- Email: [email protected]
MORPHINE-6-GLUCURONIDE, AN IMPORTANT FACTOR IN INTERPRETING MORPHINE RADIOIMMUNOASSAYS
210 no significant effect on the apparent volume of distribution of antipyrine, but caused a significant prolongation of the mean
had
biological half-life of antipyrine from 11-4±2-33 h to 14-7±3’11 h (p=0’004). Contrary to previous findings,we have thus demonstrated that ketoconazole competitively inhibits the oxidative metabolism of antipyrine, a mechanism which probably underlies the reported ketoconazole-cyclosporin interaction in organ transplant patients. Department of Pharmaceutics, School
of Pharmacy, University of Pittsburgh, Pittsburgh, Pennsylvania, USA Clinical Pharmacokmetics Laboratory, School of Pharmacy, Texas Southern University, Houston, Texas 77004, USA; and University of Texas Medical School, Houston
ANIL P. D’MELLO MARTIN J. D’SOUZA
THEODORE R. BATES
1 Niemegeers CJE, Levron JCI, Awouters F, Janssen PAJ. Inhibition and induction of microsomal enzymes in the rat: a comparative study of four antimycotics: Miconazole, econazole, clotrimazole, and ketoconazole. Arch Int Pharmacodyn 1981; 251: 26. 2. Maurer G, Loosli HR, Schreier E, Keller B. Disposition of cyclosporin in several animal species and man I: Structural elucidation of its metabolites. Drug Metab Disposition 1984; 12: 120-26. 3. Danhof M, Breimer DD Studies on the different metabolic pathways of antipyrine in man I: Oral administration of 250, 500, and 1000 mg to healthy volunteers. Br J Clin Pharmacol 1979; 8: 529-37. 4. Daneshmend TK, Warnock DW, Ene MD, et al. Multiple dose pharmacokinetics of ketoconazole and their effects on antipyrine kinetics in man. J Antimicrob Chemother 1983; 12: 185-88. 5. Ferguson RM, Sutherland DER, Simmons RL, Najarian JS. Ketoconazole cyclosporin metabolism and renal transplantation. Lancet 1982; ii: 882-83. 6. Dieperink H, Moller J. Ketoconazole and cyclosporin. Lancet 1982; ii: 1217. 7 Morgenstern GR, Powles R, Robinson B, McElwain TJ. Cyclosporin interaction with ketoconazole and melphalan. Lancet 1982, ii: 1342. 8. Danhof M, DeGroot-van der Vis E, Breimer DD. Assay of antipyrine and its primary metabolites in plasma, saliva, and urine by high performance liquid chromatography and some preliminary results in man. Pharmacology 1979; 18: 210-23
INDICATION FOR THERAPY IN MULTIPLE MYELOMA: SHOULD IT BE STAGE OR STABILITY?
SIR,-Before the introduction of melphalan for the treatment of myeloma in 1958 the median survival of patients with multiple myeloma was less than 12 months. Melphalan-induced responses with or without prednisone became accepted as a consistent and I reproducible effect in about 50% of patients. No other combination of drugs has emerged in more than 25 years as clearly superior to this alkylating agent (or cyclophosphamide) and prednisone. An important factor in predicting survival has been the delineation of prognostic factors present at diagnosis. Renal function, performance status, and haemoglobin were all clearly identified as such factors by the Medical Research Council studies in myelomatosis2and by similar studies in the USA.This led to the clinical staging system of Durie and Salmon4which was initially correlated with tumour cell mass, based on in-vitro myeloma cell immunoglobulin secretion. Despite widespread use of this staging system, there is not universal agreement on its ability to be correlated with tumour cell mass5 or on its value in determining prognosis in the absence of renal failure. Stage IIIB has a grave prognosis but Durie’s data show that there is no difference between stage IA and IIA, and IIIA is only marginally inferior in prognosis to IIA.6 Survival differences between IA, IIA, and IIIA have not been found in other large cooperative studies.Staging itself is inadequate as a guide to chemotherapy because it takes no account of the biology of the tumour, as reflected in its labelling index,6 or the effects that the host’s immune system may have in modulating the neoplastic clone.8 Furthermore, because multiple myeloma does not have a contiguous method of spread and therapy does not change with stage, the value of stage above the simple prognostic indicators such as renal failure are doubtful. Furthermore, some patients show little fall in paraprotein levels (non-responders), yet6 have long survival despite minimal cell kill with chemotherapy.6 Several studies have suggested that rapid tumour regression is associated with short survival. 6, Interim data from the fourth MRC
