9 Hormone production by tumours: Biological and clinical aspects

9 Hormone Production by Tumours: Biological and Clinical Aspects ANDREE DE BUSTROS STEPHEN B. BAYLIN

The purpose of this review is not to present an extensive discussion or an exhaustive classification of hormonal syndromes associated with neoplasia. The reader is referred to several excellent publications dealing with this subject (Rees, 1975; Blackman, Rosen and Weintraub, 1978 ; Imura, 1980; Odell and Wolfsen , 1982) . Rather , in this review, we will try to (1) briefly summarize the well-defined clinical syndromes associated with hormone production by tumours and broadly outline a practical diagnostic and therapeutic approach to these entities; (2) summarize the current understanding of hormone production by tumours , with emphasis on events in the biosynthesis of these hormones in relation to malignant transformation and status of cellular differentiation of tumours producing hormones; and (3) give a perspective on ongoing and future directions of research on the problem of 'aberrant' production of hormones by cancer cells. We will stress the importance of such research for the fundamental understanding of regulation of gene expression , the relevance of hormonal tumour products as biological markers for tumour progression in the host and the crucial role that hormones play in cell to cell communication and possibly in the regulation of cell growth. Perhaps, most importantly , we will stress the concept that hormone production by tumours may not simply be a byproduct of genetic abnormalities in neoplastic cells but may indeed be instrumental in driving the neoplastic process. HISTORICAL PERSPECTIVE AND CHANGING CONCEPTS Liddle first used the term 'ecto pic' hormone production in conjunction with the elaboration , by neoplasms, of hormones that are usually not produced by the tissues of origin of these neoplasms (Liddle et al, 1969). While this term remains useful in terms of referring to the clinical manifestations of hormonal production by tumours (rectopic' hormone Clinics in Endocrinology and M etabolism -Vol. 14. No. I. Februar y 1985

221

222

A. DE BUSTROS AND S. B. BA YLIN

syndromes), it has now become apparent that the types of normal cells capable of hormone production are far more widespread than originally thought. It is now well established that polypeptides are present in sites other than the specialized glandular tissue responsible for their highest concentrations. Somatostatin, calcitonin, vasoactive intestinal peptide (VIP) , as well as several other peptides have been detected in the brain, using sensitive immunocytochemical techniques (Brownstein et ai , 1975; Said and Rosenberg, 1976; Fisher et ai , 1981). Immunoreactive insulin, growth hormone, corticotrophin (ACfH) and corticotrophin-releasing factor (CRF) have been found in a variety of normal tissues (Rosenzwe ig et ai, 1980; Kyle et ai, 1981; Saito et ai , 1983; Suda et ai , 1984). The trophoblastic cell glycoprotein product, human chorionic gonadotrophin (hCG) , is detectable in normal human serum, testes, and other normal tissues (Braunstein et ai, 1975; Yoshimoto et ai, 1977; Borkowski and Muquardt, 1979). Whereas most of the preceding data has been derived using immunological techniques, more sensitive and specific DNA-RNA hybridization assays are increasingly being used to detect the presence of hormones in normal tissues. For example, messenger RNA (mRNA) for the ACTH precursor molecule, proopiomelanocortin (POMC) , has been found in the Leydig cells of adult rat testis (Pintar et al , 1984). It is also fascinating that small polypeptide hormones classically considered only for their endocrine role in complex multicellular organisms have now also been found in primitive organisms such as protozoa. In these cells , the identified peptides are presumed to act locally, and thus playa role in the regulation of cell growth and differentiation (Sporn and Todaro, 1980; Kolata, 1982). Such cellular self-regulation ('autocrine' secretion pattern), as well as control of adjacent cells by diffusion of regulatory substances through the extracellular space ('paracrine' secretion pattern) , must also be at play in multicellular organisms, especially in early embryonic life, where dramatic rates of growth are seen , long before the development of the complex circulatory and endocrine systems. One then must place the question of peptide hormones in tumour cells into perspective for the above findings for normal cells. It is tempting to speculate that malignancy is a state of 'aberrant' cellular differentiation, with many cellular events ongoing which are operative in early embryonic life , and that the production of hormones by tumours could be tightly linked to neoplastic transformation and to the degree of differentiation of the cells giving rise to tumours. INCIDENCE OF ECTOPIC HORMONE SYNDROMES AND CRITERIA FOR DIAGNOSIS The incidence of hormonal production by neoplasms is very difficult to assess . Not all hormones produced by tumours produce clinical symptoms , either because they are released in the blood in a biologically inactive form, or because their levels are not high enough to produce distant

HORMONE PRODUCTION BY TUMOURS

223

manifestations. For instance, the biologically inactive ACTH precursor (big or pro-ACTH) has been detected in the sera of a large number of patients with lung cancer who do not suffer from Cushing's syndrome (Ratcliffe et al , 1972; Gewirtz and Yalow, 1974; Wolfsen and Odell, 1979). One then wonders how many hormones are produced in an asymptomatic manner by tumours, since an endocrine evaluation is rarely carried out in cancer patients who have no endocrine manifestations. Radioimmunoassays, which are still the traditional means of hormonal measurements, are far from perfect. The classic criteria for the demonstration of hormone synthesis by tumours, such as the presence of an A-V gradient for a given hormone across the tumour or the detection of immunoreactivity for such a hormone in tumour tissues, depend on these immunological techniques, which can suffer from lack of specificity. For example, conclusive evidence for parathyroid hormone (PTH) production by tumours in patients with cancer-related hypercalcaemia has been difficult to obtain, largely because of the technical limitations of PTH radioimmunoassays (Habener and Segre, 1979). The evolution of the techniques of molecular biology should provide, in the near future, a new perspective to the problem of hormone production by tumours. The advent of recombinant DNA techniques has allowed the characterization of many peptide hormone genes (Table 1). Using cloned cDNA or genomic DNA fragments and highly sensitive and specific DNA-RNA hybridization assays, one can determine the level of a particular peptide hormone mRNA species in tumours. Furthermore, the potentially powerful technique of in situ hybridization can be used to study, in tumours, the distribution of a specific mRNA species at the cellular level. and thus could help characterize the specific cell types involved in hormonal production.

Table l. Partial list of characterized hormone genes. 1. Corticotrophin-releasing factor (Furutani et al , 1983; Shibahara et al , 1983)

2. 3. 4. 5. 6. 7. 8. 9. 1(1.

11. 12. 13. 14. 15. 16.

Growth hormone-releasing factor (Gubler et al, 1983; Mayo et al, 1983) Somatostatin (Shen et al , 1982) Glucagon (Bell et al 1983; Lund et al, 1983) Gastrin (Ito et al , 1984; Wiborg et al, 1984) Calcitonin (Rosenfeld et al , 1983; Nelkin et al , 1984; Steenbergh et al , 1984b) Vasoactive intestinal peptide (VIP) (Tatemoto and Mutt. 1981) Insulin (Bell et al. 1980) Gastrin-releasing peptide (GRP) (bombesin) (Spindel et al , 1984) Vasopressin-oxytocin (Land et al , 1982; Richter. 1983) Proopiomelanocortin (POMe) (Nakanishi et al, 1979; Takahashi et al, 1981; Whitfeld et al, 1982) Pituitary glycoprotein hormones (LH, FSH. TSH) (Chin et ai, 1981, 1983; Counis et al , 1982; Gurr et al. 1983) Prolactin (Cooke et al. 1981) Growth hormone-chorionic somatomammotrophin (Fiddes et al, 1979; Seeburg, 1982) Human chorionic gonadotrophin (Fiddes and Goodman, 1979; Boothby et al , 1981; Policastro et al, 1983) Parathyroid hormone (Vasicek et al , 1983)

224

A. DE BUSTROS AND S. B. BA YLIN

BIOLOGICAL BASIS OF HORMONAL PRODUCTION BY TUMOURS

Several theories have been proposed in an attempt to explain the production of hormones by tumours, and several classifications have been put forward based on the histology and embryogenesis of tumour tissues. The most frequently cited concept is that of the 'APUD' (amine precursor uptake and decarboxylation) endocrine cell of origin and its relationship to biologically active small polypeptide hormones (Pearse, 1980). This cell system has been claimed to originate in the embryological neural crest. However, it is clear that hormone production is by no means restricted to tumours of ectodermal origin (Stevens and Moore, 1983). Thus, the theory of a distinct class of cells derived from the neural crest and uniquely qualified to produce hormones seems inadequate to explain ectopic hormone production by tumours. At least some cells with APUD characteristics must be present in all tumours possessing endocrine activity secondary to small polypeptide hormones, irrespective of the embryological origin of these tumours. Production of a mature hormone with biological activity requires the presence, in a cell, of specific biochemical and structural features such as processing enzymes involved in the cleavage of inactive hormone precursors, neurosecretory granules in which mature hormones are stored, and intact secretory mechanisms, all leading to release of mature hormones into the outer cell space. Thus, introduction of a proinsulin-SV40 recombinant vector into At T-20 cells (an ACTH-secreting mouse pituitary cell line), resulted in a stably transformed cell line capable of proteolytic processing of proinsulin to insulin and of releasing insulin into the culture medium upon stimulation with secretagogues. By contrast, similarly transformed fibroblast L-cells secrete only proinsulin and their secretion rate is unaffected by secretagogues (Moore et aI, 1983). Thus, it may follow from the above discussion that the forms of a peptide hormone stored and/or secreted by tumours might be determined by the degree of endocrine differentiation in the neoplasm. An excellent example for this point is found among lung tumours. Tumours with neuroendocrine features, such as small cell carcinoma (SCC), possess APUD characteristics and are most frequently responsible for producing endocrine syndromes secondary to excessive amounts of small polypeptide hormones (Rees, 1975; Imura, 1980). One explanation for the endocrine differentiation features of SCC is that the tumour arises from a separate cell of origin in the bronchial epithelium from the other major forms of lung cancer. However, there is firm evidence that other histological types of lung neoplasms, such as squamous cell carcinomas or adenocarcinomas, can have elevated levels of hormones such as ACTH, lipotrophin and calcitonin (Gewirtz and Yalow, 1974; Silva et aI, 1976; Odell et aI, 1979; Wolfsen and Odell, 1979), and even contain cell populations with APUD features (Baylin et aI, 1980; Berger et aI, 1981). Several lines of evidence lead us and other investigators to believe that the various histological types of lung tumours may represent a spectrum of differentiation in a single cell system. In the laboratory, established lines of SCC are observed to

HORMONE PRODUCTION BY TUMOURS

225

acquire, in late passages, a distinct non-SCC phenotype and lose their APUD features (Goodwin and Baylin, 1982). Also, in some patients, several histological types of lung cancer can be found at different times during the evolution of the disease in the same tumour tissue (Brereton et al, 1978; Abeloff et aI, 1979). APUD features, then, do not necessarily reflect tumour origin from an endocrine cell, but rather may arise as a consequence of a certain direction of differentiation, and the degree of endocrine differentiation may be a reflection of the level of cell maturation along this pathway (Baylin and Mendelsohn, 1982). It thus appears that tumour cells with a high degree of APUD differentiation, such as SCC cells, are able to produce and secrete mature hormones. Some tumour cells do not evolve the machinery needed for proteolytic processing of precursor hormones and thus produce only biologically inactive hormones. Tumour cells with an even lower degree of APUD differentiation are, perhaps, only able to transcribe certain peptide hormone genes. In such tumours, the elaboration of hormones might only be detectable by a DNA-RNA hybridization assay. Finally, tumour cells completely devoid of APUD characteristics may not even transcribe polypeptide hormone genes. Tumour evolution in the host is a dynamic process and the ongoing movement of differentiation within a tumour may lead to the evolution of heterogeneous cell populations, with variable degrees of APUD differentiation. For example, ACTH production and the ectopic Cushing's syndrome are seen to evolve only with the aggressive phase of certain tumours such as lung and prostatic carcinomas (Abeloff et al, 1981; Vuitch and Mendelsohn. 1981). In medullary thyroid carcinoma (MTC), a tumour of defined cell origin (C-cell of the thyroid) and for which distinct stages of development have been defined (C-cell hyperplasia, microscopic cancer, disseminated tumour), the levels of the mature hormone calcitonin decrease while those of the enzyme, L-dopa decarboxylase (DDC)-a marker of APUD features-increase during tumour progression (Trump, Mendelsohn and Baylin, 1979; Lippman et al. 1982). In the laboratory, we have observed a similar biochemical profile (low calcitonin production, elevated DDC levels) during the exponential growth phase of human MTC cells in culture (Berger et al, 1984). Recently, a proopiomelanocortin (POMC) specific eDNA probe was used to look at expression of the POMC gene in MTC. POMC-specific mRNA was detected in metastases derived from MTC but not in primary tumours, implying that tumour progression may be accompanied by the emergence of a new cell population with distinct properties (Steenbergh et al, 1984a). The molecular basis of polypeptide hormone production by tumours is far from being understood. The regulation of gene expression in neoplasms may be as complex as the regulation of gene expression in normal tissues. Random derepression of polypeptide hormone genes in tumours does not explain the fact that certain peptides are produced more frequently than others (calcitonin, ACTH), and that certain tumours tend to be associated with specific hormones (Shields, 1977). In general, small polypeptide hormones such as ACTH and calcitonin are produced by tumours

