Medical Hypotheses I
Medical Hypohh-sar(1992) 39.366366 0L.Jn@mGroupIJKLtd1992
The Detachment
Cascade During Metastasis
C. H. VAN ASWEGEN and D. J. DU PLESSIS Department Department
of Urology, University of Pretoria, Pretoria 0001, South Africa (Correspondence to CHVA, of Urology, HF Verwoerd Hospital, Private Bag xl 69, Pretoria 0001, South Africa).
Abstract-The cause of detachment of tumour cells during metastasis is still one of the most intriguing questions of tumour propagation. A hypothesis is suggested herein for lysis of extracellular matrix that could ultimately lead to the detachment and spreading of malignant cells. According to this theory a certain optimal estrogen level initiates a series of enzymatic activations that culminate in detachment and spreading of tumour cells.
Cancer is one of the most common and troublesome health care problems in modem living. Little is known about the natural history and clinical causes of the disease and numerous questions remain unanswered. The metastatic process of cancer propagation can be summarized as follows (1): 1) After progressive growth of neoplastic cells; 2) extensive vascularization and; 3) local invasion of the host stroma by some of the tumour cells; 4) detachment and embolism ensue, followed by; 5) cell entrapment in the capillary beds of the organs and finally; 6) extravasion and; 7) proliferation within the parenchyma. One of these events, namely detachment of cells, implies breakdown of the extracellular matrix, allowing spreading of tumour cells to occur. We hypothesize that a cascade of mechanisms, initiated by estrogen, is responsible for this breakdown of the extracellular matrix, as schematically illustrated in the Figure. It is well known that estrogens are; possible carcinogens (2, 3), biphasic (4, 5), and are, at least, cofactors in carcinogenesis (6-17). On this basis we hypothesize that an optimal estrogen concentration may well initiate the detachment cascade and could exDate received Date accepted
18 June 1992 11 July 1992
PLASMINOCEN
1
,PLASMIN
I COLLACENASE 1 COLLAGEN 1 DETACHMENT OF CELLS Fig. Schematic diagram of the putative detachment cascade mechanisms resulting in breakdown of the extracellular matrix.
plain the unidentified role of estrogens in the prostate
364
365
THEDETACHMENI-CASCADEDLJRINGMETASTASIS
gland (18). Although it has been reported that some cancer tissues do not contain estrogen receptors, we consider that estrogen receptors are indeed present, but that these receptors are either occupied with estrogens (19) or are masked by sialic acid (20) at the time of assay. Estrogens, which are known to stimulate DNA and mRNA synthesis (21, 22), have a bifunctional effect on prostaglandins and peptide growth factors. Estrogen stimulates the prostaglandin F-type (23, 24) and E-type (25) which in turn stimulate urokinase (UK) activity (26). The second function of estrogens is to stimulate the production of UK via increased production of fibroblast growth factor (FGF), epidermal growth factor (EGF) and platelet derived growth facto1 (PDGF)(27-31). These growth factors are responsible for increasing the levels of UK (32-35). Pro-UK may be activated by: 1) autocatalytic intrinsic activation; 2) activation by serine proteases (plasmin, kallikrein, Factor xiii a) and/or, 3) activation by cysteine proteases (cathepsin B and L) (36). Increased UK levels and activity have been reported with various cancers (37-41) as well as a correlation between UK production and metastasis (36, 40,42, 43). Urokinase in its turn, activates collagenase by hydrolyzing plasminogen to plasmin which then activates latent collagenase (33,44,45) that degrades fibrillar collagen or basement membrane collagen (33). Plasmin also degrades both fibronectin and laminin (33). Collectively, these degradation processes lead to lysis of the extracellular matrix and metastasis of the neoplastic cells. The detachment of tumour cells is obviously a complex process, involving numerous interactions. Thus, whilst we admit that this hypothesis must be an over simplification of the real process, it does, nonetheless, illustrate a possible orchestration of estrogens with the relevant enzymes in the detachment cascade of tumour cells.
6.
7. 8.
9.
10.
11.
12.
13. 14. 15.
16.
17. 18.
19.
20.
21.
References 22. I.
