Competitive radioassays and “specific” tumor markers

Competitive

Radioassays and “Specific” Tumor Markers

By Bruce Competitive to detect and

other

endocrine teins,

radioassays ectopic

normally tissue,

have

production

polypeptides neoplasms.

particularly restricted may

serve

by

various

fetal as

used

pro-

synthesis or

is

placental

specific

and Saul W. Rosen markers male

non-

of these

whose

to

been

of hormones

Some

those

D Welntraub

true

when

incidence

specificity lished

detected

or nonpregnant

and

specificity

of these

for

cancer

relate

in the

female.

to

of the assay

serum

However,

markers

and

are

not

the

sensitivity

methods

yet

of a the their estaband

employed.

tumor

OMPEfITIVE

RADIOASSAYS have greatly enhanced the ability of the to diagnose, localize, and treat various tumors. There are numerous examples, considered in other articles of this volume, where these methods have been applied to the study of neoplasms arising from various endocrine glands. In recent years it has also been shown Lhat malignant tumors. including those arising from nonendocrine tissue, can secrete a variety ot hormones and other polypeptides not normally associated with their tissue of polypeptide syndromes.’ ’ Although these synorigin. resulting in “ectopic” dromes were first identified by the biologic effects of high concentrations ol circulating hormone, the development of competitive radioassays has permitted suspicion of neoplastic polypeptide production even before clinical recogniCon of neoplasm 43 In most cases, it has not been possible to diagnose ectopic hormone production in the earliest stages of malignancy because of the hormone’s similarity tv the product of a normal endocrine organ. ’ 3 Even when tumor production is suficlently great to cause an elevation of serum hormone concentration, it musk then be proved that the hormone is secreted from a nonendocrine source. The two best criteria are demonstration of an arteriovenous gradient across the tumor, a5 has been shown for follicle-stimulating hormone’ and parathyroid hormone.’ or demonstration of tumor production of hormone in vitro as has been shown for vasopressin.” growth hormone,’ and prolactin.‘” More often. tumor production has been inferred by demonstration of hormone in tumor by extraction or immunocytologic techniques Such studies do not actuailq prove the site of hormone synthesis, and “trapping’of serum hormones b! tumors has been claimed,” although “trapping” alone would not account for physician

Merabolmn.

Vol 22. No. 8 (August).

1913

li

I9

1120

WEINTRAUB

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persistently elevated serum hormone concentrations. Other criteria for ectopic hormone production, such as decreasing serum hormone concentrations after therapy or absence of normal homeostatic suppressive mechanisms, are less specific. In view of such difficulties in proving ectopic hormone synthesis, a search has been made for unique tumor products whose presence in peripheral.serum would be diagnostic of malignancy. Indeed, it has been shown for ACTH and parathyroid hormone (PTH) that certain antisera used in radioimmunoassay recognize differences between the normal and the ectopic hormone. In the case of ACTH, antisera were developed against the N-terminal as well as the Cterminal portions of the molecule and each was used in radioimmunoassay.‘* It was found that, unlike normal ACTH, the ectopic hormone had greater activity in the C-terminal than in the N-terminal immunoassay. Recently, it has been found that tumors secreting ectopic ACTH also secrete both normal and abnormal melanocyte-stimulating hormones (MSH),13 although it is not clear whether these hormones (or other peptides) account for the excessive reactivity in the C-terminal ACTH immunoassay, In the case of PTH, two groups have reported immunochemical differences between the normal and the ectopic hormone as demonstrated by radioimmunoassy.‘41’5 One of these groupsI demonstrated a variant of ectopic PTH with high immunoreactivity but low bioactivity, which may explain the earlier observation of neoplasms associated with elevated serum PTH but no hypercalcemia.4 The significance of these apparently abnormal ectopic ACTH-MSH and PTH variants is not yet clear and must await further purification and amino acid analyses. There is accumulating evidence for the heterogeneity of “normal” polypeptide hormones in serum,‘6J7 some of which may represent precursor forms’* as well as degraded formsI of the hormone. It is thus possible that the apparent differences between normal and ectopic hormones may be quantitative rather than qualitative, similar to the increased secretion of proinsulin of the explanation, the specificity of by certain islet-cell tumors. *O Regardless competitive radioassays has provided a means to recognize variants of hormones associated with tumors. Much of the recent work related to “specific” tumor markers has focused on fetal polypeptides, such as alpha-fetoprotein and carcinoembryonic antigen, and placental polypeptides, such as chorionic gonadotropin, placental lactogen, and placental alkaline phosphatase. If it can be documented that the synthesis of such proteins is normally restricted to embryonic life, their later detection in a male or nonpregnant female should provide a specific and sensitive marker recent introduction of competitive radiofor neoplasm. 2’ The comparatively assays in this field has provided the methodology to more fully explore the diagnostic potential of these embryonic proteins. About a decade ago, it was shown that alpha-fetoprotein (aFP), the dominant serum protein secreted by the liver early in fetal life, was present in the blood of rodents bearing chemically induced hepatomas.22 Soon after, (uFP was detected in the blood of patients with primary hepatocellular carcinoma,‘3 and blind determinations of (uFP in 308 sera demonstrated this protein in the majority of patients with. primary carcinoma of the liver and in about 250,, of