226

A. DE BUSTROS AND S. B. BA YLIN

exhibiting APUD characteristics such as small cell carcinomas of the lung and pancreatic islet cell tumours (Abe et al, 1977). By contrast, placental hormones such as human chorionic gonadotrophin hCG, tend to be produced by tumours lacking APUD characteristics, such as large cell carcinomas of the lung (Fusco and Rosen, 1966). Occasionally, a tumour has been found to produce both hormones of the APUD series (ACTH and calcitonin) and of placental origin (hCG) (Hattori et ai, 1979). The coexistence of several different hormones in the same tumour (Hirata et al, 1976; Hattori et ai, 1979) is interesting and has led some investigators to postulate the existence of a common precursor molecule for all the hormones produced by non-endocrine tumours (Lips et al, 1978). Although the characterization of hormone genes has largely disproved this theory, it is interesting to note that most peptide hormones are now known to be derived from genes that code for several other peptides (Douglass et al, 1984). Whether these genes are differentially processed in nonendocrine tumours as compared to normal or neoplastic endocrine tissues remains to be established. The widespread availability of various cloned hormone genes is allowing investigators to study the expression of these genes in tumours. So far, all the evidence points to the fact that the hormone genes expressed in tumours do not have major structural differences in coding regions from hormone genes expressed in the tissues of origin of these hormones (Boothby et al, 1981; DeBold et al, 1983). However, other structural gene changes such as DNA rearrangements, methylation, etc during malignant transformation and tumour progression must be more extensively studied. Feinberg and Vogelstein (1983) have shown that the growth hormone gene is hypomethylated in tumours as compared to normal adjacent tissue. Whether such hypomethylation correlates with increased gene expression or is simply a random event in neoplasia remains to be elucidated. ECTOPIC CUSHING'S SYNDROME

The association of Cushing's syndrome with non-endocrine tumours has been recognized for several decades, and symptoms of ACTH and cortisol excess have been described in a large variety of non-pituitary neoplasms (Meador et al, 1962; Friedman et al, 1966). However, tumours derived from APUD cells account for the vast majority of cases of ectopic Cushing's syndrome (Azzopardi and Williams, 1968). Small cell carcinoma (SCC) of the lung is associated with more than 50% of reported cases. Pancreatic islet cell tumours, carcinoid tumours of the lung and thymus, medullary thyroid carcinomas and phaeochromocytomas are responsible, in comparable percentages, for the remaining 50% of cases (Imura, 1980). The incidence of ectopic Cushing's syndrome, even in association with SCC ofthe lung, is low (3.2% to 4.8%) (Abeloff et al , 1981; Lokich, 1982). By contrast, ACTH immunoreactivity is found in the majority of lung tumour extracts and in the plasma of a large number of patients with lung cancer (Ratcliffe et al, 1972; Gewirtz and Yalow, 1974; Wolfsen and Odell,

HORMONE PRODUCTION BY TUMOURS

227

1979). This immunoreactivity is usually present in a large molecular weight form called 'big' or 'pro' ACTH, thought to be biologically inactive. Thus, ectopic ACTH production is common in patients with lung cancer, but is rarely associated with the symptomatology of Cushing's syndrome. Biological basis of the ectopic Cushing's syndrome The approach to this clinical entity reflects the dramatic evolution of medicine and endocrinology from purely descriptive disciplines to clinical sciences that rely heavily on basic research in the areas of biochemistry, immunology and, recently, molecular biology.

ACTH and POMC-derived peptides In 1928, Brown described a patient with Cushing's syndrome, and noted the existence at autopsy of an oat (small) cell carcinoma (SCC) of the lung (Brown. 1928). The precise causal relationship between the two entities was not delineated. In 1961, corticotrophic activity was measured in the plasma of two patients with cancer, using an adrenal weight maintenance assay (Christy, 1961). Shortly thereafter, an ACTH-like substance was identified in tumour tissues, thus tracing the origin of this hormonal material to a non-endocrine source (Holub and Katz, 1961). In his classic review in 1969, Liddle characterized further the syndrome of 'ectopic' ACTH production, and suggested that the 'ectopic' ACTH molecule was similar to pituitary ACTH (Liddle et al, 1969). Subsequently, several investigators attempted to characterize further the structure of the ACTH molecule produced by non-endocrine tumours (Orth et al, 1973; Lowry et al , 1976). Tumours producing ACTH immunoreactivity were shown to elaborate (3MSH immunoreactivity as well (Abe et al, 1967; Shapiro et aI, 1971; Schteingart et al, 1972). Bloomfield et al (1974) showed that (3MSH was an artefact of protein extraction techniques, and Chretien et al (1977) proposed that (3lipotrophin ((3LPH), originally characterized by Li et al (1965), could be a precursor of (3MSH and (3-endorphin. (3-Lipotrophin as well as (3-endorphin were subsequently shown to be produced by non-endocrine tumours (Bertagna et al. 1978; Odell et al, 1979). The elucidation by Eipper and Mains of the biosynthetic pathway for ACTH in the pituitary, and their demonstration of a common ACTHI(3-lipotrophin precursor, provided a unifying theory to the previous observations (Mains et al, 1977; Eipper and Mains, 1980). Further research suggested that this precursor molecule was found in non-pituitary ACTH-producing tumours as well (Bertagna et al , 1978; Orth et al , 1978). Recently, RNA extracted from both an ACTH-producing thymic carcinoid and from a pituitary gland was found to direct the synthesis in a cell-free translation system of a protein with ACTH and f:3-endorphin immunoreactivity (Tsukada et al, 1981). Hybridization of a cDNA for bovine ACTHI(3-lipotrophin (proopiomelanocortin or POMC) precursor to RNA extracted from the carcinoid tumour revealed two species of POMC-specific mRNA: a predominant species

228

A. DE BUSTROS AND S. B. BA YLIN

similar in size to pituitary POMC specific mRNA and a minor, slightly larger species. Analysis of another carcinoid tumour, using a human cDNA POMC clone, yielded similar results (DeBold et aI, 1983). These studies suggest that a similar POMC gene is expressed in both pituitary tissue and non-pituitary ACTH-producing tumours. It is unclear at present whether the POMC mRNA species detected in low amounts exclusively in the non-pituitary ACTH-producing tumours is derived from a second POMC gene or whether it represents an incompletely processed transcript of the same gene. Further research should help elucidate the regulation of the POMC gene expression both in pituitary tissue and in non-endocrine tumours where ACTH production is considered to be 'aberrant'. CRF

In 1971, Upton and Amatruda detected corticotrophin-releasing-factor (CRF)-like activity in association with ACTH, in pancreatic and lung tumours of patients with the ectopic Cushing's syndrome (Upton and Amatruda, 1971). However, the biochemical and immunological characterization of this CRF-like activity was not established until very recently, when CRF was characterized from ovine (Vale et al, 1981; Furutani et al, 1983), rat (Rivier et aI, 1983), and human (Shibahara et aI, 1983) hypothalami. The structure of human CRF has been found to be identical to rat CRF and to differ from ovine CRF by seven out of 41 amino acid residues. Using newly developed antisera to rat and human CRF, Carey et al detected CRF immunoreactivity in metastatic prostatic carcinoma cells (Carey et al, 1984). They also showed that this tumour CRF coelutes with rat and human CFR on high-pressure liquid chromatography. Thus, ectopic production of CRF is a cause of Cushing's syndrome in humans. Clinical aspects

The clinical manifestations associated with the production of ACTH and other POMC-derived peptides as well as CRF by non-pituitary tumours are important to recognize because they can contribute significantly to the morbidity and mortality of patients suffering from such tumours. It is known that, due to the rapid progression of certain cancers associated with the ectopic Cushing's syndrome, such as small cell carcinoma of the lung, some patients do not develop the phenotypic characteristics of Cushing's syndrome, but rather the metabolic abnormalities associated with cortisol and ACTH excess: metabolic alkalosis, hypokalaemia, hypertension, carbohydrate intolerance, muscle weakness, and hyperpigmentation (Urbanic and George, 1981). These metabolic derangements can be subtle and thus may be overlooked. Hence, the ectopic Cushing's syndrome may not always be recognized and its real incidence may not be appreciated. The diagnosis of Cushing's syndrome is sometimes difficult to make, especially when the symptomatology is minimal. The stress of cancer as well as the often traumatic hospital experience may lead to elevations of plasma ACTH as well as to abnormal cortisol dynamics. Urinary free

HORMONE PRODUCTION BY TUMOURS

229

cortisol measurements are seldom affected by stress and hence should be used to document the presence of hypercortisolism (Crapo, 1979). High-dose dexamethasone testing classically demonstrates lack of suppression of the levels of urinary 17-hydroxycorticosteroids, and metyrapone administration leads to no increase in these levels. In some patients suffering from carcinoid tumours and the ectopic Cushing's syndrome, steroid dynamics may be indistinguishable from those seen in pituitary Cushing's syndrome. In such patients, as well as in some cases where the diagnosis is equivocal, selective venous catheterization may be used to demonstrate ACTH or CRF production by the tumour. Treatment of the ectopic Cushing's syndrome is theoretically by treatment of the underlying tumour. This is often an unrealistic goal. In some malignancies such as small cell carcinoma of the lung, Cushing's syndrome tends to occur in the aggressive phase of the disease and may not recede even in the face of tumour response to chemotherapy (Abeloff et al, 1981). In such patients, inhibitors of cortisol production such as aminogluthetemide and metyrapone should be used to alleviate symptoms. VASOPRESSIN AND THE SYNDROME OF INAPPROPRIATE ANTIDIURESIS In 1957, Schwartz et al reported two patients with bronchogenic carcinoma who had marked hyponatraemia with persistent urinary sodium loss as well as impaired urinary dilution (Schwartz et al, 1957). They postulated that these metabolic abnormalities were due to excessive antidiuretic hormone (ADH) production. They suggested that the tumour could be inducing an inappropriate secretion of ADH either through impingement upon an intrathoracic structure such as the vagus nerve, or by invasion of a brain area involved in the regulation of ADH secretion. In the early 1960s, Amatruda et al detected by bioassay large amounts of antidiuretic activity in a tumour tissue derived from a patient with bronchogenic carcinoma and hyponatraemia, thus raising the possibility of ADH synthesis by the tumour (Amatruda et ai, 1963). Several investigators further characterized the material extracted from tumours of patients with inappropriate antidiuresis and found it to be similar to ADH biochemically and immunologically (Lipscomb et al, 1968; Vorherr et al, 1968; George et al, 1972). Biological aspects

ADH or vasopressin is synthesized in the hypothalamus as a composite precursor, provasopressin, which consists of the hormone vasopressin, its carrier protein neurophysin, and a glycoprotein. In 1977, a cell culture line of anaplastic small cell lung carcinoma producing vasopressin but not neurophysin was described (Pettengill et al, 1977). This suggested that the biosynthetic sequence of vasopressin in tumours could be different from that in the hypothalamus. However, more recently, the incorporation of

230

A . DE BUSTROS AND S. B. BA YLIN

labelled compounds into cells from a transplantable small cell carcinoma of the lung with ectopic ADH production was studied (Yamaji et al, 1981). A classic pulse chase experiment revealed that neurophysin is synthesized by post-translational processing from a glycosylated 20000 dalton protein , shown previously to be a common precursor to neurophysin and vasopressin . Thus, the biosynthetic pathway for ADH in this tumour is similar to that in the hypothalamus. Recently, the structural organization of the bovine vasopressin gene was reported (Land et al , 1982), and a single base deletion was shown to be the cause of diabetes insipidus in Brattleboro rats (Schmale and Richter, 1984). Hopefully, similar progress will soon be made in the understanding, at the molecular level , of the syndrome of inappropriate antidiuresis in tumours. Clinical aspects

Inappropriate secretion of ADH has been reported in association with a variety of tumours, including thymomas, gastrointestinal and genitourinary tumours but particularly with lung and central nervous tissue neoplasms (Zerbe et ai, 1980). The frequency of this syndrome in patients with bronchogenic carcinoma of the lung is estimated to be 1.6% (Azzopardi et al , 1970). The most common cell type of lung carcinoma associated with ADH production is small (oat) cell. The true frequency of ADH production by tumours may have been underestimated , since ADH production is not always accompanied by metabolic abnormalities. Indeed, patients with lung tumours without hyponatraemia have been shown to have higher plasma levels of ADH than normal subjects (Padfield et al, 1976). However, all patients with the syndrome of inappropriate antidiuresis do not have high levels of ADH. In fact. as reported by Zerbe et al (1980) , 80% of such patients have ADH levels comparable to healthy adults but these levels are clearly inappropriate in relation to the hypotonicity of the plasma. In their classic review, these authors classified the osmoregulatory defects seen in patients with inappropriate antidiuresis by measuring plasma ADH levels in response to an osmotic challenge. They suggested that. in addition to the secretion of ADH or ADHreleasing factors, tumours can act by resetting the osmostat and causing inappropriate ADH release from the neurohypophysis. Recently, the vasopressin-neurophysin precursor, propressophysin, has been detected in the plasma of patients with small cell carcinoma of the lung but not in the plasma of patients with inappropriate antidiuresis due to central nervous system disease, suggesting that propressophysin may be a marker for ectopic vasopressin production (Yamaji et al, 1984) . The diagnosis of inappropriate anti diuresis should be entertained in the presence of hyponatraemia (Na <130 mfiq/l), serum hypo-osmolarity «275 mOsm/kg) with hypertonic urine , and elevated urinary sodium (>20 mEq/I) . ADH radioimmunoassays have now improved. They should be used when available , and always be correlated with serum osmolarity. A saline