trial show that in the absence of Bence-Jones proteinuria, 2-year survival in myeloma is not related to the fall in paraprotein. However, it has become clear that prognosis in myeloma is related to the attainment of a kinetically characteristic "plateau" phase. Chemotherapy does not prolong this state in which the myeloma cells are kinetically hypoactive and have a low tritiated thymidine labelling index.1O Patients in plateau disease have increased suppressor T-cells and exhibit light-chain isotype suppression.1.1 Aggressive disease shows loss of light-chain isotype suppression. Furthermore the loss of the immunological findings of plateau disease often precedes changes in other facets of tumour activity, such as change in paraprotein levels, and is valuable in the clinical monitoring of patients with multiple myeloma. 12 These immunological features are found in patients who are clearly in
plateau phase at presentation.13 Potentially leukaemogenic cytotoxic therapy is not indicated in these cases or in patients who establish plateau phase after chemotherapy. These medications should be stopped as soon as plateau phase is reached since continuation may do more harm than good. We suggest that the identification of "stable" disease may allow therapy only to be introduced at an appropriate time and lessen the cumulative toxic effect of chemotherapy. Thus patients with markers of plateau phase (light-chain isotype suppression) can be recognised at presentation as well as during response to therapy and do not require chemotherapy. A measure of "stability" may be more important in determining the need for chemotherapy than "stage". DOUGLAS E. JOSHUA ALAINE WEARNE HARRY KRONENBERG
Haematology Department, Royal Prince Alfred Hospital, Camperdown, NSW 2050, Australia 1. Alexanian 2
3. 4.
R, Bergsagel DE, Migliore PJ, et al. Melphalan therapy for plasma cell myeloma. Blood 1968; 31: 1-10 MRC working party for therapeutic trials in leukaemia. Report on the first myelomatosis trial: Analysis of presenting features of prognostic importance Br J Haematol 1973; 24: 123-39. Alexanian R, Balcerzak S, Bonnett JD, et al. Prognostic factors in multiple myeloma Cancer 1975; 36: 1192-201. Durie BGM, Salmon SE. A clinical staging system for multiple myeloma. Cancer 1975, 36: 842-54
5.
6.
7 8
Cooper DA, Miller AC, Penny R Biosynthesis of immunoglobulins or human immunoproliferative diseases: Comparison of tumour cell mass in multiple myeloma measured by synthetic rate: Studies with that calculated from clinical staging systems. Acta Haematol 1982; 68: 224-36. Dune BCM, Salmon SE, Moon TE. Pretreatment tumor mass, cell kinetics and prognosis in multiple myeloma Blood 1980; 55: 364-72. Pennec Y, Mottier D, Youinou P, et al. Critical study of staging in multiple myeloma ScandJ Haematol 1983; 30: 183-90. Leonard RCF, Maclennan ICM, Smart Y, et al Light chain isotype-associated suppression of normal plasma cell numbers in patients with multiple myeloma Int J Cancer 1979; 24: 385-93.
9 Hobbs JR. Immunological classes of myelomatosis. Including data from a therapeutic trial conducted by a Medical Research Council working party. Br J Haematol 1969,
16: 607. 10 11
Durie BGM, Russell DH, Salmon SE. Reappraisal of plateau phase in myeloma. Lancet 1980, ii: 65-67 Wearne A, Joshua DE, Kronenberg H. Light chain isotype associated suppression of surface
immunoglobulin
expression
on
peripheral
blood
lymphocytes in myeloma
during plateau phase. Br J Haematol 1984, 58: 483-89. 12. Wearne A, Joshua DE, Kronenberg H. Monitoring myeloma: light chain isotype suppression: a new parameter Aust NZ J Med (in press). 13. Alexanian R Localised and indolent
myeloma.
Blood 1980; 56: 521-25.