COMPETITIVE

RADIOASSAYS

AND

“SPECIFIC”

TUMOR

MARKERS

1121

patients with testicular tumor, but in no cases of metastases to liver, a variety of other tumors, or liver diseases other than tumor (cirrhosis, hepatitis, etc.).24 In all these publications, the detection of aFP involved double well immunodiffusion in agar, a method whose sensitivity is probably limited to about 10,000 ng/ml. With the use of quantitative radial immunodiffusion or counterimmunoelectrophoresis, L~FP could be detected down to concentrations of 250 “g/ml, and figures for its “incidence” in sera of patients with primary hepatocellular carcinoma had to be revised upward.25 Furthermore, it could now be shown that cvFP was detectable in many normal second and third trimester pregnancy sera and transiently in serum samples from some patients with acute viral hepatitis.26 Using highly purified aFP, one group has recently developed a specific radioimmunoassay sensitive to 0.25 ng/ml. 27,28These workers have detected (uFP in all normal sera examined, at concentrations ranging from 4410.5 ng/m129~‘” and point out the incomplete suppression in the adult of the fetal gene responsible for its production. Thus, the early claims for production of aFP only by hepatocellular (or certain testicular) carcinomas, already under assault by an enhanced methodology at the 100 ng/ml level, break down completely under the sensitive scrutiny afforded by the methodology of competitive radioassay. What seemed to be a qualitative marker for neoplasm is seen to be a marker however usefulLonly in the quantitative sense. The carcinoembryonic antigen (CEA), a glycoprotein later found to be present in fetal digestive tissues, was first discovered by absorbing antisera to extracts of human colon adenocarcinoma with identical extracts of adjacent normal colon from the same patient.3’ Analysis of extracts from other digestive tract epithelial neoplasms with this same anti-CEA serum revealed that CEA was found generally in GI carcinomas derived from entoderm but not in other carcinomas~-nor in any normal adult tissues, including placenta. In fact. CEA seemed to be present normally only in extracts of fetal gut, liver and pancreas between 2 and 6 mo of gestation.3’ Partial purification of CEA was soon followed by development of a radioimmunoassay capable of detecting 2.5 ng CEA/ml in extracts of serum.33 At first, the serum data seemed to substantiate the tissue results, showing levels of CEA exceeding 2.5 ng/ml only in cases of entodermally derived epithelial GI cancer. More recently, others using the same labeled CEA and anti-CEA have found elevations in serum from some patients with severe alcoholic liver disease and some with severe renal disease. 34Now the original group has reported finding CEA in colitis apparently unaccompanied by carcinoma, as well as in some carcinomas originating outside the GI tract. 35 Neither of these laboratories detected CEA in any of 67 normal sera at the 2.5 ng/ml level of sensitivity. However, CEA has been recently detected in normal colon.36 and after extraction of a large pool, CEA was demonstrated in normal plasma at a concentration of about 1 ng/ml. 37 Although CEA does not meet the criteria for a specific cancer marker, its quantitation in the serum of patients with cola-rectal carcinomas is already a major advance in the early detection and management of these neoplasms. In contrast to the fetal proteins, studies of placental polypeptides produced