HORMONE PRODUCTION BY TUMOURS

231

loading test is a helpful adjunct to diagnosis, and helps to define subgroups of patients with different osmoregulatory defects (i.e. reset osmostat versus erratic, non-suppressible ADH production) (Zerbe et al , 1980). Inappropriate antidiuresis and water intoxication can sometimes be alleviated by treatment of the tumour by surgery, radiotherapy or chemotherapy. However, this is rarely possible and water restriction is usually necessary. Correction of hyponatraemia is slow and difficult to maintain chronically. Intravenous saline has to be given for patients with severe hyponatraemia and life-threatening neurological complications. Demeclocycline, a drug that counteracts the effects of ADH at the tubular level, in doses of 600 mg to 1200 mg daily, appears to be the safest and most effective treatment of chronic hyponatraemia (Cherrill et al , 1975; Forrest et al, 1978). TUMOUR-RELATED HYPERCALCAEMIA Hypercalcaemia is probably the most common endocrine manifestation of malignancy. The frequency of tumour-related hypercalcaemia is probably similar to that of primary hyperparathyroidism in the general population. However, in hospitalized patients, hypercalcaemia is most commonly found to be secondary to cancer (Fisken et al, 1981). Hypercalcaemia in malignancies mayor may not be associated with skeletal metastasis. Some tumours, such as breast cancer, are thought to produce hypercalcaemia by direct resorption of bone by tumour cells or by osteolytic factors released locally either by tumour cells or by activated immune cells (Eilon and Mundy, 1978). By contrast, some tumours, typically squamous carcinoma of the lung and renal cell carcinoma, are believed to cause hypercalcaemia by releasing bone-resorbing humoral substances (Bockman, 1980; Mundy and Martin, 1982). The nature of these factors is controversial. Some of the recent developments in this area of research have been reviewed extensively elsewhere (Mundy et al, 1984) and are summarized in the following section. Biological basis of tumour-related hypercalcaemia Fuller Albright (Case Records of the Massachusetts General Hospital, 1941), in discussing a patient with renal cell carcinoma and hypercalcaemia, first suggested that non-parathyroid tumours could cause hypercalcaemia by secreting parathyroid hormone (PTH). Subsequently, immunoreactive PTH was indeed identified in tumour extracts and sera of patients with hypercalcaemia (Sherwood et al, 1967; Roof et al, 1971). Further studies, however, yielded conflicting data due to (1) the heterogeneous populations of circulating PTH fragments; (2) the poor correlation between PTH levels measured by different assays; and (3) the assumption by some investigators that detectable PTH immunoreactivity in malignancies with hypercalcaemia was definitive evidence for 'ectopic' PTH secretion (Benson et al , 1974; Raisz et al, 1979). Recently, using

232

A. DE BUSTROS AND S. B . BA YLIN

cloned PTH DNA in a DNA-RNA hybridization assay, Simpson et al failed to detect PTH mRNA in a series of human tumours, some derived from patients with hypercalcaemia (Simpson et ai, 1983). This study provides convincing evidence for the lack of synthesis of PTH by these tumours . However, a more comprehensive survey of tumours by such hybridization analysis is needed before ruling out completely the contribution of PTH to the hypercalcaemia of malignancy .

PTH-like substances Biochemical evaluation of patients with cancer and hypercalcaemia by Stewart et al revealed a distinct group of patients who had elevated nephrogenous cyclic AMP excretion, and absent serum PTH immunoreactivity (Stewart et aI, 1980). Moreover, these patients had lower plasma levels of 1,25-dihydroxy-vitamin D and higher fasting calcium excretion than patients with primary hyperparathyroidism. These findings led to the suggestion that tumours produce factors that act like PTH in certain respects, but fail to react with PTH antisera. Recently, such tumours have been shown to exhibit in vitro adenylate cyclase stimulating activity due to specific binding to renal receptors (Stewart et al, 1983; Strewler et al, 1983). This suggests that tumours do indeed produce substances that mimic the action of PTH on the kidney . These substances could account for the majority of tumour hypercalcaemia originally thought to be secondary to PTH. There may be enough similarity in immunogenicity between these factors and PTH to account for some of the positive PTH radioimmunoassay data reported in some series. Characterization of such substances and the clarification of their relationship to hypercalcaemia will require further investigation. Prostaglandins (PGs) Prostaglandins (PGs) have been implicated in the pathogenesis of tumour-related hypercalcaemia, and data derived from well-studied animal models have provided solid support for the role of PGs in the pathogenesis of hypercalcaemia in malignancy (Tashjian et al, 1972; Tashjian, 1975). In an extensive survey of patients with cancer, Seyberth et al found higher levels of urinary PG metabolites in hypercalcaemic than normocalcaemic patients (Seyberth et aI, 1975). In this study, as well as in isolated reports of patients in the literature, levels of PGs and calcium have been shown to decrease simultaneously in response to indomethacin administration (Brereton et ai, 1974; Seyberth et al, 1975). However, further clinical experience with this drug has proven to be disappointing. Hence, it seems that PGs mediate hypercalcaemia only in a small group of patients with cancer.

Transforming growth factors A new clue to the aetiology of hypercalcaemia in cancers has come from the realization that transforming growth factors , a family of polypeptides

233

HORMONE PRODUCTION BY TUMOURS

produced by tumours that can cause normal cells to assume a malignant phenotype, have bone-resorbing activity (see review by Mundy et ai, 1984). For example, extracts of a transplantable rat Leydig cell tumour, which is known to elicit hypercalcaemia in the absence of bone metastasis, are shown to contain a bone-resorbing factor that copurifies with a tumour-derived transforming growth factor (Ibbotson et al, 1983). These exciting data will certainly stimulate further research directed towards the identification of such bone-resorbing growth factors in human tumours.

Immune cell products Osteoclast-activating factor (OAF), released in vitro by myeloma cells and able to stimulate osteoclastic bone resorption, is similar to a family of proteins (Iymphokines) secreted by normal or activated lymphoid cells (Mundy et ai, 1974). Similarly, interleukin I, a monocyte product, has been shown to exhibit bone-resorbing activity (Gowen et ai, 1983). In the newly recognized syndrome of leukaemic T-cell lymphoma, Iymphokines are produced in vitro by T cells transformed by the T-cell leukaemialymphoma virus (Salahuddin et al , 1984), and are thought to account for the very high incidence of hypercalcaemia (Grossman et al, 1981).

Vitamin D The role of vitamin D in tumour-related hypercalcaemia is not clearly established. Recently, Breslau et al found elevated plasma levels of 1,25-dihydroxy-vitamin D in three patients with lymphoma (Breslau et ai, 1984). These high levels occurred in the face of renal impairment and PTH suppression, and fell in response to glucocorticoid therapy-

Conclusion It is thus clear that several pathophysiological pathways (Table 2) can cause hypercalcaemia in malignancy. Although these pathways have been arbitrarily categorized into (1) local, due to invasion of bone by tumour cells, and (2) humoral, due to release by tumours of osteolytic factors, the distinction between these two categories is not always easy to make. Bone metastasis may not always be demonstrable and, in certain cases, more than one mechanism may be at play. For example, in certain haernatologiTable 2. Mediators of humoral hypercalcaemia. Tumours with bone metastasis

Breast, lung, pancreas

Direct or local osteolytic factors

Tumours without bone metastasis

Squamous lung cancer, renal cell carcinoma, pancreas, ovary

PTH? Prostaglandins? PTH-like factors, transforming growth factors

Haematological malignancies Multiple myeloma, lymphomas

Lymphokines (OAF), 1.25-dihydroxy-vitamin D

234

A. DE BUSTROS AND S. B. BA YLIN

cal malignancies, tumour cells actually invade bone, but are also shown to release bone-resorbing factors such as OAF locally. What mechanisms are operating may be intimately linked to the biology of the underlying tumour. The appreciation that tumour-derived growth factors and interleukins have bone-resorbing activity makes it likely that hypercalcaemia is tightly linked to the neoplastic process and osteolysis may actually playa role in the establishment and progression of some forms of cancer. Diagnosis and treatment In most cases of hypercalcaemia and malignancy, the tumours are clinically obvious. Problems in diagnosis may arise when PTH levels are found to be elevated, thus suggesting the presence of coincident primary hyperparathyroidism. In general, however, PTH levels tend to be distinctly higher for a given calcium level in patients with primary hyperparathyroidism. Also, coexistent primary hyperparathyroidism should be seriously considered if the hypercalcaemia is long-standing or if the patient has kidney stones or subperiosteal bone resorption. If the diagnosis of primary hyperparathyroidism is made, then neck exploration, in certain patients, may significantly diminish the morbidity due to hypercalcaemia in a patient with cancer. Guidelines for the treatment of hypercalcaemia are presented in Table 3. A detailed approach to this problem is found in two excellent reviews (Mundy et al, 1983; Stewart, 1983). Initial treatment of patients with severe hypercalcaemia is by intravenous hydration with saline. Addition of furosemide may lead to a more vigorous diuresis and enhance calcium clearance. Such a regimen will lower serum calcium within a few hours. Attention should be directed, however, to prompt electrolyte replacement. Other agents administered parenterally, such as mithramycin and calcitonin, may lower serum calcium within one day. Mithramycin, a cytotoxic antibiotic that inhibits bone resorption, is almost invariably effective. However, its potential renal, hepatic and haematological toxicity, as well as its mode of administration, limit its use to emergency situations and to hospitalized patients. Calcitonin, also an inhibitor of bone resorption, is occasionally effective. However, its hypocalcaemic effect is

Table 3. Treatment of hypercalcaemia. Acute

Normal saline Furosemide Mithramycin Calcitonin

llq.4-6h 40-80 mg q. 4-6 h 15-25 ug/kg intravenously 100--200 MRC subcutaneously q.12h

Chronic

Prednisone Oral phosphates Indomethacin Diphosphonates

60-80 mg daily 1-3 gdaily 75-150 mg daily Investigational

HORMONE PRODUCTION BY TUMOURS

235

transient. Some investigators have used glucocorticoids in combination with calcitonin, and have found that the hypocalcaemic effect of this drug combination is prolonged (Binstock and Mundy, 1980). The long-term treatment of hypercalcaemia of malignancy is frustrating. So far, no single pharmacological agent has been found to be completely effective. Glucocorticoids are widely used but are not uniformly effective. Prostaglandin synthetase inhibitors such as indomethacin are similarly rarely effective. Oral phosphates are used successfully provided renal function is normal. Their usefulness is limited by their side-effects (diarrhoea) and the potential risk of extraskeletal calcifications. Diphosphonates and, in particular, the newer agents, such as aminohydroxypropane diphosphonate (APO), that do not inhibit bone mineralization in contrast to the older agents, are promising and deserve further investigation, alone or in combination with other agents such as oral phosphates (Mundy et al , 1983).

GROWTH HORMONE EXCESS AND ACROMEGALY Well-documented instances of acromegaly have occurred in patients with carcinoid tumours. and this complex may have several explanations. A carcinoid tumour and a growth hormone (GH)-producing pituitary tumour could be part of a multiple endocrine neoplasia syndrome (MEN I). Alternatively, the carcinoid tumour could elaborate GH. However, GH immunoreactivity has rarely been demonstrated in the tumour extracts from patients with carcinoid tumours (Leveston et ai, 1981). On the other hand, GH has been found, in the absence of acromegaly, in cancers of the lung, stomach, ovary and breast (Beck and Burger, 1972; Kaganowicz et al , 1979). Interestingly, in some patients with a carcinoid tumour, acromegaly and an enlarged pituitary, the pituitary enlargement has been shown to regress along with the acromegalic features following resection of the carcinoid tumour (Scheithauer et al , 1984). This series of events suggests that carcinoid tumours can produce GH-releasing substances. Indeed, GH-releasing factor (GRF) has been purified from carcinoid and pancreatic islet cell tumours (Frohman et al , 1980). Recent studies suggest that GRF is detectable by radioimmunoassay in a large number of small cell carcinomas of the lung, carcinoid and pancreatic islet cell tumours, as well as in a substantial number of phaeochromocytomas, neuroblastic tumours and medullary thyroid carcinomas (Abe et al, 1984; Frohman, 1984). The structure of human ectopic GRF has been determined from pancreatic tumours removed from acromegalic individuals (Guillemin et al , 1982; Rivier et al, 1982). Cloned DNA complementary to GRF mRNA is now available (Gubler et al, 1983; Mayo et al, 1983), and currently being used in RNA-DNA hybridization analysis to study the expression of the GRF gene by normal tissues and by tumours. Human hypothalamic GRF has not yet been identified but evidence points to the fact that it may be similar if not identical to ectopic GRF.