Fidler 1 J. Critical factors in the biology of human cancer metastasis: Twenty-eight G.H.A. Clowes memorial award lecture. Cancer Res 50: 6130-6138. 1990. 2. Van Aswegen C H. Estradiol and catecholestradiols as passible genotoxic carcinogens. Clin Physiol Biochem 7: 34-39, 1989. 3. Nieuwoudt L B. van Aswegen C H, B&&man H I, van der Merwe C A, du Plessis D J. Correlation between the macromolecular effects of estradiol and catecholestradiols and the total prostatic catecholestrogen concentration. Clin Physiol Biochem 8: 231-237, 1990. 4. Lippman M, Bolan G, Huff K. ‘Ihe effects of estrogens and antiestrogens on hormone-responsive human breast cancer in long-term tissue culture. Cancer Res 36: 4595-4601. 1976. 5. Soto A M, Sonnenschein C. The role of estrogens at the
23.
24.
25.
26. 27.
proliferation of human breast tumor cells (MCF-7). J Steroid Biochem 23: 87-94. 1985. Gray L A, Cristopherson W M, Hoover R N. Estrogens and endometrial carcinoma. J Obstetr and Gynecol 49: 385-389.1972. Lipsett M B. Interaction of drugs, hormones and nutrition in the causes of cancer. Cancer 43: 19761981, 1979. Le Quesne P W, AMel-Baky S, Durga A V. Active nonaromatic intermediates in the conversion of steroidal estrogens into catechol estrogens. Biochemistry 25: 2065-2072, 1986. La Vecchia C, Decarli A, Parazzini F, Gentile A. Liberati C. Non-contraceptive oestrogens and the risk of breast cancer in woman. Int J Cancer 38: 853-858, 1986. Juriansz R L, Huseby R A, Wilcox R B. Interactions of putative estrogens with the intercellular receptor complex in mouse leydig cells: Relationship to pretteoplastic hyperplasia. Cancer Res 48: 14-18. 1988. Miller W R, O’Neill J S. The significance of steroid metabolism in human cancer. J Steroid Biochem Molec Biol 37: 317-325, 1990. Ebeling K. Nischan P, Schindler C H. Use of oral contraceptives and risk of invasive cervical cancer in previously screened woman. Int J Cancer 39: 427-430. 1987. Longman S M, Buehring G C. Oral contraceptives and breast cancer. Cancer 59: 281-287. 1987. Jick S S, Walker A M, Stergachis A, Jick H. Oral contraceptives and breast cancer. Br J Cancer 59: 618-621, 1989. Vessey M P. McPherson K, Villard-Mackintosh L. Yeates D. Oral contraceptives and breast cancer: latest findings in a large cohort study. Br J Cancer 59: 613-617, 1989. Hildensheim A, Reeves W C. Brintat L A. Lavary C, Brenes M, De La Guardia M, Godoy J, Rawls W E. Association of oral contraceptive use and human papillomaviruses in invasive cervical cancers. Int J Cancer 45: 860-864, 1990. Chilvers C E D. Deacon J M. Oral contraceptives and breast cancer. Br J Cancer 61: l-4, 1990. Griffiths K, Eaton C L. Harper M E, Peeling B, Davies P. Steroid hormones and the pathogenesis of benign prostatic hyperplasia. Eur Ural 20 (suppl): 68-77, 1991. McGuire W L, Chamness G C, Costlow M E, Shepherd R E. Hormone dependence in breast cancer. Metabolism 23: 75-100, 1974. Van Aswegen C H, van Rensburg H G J, Becker P J, Wittliff J L, du Plessis D J. Influence of sialic acid on the binding activity of estrogen receptors. Clin Phys Biochem 8: 169-178, 1990. Aitken S C, Lippman M E, Kasid A. Schoenberg D R. Relationship between the expression of estrogen-regulated genes and estrogen-stimulated proliferation of MCF-7 mammary tumor cells. Cancer Res 45: 2608-2615, 1985. Bigsby R M. Cunha G R. Estrogen stimulation of deoxyribonucleic acid synthesis in uterine epithelial cells which lack estrogen receptors. Endocrinology 119: 390-396, 19% Ham E A, Cirollo V J, Zanetti M E. Kuehl F A. Estrogen directed synthesis of specific prostaglandins in uterus. Rot Nat Acad Sci USA 72: 1420-1424, 1975. Orlicky D J, Lieberman R, Gerschersat L E. A role for prostaglandins in estrogen growth regulation. Medical Hypotheses 25: l-5, 1988. Katayama S. Lee J B. Estradiol stimulates rat renopapillary prostaglandin Ez (PGE& but not PGF2, biosynthesis. Endocrinology 117: 656-661, 1985. Strickland S. Beers W H. Studies on the role of plasminogen activator in ovulation. J Biol Chem 251: 5694-5702. 1976. Lippman M E, Dickson R B, Kasid A et al. Autocrine and paracrine growth regulation of human breast cancer. J Steroid
366
28. 29.