MORPHINE-6-GLUCURONIDE, AN IMPORTANT FACTOR IN INTERPRETING MORPHINE RADIOIMMUNOASSAYS
SIR,-Dr Joel and colleagues (May 11, p 1099) have drawn attention to the potential importance of morphine-6-glucuronide (M-6-G) to the clinical pharmacology of morphine. M-6-G has and’is present greater analgesic activity than the parent in plasma in concentrations that exceed those of morphine itself." Radioimmunoassays (RIA) are often used to measure plasma morphine concentrations, but the contribution of M-6-G to the results obtained has not been considered. The specificity of an antiserum is profoundly influenced by the structure of the immunogen used to elicit the antibodies.3 Antisera raised to immunogens prepared through the 0-6 position of
compound
morphine generally display low percentage cross-reactivity with morphine-3-glucuronide (M-3-G), the major inactive metabolite of morphineIt is likely, though, that these antisera will cross-react
211 much greater extent with M-6-G. The cross-reaction of M-6-G (kindly supplied by Dr S. Joel and Dr M. Slevin) with two morphine antisera is described here. Antiserum G/G/1was raised in a goat in to a
response
conjugate
to
6-succinylmorphine/bovine-serum-albumin (BSA)
and
G/S/7
was
raised
in
a
sheep
to
an
N-succinylnormorphine-BSA conjugate.6 Cross-reaction at 50% of zero binding of both M-6-G and M-3-G has: morphine M-3-G M-6-G
G/G/7 100% 2-52% >100%
G/S/7 100% 0-011% 0 013070
The effect of these cross-reactions on the measurement of morphine can be illustrated by using normal drug-free plasma spiked with a known amount of morphine (100 ng/ml) and increasing amounts of either M-3-G or M-6-G. With the more specific G/S/7 antiserum, the measurement of morphine is unaffected even when the metabolites are present at concentrations more than 1000 times that of morphine. In contrast, with the goat antiserum overestimation of morphine occurs in the presence of a 10-20 fold excess of M-3-G. M-6-G causes overestimation of morphine even when it is present at only half the concentration of
morphine. We believe, therefore, that for pharmacokinetic studies, antisera raised in response to immunogens prepared through the cyclic N position are superior reagents for the specific measurement of morphine itself. RIA results obtained with antisera raised to morphine-O-6 immunogens should be interpreted with the understanding that they probably reflect the levels of both morphine and M-6-G. Department of Biochemistry, University of Surrey, Guildford, Surrey GU2 5XH
G. WYNNE AHERNE P. LITTLETON
1 Shirmomura K, Kamato O, Ueki S, 2.
et al. Analgesic effects of morphine glucuronides. Tohoku J Exp Med 1971; 105: 45-52. Svensson JO, Rane A, Sawe J, Sjöquist F. Determination of morphine, morphine-3glucuronide and (tentatively) morphine-6-glucuronide in plasma and urine using ion-pair high performance liquid chromatography. J Chromatogr 1982; 230:
427-32
Findlay JWA, Butz RF, Jones EC. Relationships between
immunogen structure and Clin Chem 1981; 27: 1524-35. 4. Aherne GW, Piall EM, Robinson JD, Morris BA, Marks V Two applications of a radioimmunoassay for morphine. In. Pasternak CA, ed. Radioimmunoassay in clinical biochemistry. London Heydon & Son, 1975: 81-91. 5 Moore RA, Baldwin D, Allen M, Watson P, Bulhngham RES, MacQuay HJ Sensitive and specific morphine radioimmunoassay with iodine label. Pharmacokinetics of morphine in man after intravenous administration. Ann Clin Biochem 1984; 21: 318-26. 6 Morris BA, Robinson JD, Piall E, Aherne GW, Marks V. Development of a radioimmunoassay for morphine having minimal cross-reactivity with codeine. J Endocrinol 1974; 64: 6-7. 3.
antisera
Our observations indicate that the elimination rate of morphine is similar in uraemic patients and in patients with normal kidney function.The most striking but also expected differences are the high plasma levels and long half-life of M-3-G in the uraemic subjects. Our results accord with those of Aitkenhead and coworkers,3who also used HPLC. Like us they found considerable differences in morphine kinetics between patients. However, the mean values of the terminal elimination half-life and the total body clearance were similar in patients and in controls. They concluded that the elimination of unchanged morphine is not impaired in patients with chronic renal failure although M-3-G and M-6-G might accumulate. Our data show that there is considerable accumulation of glucuronidated metabolites. The discrepancy between Ball and colleagues’ results and Aitkenhead’s and ours probably lies in the specificity of the analytical methods used.4The radioimmunoassay is non-specific for morphine since the antisera to some extent detects the glucuronides as well.5-7 Cross-reactivity with as little as 1% of M-3-G is likely to lead to sizeable errors in estimation of the plasma concentrations of morphine in patients with renal failure. The degree of cross-reactivity with M-6-G is not
known.8
The clinical importance of the high concentrations of morphine in uraemic patients is not known. So far there is no evidence that M-3-G has pharmacological activity, but M-6-G may have an analgesic effect of its own9and this may contribute to the i reported increased sensitivity to morphine in uraemic patients. 10, II Other contributing factors may be enterohepatic recirculation of morphine from intestinal hydrolysis of the glucuronides or alterations in drug response. The metabolism of morphine in patients with kidney disease is not significantly impaired but it remains to be seen whether uraemic patients have, nevertheless, altered sensitivity to morphine. If they do, pharmacokinetic and pharmacodynamic reasons will have to be
glucuronides
explored. Department of Clinical Pharmacology, Huddinge University Hospital,
J. SAWE J. O. SVENSSON
S-141 86
I. ODAR-CEDERLOF
1. Svensson
JO, Rane A, Säwe J, Sjöqvist F. Determination of morphine, morphine-3glucuronide, and (tentatively) morphine-6-glucuronide in plasma and urine using ion-pair high performance liquid chromatography. J Chromatogr 1982; 230:
specificity in the narcotic alkaloid series.