236

A . DE BUSTROS AND S. B. BA YLIN

PROLACTIN EXCESS AND GALACTORRHOEA

In 1971, Turkington described a patient with undifferentiated bronchogenic carcinoma and another patient with hypernephroma , who had ele vated serum levels of prolactin (Turkington , 1971). Prolactin levels declined significantly following irradiation of the lung cancer and surgical excision of the hypernephroma . Also , when maintained in culture, the hypernephroma cells secreted prolactin actively into the culture medium, thus providing evidence for prolactin production by this tumour. Subsequently , investigators examined the frequency of prolactin production by tumours , and reported an elevated incidence of hyperprolactinaemia in certain tumours (33 % of bronchogenic carcinomas) (D avis et al , 1979) . However, when known mechanisms leading to hypcrprolactinaemia (drugs, chest wall irritation, etc.) have been excluded , 'ectopic' prolactin production seems to be a rare event. In the largest series reported by Molitch et al , only two out of 215 patients with various malignancies had clearly elevated prolactin levels (Molitch et al, 1981) . By contrast, when malignant cell lines are examined, the frequency of prolactin production is 25% (Rosen et al, 1980). Interestingly, in such cell lines, prolactin is not detected in the culture medium, suggesting that the cells' enzymatic machinery is unable to process the hormone to the mature form for secretion into the medium. TUMOUR HYPOGLYCAEMIA

Hypoglycaemia has been described in association with non-islet cell tumours such as hepatomas and fibrosarcomas as early as the late 1920s (Elliott, 1929; Doege, 1930) . Subsequently, it was reported in conjunction with a large variety of tumours . mostly mesenchymal tumours, adrenocortical carcinomas and liver tumours (see reviews by Unger, 1966, and Kahn , 1980). Until now, the pathogenesis of this metabolic abnormality in tumours remains poorly understood. Biological aspects

Several mechanisms have been proposed to explain the aetiology of tumour hypoglycaemia (Unger, 1966; Kahn, 1980). Excessive glucose utilization by tumours and failure of compensatory hyperglycaemic mechanisms in patients with malignancies are two of the well-accepted non-humoral aetiologies of tumour hypoglycaemia. Insulin production by non-islet cell tumours has mostly been detected by bioa ssays and definite proof of true insulin production by these tumours has not been obtained, as later studies failed to confirm those early observations (Skrabanek and Powell , 1978) . By contrast, factors with insulin-like biological activity have been strongly implicated in the pathogenesis of hypoglycaemia in tumours (Megyesi et al , 1974; Hyodo et al , 1977; Plovnick et al, 1979) . These

HORMONE PRODUCTION BY TUMOURS

237

factors are referred to as non-suppressible insulin-like activity (NSILA) bec ause they do not cross-react with insulin antibodies in insulin radioimmunoassays . These factors are believed to belong to a heterogeneous family of compounds that includes insulin-like growth factors I and II (IGF I and IGF II). The development of specific radioimmunoassays and radioreceptor assays for these substances has helped to some extent to clarify their role in tumour hypoglycaemia . However, the ultimate proof for production of such substances by tumours will be obtained by the demonstration of specific mRNAs for these substances in tumour extracts. The recent characterization of the IGF I and IGF II genes (Bell et ai , 1984; Brissenden et ai, 1984; Dull et ai, 1984; Tricoli et ai , 1984) suggests that these studies will be performed within a very short period of time. Clinical aspects Tumours associated with hypoglycaemia are usually very large and present as mass lesions. They are usually beyond resection and thus palliative treatment is all that can be offered to patients with tumour hypoglycaemia . An adequate caloric intake and constant glucose supplementation are often all that is needed. Glucagon and glucocorticoids are sometimes of transient benefit. ECTOPIC PRODUCTION OF HUMAN CHORIONIC GONADOTROPHIN (hCG) BY NEOPLASMS Sexual precocity in children and gynaecomastia in adult males have long been known to occur in association with neoplasms , especially liver and lung tumours (Reeves et ai , 1959; Fusco and Rosen, 1966; McArthur et ai , 1973) . The source of the excessive gonadotrophic activity was traced to the tumour tissue as early as 1959 (Reeves et ai , 1959). However , the methods used then and during the following decade were unable to differentiate between pituitary and placental gonadotrophins . The development of specific radioimmunoassays for the various human glycoprotein trophic hormones (TSH, FSH , LH and hCG) has led to the appreciation that hCG is almost exclusively the source of the enhanced gonadotrophic activity that accompanies certain non-trophoblastic tumours (Vaitukaitis et al, 1972; Braunstein et al, 1973). Furthermore, the isolated or unbalanced production of the hCG-specific l3-subunit and especially of the a-subunit, thought to be common to all human glycoprotein hormones, was soon shown to be prevalent in patients with various forms of malignancies (Weintraub and Rosen, 1973; Rosen and Weintraub, 1974; Rosen et al , 1975; Kourides and Schorr-Toshav, 1981) . Biological aspects Recent data about the biosynthesis of hCG by tumours has provided further insight into relationships between gene expression and neoplasia.

238

A. DE BUSTROS AND S. B. BAYLIN

Several cell lines derived from human tumours have been shown to produce hCG and/or one of its subunits in culture (Tashjian et ai, 1973; Lieblich et ai, 1976). The HeLa cell line, derived from a carcinoma of the cervix, is a well-studied example of such tumours. Sodium butyrate induces synthesis of hCG and the a-subunit in this tumour cell line but represses it in a trophoblastic cell line derived from a choriocarcinoma (Chou et ai, 1977). This and other studies by Hussa et al (1978) would suggest that the regulation of hCG production is different in ectopic versus eutopic sources ofthis hormone. However, when the copy number and the structure of the a-subunit gene, including restriction enzyme site polymorphisms, were analysed in these two types of tumours, no differences were noted (Boothby et ai, 1981). Thus, ectopic production of the a-subunit does not seem to be the result of DNA structural rearrangements of the a-subunit gene. More recently, further analysis of the a-subunit gene has suggested the presence of a link between expression of this gene and tumorigenicity. Restriction enzyme patterns were compared in normal placenta, hydatidiform moles and choriocarcinomas. An uncommon DNA polymorphism pattern, homozygosity for the absence of an EcoRI site and the presence of a HindIII site, predominated in choriocarcinoma (Hoshina et ai, 1984). These data raise the possibility that a particular DNA rearrangement at the a-subunit locus predisposes to malignancy. Also, when He La cells are fused with normal fibroblasts, expression of the hCG a-subunit in the hybrids correlates specifically with the tumorigenicity of these hybrids (Stanbridge et ai, 1982). One wonders whether the hCG a-subunit is involved in cell growth regulation or whether a certain DNA rearrangement at the hCG a-subunit locus involves closely linked genes that are more directly implicated in the malignant process, similar to that seen with the myc oncogene/immunoglobulin loci (Dalla-Favera et ai, 1982; Taub et ai, 1982). Recently, the messenger RNAs encoding the a- and 13-subunits of human choriogonadotrophin have been identified (Daniels-McQueen et ai, 1978) and the hCG 13-subunit gene cluster was localized to chromosome 19 while the a-subunit gene was found to be located on chromosome 6 (Naylor et ai, 1983). Further characterization of the genomic organization of these genes and their linkage patterns will undoubtedly provide more insight into the problem of ectopic hCG production. Clinical aspects

The frequency and types of tumours associated with elevated serum hCG levels were assessed in a survey of a large number of patients with various types of malignancies, using a specific hCG radioimmunoassay (Braunstein et ai, 1973). As expected, the incidence of elevated serum hCG levels was very high in trophoblastic tumours (100% of choriocarcinomas, 56% of testicular embryonal carcinomas, 37.5% of seminomas). By contrast, only in 11% of the non-trophoblastic tumours were serum levels of hCG

HORMONE PROD U CTI ON BY TUMOURS

239

detectable by radioimmunoassay. These tumours include pancreatic neoplasms , gastric carcinomas, hepatomas, breast carcinomas , melanomas and bronchogenic carcinomas. In a study of several patients with pancreatic islet-cell tumours, Kahn et al found that up to two-thirds of patients with functioning malignant tumours had elevated serum levels of hCG or one of its subunits, most commonly the a-subunit (Kahn et ai, 1977). By contrast, none of the patients with benign tumours had detectable levels of hCG or one of its subunits in their serum. These data led to the suggestion that hCG and its subunits could be used as specific markers for malignant islet-cell tumours. Patients with islet-cell cancers and carcinoids have the highest incidence of elevated serum a-subunit. Elevated serum a-subunit is also found in a small percentage of patients with lung cancers (5%) and patients with non-islet cell gastrointestinal tract malignancies (4%) (Kourides and Schorr-Toshav, 1981) . However, in a large survey of patients with a variety of benign and malignant disorders, Braunstein et al concluded that a-subunit was not a useful marker for cancer screening or for monitoring the course of patients with the vast majority of cancers (Braunstein et al, 1979). One has also to exercise caution in interpreting the significance of the ' hCG -like material' that is often detected in tumour tissues. Yoshimoto et al (1979) identified hCG immunoreactivity in extracts of all normal human tissues and all human cancers studied . This hCG molecule was less glycosylated than placental hCG. Since glycosylation has been shown to prolong the half-life of hCG, these authors postulated that those cancers associated with detectable hCG in blood must produce carbohydrate-rich hCG . In conclusion, although the production of a placental product by non-trophoblastic tumours raises interesting questions in tumour biology, the practical implications of such a phenomenon for tumour diagnosis appear to be limited . Only rare patients develop signs and symptoms of excessive heG production by tumours. However, tumours associated with precocious puberty in children and gynaecomastia in adult males are usually far advanced and respond poorly to treatment. Hence, there is usually little place for therapy directed at the endocrine manifestations of hCG exce ss. CALCITONIN Calcitonin is normally produced by the C-cells of the thyroid gland. Its levels are greatly increased in the serum of patients with medullary thyroid carcinoma (MTC). However, it is now apparent that calcitonin is not an exclusive marker of MTC and that elevated calcitonin levels can be seen in other neoplasms (Coombes et al, 1974; Milhaud et al , 1974; Schwartz et ai, 1979). The reported incidence of calcitonin production by tumours is variable. In one study , up to 84% of patients with extensive small cell carcinomas (SCC) of the lung had increased circulating levels of calcitonin immunoreactivity (Wallach et al , 1981). In some series, a significant

240

A . DE BUSTROS AND S. B. BAYLIN

number of tumours, such as carcinomas of the breast (38%), pancreas (42%) , stomach (30%) and colon (24%), have been found to contain calcitonin immunoreactivity (Schwartz et ai , 1979). Biological aspects

The recognition of calcitonin production by tumours, especially MTC, has led to exciting observations relating to the biosynthesis of this peptide and to interesting relationships between the biology of the tumours involved and calcitonin production. As are other small bioactive peptides, calcitonin is originally synthesized in the form of a large polypeptide precursor. In contrast to small molecular weight forms of CT found in MTC, the predominant immunoreactive calcitonin forms detected in lung tumours are large calcitonin moieties, presumably precursor forms (Becker et ai, 1978). Not all of the high molecular weight fractions are well recognized by some calcitonin antibodies used in radioimmunoassays. Failure to detect calcitonin immunoreactivity in certain tumours may thus be due to the heterogeneity of the calcitonin species produced. Conversely, spurious elevations of plasma calcitonin can occur, apparently due to artefacts affecting the radioimmunoassay. For example, Roos et al reported that heating the plasma, presumably destroying protease activity, led to normalization of the previously elevated calcitonin levels in plasma from patients with squamous and anaplastic lung carcinomas but not small cell or adenocarcinomas (Roos et aI, 1980). Caution thus has be exercised in the interpretation of plasma immunoreactive calcitonin levels. Roos's observations also suggest that only certain types of lung cancers (small cell and adenocarcinomas) produce calcitonin. Indeed, calcitonin is sometimes thought to be exclusively produced by tumours derived from cells with endocrine features . With respect to relationships between tumour types, cr production, and other features of endocrine cell differentiation, data from our own laboratory suggest that true differences in the frequency of the various markers of endocrine activity among the various histological types of lung tumours exist, but that these differences are quantitative and not qualitative ones. Thus , endocrine tumour-related markers such as L-dopa decarboxylase, histaminase and l3-endorphin are generally highest in SCC tumour tissues. The distribution of calcitonin is more uniform. Calcitonin was extracted from 54% of sec, 50% of adenocarcinomas, 55% of large cell undifferentiated tumours, and 33% of squamous cell carcinomas (Berger et al, 1981). The simultaneous appearance of high levels of two or more endocrine markers favoured SCC in this study. Thus, we believe that endocrine-related properties occur throughout the spectrum of lung cancer and reflect a continuum of 'A PU D ' differentiation within a common cell lineage (Baylin and Mendelsohn, 1980; Baylin and Mendelsohn , 1982). The recent availability of the cloned calcitonin gene has allowed the study of calcitonin production by tumours at the mRNA level. Also, characterization of the calcitonin gene structure in rat and more recently in

HORMONE PRODUCTION BY TUMOURS

241

man has provided tremendous insight into the regulation of the expression of this gene in various tissues. The rat calcitonin gene comprises several exons or coding domains (Rosenfeld et aI, 1983). By alternative splicing of the primary RNA transcript from this gene, two distinct mRNA molecules are produced. One mRNA encodes calcitonin and a second mRNA encodes a peptide referred to as calcitonin-gene-related peptide (CGRP). The expression of these peptides is tissue-specific. Thus, calcitonin is expressed in the thyroid while CGRP appears to predominate in the central nervous system. There is recent evidence that the human calcitonin gene is similarly organized and that alternative processing of this gene can occur in human cells (Nelkin et al, 1984; Steenbergh et aI, 1984b). In our laboratory, we have detected calcitonin and CGRP mRNA in several human lung tumour cell lines, including small cell, large cell, adenocarcinomas and squamous carcinomas (Nelkin et aI, 1984). The ratio of CGRP to calcitonin mRNA seems to be higher in most of these cell lines than in a human MTC cell line. This suggests differential processing of the calcitonin gene in these tissues. Moreover, we also find that the sizes of those mRNA species are larger in lung carcinomas than in MTC. These initial findings concerning CT mRNA in non-thyroidal tissue and tumours may prove helpful in the understanding of ectopic hormone production in general. They also suggest that CGRP could be used as an additional marker for lung cancer. In our laboratory, we are currently interested in the chemical manipulation of the various lung carcinoma cell lines available to us, as well as of the human MTC cell culture line (de Bustros et aI, 1985), in order to see whether the regulation of the calcitonin gene expression differs in 'ectopic' versus 'eutopic' sources of calcitonin. The relationships of gene expression to tumour differentiation and progression could thus be examined, as well as the possible role of calcitonin and CGRP in the regulation of tumour growth. Clinical aspects Calcitonin production by non-MTC tumours tends to be asymptomatic. The levels of calcitonin are usually not high enough in the blood to cause flushing, diarrhoea or any of the symptoms that are often associated with hypercalcitoninaemia in MTC. Calcitonin levels have been used to monitor tumour burden and disease activity in small cell carcinoma of the lung (Silva et aI, 1979; Wallach et aI, 1981; Cate et al, 1984). Although calcitonin is occasionally useful as a tumour marker, one has to be careful about the fact, as we have discussed previously, that neoplasia is a dynamic process and that events in tumour progression may lead to a change in the status of differentiation of tumour cells, such that endocrine activity is no longer present in these cells (Baylin et ai, 1978). It is interesting to note that, even in MTC, plasma calcitonin is not always a useful tumour marker. Patients with disseminated disease do not always have an increase in plasma calcitonin in proportion to their tumour mass (Trump et ai, 1979). In fact, analysis of tumour tissues from such