30.
3 1. 32.
33.
34.
35.
36.
MEDICALHYPOTHESES
B&hem 24: 147-154, 1986. Fisher D A, Salids E C, Barajas L. Epidermal growth factor and the kidney. Amur Rev Physio151: 6730. 1989. Tuomela T. Viinikka L, Perheentupa J. Effects of estradiol and progesterone on epidermal growth factor concentration in plasma, bile, urine submandibular gland and kidney of mouse. Horm Res 31: 143-147. 1989. Tuomela T. Miettinen P. Pesonen K, Viinikka L. Perheentupa J. EpidermaI growth factor in mice: Effects of estradiol, testosterone and dexamethasar. Acta Endocrinol 123: 211-217, 1990. Bosch R J L H. Patbogenesis of benign prostatic hyperplasia. Euro Urol 20 (suppl): 27-30, 1991. Lee L. Weinstein I B. Epidermal growth factor, like pharbol estres, induces plasminogen activator in HeLa cells. Nature 274: 69-97.1978. Ttyggvasort K, Hoyhtya M, Salo T. Proteolytic degradation of extracellular matrix in tumor invasion. Biochem Biophys Acta 907: 191-217, 1987. Isacchi A, Bergatsoni L, Statuto M et al. A mutant of basic fib&last growth factor that has lost the ability to stimulate plasminogen activator synthesis in endothehal cells. ~369 in The Fibroblast Growth Factor Family. Ann NY Acad Sci, vol 638. (A Baird, M Klagsbrum, eds) NY Acad Sci, 1991. Presta M, Maier J A M, Ragnotti G. The mitogenic signalling pathway but not the plasminogen activator-inducing pathway of basic fibroblast growth factor is mediated through protein kinase C in fetal bovine aortic endothelial cells. J Cell Biol 109: 1877-1884, 1989. Goretxki L. Schmitt M, Mann K et al. Effective activation of the proenzyme form of the urokinase-type plasminogen acti-
37.
38.
39.
40.
41.
42. 43.
44. 45.
vator (pro-uPA) by the cysteine protease cathepsin L. FEBS 297: 112-118. 1992. Markus G, Takita H, Camiolo S M, Corasanti J G, Evers J L, Hobika G H. Content and characterization of plasminogen activators in human lung tumors and normal lung tissue. Cancer Res 40: 841-848, 1980. Mira-y-Lopez R. Reich E, Stolfi R L. Martin D S, Ossowski L. Coordinate inhibition of plasminogen activator and tumor growth by hydrocortisone in mwse mammary carcinoma. Cancer Res 45: 2270-2276, 1985. De Bruin R A F, Griftioen G, Verspaget H W et al. Plasminogen activator profiles in neoplastic tissues of the human colon. Cancer Res 48: 4520-4524, 1988. Pereyra-Alfarso S. Haedo A, Joffe E B. Correlation between urokinase-type plasminogen activator production and the metastasizing ability of two murine mammary adenocarcinanas. lnt J Cancer 42: 59-63, 1988. Duffy M J, O’Grady P. Devaney D, O’Siorain L. Fennelly J J, Lijnen H R. Tissue-type plasminogen activator, a new prognostic marker in breast cancer. Cancer Res 48: 1348-1349, 1988. Saksela 0. Plasminogen activation and regulation of pericellular proteolysis. Biochem Biophys Acta 823: 35-65, 1985. Zucker S. A critical appraisal of the role of proteolytic enzymes in cancer invasion: emphasis on tumor surface proteinases. Cancer Inv 6: 219-231, 1988. Ossowski L. Reich E. Antibodies to plasminogen activator inhibit human tumor metastasis. Cell 35: 611-619, 1983. Duffy M J, O’Grady P Plasminogen activator and cancer. Eur J Cancer Chn Oncol 20: 577-582, 1984.