KINETICS OF MORPHINE IN PATIENTS WITH RENAL FAILURE
SIR,-Dr Ball and co-workers (April 4, p 784) discuss the use of morphine in renal failure in the light of data on morphine kinetics obtained by radioimmunoassay. Using a specific high performance liquid chromatographic (HPLC) method’ we have measured the concentrations of morphine and morphine-3 (M-3-G) and morphine-6 (M-6-G) glucuronides in plasma from patients with renal failure (serum creatinine 220-952 pmol/1). Seven patients received a single 4 mg intravenous dose of morphine hydrochloride and drug concentrations were measured over 96-120 h. The lower limit of detection of morphine and M-3-G is 4 nmol/1.The apparent elimination half-life varied between 0 -6and 4’ 0 h (mean 2 -5), the volume of distribution between 11and 6 -33 1/kg (mean 4 - 3), and total plasma clearance between 9 - 0 and 31 . 3 ml/min/kg (mean 19 - 8). The corresponding mean values in patients with normal kidney function are 1 -7h, 4 - 0 lIkg, and 28 ml/min/kg.2 The concentrations of the glucuronidate metabolites rapidly rose to exceed those of morphine in all patients and then remained high throughout the observation period. The elimination half-life of the main metabolite (M-3-G) varied between 14’5 5 and 66.1 1 h (mean 41-0). These values are distinctly different from those obtained in patients with normal kidney function,(range 2’ 4—6 -7 h, mean 4 -0; unpublished).
Huddinge, Sweden
427-32 2. Säwe J, Kager L, Svensson JO, Rane A. Oral morphine in cancer patients in vivo kinetics and in vitro hepatic glucuronidation. Br J Clin Pharmacol 1985; 19: 495-501. 3. Aitkenhead AR, Vater M, Achola K, Cooper CMS, Smith G. Pharmacokinetics of single-dose iv morphine in normal volunteers and patients with endstage renal failure Br J Anaesth 1984; 56: 813-19.
4. Moore RA, Baldwin D, Allen M, Watson P, Bullingham RES, MacQuay HJ. Sensitive and specific morphine radioimmunoassay with iodine label. Pharmacokinetics of morphine in man after intravenous administration. Ann Clin Biochem 1984; 21: 318-25. 5. Aherne GW. The specificity of
morphine radioimmunoassays Roy Soc Med Int Congr Symp Ser 1983; 58: 21-26. 6. Grabinski PY, Kaiko RF, Walsh TD, Foley KH, Houde RW. Morphine radioimmunoassay specificity before and after extraction of plasma and cerebrospinal fluid. Pharmacol Sci 1983, 72: 27-30. 7 Hanks GW, Aherne GW. Morphine metabolism. Does the renal hypothesis hold 8 9
water? Lancet 1985, i: 221. Joel SP, Osborne RJ, Nixon NS, Slevin ML Morphine-6-glucuronide, an important metabolite Lancet 1985; i: 1099-100 Yosimura H, Ida S, Oguri F, Tsukamoto H Biochemical basis for analgesic activity of morphine-6-glucuronide I. Penetration of morphine-6-glucuronide in the brain of Biochem Pharmacol 1973, 22: 1423-30. HF, Dieppa RA, Taylor P. Narcotic analgesics in
rats.
10. Don
anuric
patients. Anesthesiology
1975; 42: 745. 11.
Reynard CFB, Twycross RG.
Metabolism of narcotics. Br MedJ
1984, 288:
860.
