounts for the initial rise in blood pressure and leads to perfusion of tissues in excess of their metabolic needs. Thus active tension of vascular smooth muscle increases via mechanisms similar to those involved in normal short-term autoregulation of peripheral blood flow.4 As a result, there is a compensatory rise in total peripheral resistance, which ultimately restores cardiac output to a value close to the initial level. This theory has staunch advocatess and fierce opponents.b6 Swedish workers7 have now studied the haemodynamics of young men with mild hypertension, as measured by sphygmomanometer, and of control subjects. The experiments were repeated in many of the men five years later. At the initial examination the heart rate was slightly increased and the cardiac index was significantly raised in the mildly hypertensive group. Direct arterial blood pressures were normal. Five years later heart rate and cardiac index had fallen in both groups whereas direct blood pressure and total peripheral resistance had not changed. Initially the individuals with hypertension were subdivided according to their cardiac index into hyperkinetic (cardiac index 3-86 1/min X mz) and normokinetic groups. Five years later the researchers found that the fall in cardiac index in the hypertensive group as a whole was entirely attributable to a decrease in the hyperkinetic patients. However, this subgroup showed no increased resistance to blood flow at maximum vasodilatation-ie, the experiment failed to show that the fall in cardiac output had been accompanied by structural adaptation in the resistance vasculature. No individual had manifested hypertension that required treatment. Although the follow-up may have been too short to show that sustained hypertension develops in individuals with a hyperkinetic circulation, no rise at all was seen over five years. Moreover, whilst the haemodynamic profile differed between the two subgroups of mildly hypertensive subjects, neither showed evidence of vascular remodelling. Other studies have shown that tissue oxygen uptake is increased during rest in individuals with increased cardiac output, and when cardiac output is plotted against oxygen
period between tests is not stipulated but is "a matter for discussion" by the two doctors diagnosing brainstem death.13 Donor numbers could probably be increased if the interval between the tests of brainstem function were shortened in potential donors who were cardiovascularly unstable. Sometimes the circulation can be improved by use of antidiuretic agents and hormones in addition to the more usual methods. Intensive care units should be able to obtain advice on these manoeuvres from the transplant coordinators. If the circulation collapses and asystole develops in the potential donor, it may still be possible to salvage the organs if the transplant team is on hand. "1n the English audit, five successful donations occurred under these circumstances.S Finally, there is a large group of patients who die in hospital with a possible diagnosis of brainstem death, in whom formal testing of brainstem function is never considered. In the English audit, such patients comprised 26% of those with possible brainstem death who died in intensive care units. Additionally, the Welsh audit has pointed to the existence of a large number of potentially suitable patients who die, unventilated, outside intensive care units. If such patients were to have become donors (which may well have been their wish), they would have needed to be ventilated and formally tested for brainstem death as already mentioned. In most hospitals the intensive care units are very busy and would have great difficulty in taking on this task. Ventilation, however, could take place in a high-dependency unit or even in a cubicle on a general ward. Since the potential for increasing donor numbers is greatest in this area, acute hospitals should be provided with funds to increase their facilities for ventilating patients. How then might the numbers work out? Improved measures for identifying and obtaining donors from intensive care units could raise the annual donation rate by 3 donors pmp. A rise of this order, albeit brief, is often observed when there has been nationwide publicity on the need for donor organs.14 The use of potential .donors dying on medical wards might provide another 5 donors pmp and this figure, when added to the current rate of donation, would give a total of 22 donors or 44 kidneys pmp. Such a figure is not too far removed from the perceived annual need of 48 kidneys pmp as calculated earlier. There is still a problem of finding kidneys for the large pool of patients already on the transplant waiting list. Some of this shortfall could be made up by increasing the use of
living related kidney donors. Raising the rate of kidney donation would also provide more hearts, heart-lungs, and livers for transplantation. The need for these organs is as great as for kidneys; at the end of October, 1989, 232 patients were waiting for heart transplants, 213 for heart-lungs, and 47 for liver transplants.4 The prospects for transplantation in the 1990s are not as bleak as the figures would at first suggest and there are practical ways by which donor numbers can be increased. Acute hospitals should look to how this