1122

WEINTRAUR

AND

ROSEN

by tumors have suggested that they may be specific cancer markers. It should be emphasized, however, that this apparent specificity has not yet been subjected to the scrutiny already applied to the fetoproteins. In particular, too few control sera have been examined for placental proteins, especially from nonneoplastic conditions associated with tissue regeneration, impaired metabolic clearance, and virus infection. The need to document cancer specificity seems especially warranted in light of the recent demonstration of placental lactogen in apparently normal human testis.38 Human chorionic gonadotropin (HCG) has served as a cancer marker in the diagnosis of trophoblastic neoplasms for over a decade.?’ 4’ In most of the early studies, the gonadotropin was measured by a urinary bioassay that did not distinguish among HCG, follicle-stimulating hormone (FSH), luteinizing hormone (LH), and tumor gonadotropin. However, since these neoplasms are often capable of producing gonadotropin in amounts far greater than the pituitary, quantitative bioassay has been useful in tumor diagnosis and in following the response to surgical and pharmacologic therapy. The development of gonadotropin radioimmunoassays 42,43has provided a more sensitive, specific. and convenient diagnostic method for these trophoblastic neoplasms.44 Although the conventional radioimmunoassays do not distinguish HCG from LH, recent immunoassays have made this distinction possible and promise to further enhance diagnostic capabilities. In two laboratories, such assays were developed by screening a number of anti-HCG sera for specificity.4”*4h Others have developed an HCG-beta subunit immunoassay, which recognizes intact HCG and its beta subunit, but not LH.47 Finally. another group has developed radioimmunoassay, which displays a paradoxic increase in a “cooperative” tracer binding with intact HCG but not with HCG subunits or LH.48 Gonadotropin production is not limited to trophoblastic neoplasms but has also been found with a number of nontrophoblastic neoplasms,49.50 particularly and bronchogenic carcinoma.5.b,5’ When studied, the hepatob1astoma5’~52 hormones have all proved to be similar to HCG or LH except for one case where there was evidence for a second FSH-like gonadotropin.’ A recent retrospective study of 405 cancer sera with the HCG-beta subunit immunoassay with a variety of nontrophoblastic, identified HCG in over 79, of patients nongonadal neoplasms. 54 It is not yet clear whether the ectopic and trophoblastic tumor gonadotropins differ from normal HCG, as even specific imamong them. Although such a dismunoassays 48s4 have not yet distinguished tinction is not necessary for cancer diagnosis in a male or nonpregnant female, it could be invaluable in a pregnant female for differentiation of normal and neoplastic trophoblast. The concept of specific tumor markers has been extended to the subunits of polypeptide hormones. It has recently been recognized that HCG shares a similar, if not identical, alpha subunit with the other glycoprotein trophic hormones (LH, FSH, TSH), but has a unique beta subunit.s’ Radioimmunoassays have been developed that recognize both the free alpha and beta subunits of HCG, although these assays are not yet totally specific and show some cross-reaction with intact HCG.56 As a result, the recognition of free subunits in pregnancy serum and urine has required separation of the large amount of

COMPETITIVE

RADIOASSAYS

AND “SPECIFIC”