DENGUE IN NORTH-EAST AFRICA
SIR,-Investigations have been carried out on outbreaks of fever in Port Sudan and in Mogadishu (in Somalia). Both of these cities were found to be permissive to dengue transmission with Aedes aegypti house indices of greater than 50% (unpublished). In January, 1984, studies were undertaken in Port Sudan to investigate the cause of perennial epidemics of fever (treated as malaria). To date, using mammalian and mosquito cell lines, viruses have been isolated from twenty patients, and seventeen of these were identified as dengue type 2 and one was identified as dengue type 1 by fluorescent antibody studies with type-specific
had
biological half-life of antipyrine from 11-4±2-33 h to 14-7±3’11 h (p=0’004). Contrary to previous findings,we have thus demonstrated that ketoconazole competitively inhibits the oxidative metabolism of antipyrine, a mechanism which probably underlies the reported ketoconazole-cyclosporin interaction in organ transplant patients. Department of Pharmaceutics, School
of Pharmacy, University of Pittsburgh, Pittsburgh, Pennsylvania, USA Clinical Pharmacokmetics Laboratory, School of Pharmacy, Texas Southern University, Houston, Texas 77004, USA; and University of Texas Medical School, Houston
ANIL P. D’MELLO MARTIN J. D’SOUZA
THEODORE R. BATES
1 Niemegeers CJE, Levron JCI, Awouters F, Janssen PAJ. Inhibition and induction of microsomal enzymes in the rat: a comparative study of four antimycotics: Miconazole, econazole, clotrimazole, and ketoconazole. Arch Int Pharmacodyn 1981; 251: 26. 2. Maurer G, Loosli HR, Schreier E, Keller B. Disposition of cyclosporin in several animal species and man I: Structural elucidation of its metabolites. Drug Metab Disposition 1984; 12: 120-26. 3. Danhof M, Breimer DD Studies on the different metabolic pathways of antipyrine in man I: Oral administration of 250, 500, and 1000 mg to healthy volunteers. Br J Clin Pharmacol 1979; 8: 529-37. 4. Daneshmend TK, Warnock DW, Ene MD, et al. Multiple dose pharmacokinetics of ketoconazole and their effects on antipyrine kinetics in man. J Antimicrob Chemother 1983; 12: 185-88. 5. Ferguson RM, Sutherland DER, Simmons RL, Najarian JS. Ketoconazole cyclosporin metabolism and renal transplantation. Lancet 1982; ii: 882-83. 6. Dieperink H, Moller J. Ketoconazole and cyclosporin. Lancet 1982; ii: 1217. 7 Morgenstern GR, Powles R, Robinson B, McElwain TJ. Cyclosporin interaction with ketoconazole and melphalan. Lancet 1982, ii: 1342. 8. Danhof M, DeGroot-van der Vis E, Breimer DD. Assay of antipyrine and its primary metabolites in plasma, saliva, and urine by high performance liquid chromatography and some preliminary results in man. Pharmacology 1979; 18: 210-23
INDICATION FOR THERAPY IN MULTIPLE MYELOMA: SHOULD IT BE STAGE OR STABILITY?
SIR,-Before the introduction of melphalan for the treatment of myeloma in 1958 the median survival of patients with multiple myeloma was less than 12 months. Melphalan-induced responses with or without prednisone became accepted as a consistent and I reproducible effect in about 50% of patients. No other combination of drugs has emerged in more than 25 years as clearly superior to this alkylating agent (or cyclophosphamide) and prednisone. An important factor in predicting survival has been the delineation of prognostic factors present at diagnosis. Renal function, performance status, and haemoglobin were all clearly identified as such factors by the Medical Research Council studies in myelomatosis2and by similar studies in the USA.This led to the clinical staging system of Durie and Salmon4which was initially correlated with tumour cell mass, based on in-vitro myeloma cell immunoglobulin secretion. Despite widespread use of this staging system, there is not universal agreement on its ability to be correlated with tumour cell mass5 or on its value in determining prognosis in the absence of renal failure. Stage IIIB has a grave prognosis but Durie’s data show that there is no difference between stage IA and IIA, and IIIA is only marginally inferior in prognosis to IIA.6 Survival differences between IA, IIA, and IIIA have not been found in other large cooperative studies.Staging itself is inadequate as a guide to chemotherapy because it takes no account of the biology of the tumour, as reflected in its labelling index,6 or the effects that the host’s immune system may have in modulating the neoplastic clone.8 Furthermore, because multiple myeloma does not have a contiguous method of spread and therapy does not change with stage, the value of stage above the simple prognostic indicators such as renal failure are doubtful. Furthermore, some patients show little fall in paraprotein levels (non-responders), yet6 have long survival despite minimal cell kill with chemotherapy.6 Several studies have suggested that rapid tumour regression is associated with short survival. 6, Interim data from the fourth MRC