242

A . DE BUSTROS AND S. B. BAYLIN

patients reveals a significant cellular heterogeneity for calcitonin by immunohistochemistry, as well as a lower total tissue calcitonin content than in patients with indolent disease (Baylin and Mendelsohn, 1982; Lippman et aI, 1982; Saad et aI, 1984). It is tempting to speculate that selective RNA splicing occurs during tumour progression such that calcitonin expression is decreased while CGRP expression is increased. The availability of calcitonin and CGRPspecific cloned DNA should allow the investigation of such a possibility, both in MTC and in lung tumours , during the various stages of tumour progression . NEUROENDOCRINE PEPTIDES In the past two decades, a great number of small peptides that function as neuromodulators in the brain and/or hormones in the circulation has been discovered. These peptides are found in a variety of tissues. Thus, the term 'ectopic' is particularly inappropriate when referring to the production by tumours of these peptides. The advent of recombinant DNA techniques has allowed the characterization of the genes coding for these peptides. Sequencing these genes has led to the determination of the amino acid sequence of the corresponding proteins and to the realization that a similar biosynthetic pathway exists for these peptides. Thus, ACTH, ADH, somatostatin , gluc agon , gastrin , vasointestinal peptide (VIP) , calcitonin, etc. are all derived from large precursor molecules (Douglass et ai, 1984). These precursors can be the source of more than one peptide and thus generate a tremendous diversity of biological activity. Tissue-specific processing of these precursors or of their corresponding genes appears to be yet another source of complexity in the biosynthesis of these peptides. For example, processing of the proopiomelanocortin molecule is different in the anterior and neurointermediate lobes of the pituitary (Eipper and Mains, 1980). Also, the calcitonin/CGRP gene is differentially expressed in the thyroid and in the hypothalamus (Rosenfeld et ai, 1983). In tumours, preliminary observations from our laboratory and others (see discussion of calcitonin and ectopic Cushing's syndrome) indicate that the transcriptional and translational processing of peptides in tumours can be different from these processes in normal tissues. Specific peptides The production of some of these peptides such as calcitonin , ACTH, ADH, GHRH , CRF, has been mentioned earlier in this review, in specific sections. Immunoreactivity for several other peptides , such as glucagon, somatostatin, enkephalin , pancreatic polypeptide (PP) , etc. , has been detected in a large number of tumours, mostly carcinoids (Alumets et aI, 1981; Sporrong et ai , 1982; Yang et aI, 1983) . Gastrin-releasing peptide (GRP), which is closely related to the amphibian bombesin molecule, is

HORMONE PRODUCTION BY TUMOURS

243

currently generating interest as a tumour marker for certain tumours, including lung cancers (Moody et ai, 1981; Erisman et aI, 1982; Abe et aI, 1984; Matsubayashi et al, 1984). Elevated vasointestinal peptide (VIP) levels have been demonstrated in plasma and tumour extracts from patients with the watery-diarrhoea syndrome and a variety of tumours, including bronchogenic carcinomas and phaeochromocytomas (Said and Faloona, 1975; Said, 1976). Thus, the presence of the watery-diarrhoea syndrome should always lead to a careful search for pancreatic as well as non-pancreatic tumours. OTHER RARE MANIFESTATIONS OF NEOPLASIA Erythrocytosis

Erythrocytosis has been reported in association with cerebellar haemangiomas, liver tumours and hypernephromas, and has been attributed to the production of erythropoietin or erythropoietin-stimulating substances by these tumours (Hammond and Winnick, 1974). These substances have been mostly measured by bioassays, because of the unavailability, until recently, of purified erythropoietin suitable for radiolabelling and use in a radioimmunoassay (Miyake et al, 1977; Sherwood and Goldwasser, 1979). However, so far, the availability of purified erythropoietin is restricted, and a better understanding of the paraneoplastic syndrome of erythrocytosis will have to await cloning of the erythropoietin gene and the study of its expression by tumours. Hypophosphataemia Profound hypophosphataemia has been described with mesenchymal tumours (Salassa et al, 1970; Drezner and Feinglos, 1977). It is believed to be due to the production by these tumours of a substance that promotes phosphate loss in the kidney and possibly impairs the renal lahydroxylation of 25-hydroxy-vitamin D. Muscle weakness and osteomalacia are the predominant manifestations of this entity and respond partially to phosphorus and 1,25-dihydroxy-vitamin D supplementation. Surgical resection of the tumour, which can be very small, often leads to a complete cure. Hyperthyroidism

Hyperthyroidism can occasionally occur with hydatidiform moles and choriocarcinomas. It is believed that the hyperthyroid state in these patients is due to the TSH-like activity of the hCG molecule produced by these tumours (Nisula and Ketelslegers, 1974). Characteristically, these patients have goitres, increased radioactive iodine uptake in the neck, but no ophthalmopathy or any other of the stigmata of Graves' disease. Recently, immunoreactive TRH (thyrotrophin-releasing hormone) has

244

A . DE BUSTROS AND S. B. BA YLiN

been identified in human neoplasms (Wilber and Spinella, 1984). However, the occurrence of thyrotoxicosis with such tumours is unlikely because very slight elevations in serum thyroid hormones can completely inhibit pituitary TSH responses to TRH. CONCLUSION In this review , we have tried to summarize some of the recent biological and clinical developments in the field of hormone production by neoplasms. While it is clear that no unifying concept can explain all the facts about ectopic hormone production presented in this review, the perspective with which the problem is now approached reflects the major recent advances in molecular and cellular biology. It seems impossible at present to separate the problem of hormone production by neoplasms from the broader perspective of mechanisms underlying the processes of gene expression, regulation of normal cell differentiation, and cellular transformation. From the information currently at hand , we suggest the following overall conclusions concerning the relationships between tumours and the production of hormones. Investigators working in this field are now asking questions about the molecular basis of aberrant hormone production by neoplasms. Increasingly sophisticated techniques of peptide and nucleic acid analysis, plus the use of complex cell culture systems, have provided characterization of hormones at the peptide level, and led to the delineation of the multiple steps involved in the biosynthesis of hormones in normal endocrine cells as well as in tumour cells . The entire structure of numerous hormones, including those present in minute amounts in various tissues, has been determined. The application of recombinant DNA techniques to the study of peptide gene expression has led to the discovery of peptides with , as yet, no known function . It will be exciting to define the actions of such peptides and their role in mediating specific cellular responses, especially in tumour cells. The understanding of events in the regulation of hormone gene expression in tumours as well as in normal tissues is now being actively pursued, with emphasis on DNA rearrangements , restriction enzyme site polymorphisms, methylation patterns, as well as the identification of DNA sequences acting as promoters or enhancers and of regulatory events for transcription and translation. Manipulation of cells in culture is also used to examine the process of gene expression by tumours. For example, deletion of specific DNA sequences from a given gene followed by transfer of such a gene into defined cells in culture is likely to provide , in the near future , clues about the specific nucleotide sequences involved in transcriptional control , as well as those regulating proteolytic processing of hormones in different tissues . More emphasis is being placed upon the potential functional role of hormone production by neoplastic cells. Neoplasia is classically thought of as a disorder of cell growth and cell-to-cell communication . Several lines of

HORMONE PRODUCTION BY TUMOURS

245

evidence suggest that the various hormones produced by tumours may be involved in the aberrant growth of cancer cells. For example, some of the growth factors produced by tumours are homologous to some of the recently recognized oncogenes . Thus, platelet-derived growth factor (PDGF), a potent mitogen for mesenchymal cells , seems to be identical to the product of the oncogene, V-sis (Doolittle et al, 1983; Waterfield et al, 1983). Also, the receptor for epidermal growth factor (EGF) is structurally related to the transforming protein encoded by another oncogene V-erb-B (Downward et ai, 1984) . The growth factor IGF II gene has now been localized to chromosome 11, next to the proto-oncogene c-Ha-ras 1, while the gene for IGF 1 has been mapped to chromosome 12, next to c-Ha-ras 2, implying a functional relationship between genes from the ras family and genes from the insulin family, in malignant transformation (Brissenden et ai, 1984; Tricoli et al, 1984). Blalock and Smith (1980) have demonstrated antigenic and structural similarities between ACTH, endorphins and interferon, and have suggested that human leucocyte interferon may be a precursor to ACTH and endorphins. The production by the immune system of neuroendocrine hormones may represent a mechanism by which signals are conveyed to key endocrine organs, such as the adrenal glands, during infection or neoplasia . The high frequency with which ACTH is produced in tumours would also suggest that the gene coding for ACTH is linked to one or more genes intimately involved in the establishment and progression of the malignant process, such as oncogenes. One can thus imagine a situation where the expression of combinations of genes could be simultaneously increased in a tumour by a common promoter or enhancer. Combinations of hormones with potential for interaction and a capability for response to multiple peptides also exist in tumours. Classic studies have demonstrated the existence of 'aberrant' receptors in certain endocrine tumours. For example , FSH receptors have been found in adrenocortical carcinomas (Schorr et al, 1972). Moreover , the adenyl cyclase system of such tumours responds well to stimulation by FSH. Such findings indicate that tumour cells possess regulatory mechanisms different from those of normal cells . The presence of certain functional pairs of hormones within the same tumour, such as ACTH and CRF, LH and LHRH (Upton and Amatruda, 1971; Suda et ai, 1977; Hashimoto et ai, 1980; Wahlstrom and Seppala, 1981), also suggests the existence of an autonomous regulatory system within these cells. Further dissection of the association between peptide hormone elaboration and tumours could lead to improved therapeutic modalities for cancer. We have stressed in this review that the expression of peptide hormone genes may be intimately linked to the neoplastic process itself in several ways. First, such gene expression may mark cellular events in early steps of neoplastic transformation. If so, identification of regulatory mechanisms for expression of peptide hormone genes could elucidate new target points for manipulating cancer cells . Second , we have outlined relationships between the status of differentiation for cell populations in common neoplasms such as lung cancer and the degree of endocrine differentiation. It is now a much sought-after goal to use agents which alter the

246

A . DE BUSTROS AND S. B. BA YLiN

differentiation status of cells to treat different forms of tumours. Endocrine biochemistry could be a potential monitor for the effects of such drugs. Finally, we have discussed the potential autocrine and paracrine roles that peptide hormones could play in tumours. Altering the levels of such hormones, through changing the differentiation status of the cells and/or directly blocking hormonal effects with antibodies to the hormones themselves or their receptors, could potentially influence neoplastic cell growth. It seems likely that the next years of research into the mechanisms underlying 'aberrant' hormone production by tumours will include a focus on these possibilities for cancer therapy .

REFERENCES Abe K, Nicholson WE, Liddle GW, Island DP & Orth DN (1967) Radioimmunoassay of fl-MSH in human plasma and tissues. Journal of Clinical Investigation 46: 1609-1616. Abe K, Adachi I, Miyakawa S et al (1977) Production of calcitonin, adrenocorticotropic hormone and beta-melanocyte-stimulating hormone in tumors derived from amine precursor uptake and decarboxylation cells . Cancer Research 37: 4190-4194. Abe K , Yamaguchi K & Adachi I (1984) Production of gastrin-releasing peptide (bombesin) by tumors. Abstracts, 7th International Congress of Endocrinology, Quebec City, Canada, S], p 17. Abeloff MD , Eggleston lC, Mendelsohn et al (1979) Changes in morphologic and biochemical characteristics of small cell carcinoma of the lung-a clinicopathologic study. American Journal of Medicine 66: 757-764. Abeloff MD, Trump, DL & Baylin SB (1981) Ectopic adrenocorticotrophic (ACTH) synd ro me and small cell carcinoma of the lung-assessment of clinical implications in patients on combination chemotherapy. Cancer 48: 1082-1087. Alumets J , Aim P , Falkmer S et al (1981) Immunohistochemical evidence of peptide hormones in endocrine tumors of the rectum. Cancer 48: 2409-2415 . Amatruda IT Jr , Mulrow Pf , Gallagher JC & Sawyer WH (1963) Carcinoma of the lung with inappropriate antidiuresis. Demonstration of antidiuretic-hormone-like activity in tumor extracts. New England Journal of Medicine 269: 544-549. Azzopardi JG & Williams ED (1968) Pathology of ' nonendocr ine' tumors associated with Cushing's synd ro me . Cancer 22: 274-286 . Azzopardi JG, Freeman E & Poole G (1970) Endocrine and metabolic disorders in bronchial carcinoma. British Medical Journal Iv: 528-529. Baylin SB & Mendelsohn G (1980) Ectopic (inappropriate) hormone production by tumors: mechanisms involved and the biological and clinical implications. Endocrine Reviews 1: 45-77. Baylin SB & Mendelsohn G (1982) Medullary thyroid carcinoma: a model for the study of tumor progression and cell heterogeneity. In Owens AH Jr, Co ffey DS, & Baylin SB (eds) Tumor Cell Heterogeneity-Origins and Implications , pp 9-27. New York: Academic Press. Bayl in SB, Weisburger WR, Eggleston JC et al (1978) Variable content of histaminase, L-dopa decarboxylase and calcitonin in small cell carcinoma of the lung. New England Journal of Medicine 299: 105-110. Baylin SB , Abeloff MD, Goodwin G , Carney DN & Gazdar AF (1980) Activities of L-do pa decarboxylase and diamine oxida se (h istaminase) in human lung can cers and decarboxylase as a marker for small (oat) cell cancer in cell culture. Cancer Research 40: 1990-1994. Beck C & Burger HG (1972) Evidence for the presence of immunoreactive growth hormone in cancers of the lung and stomach . Cancer 30: 75-79 . Becker KL , Snider RH , Silva OL & Moore CF (1978) Calcitonin heterogeneity in lung cancer and medullary thyroid cancer. A cta Endocrinologica 89: 89-99 .