TUMOR

MARKERS

1123

intact HCG by gel chromatography prior to assay.” It is not yet clear what proportion of the subunits were secreted by trophoblast and what proportion derived from peripheral degradation of intact hormone. In contrast. the isolated production of HCG-beta subunit has been found in a patient with pancreatic carcinoma. 58.5yThe beta subunit was identified in serum and tumor extracts, although there was no evidence for production of intact HCG or its alpha subunit. This case of isolated production of HCG-beta suggests that there will be other examples of tumors producing free subunits of glycoprotein hormones, analogous to unbalanced immunoglobulin chain synthesis in certain forms of myeloma. Human placental lactogen (HPL. chorionic somatomammotropin) is another unique placental hormone, which shares biologic and immunologic features specific radiowith pituitary growth hormone. 6” Despite these similarities, immunoassays have been developed6’ 63and have permitted use of the hormone as a cancer marker. In contrast to HCG, HPL production by trophoblastic neoplasms was found to be much less efficient than by normal trophoblast.6’.6’ However. recent advances have substantially increased the sensitivity of the HPL radioimmunoassay and have enhanced the hormone’s diagnostic useful” In particular, affinity chromaness. particularly after tumor chemotherapy. tography has permitted isolation of high affinity antibody populations from individual anti-HPL serab6 as well as isolation and concentration of HPL from serum.h7 Moreover, the very inefficiency of HPL production by trophohlastic neoplasms is of diagnostic usefulness, and low serum HPL/HCG ratios have been used to distinguish neoplastic from normal trophoblast.h5 Just as with HCG, ectopic HPL production has been detected in nontrophoblastic, nongonadal neoplasms. In one retrospective survey of 128 cancer sera and 80 control sera. HPL was detected in over V’,, of the cancer sera but in none of the controls.2’ The hormone was associated with a variety of tumors, particularly carcinomas of the lung. In some of these cancer cases, HPL was not detected in unconcentrated sera but was detected after 500-fold concentration by athnity chromatography. Control sera similarly concentrated contained no detectable HPL (< 2 pg/ml). This again emphasizes that the incidence of detection of a cancer marker is proportional to the sensitivity of the method employed. A unique placental isoenzyme of alkaline phosphatase (PAP) has been identified by its distinctive electrophoretic, biochemical, and immunologic properties.“x PAP has also been shown to be a cancer marker and has been associated with a variety of neoplasms, particularly carcinomas of the lung. In one retrospective study of almost 600 cancer sera, PAP was detected in about 4”,, of nontumors. ” These workers were able to detect ectopic trophoblastic, nongonadal PAP even when total serum alkaline phosphatase activity was normal, and they found that changes in serum PAP appeared to correlate with the clinical response to chemotherapy. Recently, a PAP radioimmunoassay has been devcloped in another laboratory. but these workers were unable to find a single case of ectopic PAP in a retrospective survey of 100 cancer sera.7” Additional tumor-associated polypeptides,” 77 not yet fully characterized. are certain to play an important role in future cancer diagnosis. It will almost

1124

WEINTRAUB

AND ROSEN

surely be necessary to develop specific and sensitive competitive radioassays before their diagnostic potential can be fully explored. Although a single neoplasm can produce multiple tumor markers, ‘* the production of these polypeptides is usually discordant. 79 Thus, the use of assays for several such proteins should increase the incidence of early cancer detection. However, the true incidence of these markers and their specificity for cancer remain moot, partly because of differences in methodology and differences in patient populations studied. What is needed are prospective, unbiased studies of sera from cancer patients and appropriate control groups, using radioassays of high sensitivity and specificity. Once the true incidence and specificity of cancer detection with various serum markers has been established, it may be possible to make rational decisions about mass screening of various patient groups according to the principles of cost/effectiveness. In addition, such radioassays can also provide assistance in tumor localization, as has already been demonstrated by venous sampling studies in cases of endocrine neoplasms.80 The existence of ectopic polypeptide production has implications beyond the realm of cancer detection. To account for this phenomenon, it has been widely proposed that neoplastic transformation is associated with derepression of the genome, resulting in transcription of previously repressed, but normal DNA. Such a mechanism predicts “normal” tumor polypeptides, but the detection of “abnormal” tumor products by competitive radioassays challenges the simple derepression hypothesis and raises the possibility of other derangements in DNA transcription as well as in translation. However, as discussed above, these apparently abnormal proteins may actually be normal precursor OJ subunit forms, whose conversion to the normal protein may be impaired-such as by the absence of a specific converting enzyme. If so, the derepression hypothesis would still be tenable, and recognition of such precursors and subunits could provide valuable insight into the mechanism of normal protein synthesis. REFERENCES I. Bower BF, Gordan GS: Hormonal effects of nonendocrine tumors. Ann Rev Med 16:83. 1965 2. Lipsett MB: Humoral syndromes associated with cancer. Cancer Res 25: 1068, 1965 3. Liddle GW, Nicholson WE, Island DP, et al; Clinical and laboratory studies of ectopic humoral syndromes. Recent Prog Horm Res 25283. 1969 4. Berson SA, Yalow RS: Parathyroid hormone in plasma in adenomatous hyperparaand bronchogenic carcithyroidism, uremia, noma. Science 154907, 1966 5. Rosen SW, Becker CE. Schlaff S. et al: Ectopic gonadotropin production before clinical recognition of bronchogenic carcinoma. N Engl J Med 279:640, 1968 6. Faiman C. Colwell JA. Ryan RJ. et al: Gonadotropin secretion from a bronchogenic carcinoma. N Engl .I Med 277: 1395, 1967