trial show that in the absence of Bence-Jones proteinuria, 2-year survival in myeloma is not related to the fall in paraprotein. However, it has become clear that prognosis in myeloma is related to the attainment of a kinetically characteristic "plateau" phase. Chemotherapy does not prolong this state in which the myeloma cells are kinetically hypoactive and have a low tritiated thymidine labelling index.1O Patients in plateau disease have increased suppressor T-cells and exhibit light-chain isotype suppression.1.1 Aggressive disease shows loss of light-chain isotype suppression. Furthermore the loss of the immunological findings of plateau disease often precedes changes in other facets of tumour activity, such as change in paraprotein levels, and is valuable in the clinical monitoring of patients with multiple myeloma. 12 These immunological features are found in patients who are clearly in
plateau phase at presentation.13 Potentially leukaemogenic cytotoxic therapy is not indicated in these cases or in patients who establish plateau phase after chemotherapy. These medications should be stopped as soon as plateau phase is reached since continuation may do more harm than good. We suggest that the identification of "stable" disease may allow therapy only to be introduced at an appropriate time and lessen the cumulative toxic effect of chemotherapy. Thus patients with markers of plateau phase (light-chain isotype suppression) can be recognised at presentation as well as during response to therapy and do not require chemotherapy. A measure of "stability" may be more important in determining the need for chemotherapy than "stage". DOUGLAS E. JOSHUA ALAINE WEARNE HARRY KRONENBERG
Haematology Department, Royal Prince Alfred Hospital, Camperdown, NSW 2050, Australia 1. Alexanian 2
3. 4.
R, Bergsagel DE, Migliore PJ, et al. Melphalan therapy for plasma cell myeloma. Blood 1968; 31: 1-10 MRC working party for therapeutic trials in leukaemia. Report on the first myelomatosis trial: Analysis of presenting features of prognostic importance Br J Haematol 1973; 24: 123-39. Alexanian R, Balcerzak S, Bonnett JD, et al. Prognostic factors in multiple myeloma Cancer 1975; 36: 1192-201. Durie BGM, Salmon SE. A clinical staging system for multiple myeloma. Cancer 1975, 36: 842-54
5.
6.
7 8
Cooper DA, Miller AC, Penny R Biosynthesis of immunoglobulins or human immunoproliferative diseases: Comparison of tumour cell mass in multiple myeloma measured by synthetic rate: Studies with that calculated from clinical staging systems. Acta Haematol 1982; 68: 224-36. Dune BCM, Salmon SE, Moon TE. Pretreatment tumor mass, cell kinetics and prognosis in multiple myeloma Blood 1980; 55: 364-72. Pennec Y, Mottier D, Youinou P, et al. Critical study of staging in multiple myeloma ScandJ Haematol 1983; 30: 183-90. Leonard RCF, Maclennan ICM, Smart Y, et al Light chain isotype-associated suppression of normal plasma cell numbers in patients with multiple myeloma Int J Cancer 1979; 24: 385-93.
9 Hobbs JR. Immunological classes of myelomatosis. Including data from a therapeutic trial conducted by a Medical Research Council working party. Br J Haematol 1969,
16: 607. 10 11
Durie BGM, Russell DH, Salmon SE. Reappraisal of plateau phase in myeloma. Lancet 1980, ii: 65-67 Wearne A, Joshua DE, Kronenberg H. Light chain isotype associated suppression of surface
immunoglobulin
expression
on
peripheral
blood
lymphocytes in myeloma
during plateau phase. Br J Haematol 1984, 58: 483-89. 12. Wearne A, Joshua DE, Kronenberg H. Monitoring myeloma: light chain isotype suppression: a new parameter Aust NZ J Med (in press). 13. Alexanian R Localised and indolent
myeloma.
Blood 1980; 56: 521-25.