HORMONE PRODUCTION BY TUMOURS

247

Bell GI, Pictet RL, Rutter WJ et al (1980) Sequence of the human insulin gene. Nature 284: 26-32. Bell GI, Santerre RF & Mullenbach GT (1983) Hamster preproglucagon contains the sequence of glucagon and two related peptides. Nature 302: 716-718. Bell GI, Merryweather JP, Sanchez-Pescador et al (1984) Sequence of a cDNA clone encoding human preproinsulin-like growth factor II. Nature 319: 775-777. Benson RC Jr, Riggs BL, Pickard BM & Arnaud CD (1974) Radioimmunoassay of parathyroid hormone in hypercalcemic patients with malignant disease. American Journal of Medicine, 56: 821-826. Berger CL, Goodwin G, Mendelsohn Get al (1981) Endocrine-related biochemistry in the spectrum of human lung carcinoma. Journal of Endocrinology and Metabolism 53: 422-429. Berger CL, deBustros A, Roos BA et al (1984) Human medullary thyroid carcinoma in culture provides a model relating growth dynamics, endocrine cell differentiation and tumor progression. Journal of Clinical Endocrinology and Metabolism 59: 338-343. Bertagna XY, Nicholson WE, Sorenson GD et al (1978) Corticotropin, lipotropin and I:\-endorphin production by a human non-pituitary tumor in culture: evidence for a common precursor. Proceedings of the National Academy of Sciences, USA 75: 5160-5164. Binstock ML & Mundy GR (1980) Effects of calcitonin and glucocorticoids in combination on the hypercalcemia of malignancy. Annals of Internal Medicine 93: 269-272. Blackman MR, Rosen SW & Weintraub BD (1978) Ectopic hormones. Advances in Internal Medicine 23: 85-113. Blalock JE & Smith EM (1980) Human leukocyte interferon: structural and biological relatedness to adrenocorticotropic hormones and endorphins. Proceedings of the National Academy of Sciences, USA 77: 5972-5974. Bloomfield GA, Scott AP, Lowry PJ, Gilkes JJH & Rees LH (1974) A reappraisal of human I:\-MSH. Nature 252: 492-493. Bockman RS (1980) Hypercalcaemia in malignancy. Clinics in Endocrinology and Metabolism 9: 317-333. Boothby M, Ruddon RW, Anderson C, McWilliams D & Boime I (1981) A single gonadotropin a-subunit gene in normal tissue and tumor-derived cell lines. Journal of Biological Chemistry 256: 5121-5127. Borkowski A, & Muquardt C (1979) Human chorionic gonadotropin in the plasma of normal, nonpregnant subjects. New England Journal of Medicine 301: 298-302. Braunstein GO, Vaitukaitis JL, Carbone PP & Ross GT (1973) Ectopic production of human chorionic gonadotropin by neoplasms. Annals of Internal Medicine 78: 39-45. Braunstein GD, Rasor JL & Wade ME (1975) Presence in normal human testes of a chorionic-gonadotropin-like substance distinct from human luteinizing hormone. New England Journal of Medicine 293: 1339-1343. Braunstem GD, Forsythe AB, Rasor JL et al (1979) Serum glycoprotein hormone alpha subunit levels in patients with cancer. Cancer 44: 1644-1651. Brereton HD, Halushka PV, Alexander RW et al (1974) Indomethacin-responsive hypercalcemia in a patient with renal-cell carcinoma. New England Journal of Medicine 291: 83-85. Brereton HD, Mathews MM, Costa J, Kent CH & Johnson RE (1978) Mixed anaplastic small-cell and squamous-cell carcinoma of the lung. Annals of Internal Medicine 88: 805-806. Breslau NA, McGuire JL, Zerwekh JE, Frenkel EP & Pak CYC (1984) Hypercalcemia associated with increased serum calcitriollevels in three patients with lymphoma. Annals of Internal Medicine 100: 1-7. Brissenden JE, Ullrich A & Francke U (1984) Human chromosomal mapping of genes for insulin-like growth factors I and II and epidermal growth factor. Nature 319: 781-784. Brown WH (1928) A case of pluriglandular syndrome: 'diabetes of bearded women'. Lancet ii: 1022-1023. Brownstein M, Arimura A, Sato H, Schally AV & Kizer JS (1975) The regional distribution of somatostatin in the rat brain. Endocrinology 96: 1456-1461.

248

A. DE BUSTROS AND S . B. BA YLIN

Carey RM . Varma SK, Drake CR Jr et al (1984) Ectopic secretion of corticotropin-releasing factor as a cause of Cushing's syndrome: a clinical, morphological and biochemical study. New England Journal of Medicine 311: 13-20 . Case Records of the Massachusetts General Hospital (1941) New England Journal of Medicine 225: 789-791. Cate CC, Douple EB, Andrews KM et al (1984) Calcitonin as an indic ator of the response of human small cell carcinoma of the lung cells to drugs and radiation . Cancer Research 44: 949-954. Cherril DA , Stote RM, Birge JR & Singer I (1975) Derneclocycline treatment in the syndrome of inappropriate ant idiuretic hormone secretion. Annals of Internal Medicine 83: 654-656. Chin WW . Kronenberg HM , Dee PC, MaloofF & Habener JF (1981) Nucleotide sequence of the mRN A encoding the pre-a-subunit of mouse thyrotropin . Proceedings ofthe National Academy of Sciences 78: 5329-5333 . Chin WW, Godine JE. Klein DR et al (1983) Nucleotide sequence of the cDNA encoding the precursor of the ~-subunit of rat lutropin. Proceedings of the National Academy of Sciences, USA 80: 4649-4653. Chou JY, Robinson JC & Wang S-S (1977) Effects of sodium butyrate on synthesis of human chorionic gonadotrophin in trophoblastic and non-trophoblastic tumours. Nature 268: 543-544. Chretien M, Seidah NG, Benjannet S et al (1977) A ~-LPH precursor model: recent developments concerning morphine-like substances. Annals of the New York Academy of Sciences 297: 84-107. Christy NP (1961) Adrenocorticotrophic activity in the plasma of patients with Cushing's syndrome associated with pulmonary neoplasms. Lancet i: 85-86. Cooke NE, Coit D . Shine J , Baxter JD & Martial JA (1981) Human prolactin . eDNA structural analysis and evolutionary comparisons. Journal of Biologi cal Chemistry 256: 4007-4016. Coombes RC . Hillyard C, Greenberg PB & Macintyre I (1974) Plasma-immunoreactivecalcitonin in patients with non-thyroid tumours. Lancet i: 1080-1083 . Counis R, Corbani M, Poissonier M & Jutisz M (1982) Characterization of the precursors of a and 13 subunits of follitropin following cell-free translation of rat and ovine pituitary mRNAs. Biochemical and Biophysical Research Communications 107: 998·1005. Crapo L (1979) Cushing's syndrome: a review of diagnostic tests. Metabolism 28: 955-977 . Dalla-Favera R , Bregni M , Erikson J et al (1982) Human c-myc oncogene is located on the region of chromosome 8 that is translocated in Burkitt lymphoma cells. Proceedings of the National Academy of Sciences USA 79: 7824-7827. Daniels-McOueen S. McWilliams D , Birken S et al (1978) Identification of mRNAs encoding the a and 13 subunits of human choriogonadotropin. Journal of Biological Chemistry 253: 7109-7114 . Davis S, Proper S, May PB & Ertel NH (1979) Elevated prolactin levels in bronchogenic carcinoma. Cancer 44: 676-679 . DeBold CR, Schworer ME, Connor TB, Bird RE & Orth DN (1983) Ectopic pro-opiolipomelanocortin: sequence of cDNA coding for l3-melanocyte-stimulating hormone and l3-endorphin. Science 220: 721-723 . de Bustros A, Baylin SB, Berger CL et al (1985) Phorbol esters increase calcitonin gene transcription and decrease c-MYC mRNA levels in cultured human medullary thyroid carcinoma. Journal of Biological Chemistry 260: 98-104. Doege KW (1930) Fibrosarcoma of the mediastinum. Annals of Surgery 92: 955-960. Doolittle RF, Hunkapiller MW, Hood LE et al (1983) Simian sarcoma viru s one gene, v-sis, is derived from the gene (or genes) encoding a platelet-derived growth factor. Science 221: 275-277 . Douglass J . Civelli 0 & Herbert E (1984) . Pol yprotein gene expression : generation of diversity of neuroendocrine peptides. Annual Review of Biochemistry 53: 665-715. Downward J , Yarden Y, Maye s E et al (1984) Close similarity of epidermal growth factor receptor and v-erb -B oncogene protein sequences. Nature 307: 521-527. Drezner MK & Feinglos MN (1977) Osteomalacia due to la,25-d ihydroxycholecalciferol deficiency. Association with a giant tumor of bone . Journal of Clinical Investigation 60: 1046-1053 .

HORMONE PRODUCTION BY TUMOURS

249

Dull TJ, Gray A, Hayflick lS & Ullrich A (1984) Insulin-like growth factor II precursor gene organization in relation to insulin gene family. Nature 310: 777-781. Eilon G & Mundy GR (1978) Direct resorption of bone by human breast cancer cells in vitro. Nature 276: 726-728. Eipper BA & Mains RE (1980) Structure and biosynthesis of pro-adrenocorticotropin/endorphin and related peptides. Endocrine Review I: 1-27. Elliott CA (1929) Hepatogenic hypoglycemia associated with primary liver-cell carcinoma. Transactions of the Association of American Physicians 44: 121-126. Erisman MD, Linnoila RI, Hernandez 0, DiAugustine RP & Lazarus LH (1982) Human lung small-cell carcinoma contains bombesin. Proceedings of the National Academy of Sciences USA 79: 2379-2383. Feinberg AP & Voge1stein B (1983) Hypomethylation distinguishes genes of some human cancers from their normal counterparts. Nature 301: 89-92. Fiddes lC & Goodman HM (1979) Isolation, cloning and sequence analysis of the cDNA for the a-subunit of human chorionic gonadotropin. Nature 281: 351-356. Fiddes lC, Seeburg PH, DeNoto FM et al (1979) Structure of genes for human growth hormone and chorionic somatommamotropin. Proceedings of the National Academy of Sciences USA 76: 4294-4298. Fisher lA, Tobler PH, Kaufmann M et al (1981) Calcitonin: regional distribution of the hormone and its binding sites in the human brain pituitary. Proceedings of the National Academy of Sciences USA 78: 7801-7805. Fisken RA, Heath DA, Somers S & Bold AM (1981) Hypercalcaemia in hospital patients. Clinical and diagnostic aspects. Lancet i: 202-207. Forrest IN. Cox M, Hong C et al (1978) Superiority of demeclocyline over lithium in the treatment of chronic syndrome of inappropriate secretion of antidiuretic hormone. New England Journal of Medicine 298: 173-177. Friedman M, Marshall-Jones P & Ross EJ (1966) Cushing's syndrome: adrenocortical hyperactivity secondary to neoplasms arising outside the pituitary-adrenal system. Quarterly Journal of Medicine 138: 193-214. Frohman LA (1984) Production of growth hormone-releasing factor by tumors. Abstracts, 7th International Congress of Endocrinology, Quebec City, Canada, S2, p 16. Frohman LA, Szabo M, Berelowitz M & Stachura ME (1980) Partial purification and characterization of a peptide with growth hormone-releasing activity from extrapituitary tumors in patients with acromegaly. Journal of Clinical Investigation 65: 43-54. Furutani Y, Morimoto Y, Shibahara S et al (1983) Cloning and sequence analysis of cDNA for ovine corticotropin-releasing factor precursor. Nature 301: 537-540. Fusco FD & Rosen SW (1966) Gonadotropin-producing anaplastic large-cell carcinomas of the lung. New England Journal of Medicine 275: 507-515. George 1M, Capen CC & Phillips AS (1972) Biosynthesis of vasopressin in vitro and ultrastructure of a bronchogenic carcinoma. Patient with the syndrome of inappropriate secretion of antidiuretic hormone. Journal of Clinical Investigation 51: 141-148. Gewirtz G & Yalow RS (1974) Ectopic ACTH production in carcinoma of the lung. Journal of Clinical Investigation 53: 1022-1032. Goodwin G & Baylin SB (1982) Relationships between neuroendocrine differentiation and sensitivity to "'(-radiation in culture line OH-l of human small cell lung carcinoma. Cancer Research 42: 1361-1367. Gowen M, Wood DD, Ihrie EJ, McGuire MKB & Russell RGG (1983) An interleukin-l-like factor stimulates bone resorption in vitro. Nature 306: 378-380. Grossman B, Schechter GP, Horton JE et al (1981) Hypercalcemia associated with T-cell lymphoma leukemia. American Journal of Clinical Pathology 75: 149-155. Gubler U, Monahan JJ, Lomedico PT et al (1983) Cloning and sequence analysis of cDNA for the precursor of human growth hormone-releasing factor, somatocrinin. Proceedings of the National Academy of Sciences USA 80: 4311-4314. Guillemin R, Brazeau P, Bohlen P et al (1982) Growth hormone-releasing factor from a human pancreatic tumor that caused acromegaly. Science 218: 585-587. Gurr JA, CatteralllF & Kourides IA (1983) Cloning of cDNA encoding the pre-B subunit of mouse thyrotropin. Proceedings ofthe National Academy ofSciences USA 80: 2122-2126.