Medical Hypohh-sar(1992) 39.366366 0L.Jn@mGroupIJKLtd1992
The Detachment
Cascade During Metastasis
C. H. VAN ASWEGEN and D. J. DU PLESSIS Department Department
of Urology, University of Pretoria, Pretoria 0001, South Africa (Correspondence to CHVA, of Urology, HF Verwoerd Hospital, Private Bag xl 69, Pretoria 0001, South Africa).
Abstract-The cause of detachment of tumour cells during metastasis is still one of the most intriguing questions of tumour propagation. A hypothesis is suggested herein for lysis of extracellular matrix that could ultimately lead to the detachment and spreading of malignant cells. According to this theory a certain optimal estrogen level initiates a series of enzymatic activations that culminate in detachment and spreading of tumour cells.
Cancer is one of the most common and troublesome health care problems in modem living. Little is known about the natural history and clinical causes of the disease and numerous questions remain unanswered. The metastatic process of cancer propagation can be summarized as follows (1): 1) After progressive growth of neoplastic cells; 2) extensive vascularization and; 3) local invasion of the host stroma by some of the tumour cells; 4) detachment and embolism ensue, followed by; 5) cell entrapment in the capillary beds of the organs and finally; 6) extravasion and; 7) proliferation within the parenchyma. One of these events, namely detachment of cells, implies breakdown of the extracellular matrix, allowing spreading of tumour cells to occur. We hypothesize that a cascade of mechanisms, initiated by estrogen, is responsible for this breakdown of the extracellular matrix, as schematically illustrated in the Figure. It is well known that estrogens are; possible carcinogens (2, 3), biphasic (4, 5), and are, at least, cofactors in carcinogenesis (6-17). On this basis we hypothesize that an optimal estrogen concentration may well initiate the detachment cascade and could exDate received Date accepted
18 June 1992 11 July 1992
PLASMINOCEN
1
,PLASMIN
I COLLACENASE 1 COLLAGEN 1 DETACHMENT OF CELLS Fig. Schematic diagram of the putative detachment cascade mechanisms resulting in breakdown of the extracellular matrix.
plain the unidentified role of estrogens in the prostate
364
365
THEDETACHMENI-CASCADEDLJRINGMETASTASIS
gland (18). Although it has been reported that some cancer tissues do not contain estrogen receptors, we consider that estrogen receptors are indeed present, but that these receptors are either occupied with estrogens (19) or are masked by sialic acid (20) at the time of assay. Estrogens, which are known to stimulate DNA and mRNA synthesis (21, 22), have a bifunctional effect on prostaglandins and peptide growth factors. Estrogen stimulates the prostaglandin F-type (23, 24) and E-type (25) which in turn stimulate urokinase (UK) activity (26). The second function of estrogens is to stimulate the production of UK via increased production of fibroblast growth factor (FGF), epidermal growth factor (EGF) and platelet derived growth facto1 (PDGF)(27-31). These growth factors are responsible for increasing the levels of UK (32-35). Pro-UK may be activated by: 1) autocatalytic intrinsic activation; 2) activation by serine proteases (plasmin, kallikrein, Factor xiii a) and/or, 3) activation by cysteine proteases (cathepsin B and L) (36). Increased UK levels and activity have been reported with various cancers (37-41) as well as a correlation between UK production and metastasis (36, 40,42, 43). Urokinase in its turn, activates collagenase by hydrolyzing plasminogen to plasmin which then activates latent collagenase (33,44,45) that degrades fibrillar collagen or basement membrane collagen (33). Plasmin also degrades both fibronectin and laminin (33). Collectively, these degradation processes lead to lysis of the extracellular matrix and metastasis of the neoplastic cells. The detachment of tumour cells is obviously a complex process, involving numerous interactions. Thus, whilst we admit that this hypothesis must be an over simplification of the real process, it does, nonetheless, illustrate a possible orchestration of estrogens with the relevant enzymes in the detachment cascade of tumour cells.
6.
7. 8.
9.
10.
11.
12.
13. 14. 15.
16.
17. 18.
19.
20.
21.
References 22. I.