MORPHINE-6-GLUCURONIDE, AN IMPORTANT FACTOR IN INTERPRETING MORPHINE RADIOIMMUNOASSAYS
SIR,-Dr Joel and colleagues (May 11, p 1099) have drawn attention to the potential importance of morphine-6-glucuronide (M-6-G) to the clinical pharmacology of morphine. M-6-G has and’is present greater analgesic activity than the parent in plasma in concentrations that exceed those of morphine itself." Radioimmunoassays (RIA) are often used to measure plasma morphine concentrations, but the contribution of M-6-G to the results obtained has not been considered. The specificity of an antiserum is profoundly influenced by the structure of the immunogen used to elicit the antibodies.3 Antisera raised to immunogens prepared through the 0-6 position of
compound
morphine generally display low percentage cross-reactivity with morphine-3-glucuronide (M-3-G), the major inactive metabolite of morphineIt is likely, though, that these antisera will cross-react
211 much greater extent with M-6-G. The cross-reaction of M-6-G (kindly supplied by Dr S. Joel and Dr M. Slevin) with two morphine antisera is described here. Antiserum G/G/1was raised in a goat in to a
response
conjugate
to
6-succinylmorphine/bovine-serum-albumin (BSA)
and
G/S/7
was
raised
in
a
sheep
to
an
N-succinylnormorphine-BSA conjugate.6 Cross-reaction at 50% of zero binding of both M-6-G and M-3-G has: morphine M-3-G M-6-G
G/G/7 100% 2-52% >100%
G/S/7 100% 0-011% 0 013070
The effect of these cross-reactions on the measurement of morphine can be illustrated by using normal drug-free plasma spiked with a known amount of morphine (100 ng/ml) and increasing amounts of either M-3-G or M-6-G. With the more specific G/S/7 antiserum, the measurement of morphine is unaffected even when the metabolites are present at concentrations more than 1000 times that of morphine. In contrast, with the goat antiserum overestimation of morphine occurs in the presence of a 10-20 fold excess of M-3-G. M-6-G causes overestimation of morphine even when it is present at only half the concentration of
morphine. We believe, therefore, that for pharmacokinetic studies, antisera raised in response to immunogens prepared through the cyclic N position are superior reagents for the specific measurement of morphine itself. RIA results obtained with antisera raised to morphine-O-6 immunogens should be interpreted with the understanding that they probably reflect the levels of both morphine and M-6-G. Department of Biochemistry, University of Surrey, Guildford, Surrey GU2 5XH
G. WYNNE AHERNE P. LITTLETON
1 Shirmomura K, Kamato O, Ueki S, 2.
et al. Analgesic effects of morphine glucuronides. Tohoku J Exp Med 1971; 105: 45-52. Svensson JO, Rane A, Sawe J, Sjöquist F. Determination of morphine, morphine-3glucuronide and (tentatively) morphine-6-glucuronide in plasma and urine using ion-pair high performance liquid chromatography. J Chromatogr 1982; 230:
427-32
Findlay JWA, Butz RF, Jones EC. Relationships between
immunogen structure and Clin Chem 1981; 27: 1524-35. 4. Aherne GW, Piall EM, Robinson JD, Morris BA, Marks V Two applications of a radioimmunoassay for morphine. In. Pasternak CA, ed. Radioimmunoassay in clinical biochemistry. London Heydon & Son, 1975: 81-91. 5 Moore RA, Baldwin D, Allen M, Watson P, Bulhngham RES, MacQuay HJ Sensitive and specific morphine radioimmunoassay with iodine label. Pharmacokinetics of morphine in man after intravenous administration. Ann Clin Biochem 1984; 21: 318-26. 6 Morris BA, Robinson JD, Piall E, Aherne GW, Marks V. Development of a radioimmunoassay for morphine having minimal cross-reactivity with codeine. J Endocrinol 1974; 64: 6-7. 3.
antisera
Our observations indicate that the elimination rate of morphine is similar in uraemic patients and in patients with normal kidney function.The most striking but also expected differences are the high plasma levels and long half-life of M-3-G in the uraemic subjects. Our results accord with those of Aitkenhead and coworkers,3who also used HPLC. Like us they found considerable differences in morphine kinetics between patients. However, the mean values of the terminal elimination half-life and the total body clearance were similar in patients and in controls. They concluded that the elimination of unchanged morphine is not impaired in patients with chronic renal failure although M-3-G and M-6-G might accumulate. Our data show that there is considerable accumulation of glucuronidated metabolites. The discrepancy between Ball and colleagues’ results and Aitkenhead’s and ours probably lies in the specificity of the analytical methods used.4The radioimmunoassay is non-specific for morphine since the antisera to some extent detects the glucuronides as well.5-7 Cross-reactivity with as little as 1% of M-3-G is likely to lead to sizeable errors in estimation of the plasma concentrations of morphine in patients with renal failure. The degree of cross-reactivity with M-6-G is not
known.8
The clinical importance of the high concentrations of morphine in uraemic patients is not known. So far there is no evidence that M-3-G has pharmacological activity, but M-6-G may have an analgesic effect of its own9and this may contribute to the i reported increased sensitivity to morphine in uraemic patients. 10, II Other contributing factors may be enterohepatic recirculation of morphine from intestinal hydrolysis of the glucuronides or alterations in drug response. The metabolism of morphine in patients with kidney disease is not significantly impaired but it remains to be seen whether uraemic patients have, nevertheless, altered sensitivity to morphine. If they do, pharmacokinetic and pharmacodynamic reasons will have to be
glucuronides
explored. Department of Clinical Pharmacology, Huddinge University Hospital,
J. SAWE J. O. SVENSSON
S-141 86
I. ODAR-CEDERLOF
1. Svensson
JO, Rane A, Säwe J, Sjöqvist F. Determination of morphine, morphine-3glucuronide, and (tentatively) morphine-6-glucuronide in plasma and urine using ion-pair high performance liquid chromatography. J Chromatogr 1982; 230:
specificity in the narcotic alkaloid series.