7. Knill-Jones RP, Buckle RM, Parsons V, et al: Hypercalcemia and increased parathyroidhormone activity in a primary hepatoma. N Engl J Med 282:704, 1970 8. George JM, Capen CC, Phillips AS: Biosynthesis of vasopressin in vitro and ultrastructure of a bronchogenic carcinoma. J Clin Invest 51:141, 1972 9. Greenberg PB, Martin TJ, Beck C, et al: Synthesis and release of human growth hormone from lung carcinoma in cell culture. Lancet 1:350, 1972 IO. Turkington RW: Ectopic production of prolactin. N Engl J Med 285: 1455, 197 1 Il. Unger RH, Lochner JO. Eisentraut AM: Identification of insulin and glucagon in a bronchogenic metastasis. J Clin Endocrinol Metab 24:823. 1964 12. Orth DN, Island DP, Nicholson WE, et al: ACTH radioimmunoassay: Interpretation,

COMPETITIVE

comparison

RADIOASSAYS

with bmassay, and chnical

tion, tn Hayes (eds):

RL.. Goswitz

Radml\otopes

Studies. Oak Nuclear

in

Ridge,

Oak

BEP

In

Vitro

Ridge Institute

I Orth

Endocrinol

DN,

et

and pitui-

Metab

26. Smith ings First bryonic

BS, Carpenter

Some thoughts thyroid

on the nature

hormones.

Ii.

B, Fink

DJ. et al:

of ectopic

Am J Med 50:686,

Riggs BL. Arnaud Immunologic

CD,

hyperparathyroidism

of

JC, et primary

from hyperparathyroidism

due to nonparathyroid

Term

Ih.

cancer. J Clin Invest 50:

Berson SA. Yalow of

KS: Immunochemlcal

parathyroid

plasma. J Chn Endocrinol

27. Ruoslahtl

17. Sherman Circulating

BM,

Garden

insulin:

erties of “big”

The

in

28: 1037, 1968

P. Roth

J. et al.

proinsulin-like

insulin in patients

cell tumors. J Clin Invest 50:849.

a

Demonstratlcpn 28.

Ruoslahti of

prop-

withuut

islet

1971

al: Parathyrotd

hormone:

Murray

Secretion

TM,

et

and metab-

olism in vlro. Proc Nat] Acad Sci US.4 68:29X6, 1971

t,

insulin-like

P.

Sherman

component

the basal state

and

B,

Roth

of circulating

m patients

with islet cell tumors.

J Clan

J: Proinsulm

and

In

hamsters

Invest

50:2l 13.

1971

in norlnal

ductjon

BD,

of human

tropin

by

Rosen SW: Ectoplc

chorionic

Endvcrinol

Metab

.4belev (;I:

ture of tumor\

pro-

bomatomammo-

nontrophoblastic

cancers.

J

Chn

30. Ruoslahtl

Acta

Union

Int Contra

strucCancer

Increased

importance

of

detectlon patients.

in

the

the

31

Gold

absorption

techniques.

32. Gold

P,

clnoembrqonic

al:

GI.

Assecritova

Embryonal

cancer patients.

Diagnostic

serum

Kraevsky

cu-globulin

In

value. Int J Cancer

2:551. 1967 25. Alpert tu-Fetoprotein

m human

detection

in serum,

using

new

a

enterology61:137,

and

sensitive 1971

R, Schur PH. et al: hepatoma: quantitative technique.

Improved studies Gastro-

J

DMP.

‘tnd

sqbtem

IV’?

Demon5trdltr~ll human

iolon~c

tolerance

Exp SO:

and

t !I 4.2~

Med

Spccltl<

(al-

Jlgestive

1965

Krupe)

ing carcinoembrqonic

J.

antigen

t rtedm,rl; of cIrcu/rlt-

of

Proc Nat1 Acad

the

hunl~rl

SLI I. S;!

b.4

161. 1969 Kupchlk

al: Carcinoembryonlc of the colon

HZ.

antigen

and pancreas

Marcon assay

American

Cancer

Cancer

and other

study of a test for

36.