Fidler 1 J. Critical factors in the biology of human cancer metastasis: Twenty-eight G.H.A. Clowes memorial award lecture. Cancer Res 50: 6130-6138. 1990. 2. Van Aswegen C H. Estradiol and catecholestradiols as passible genotoxic carcinogens. Clin Physiol Biochem 7: 34-39, 1989. 3. Nieuwoudt L B. van Aswegen C H, B&&man H I, van der Merwe C A, du Plessis D J. Correlation between the macromolecular effects of estradiol and catecholestradiols and the total prostatic catecholestrogen concentration. Clin Physiol Biochem 8: 231-237, 1990. 4. Lippman M, Bolan G, Huff K. ‘Ihe effects of estrogens and antiestrogens on hormone-responsive human breast cancer in long-term tissue culture. Cancer Res 36: 4595-4601. 1976. 5. Soto A M, Sonnenschein C. The role of estrogens at the
23.
24.
25.
26. 27.
proliferation of human breast tumor cells (MCF-7). J Steroid Biochem 23: 87-94. 1985. Gray L A, Cristopherson W M, Hoover R N. Estrogens and endometrial carcinoma. J Obstetr and Gynecol 49: 385-389.1972. Lipsett M B. Interaction of drugs, hormones and nutrition in the causes of cancer. Cancer 43: 19761981, 1979. Le Quesne P W, AMel-Baky S, Durga A V. Active nonaromatic intermediates in the conversion of steroidal estrogens into catechol estrogens. Biochemistry 25: 2065-2072, 1986. La Vecchia C, Decarli A, Parazzini F, Gentile A. Liberati C. Non-contraceptive oestrogens and the risk of breast cancer in woman. Int J Cancer 38: 853-858, 1986. Juriansz R L, Huseby R A, Wilcox R B. Interactions of putative estrogens with the intercellular receptor complex in mouse leydig cells: Relationship to pretteoplastic hyperplasia. Cancer Res 48: 14-18. 1988. Miller W R, O’Neill J S. The significance of steroid metabolism in human cancer. J Steroid Biochem Molec Biol 37: 317-325, 1990. Ebeling K. Nischan P, Schindler C H. Use of oral contraceptives and risk of invasive cervical cancer in previously screened woman. Int J Cancer 39: 427-430. 1987. Longman S M, Buehring G C. Oral contraceptives and breast cancer. Cancer 59: 281-287. 1987. Jick S S, Walker A M, Stergachis A, Jick H. Oral contraceptives and breast cancer. Br J Cancer 59: 618-621, 1989. Vessey M P. McPherson K, Villard-Mackintosh L. Yeates D. Oral contraceptives and breast cancer: latest findings in a large cohort study. Br J Cancer 59: 613-617, 1989. Hildensheim A, Reeves W C. Brintat L A. Lavary C, Brenes M, De La Guardia M, Godoy J, Rawls W E. Association of oral contraceptive use and human papillomaviruses in invasive cervical cancers. Int J Cancer 45: 860-864, 1990. Chilvers C E D. Deacon J M. Oral contraceptives and breast cancer. Br J Cancer 61: l-4, 1990. Griffiths K, Eaton C L. Harper M E, Peeling B, Davies P. Steroid hormones and the pathogenesis of benign prostatic hyperplasia. Eur Ural 20 (suppl): 68-77, 1991. McGuire W L, Chamness G C, Costlow M E, Shepherd R E. Hormone dependence in breast cancer. Metabolism 23: 75-100, 1974. Van Aswegen C H, van Rensburg H G J, Becker P J, Wittliff J L, du Plessis D J. Influence of sialic acid on the binding activity of estrogen receptors. Clin Phys Biochem 8: 169-178, 1990. Aitken S C, Lippman M E, Kasid A. Schoenberg D R. Relationship between the expression of estrogen-regulated genes and estrogen-stimulated proliferation of MCF-7 mammary tumor cells. Cancer Res 45: 2608-2615, 1985. Bigsby R M. Cunha G R. Estrogen stimulation of deoxyribonucleic acid synthesis in uterine epithelial cells which lack estrogen receptors. Endocrinology 119: 390-396, 19% Ham E A, Cirollo V J, Zanetti M E. Kuehl F A. Estrogen directed synthesis of specific prostaglandins in uterus. Rot Nat Acad Sci USA 72: 1420-1424, 1975. Orlicky D J, Lieberman R, Gerschersat L E. A role for prostaglandins in estrogen growth regulation. Medical Hypotheses 25: l-5, 1988. Katayama S. Lee J B. Estradiol stimulates rat renopapillary prostaglandin Ez (PGE& but not PGF2, biosynthesis. Endocrinology 117: 656-661, 1985. Strickland S. Beers W H. Studies on the role of plasminogen activator in ovulation. J Biol Chem 251: 5694-5702. 1976. Lippman M E, Dickson R B, Kasid A et al. Autocrine and paracrine growth regulation of human breast cancer. J Steroid
366
28. 29.