KINETICS OF MORPHINE IN PATIENTS WITH RENAL FAILURE
SIR,-Dr Ball and co-workers (April 4, p 784) discuss the use of morphine in renal failure in the light of data on morphine kinetics obtained by radioimmunoassay. Using a specific high performance liquid chromatographic (HPLC) method’ we have measured the concentrations of morphine and morphine-3 (M-3-G) and morphine-6 (M-6-G) glucuronides in plasma from patients with renal failure (serum creatinine 220-952 pmol/1). Seven patients received a single 4 mg intravenous dose of morphine hydrochloride and drug concentrations were measured over 96-120 h. The lower limit of detection of morphine and M-3-G is 4 nmol/1.The apparent elimination half-life varied between 0 -6and 4’ 0 h (mean 2 -5), the volume of distribution between 11and 6 -33 1/kg (mean 4 - 3), and total plasma clearance between 9 - 0 and 31 . 3 ml/min/kg (mean 19 - 8). The corresponding mean values in patients with normal kidney function are 1 -7h, 4 - 0 lIkg, and 28 ml/min/kg.2 The concentrations of the glucuronidate metabolites rapidly rose to exceed those of morphine in all patients and then remained high throughout the observation period. The elimination half-life of the main metabolite (M-3-G) varied between 14’5 5 and 66.1 1 h (mean 41-0). These values are distinctly different from those obtained in patients with normal kidney function,(range 2’ 4—6 -7 h, mean 4 -0; unpublished).
Huddinge, Sweden
427-32 2. Säwe J, Kager L, Svensson JO, Rane A. Oral morphine in cancer patients in vivo kinetics and in vitro hepatic glucuronidation. Br J Clin Pharmacol 1985; 19: 495-501. 3. Aitkenhead AR, Vater M, Achola K, Cooper CMS, Smith G. Pharmacokinetics of single-dose iv morphine in normal volunteers and patients with endstage renal failure Br J Anaesth 1984; 56: 813-19.
4. Moore RA, Baldwin D, Allen M, Watson P, Bullingham RES, MacQuay HJ. Sensitive and specific morphine radioimmunoassay with iodine label. Pharmacokinetics of morphine in man after intravenous administration. Ann Clin Biochem 1984; 21: 318-25. 5. Aherne GW. The specificity of
morphine radioimmunoassays Roy Soc Med Int Congr Symp Ser 1983; 58: 21-26. 6. Grabinski PY, Kaiko RF, Walsh TD, Foley KH, Houde RW. Morphine radioimmunoassay specificity before and after extraction of plasma and cerebrospinal fluid. Pharmacol Sci 1983, 72: 27-30. 7 Hanks GW, Aherne GW. Morphine metabolism. Does the renal hypothesis hold 8 9
water? Lancet 1985, i: 221. Joel SP, Osborne RJ, Nixon NS, Slevin ML Morphine-6-glucuronide, an important metabolite Lancet 1985; i: 1099-100 Yosimura H, Ida S, Oguri F, Tsukamoto H Biochemical basis for analgesic activity of morphine-6-glucuronide I. Penetration of morphine-6-glucuronide in the brain of Biochem Pharmacol 1973, 22: 1423-30. HF, Dieppa RA, Taylor P. Narcotic analgesics in
rats.
10. Don
anuric
patients. Anesthesiology
1975; 42: 745. 11.
Reynard CFB, Twycross RG.
Metabolism of narcotics. Br MedJ
1984, 288:
860.
DENGUE IN NORTH-EAST AFRICA
SIR,-Investigations have been carried out on outbreaks of fever in Port Sudan and in Mogadishu (in Somalia). Both of these cities were found to be permissive to dengue transmission with Aedes aegypti house indices of greater than 50% (unpublished). In January, 1984, studies were undertaken in Port Sudan to investigate the cause of perennial epidemics of fever (treated as malaria). To date, using mammalian and mosquito cell lines, viruses have been isolated from twenty patients, and seventeen of these were identified as dengue type 2 and one was identified as dengue type 1 by fluorescent antibody studies with type-specific