Institute

Society.

dlgearibe

I

ot

.4

U. c:~

111 LJnct‘r

tract disorders. Am J Dig Dis 16: I. lY7

C.madu,

collaboratl\c

carcinoembryonic

antlgrn

in the sera of patients with carcinoma

Martln

I-.

human

dlgestlve

of

<‘an Med Assoc J 107 75.

Martm

related

MS:

Demonstratlorl

IO colonic

system.

Int

cancer

in

J Cancer

the

h:Ay?.

lY70 37. Chu

TM,

Demonstration E, Hershberg

,tnd

m neoplaatic

SO. et al- The radloirnmunoassa) digcstlrr

lY72

Normal

in

system. J Exp Med 122467.

of antigens IV,

735:161

M.:

Freedman

1972

1966

u-Fetoproteln

antigens of the human

ru-globulin

1226,

M:

SO:

antigens

embryospecific

hepatoma

Biotram

with hepatocellular

by immunological

the colon dnd rectum

of

vl

II.

liver disease. Lancet 2.278. P. Freedman

Diagnostic

serum

serum

cr-fetoproteln

serum. Nature

AM:

blood

Vopr Onkol

24. Abelev et

YS, Nogaller

in

proteins.

of

alpha-fetoprotein

nonneopldstic

(CtA)

19:80. I963 23. Tatarinov

of

u-fetoprotein

Sepp%% M. Pihho H. et al:

E. Sepplld

35. National

32:94. lY71 Study of the antigenic

for

E, Seppsll

human

34. MooreTL,

21. Weintraub

ot

Int J Cancer 8:283, 1971

33. Thomson

20. Garden

Studies

Development

carcino-fetal

of tumor-specific

1969 D,

M

III.

human fetuses and patients

1965

Powell

ScppBIg

comparison

cent Pro& Horm

Res 25:207,

Commission).

of ,r-fetoproteln

Studies

carcinomata

JF,

t.

radiolmmunoassa)

IX Stemer DF, Clark JL. Nolan C, et al: Proinsulin and the biosynthesis of insulin. Re19. Habener

EmOak

InformatIon

Energy

protems.

29. Ruoslahti

hormone

Metab

Technical

(Atomic

on

Cancer.

1971, p. 305

cancer

heterogeneity

in

neo-

Proceed-

and Workshop

Antigens

Division

Extensions

chemical

207’). 1971

NA,

Fetal

in

conditions.

healthy human adults. Int J Cancer X:374. I971

1971

Reynolds

difl’erentlation

para-

Alpha-fetoprotein

Conference

and

carcino-fetal

14. Roof

22

JB.

piastic and nonneoplastic

33:377.

IY71

dl

1125

MARKERS

Ridge, WE,

between ectopic MSH

J Clin

TUMOR

applica-

Murphy

Medlcme:

M, Nicholson

al: Differences MSH

FA,

Term.

Studies, 1968. p 25

13. Shapiro tary

AND “SPECIFIC

Reynoso

normal human plasma. 3X. Payne RA. lactogen 1972 39

In the

Herr/

G.

Hansen

of carcinoembryonlc

R.

Nature

Ryan male Lewis

238:152.