250

A. DE BUSTROS AND S. B. BA YLIN

Habener JF & Segre GY (1979) Parathyroid hormone radioimmunoassay. Annals of Internal Medicine 91: 782-784 . Hammond 0 & Winnick S (1974) Paraneoplastic erythrocytosis and ectopic erythropoietins. Annals of the New York Academy of Sciences 230: 219-227 . Hashimoto K, Takahara J, Ogawa N et al (1980) Adrenocorticotropin, 13-liptropin, l3-endorphin and corticotropin-releasing factor-like activity in an adrenocorticotropin-producing nephroblastoma. Journal of Clinical Endocrinology and Metabolism 50: 461-465 . Hattori M, Imura H . Matsukura S et al (1979) Multiple-hormone producing lung carcinoma. Cancer 43: 2429-2437. Hirata Y. Matsukara S. Imura H et al (1976) Two cases of multiple hormone-producing small cell carcinoma of the lung. Coexistence of tumor ADH, ACTH and I3MSH. Cancer 38: 2575-2582. Holub DA & Katz FH (1961) A possible etiological link between Cushing's syndrome and visceral malignancy. Clinical Research 9: 194. Hoshina M, Boothby MR, Hussa RD et al (1984) Segregation patterns of polymorphic restriction sites of the gene encoding the a subunit of human chorionic gonadotropin in trophoblastic disease. Proceedings of the National Academy of Sciences USA 81: 2504-2507. Hussa RO. Pattillo RA. Ruckert ACF & Scheuermann KW (1978) Effect of butyrate and dibutyryl cyclic AMP on hCG-secreting trophoblastic and non-trophoblastic cells. Journal of Clinical Endocrinology and Metabolism 46: 69-76. Hyodo T. Megyesi K. Kahn CR. McLean JP & Friesen HG (1977) Adrenocortical carcinoma and hypoglycemia: evidence for production of nonsuppressible insulin-like activity by the tumor. Journal of Clinical Endocrinology and Metabolism 44: 1175-1184. Ibbotson KO, D'Souza SM, Ng KW et al (1983) Tumor-derived growth factor increases bone resorption in a tumor associated with humoral hypercalcemia of malignancy. Science 221: 1292-1294. Imura H (1980) Ectopic hormone syndromes. Clinics in Endocrinology and Metabolism 9: 235-260. Ito R, Sato K, Helmer T, Jay G & Agarwal K (1984) Structural analysis of the gene encoding human gastrin : the large intron contains an alu sequence . Proceedings of the National Academy of Sciences USA 81: 4662-4666. Kaganowicz A, Farkouh N, Frantz AG & Blaustein AU (1979) Ectopic human growth hormone in ovaries and breast cancer. Journal of Clinical Endocrinology and Metabolism 48: 5-8. Kahn CR (1980) The riddle of tumour hypoglycaemia revisited. Clinics in Endocrinology and Metabolism 9: 335-360 . Kahn CR, Rosen SW. Weintraub BO, Fajans SS & Gorden P (1977) Ectopic production of chorionic gonadotropin and its subunit by islet-cell tumors: a specific marker for malignancy. New England Journal of Medicine 297: 565-569. Kolata G (1982) New theory of hormones proposed. Science 215: 1383-1384. Kourides IA & Schorr-Toshav NL (1981) Alpha subunit of the glycoprotein hormones: secretion by human malignancies. Clinical Bulletin 11: 106-109. Kyle CY. Evans MC & Odell WD (1981) Growth hormone-like material in normal human tissues. Journal of Clinical Endocrinology and Metabolism 53: 1138-1144. Land H, Schutz G, Schmale H & Richter 0 (1982) Nucleotide sequence of cloned cDNA encoding bovine arginine vasopressin-neurophysin II precursor. Nature 295: 299-303. Levcston SA. McKeel DW Jr, Buckley PJ et al (1981) Acromegaly and Cushing's syndrome associated with a foregut carcinoid tumor. Journal of Clinical Endocrinology and Metabolism 53: 682-689. Li CH , Barnafi 10, Chretien M & Chung 0 (1965) Isolation and amino-acid sequence of I3-LPH from sheep pituitary glands. Nature 208: 1093-1094. Liddle GW. Nicholson WE, Island DP ct al (1969) Clinical and laboratory studies of ectopic humoral syndromes. Recent Progress in Hormonal Research 25: 283-314 . Lieblich JM , Weintraub BD , Rosen SW . Chou J'Y & Robinson JC (1976) Hela cells secrete the a subunit of glycoprotein tropic hormones. Nature 260: 530-532.

HORMONE PRODUCTION BY TUMOURS

251

Lippman SM, Mendelsohn G, Trump DL, Wells SA Jr & Baylin 5B (1982) The prognostic and biological significance of cellular heterogeneity in medullary thyroid carcinoma: a study of calcitonin, L-dopa decarboxylase and histaminase. Journal of Clinical Endocrinology and Metabolism 54: 233-240. Lips CJM, Van der Sluys Veer J, Van der Donk JA, Van Dam RH & Hackeng WHL (1978) Common precursor molecule as origin for the ectopic-hormone-producing-tumour syndrome. Lancet i: 16-18. Lipscomb HS, Wilson C, Retiene K, Matsen F & Ward DN (1968) The syndrome of inappropriate secretion of antidiuretic hormone: a case report and characterization of an antidiuretic hormone-like material isolated from an oat cell carcinoma of the lung. Cancer Research 28: 378-383. Lokich JJ (1982) The frequency and clinical biology of the ectopic hormone syndromes of small cell carcinoma. Cancer 50: 2111-2114. Lowry PJ, Rees LH, Tomlin S, Gillies G & Landon J (1976) Chemical characterization of ectopic ACTH purified from a malignant thymic carcinoid tumor. Journal of Clinical Endocrinology and Metabolism 43: 831-835. Lund PK, Goodman RH, Montminy MR, Dee PC & Habener JF (1983) Anglerfish islet pre-proglucagon II. Nucleotide and corresponding amino acid sequence of the cDNA. Journal of Biological Chemistry 258: 3280-3284. Mains RE, Eipper BA & Ling N (1977) Common precursor to corticotrophins and endorphins. Proceedings of the National Academy of Sciences USA 74: 3014-3018. Matsubayashi S, Yanaihara C, Ohkubo M et al (1984) Gastrin-releasing peptide immunoreactivity in medullary thyroid carcinoma. Cancer 53: 2472-2477. Mayo KE, Vale W, Rivier J, Rosenfeld G & Evans RM (1983) Expression-cloning and sequencing of a cDNA encoding human growth hormone-releasing factor. Nature 306: 86-88. McArthur JW, Toll GD, Russfield AB et al (1973) Sexual precocity attributable to ectopic gonadotropin secretion by hepatoblastoma. American Journal of Medicine 54: 390-403. Meador CK, Liddle GW, Island DP et al (1962) Cause of Cushing's syndrome in patients with tumors arising from 'non-endocrine' tissue. Journal of Clinical Endocrinology and Metabolism 22: 693-703. Megyesi K, Kahn CR, Roth J & Gorden P (1974) Hypoglycemia in association with extrapancreatic tumors: demonstration of elevated plasma NSILA-S by a new radioreceptor assay. Journal of Clinical Endocrinology and Metabolism 38: 931-934. Milhaud G, Calmette C, Taboulet J, Julienne A & Moukhtar M5 (1974) Hypersecretion of calcitonin in neoplastic conditions. Lancet i: 462-463. Miyake T, Kung C-KH & Goldwasser E (1977) Purification of human erythropoietin. Journal of Biological Chemistry 252: 5558-5564. Molitch ME, Schwartz S & Mukherji B (1981) Is prolactin secreted ectopically? American Journal of Medicine 70: 803-807. Moody TW, Pert CB. Gazdar AF, Carney DN & Minna JD (1981) High levels of intracellular bombesin characterize human small-cell lung carcinoma. Science 214: 1246-1248. Moore H-PH, Walker MD, Lee F & Kelly RB (1983) Expressing a human proinsulin cDNA in a mouse ACTH-secreting cell. Intracellular storage, proteolytic processing, and secretion on stimulation. Cell 35: 531-538. Mundy GR & Martin TJ (1982) The hypercalcemia of malignancy: pathogenesis and management. Metabolism 31: 1247-1277. Mundy GR, Raisz LG, Cooper RA, Schechter GP & Salmon SE (1974) Evidence for the secretion of an osteoclast-stimulating factor in myeloma. New England Journal of Medicine 291: 1041-1046. Mundy GR, Wilkinson R & Heath DA (1983) Comparative study of available medical therapy for hypercalcemia of malignancy. American Journal of Medicine 74: 421-432. Mundy GR, Ibbotson KJ, D'Souza SM et al (1984) The hypercalcemia of cancer. Clinical implications and pathogenic mechanisms. New England Journal of Medicine 310: 1718-1727. Nakanishi S, Inoue A, Kita T et al (1979) Nucleotide sequence of cloned cDNA for bovine corticotropin-Bdiptropin precursor. Nature 278: 423-427.

252

A. DE BUSTROS AND S. B. BA YLIN

Naylor SL, Chin WW, Goodman HM et al (1983) Chromosome assignment of genes encoding the a and I) subunits of glycoprotein hormones in man and mouse. Somatic Cell Genetics 9: 757-770. Nelkin BD, Rosenfeld KI, de Bustros A et al (1984) Structure and expression of a gene encoding human calcitonin and calcitonin gene related peptide. Biochemical and Biophysical Research Communications 123: 648-655. Nisula BC & Ketelslegers J-M (1974) Thyroid-stimulating activity and chorionic gonadotropin. Journal of Clinical Investigation 54: 494-499. Odell WD & Wolfsen AR (1982) Humoral syndromes associated with cancer: ectopic hormone production. Progress in Clinical Cancer 8: 57-74. Odell WD, Wolfsen AR, Bachelot I & Hirose FM (1979) Ectopic production of Iipotropin by cancer. American Journal of Medicine 66: 631-638. Orth DN, Nicholson WE, Mitchell WM, Island DP & Liddle GW (1973) Biological and immunological characterization and physical separation of ACTH and ACTH fragments in the ectopic ACTH syndrome. Journal of Clinical Investigation 52: 1756-1769. Orth DN, Guillemin R, Ling N & Nicholson WE (1978) Immunoreactive endorphins, lipotropins, and corticotropins in a human nonpituitary tumor: evidence for a common precursor. Journal of Clinical Endocrinology and Metabolism 46: 849-852. Padfield PL, Morton JJ, Brown JJ et al (1976) Plasma arginine vasopressin in the syndrome of antidiuretic hormone excess associated with bronchogenic carcinoma. American Journal of Medicine 61: 825-831. Pearse AGE (1980) The APUD concept and hormone production. Clinics in Endocrinology and Metabolism 9: 211-222. Pettengill OS, Faulkner CS, Wurster-Hill DH et al (1977) Isolation and characterization of a hormone-producing cell line from human small cell anaplastic carcinoma of the lung. Journal of the National Cancer Institute 58: 511-518. Pintar JE, Schachter BS, Herman AB, Durgerian S & Krieger DT (1984) Characterization and localization of proopiomelanocortin messenger RNA in the adult rat testis. Science 225: 632-634. Plovnick H, Ruderman NB, Aoki T, Chideckel EW & Poffenbarger PL (1979) Non-f-eel! tumor hypoglycemia associated with increased nonsuppressible insulin-like protein (NSILP). American Journal of Medicine 66: 154-159. Policastro P, Ovitt CE, Hoshina M et al (1983) The f3 subunit of human chorionic gonadotropin is encoded by multiple genes. Journal of Biological Chemistry 258: 11492-11499. Raisz LG, Yajnik CH, Bockman RS & Bower BF (1979) Comparison of commercially available parathyroid hormone immunoassays in the differential diagnosis of hypercalcemia due to primary hyperparathyroidism or malignancy. Annals of Internal Medicine 91: 739-740. Ratcliffe JG, Knight RA, Besser GM, Landon J & Stansfield AG (1972) Tumor and plasma ACTH concentrations in patients with and without the ectopic ACTH syndrome. Clinical Endocrinology I: 27-44. Rees LH (1975) The biosynthesis of hormones by non-endocrine tumours-a review. Journfl of Endocrinology 67: 143-175. ' Reeves RL, Tesluk H & Harrison CE (1959) Precocious puberty associated with hepatoma. Journal of Clinical Endocrinology and Metabolism 19: 1651-1660. Richter D (1983) Synthesis, processing, and gene structure of vasopressin and oxytocin. Progress in Nucleic Acid Research and Molecular Biology 30: 245-266. Rivier J, Spiess J, Thorner M & Vale W (1982) Characterization of a growth-hormone releasing factor from a human pancreatic islet tumor. Nature 300: 276-278. Rivier J, Spiess J & Vale W (1983) Characterization of rat hypothalamic corticotropinreleasing factor. Proceedings of the National Academy of Sciences USA 80: 4851-4855. Roof BS, Carpenter B, Fink DJ & Gordan GS (1971) Some thoughts on the nature of ectopic parathyroid hormones. American Journal of Medicine 50: 686-691. Roos BA, Lindall AW, Baylin SB et al (1980) Plasma immunoreactive calcitonin in lung cancer. Journal of Clinical Endocrinology and Metabolism 50: 659-666.