goal can be achieved in their own locality. Kingdom Transplant Service 1989 Annual Report. Obtainable from UKTS, Southmead Hospital, Bristol BS10 5NB. 2. Smith WGJ, Cohen DR, Asscher AW. Evaluation of renal services m Wales 1989. Report to the Welsh Office. 3. Jennett B. Brain death. Br J Anaesth 1981; 53: 1111-19. 4. Data from United Kingdom Transplant Service, Southmead Hospital, Bristol BS10 5NB. 5. Gore SM, Hinds CJ, Rutherford AJ. Organ donation from intensive care units in England. Br Med J 1989; 299: 1193-97. 6. Collins C. Organs for transplantation. Br Med J 1989; 299: 1463. 7. Combined report on regular dialysis and transplantation in Europe XIX. EDTA, 1988. 8. Vanrenterghem Y, Waer M, Roels T, et al. Shortage of kidneys, a solvable problem? The Leuven experience. Clinical transplants 1988. In: Terasaki P, ed. Los Angeles: UCLA Tissue Typing Laboratory, 1989. 9. Eurotransplant Newsletter no 69, November, 1989. 10. Road accidents. Great Britain 1988—The Casualty Report. London: HM Stationery Office, 1989. 11. Bodenham A, Berridge JC, Park GR. Brain stem death and organ donation. Br Med J 1989; 299: 1009-10. 12. Jennett B, Gentleman D. Brain stem death and organ donation. Br Med J 1989; 299: 1398-99. 13. Cadaveric organs for tranpslantation. A code of practice including the diagnosis of brain death. London: Department of Health, 1983. 14. United Kingdom Transplant Service bulletin no 68, 1989. Obtainable from UKTS, Southmead Hospital, Bristol BS10 5NR. 1. United
CHEMOTHERAPY: TOPOISOMERASES AS TARGETS An obvious goal of molecular studies in oncology is the identification of biochemical processes so characteristic of malignant cells that it will be possible to design and develop drugs with specific and predictable anti-cancer effects. This may be some way off in practice; most of the drugs in use today have evolved through empirical, even serendipitous, observation followed by chemical manipulation of a parent molecule and systematic screening until the most useful analogues have been identified. Somewhere between these two approaches comes the detailed study of compounds of proven value in cancer chemotherapy. The answers to the question "How do they work?" can be remarkably illuminating. Thus several drugs formerly regarded as
DNA-intercalating or chromosome-fragmenting agents are now known to act as topoisomerase "poisons". Two classes of DNA topoisomerase (types I and II) are recognised in mammalian cells and, in man at least, each is probably encoded by a single gene.1.2 The function of these enzyme systems appears to be to facilitate the
relaxation,
unwinding, controlled cleavage, and rejoining of the DNA helix during replication and transcription. DNA is such a large and unwieldy molecule that without topoisomerases it would be unable to participate in many biochemical reactions and would degenerate into an irretrievable tangle. Not surprisingly, topoisomerase activity tends to be high in cells that are metabolically very active, especially in those from rapidly dividing tissues. While much remains to be learned about the details of topoisomerase activity and the differences between types I and II, it seems clear that both are required for long-term cell survival. Topoisomerase I can cleave only a single DNA strand whereas type II cleaves both strands of the helix. The enzyme forms a protein bridge across the ends of the divided DNA molecule until continuity is restored, but topoisomerase poisons stabilise the DNA/protein complex so that the normally process of strand division, disentangling, and rejoining is
rapid
83
arrested at mid-stage. The mid-stage arrest appears to activate endogenous nucleases so that the cell does not merely stop growing-its DNA is degraded and it dies. The semisynthetic podophyllotoxin derivative etoposide (VP16) and its close analogue, teniposide, bind to the DNA/topoisomerase II complexes, as do the intercalating agents doxorubicin, actinomycin-D, mitoxantrone, bisantrene, and amsacrine (M-AMSA). Much, if not all, of their cytotoxic activity is attributable to their specific interactions with topoisomerase 11.3 None appears to bind to topoisomerase I, but this enzyme is the specific target for a plant alkaloid, 20(S)-camptothecin, which was investigated some twenty years ago but did not progress beyond phase I clinical trials as an anti-cancer drug."’’’ With the discovery of its mode of action, there has been renewed interest in this agent and its analogues6 and Giovanella and colleagues7 now suggest that these compounds may have a place in the chemotherapy of gastrointestinal tumours. Three human colon cancer cell lines, which could be propagated as subcutaneous tumour xenografts in nude mice, were shown to be much more sensitive to 9-amino camptothecin (9-AC) than to 5-FU, doxorubicin, or several other cytotoxic drugs. When each was tested as a single agent, only the
camptothecin-derivative was able to inhibit tumour growth completely and to induce total regression of an established xenograft at drug doses that were well tolerated by the animals. Whilst a single experiment of this type does not necessarily herald a new dawn for the clinical management of gastrointestinal tract cancer, efficacy in xenograft systems has generally proved to be a sound guide to drug sensitivity of tumours8 and there were some indications of response in patients with advanced gastrointestinal cancer in the early trials of carnptothecin.4,S That topoisomerases I and II can be "poisoned" independently of each other will undoubtedly lead to a much improved understanding of the distinct functions of the two enzyme systems in normal cell physiology. Selective resistance to inhibitors of both topoisomerases has been recorded and will obviously limit the value, for example, of 9-AC as a single agent. However, the scope for combinations of this drug with one or more of the topoisomerase II inhibitors remains to be explored. There are indications that topoisomerase levels are not only high but also that their regulation is qualitatively abnormal in tumour cells.3 If this finding can be confirmed, it may be possible to exploit the differences to increase the therapeutic index of
topoisomerase poisons. Doxorubicin and, to a far lesser extent, etoposide, have been of some value in the chemotherapy of gastrointestinal tract cancer;9.10 however, in general, results in this common group of diseases leave much to be desired. Recent advances, especially in the development of drug delivery systems, offer some grounds for optimism, but fresh studies on the camptothecins are to be welcomed not only for their potential as additions to the therapeutic arsenal but also for the insights they can provide into the molecular pathology of cancer.