30.
3 1. 32.
33.
34.
35.
36.
MEDICALHYPOTHESES
B&hem 24: 147-154, 1986. Fisher D A, Salids E C, Barajas L. Epidermal growth factor and the kidney. Amur Rev Physio151: 6730. 1989. Tuomela T. Viinikka L, Perheentupa J. Effects of estradiol and progesterone on epidermal growth factor concentration in plasma, bile, urine submandibular gland and kidney of mouse. Horm Res 31: 143-147. 1989. Tuomela T. Miettinen P. Pesonen K, Viinikka L. Perheentupa J. EpidermaI growth factor in mice: Effects of estradiol, testosterone and dexamethasar. Acta Endocrinol 123: 211-217, 1990. Bosch R J L H. Patbogenesis of benign prostatic hyperplasia. Euro Urol 20 (suppl): 27-30, 1991. Lee L. Weinstein I B. Epidermal growth factor, like pharbol estres, induces plasminogen activator in HeLa cells. Nature 274: 69-97.1978. Ttyggvasort K, Hoyhtya M, Salo T. Proteolytic degradation of extracellular matrix in tumor invasion. Biochem Biophys Acta 907: 191-217, 1987. Isacchi A, Bergatsoni L, Statuto M et al. A mutant of basic fib&last growth factor that has lost the ability to stimulate plasminogen activator synthesis in endothehal cells. ~369 in The Fibroblast Growth Factor Family. Ann NY Acad Sci, vol 638. (A Baird, M Klagsbrum, eds) NY Acad Sci, 1991. Presta M, Maier J A M, Ragnotti G. The mitogenic signalling pathway but not the plasminogen activator-inducing pathway of basic fibroblast growth factor is mediated through protein kinase C in fetal bovine aortic endothelial cells. J Cell Biol 109: 1877-1884, 1989. Goretxki L. Schmitt M, Mann K et al. Effective activation of the proenzyme form of the urokinase-type plasminogen acti-
37.
38.
39.
40.
41.
42. 43.
44. 45.
vator (pro-uPA) by the cysteine protease cathepsin L. FEBS 297: 112-118. 1992. Markus G, Takita H, Camiolo S M, Corasanti J G, Evers J L, Hobika G H. Content and characterization of plasminogen activators in human lung tumors and normal lung tissue. Cancer Res 40: 841-848, 1980. Mira-y-Lopez R. Reich E, Stolfi R L. Martin D S, Ossowski L. Coordinate inhibition of plasminogen activator and tumor growth by hydrocortisone in mwse mammary carcinoma. Cancer Res 45: 2270-2276, 1985. De Bruin R A F, Griftioen G, Verspaget H W et al. Plasminogen activator profiles in neoplastic tissues of the human colon. Cancer Res 48: 4520-4524, 1988. Pereyra-Alfarso S. Haedo A, Joffe E B. Correlation between urokinase-type plasminogen activator production and the metastasizing ability of two murine mammary adenocarcinanas. lnt J Cancer 42: 59-63, 1988. Duffy M J, O’Grady P. Devaney D, O’Siorain L. Fennelly J J, Lijnen H R. Tissue-type plasminogen activator, a new prognostic marker in breast cancer. Cancer Res 48: 1348-1349, 1988. Saksela 0. Plasminogen activation and regulation of pericellular proteolysis. Biochem Biophys Acta 823: 35-65, 1985. Zucker S. A critical appraisal of the role of proteolytic enzymes in cancer invasion: emphasis on tumor surface proteinases. Cancer Inv 6: 219-231, 1988. Ossowski L. Reich E. Antibodies to plasminogen activator inhibit human tumor metastasis. Cell 35: 611-619, 1983. Duffy M J, O’Grady P Plasminogen activator and cancer. Eur J Cancer Chn Oncol 20: 577-582, 1984.