HORMONE PRODUCTION BY TUMOURS

253

Rosen SW & Weintraub BD (1974) Ectopic production of the isolated alpha subunit of the glycoprotein hormones: a quantitative marker in certain cases of cancer. New England Journal of Medicine 290: 1441-1447. Rosen SW, Weintraub BD, Vaitukaitis JL et al (1975) Placental proteins and their subunits as tumor markers. Annals in Internal Medicine 82: 71-83. Rosen SW, Weintraub BD & Aaronson SW (1980) Nonrandom ectopic protein production by malignant cells: direct evidence in vitro. Journal of Clinical Endocrinology and Metabolism 50: 834-841. Rosenfeld MG, Mermod J-J, Amara SG et al (1983) Production of a novel neuropeptide encoded by the calcitonin gene via tissue-specific RNA processing. Nature 304: 129-135. Rosenzweig JL, Havrankova J, Lesniak MA, Brownstein M & Roth J (1980) Insulin is ubiquitous in extra pancreatic tissues of rats and humans. Proceedings of the National Academy of Sciences USA 77: 572-576. Saad MP, Ordonez NG, Guido JJ & Samaan NA (1984) The prognostic value of calcitonin immunostaining in medullary carcinoma of the thyroid. Journal of Clinical Endocrinology and Metabolism 59: 850-856. Said SI (1976) Evidence for secretion of vasoactive intestinal peptide by tumours of pancreas, adrenal medulla, thyroid and lung: support for the unifying APUD concept. Clinical Endocrinology 5: 201S-204S. Said S & Faloona GR (1975) Elevated plasma and tissue levels of vasoactive intestinal polypeptide in the watery-diarrhea syndrome due to pancreatic, bronchogenic and other tumors. New England Journal of Medicine 293: 155-160. Said S1 & Rosenberg RN (1976) Vasoactive intestinal polypeptide: abundant immunoreactivity in neural cell lines and normal nervous tissue. Science 192: 907-908. Saito E, Iwasa S & Odell WD (1983) Widespread presence of large molecular weight adrenocorticotropin-like substances in normal rat extrapituitary tissues. Endocrinology 113: 1010-1019. Salahuddin SZ, Markham PD, Lindner SG et a1 (1984) Lymphokine production by cultured human T cells transformed by human T-cell leukemia-lymphoma virus-I. Science 223: 703-707. Salassa RM, Jowsey J & Arnaud CD (1970) Hypophosphatemic osteomalacia associated with 'nonendocrine tumors. New England Journal of Medicine 283: 65-70. Schmale H & Richter D (1984) Single base deletion in the vasop.ressin ~ene is the cause of diabetes insipidus in Brattleboro rats. Nature 308: 705-709. Schorr I, Hinshaw TH, Cooper MA, Mahaffee D & Ney RL (1972) Adenyl cyclase hormone responses of certain human endocrine tumors. Journal of Clinical Endocrinology and Metabolism 34: 447-451. Scheithauer BW, Carpenter PC, Bloch B & Brazeau P (1984) Ectopic secretion of a growth hormone-releasing factor. Report of a case of acromegaly with bronchial carcinoid tumor. American Journal of Medicine 76: 605-616. Schteingart DE, Conn JW, Orth DN, Harrison TS, Fox JE & Bookstein JJ (1972) Secretion of ACTH and ~MSH by an adrenal medullary paraganglioma. Journal of Clinical Endocrinology and Metabolism 34: 676-683. Schwartz WB, Bennett W, Curelop S & Bartter FC (1957) A syndrome of renal sodium loss and hyponatremia probably resulting from inappropriate secretion of antidiuretic hormone. American Journal of Medicine 23: 529-542. Schwartz KE, Wolfsen AR, Forster B & Odell WD (1979) Calcitonin in nonthyroidal cancer. Journal of Clinical Endocrinology and Metabolism 49: 438-444. See burg PH (1982) The human growth hormone gene family: nucleotide sequences show recent divergence and predict a new polypeptide hormone. DNA 1: 239-249. Seyberth HW, Segre GV, Morgan JL et al (1975) Prostaglandins as mediators of hypercalcemia associated with certain types of cancer. New England Journal of Medicine 293: 1278-1283. Shapiro M, Nicholson WE, Orth DN, Mitchell WM & Liddle GW (1971) Differences between ectopic MSH and pituitary MSH. Journal of Clinical Endocrinology and Metabolism 33: 377-381. Shen L-P, Pictet RL & Rutter WJ (1982) Human somatostatin I: sequence of the cDNA. Proceedings of the National Academy of Sciences USA 79: 4575-4579.

254

A . DE BUSTROS AND S. B. BA YLIN

Sherwood JB & Goldwasser E (1979) A radioimmunoassay for erythropoietin. Blood 54: 885-893. Sherwood LM, O'Riordan JLH , Aurbach GD & Potts JT Jr (1967) Production of parathyroid hormone by nonparathyroid tumors. Journal of Clinical Endocrinology and Metabolism 27: 140-146. Shibahara S, Morimoto Y, Furutani Y et al (1983) Isolation and sequence-analysis of the human corticotropin-releasing factor precursor gene . EMBO Journal 2: 775-779. Shields R (1977) Gene depression in tumours. Nature 269: 752-753. Silva OL, Becker KL , Primack A , Dopprnan JL & Snider RH (1976) Increased serum calcitonin levels in bronchogenic cancer. Chest 69: 495-499 . Silva OL, Broder LE, Doppman JL et al (1979) Calcitonin as a marker for bronchogenic cancer. Cancer 44: 680-684 . Simpson EL, Mundy GR, D 'Souza SM et al (1983) Absence of parathyroid hormone messenger RNA in non parathyroid tumors associated with hypercalcemia. New England Journal of Medicine 309: 325-330. Skrabanek P & Powell D (1978) Ectopic insulin and Occam's razor: re appraisal of the riddle of tumour hypoglycaemia. Clinical Endocrinology 9: 141-154. Spindel ER, Chin WW, Price J et al (1984) Cloning and characterization of cDNAs encoding human gastrin-relating peptide. Proceedings of the National Academy of Sciences USA 81: 5699-5703. Sporn MB & Todaro GJ (1980) Autocrine secretion and malignant transformation of cells. New England Journal of Medicine 303: 878-880. Sporrong B, Falkmer S, Robboy S et al (1982) Neurohormonal peptides in ovarian carcinoids. An immunohistochemical study of 81 primary carcinoids and of intraovarian metastasis from six mid-gut carcinoids. Cancer 49: 68-74. Stan bridge EJ , Rosen SW & Sussman HH (1982) Expression of the a subunit of human chorionic gonadotropin is specifically correlated with tumorigenic expression in human cell hybrids. Proceedings of the National Academy of Sciences USA 79: 6242-6245 . Steenbergh PH, H6ppener JWM , Zandberg Jet al (1984a) Expression of the proopiomclanocortin gene in human medullary thyroid carcinoma . Journal of Clinical Endocrinology and Metabolism 58: 904-908 . Steenbergh PH, Hoppener JWM, Zandberg Jet al (1984b) Calcitonin gene related peptide coding sequence is conserved in the human genome and is expressed in medullary thyroid carcinoma. Journal of Endocrinology and Metabolism 59: 358-360 . Stevens RE & Moore GE (1983) Inadequacy of APUD concept in explaining production of peptide hormones by tumours. Lancet i: 118-119. Stewart AF (1983) Therapy of malignancy-associated hypercalcemia : 1983. American Journal of Medicine : 74: 475-480. Stewart AF, Horst R, Deftos U et al (1980) Biochemical evaluation of patients with cancer-associated hypercalcemia . Evidence for humoral and nonhumoral groups. New England Journal of Medicine 303: 1377-1383. Stewart AF, Insogna KL, Goltzman D & Broadus AE (1983) Identification of adenylate cyclase-simulating activity and cytochemical glucose-6-phosphate dehydrogenasesimulating activity in extracts of tumors from patients with humoral hypercalcemia of malignancy. Proceedings of the National Academy of Sciences USA 80: 1454-1458. Strewlcr GJ, Williams RD & Nissenson RA (1983) Human renal carcinoma cells produce hypercalcemia in the nude mouse and a novel protein recognized by parathyroid hormone receptors. Journal of Clinical Investigation 71: 769-774 . Sud a T, Demura H, Demura R et al (1977) Corticotropin-releasing factor-like activity in ACTH producing tumors . Journal of Clinical Endocrinology and Metabolism 44: 440-446. Sud a T, Tomori N, Tozawa F et al (1984) Immunoreactive corticotropin and corticotropinreleasing factor in human hypothalamus, adrenal , lung cancer and pheochromocytoma . Journal of Clinical Endocrinology and Metabolism 58: 919-924 . Takahashi H, Teranishi Y, Nakanishi S & Numa S (1981) Isolation and structural organization of the human corticotropin-fjlipotropin precursor gene . FEBS Letters 135: 97-102 . Tarnai S, Kameya T, Yamaguchi K et al (1983) Peripheral lung carcinoid tumor producing predominantly gastrin-releasing peptide (GRP) . Cancer 52: 273-281.

HORMONE PRODUCTION BY TUMOURS

255

Tashjian AH Jr (1975) Prostaglandins, hypercalcemia and cancer. New England Journal of Medicine 293: 1317-1318. Tashjian AH Jr, Voelkel EF, Levine L & Goldhaber P (1972) Evidence that the bone resorption stimulating factor produced by mouse fibrosarcoma cells is prostaglandin E 2 : a new model for the hypercalcemia of cancer. Journal of Experimental Medicine 136: 1329-1343. Tashjian AH Jr, Weintraub BD, Barowsky NJ, Rabson AS & Rosen SW (1973) Subunits of human chorionic gonadotropin: unbalanced synthesis and secretion by clonal cell strains derived from a bronchogenic carcinoma. Proceedings of the National Academy of Sciences USA 70: 1419-1422. Tatemoto K & Mutt V (1981) Isolation and characterization of the intestinal peptide porcine PHI (PHI-27), a new member of the glucagon-secretin family. Proceedings of the National Academy of Sciences USA 78: 6603-6607. Taub R, Kirsch I, Morton C et al (1982) Translocation of the c-myc gene into the immunoglobulin heavy chain locus in human Burkitt lymphoma and murine plasmacytoma cells. Proceedings of the National Academy of Sciences USA 79: 7837-7841. Tricoli JV, Rail LB, Scott J, Bell GI & Shows TB (1984) Localization of insulin-like growth factor genes to human chromosomes 11 and 12. Nature 310: 784-786. Trump DL, Mendelsohn G & Baylin SB (1979) Discordance between plasma calcitonin and tumor-cell mass in medullary thyroid carcinoma. New England Journal of Medicine JOI: 253-255. Tsukada T, Nakai Y, Jingami H et al (1981) Identification of the mRNA coding for the ACTH-f3-lipotropin precursor in a human ectopic ACTH-producing tumor. Biochemical and Biophysical Research Communications 98: 535-540. Turkington RW (1971) Ectopic production of prolactin. New England Journal of Medicine 285: 1455-1458. Unger RH (1966) The riddle of tumor hypoglycemia. American Journal of Medicine 40: 325-330. Upton GV & Amatruda IT (1971) Evidence for the presence of tumor peptides with corticotropin-releasing factor-like activity in the ectopic ACTH syndrome. New England Journal of Medicine 285: 419-424. Urbanic RC & George JM (1981) Cushing's disease-18 years' experience. Medicine 60: 14-35. Vaitukaitis JL, Braunstein GD & Ross GT (1972) A radioimmunoassay which specifically measures human chorionic gonadotropin in the presence of human luteinizing hormone. American Journal of Obstetrics and Gynecology 113: 751-758. Vale W, Spiess J, Rivier C & Rivier J (1981) Characterization of a 41-residue ovine hypothalamic peptide that stimulates secretion of corticotropin and f3-endorphin. Science 213: 1394-1397. Vasicek TJ, McDevitt BE, Freeman MW et al (1983) Nucleotide sequence of the human parathyroid hormone gene. Proceedings of the National Academy of Sciences USA 80: 2127-2131. Vorherr H, Massry SG, Utiger RD & Kleeman CR (1968) Antidiuretic principle in malignant tumor extracts from patients with inappropriate ADH syndrome. Journal of Endocrinology and Metabolism 28: 162-168. Vuitch MF & Mendelsohn G (1981) Relationship of ectopic ACTH production to tumor differentiation: a morphologic and immunohistochemical study of prostatic carcinoma with Cushing's syndrome. Cancer 47: 296-299. Wahlstrom TC & Seppala M (1981) Immunological evidence for the occurrence of luteinizing hormone-releasing factor and the a subunit of glycoprotein hormones in carcinoid tumors. Journal of Clinical Endocrinology and Metabolism 53: 209-212. Wallach SR, Royston I, Taetle R, Wohl H & Deftos LJ (1981) Plasma calcitonin as a marker of disease activity in patients with small cell carcinoma of the lung. Journal of Clinical Endocrinology and Metabolism 53: 602-606. Waterfield MD, Scrace GT, Whittle N et al (1983) Platelet-derived growth factor is structurally related to the putative transforming protein p 28'" of simian sarcoma virus. Nature 304: 35-39. Weintraub BD & Rosen SW (1973) Ectopic production of the isolated beta subunit of human chorionic gonadotropin. Journal of Clinical Investigation 52: 3135-3142.

256

A. DE BUSTROS AND S. B. BA YLIN

Whitfeld PL, Seeburgh PH & Shine J (1982) The human pro-opiomelanocortin gene: organization, sequence and interspersion with repetitive DNA . DNA 1: 133-143. Wiborg 0, Berglund L, Boel et al (1984) Structure of a human gastrin gene. Proceedings of the National Academy of Sciences USA 81: 1067-1069. Wilber JF & Spinella P (1984) Identification of immunoreactive thyrotropin-releasing hormone in human neoplasia . Journal of Clinical Endocrinology and Metabolism 59: 432-435 . Wolfsen AR & Odell WD (1979) ProACTH: use for early detection of lung cancer . American Journal of Medicine 66: 765-772. Yamaji T, Ishibashi M, Katayama S et al (1981) Neurophysin biosynthesis in vitro in oat cell carcinoma of the lung with ectopic vasopressin production . Journal of Clinical Investigation 68: 1441-1449. Yamaji T, Ishibashi M & Hori T (1984) Propressophysin in human blood : a possible marker of ectopic vasopressin production. Journal of Endocrinology and Metabolism 59: 505·512. Yang K, Ulich T, Taylor I, Cheng L & Lewin KJ (1983) Pulmonary carcinoids . Immunohistochemical demonstration of brain-gut peptides . Cancer 52: 819-823. Yoshimoto Y, Wolfsen AR & Odell WD (1977) Human chorionic gonadotropin-like substance in nonendocrine tissues of normal subjects. Science 197: 575-577. Yoshimoto Y, Wolfsen AR & Odell WD (1979) Glycosylation, a variable in the production of hCG by cancers. American Journal of Medicine 67: 414-420 . Zerbe R, Stropes L & Robertson G (1980) Vasopressin function in the syndrome of inappropriate antidiuresis. Annual Review of Medicine 31: 315-327.