1 Wang JC. DNA topoisomerases. Ann Rev Biochem 1985; 54: 665-97. 2. Futcher B. Supercoiling and transcription or vice versa. Trends Genet
1988; 4: 271-72. 3 Liu LF. DNA
topoisomerase poisons as anti-tumor drugs. Ann Rev Biochem 1989; 58: 351-75. 4 Gottlieb JA, Guarino AM, Call JB, Oliviero VT, Block JB. Preliminary pharmacological and clinical evaluation of camptothecin sodium (NSC-100880). Cancer Chemother Rep 1970; 54: 470.
Muggia FM, Creaven PJ, Hansen HH, Cohen MH, Selaway OS. Phase I clinical trials of weekly and daily treatment with camptothecin (NSC-100880): correlation with preclincial studies. Cancer Chemother Rep 1972; 56: 515-21. 6. Hsiang Y-H, Liu LF, Wall ME, et al. DNA topoisomerase I- mediated DNA cleavage and cytotoxicity of camptothecin analogues. Cancer Res 5.
1989; 49: 4385-89. 7. Giovanella BC, Stehlin
JS, Wall ME, et al. DNA topoisomerase-1targeted chemotherapy of human colon cancer in xenografts. Science 1989; 246: 1046-48. 8. Steel GG, Peckham MJ. Human tumour xenografts: a critical appraisal. Br J Cancer 1980; 41 (suppl): 133-41. 9. Cunningham D. Cytotoxic drugs for gastric and colorectal cancer. 10.
Br Med J 1989; 299: 1479-80. Fleming RA, Miller AA, Stewart CF. Etoposide:
an
update.
Clin
Pharmacol 1989; 8: 274-93.
HAEMODYNAMICS OF HYPERTENSION In established hypertension the high blood pressure is maintained by an increase in total peripheral resistance.But what initiates essential hypertension? Increased cardiac output has been reported in young hypertensive patients with normal calculated peripheral resistance.2.3 Although such subjects may well be a heterogeneous group, the autoregulation theory of hypertension has been proposed-the high cardiac output accounts for the initial rise in blood pressure and leads to perfusion of tissues in excess of their metabolic needs. Thus active tension of vascular smooth muscle increases via mechanisms similar to those involved in normal short-term autoregulation of peripheral blood flow.4 As a result, there is a compensatory rise in total peripheral resistance, which ultimately restores cardiac output to a value close to the initial level. This theory has staunch advocatess and fierce opponents.b6 Swedish workers7 have now studied the haemodynamics of young men with mild hypertension, as measured by sphygmomanometer, and of control subjects. The experiments were repeated in many of the men five years later. At the initial examination the heart rate was slightly increased and the cardiac index was significantly raised in the mildly hypertensive group. Direct arterial blood pressures were normal. Five years later heart rate and cardiac index had fallen in both groups whereas direct blood pressure and total peripheral resistance had not changed. Initially the individuals with hypertension were subdivided according to their cardiac index into hyperkinetic (cardiac index 3-86 1/min X mz) and normokinetic groups. Five years later the researchers found that the fall in cardiac index in the hypertensive group as a whole was entirely attributable to a decrease in the hyperkinetic patients. However, this subgroup showed no increased resistance to blood flow at maximum vasodilatation-ie, the experiment failed to show that the fall in cardiac output had been accompanied by structural adaptation in the resistance vasculature. No individual had manifested hypertension that required treatment. Although the follow-up may have been too short to show that sustained hypertension develops in individuals with a hyperkinetic circulation, no rise at all was seen over five years. Moreover, whilst the haemodynamic profile differed between the two subgroups of mildly hypertensive subjects, neither showed evidence of vascular remodelling. Other studies have shown that tissue oxygen uptake is increased during rest in individuals with increased cardiac output, and when cardiac output is plotted against oxygen