RJ: Humarl

Lipsett

In

1977 placentitl

subJec1. J Ural JL,

t1.1

antigen

MB-

tcj7:YY. l-ire

1126

years’ experience with chemotherapy of meta. static choriocarcinoma and related trophoblastic tumors in women. Am J Obstet Gynecol 82:631. 1961 40. Ross CT, Goldstein DP. Hertz R, et al: Sequential use of methotrexate and actinomycin D in the treatment of metastatic choriocarcinema and related trophoblastic diseases in women. Am J Obstet Gynecol93:223, 1965 41. Hertz R: Biologic aspects of gestational neoplasms derived from trophoblast. Ann NY Acad Sci 172:279. 1971 42. Midgley AR Jr: Radioimmunoassay: A method for human chorionic gonadotropin and human luteinizing hormone. Endocrinology 79:10, 1966 43. Ode11 WD, Ross CT, Rayford PL: Radioimmunoassay for luteinizing hormone in human plasma or serum: Physiologic studies. J Clin Invest 46248, 1967 44. Yen SSC. Pearson OH, Rankin JS: Radioimmunoassay of serum chorionic gonadotropin and placental lactogen in trophoblastic disease. Obstet Gynecol 32:86, 1968 45. Wide L: Early diagnosis of pregnancy. Lancet 2:863, 1969 46. Franchimont P: A study of the crossreaction between human chorionic and pituitary luteinizing hormone (HCG and HLH). Eur J Clin Invest 1:65, 1970 47. Vaitukaitis JL, Braunstein CD. Ross CT: A radioimmunoassay which specifically measures human chorionic gonadotropin in the presence of human luteinizing hormone. Am J Obstet Gynecol l13:751, 1972 48. Weintraub BD, Rosen SW. McCammon JA, et al: Apparent cooperativity in radioimmunassay of human chorionic gonadotropin. Endocrinology 92: 1250, 1973 49. McArthur JW: Para-endocrine phenomena in obstetrics and gynecology, in Meigs JV. Sturgis SH (eds): Progress in Gynecology, vol 4. New York, Grune & Stratton, 1963. p 146 SO. Case records of the Massachusetts General Hospital (Case 13-1972). N Engl J Med 286:713. 1972 51. Hung W. Blizzard RM, Migeon CJ. et al: Precocious puberty in a boy with hepatoma and circulating gonadotropin. J Pediatr 63:895. 1963 52. Root AW, Bongiovanni AM, Eberlein WR: A testicular ~interstitial-cell-stimulating gonadotropin in a child with hepatoblastoma and sexual precocity. J Clin Endocrinol Metab 28:1317. 1968

WEINTRAUB

AND

ROSEN

53. Fusco FD. Rosen SW: Gonadotropinproducing anaplastic large-cell carcinomas of the lung. N Engl J Med 275:507, 1966 54. Braunstein CD, Vaitukaitis JL, Carbone PP. et al: Ectopic production of human chorionic gonadotropin by neoplasms. Ann Intern Med 78:39, 1973 55. Pierce JG: Eli Lilly Lecture: The subunits of pituitary thyrotropin-their relationship to other glycoprotein hormones. Endocrinology 89: 133 I, 197 I 56. Vaitukaitis J, Robbins JB, Nieschlag E. et al: A method for producing specific antisera with small doses of immunogen. J Clin Endocrinol Metab 33:988. 1971 57. Franchimont P, Reuter A: Evidence of alpha and beta subunits of HCG in serum and urine of pregnant women. in Proceedings of the Second International Symposium on Protein and Polypeptide Hormones, in press 58. Weintraub BD. Kadesky YM. Rosen SW: Ectopic production of human chorionic gonadotropin (HCG) and its free beta subunit (HCG-beta). Clin Res 20:444, 1972 59. Rosen SW, Weintraub BD: Gonadotropins and placental proteins produced by human tumors, in Symposium on Endocrinology of Tumors, Fourth International Congress of Endocrinology. Washington, D.C., Excerpta Medica, 1972, in press 60. Josimovich JB, MacLaren JA: Presence in the human placenta and term serum of a highly lactogenic substance immunologically related to pituitary growth hormone. Endocrinology 71:209. 1962 61. Grumbach MM, Kaplan SL: On the placental origin and purification of chorionic “growth hormone-prolactin” and its immunoassay in pregnancy. Trans NY Acad Sci 27: 167, 1964 62. Frantz AC, Rabkin MT, Friesen H: Human placental lactogen in choriocarcinoma of the male. Measurement by radioimmunoassay. J Clin Endocrinol Metab 25: 1136, 1965 63. Beck P. Parker ML, Daughaday WH: Radioimmunologic measurement of human placental lactogen in plasma by a double antibody method during normal and diabetic pregnancies. J Clin Endocrinol Metab 25:1457, 1965 64. Samaan N, Yen SSC, Friesen H, et al: Serum placental lactogen levels during pregnancy and in trophoblastic disease. J Clin Endocrinol Metab 26: 1303, 1966 65. Saxena B. Goldstein DP, Emerson K, et al: Serum placental lactogen levels in patients

COMPETITIVE

with

RADIOASSAYS

molar

pregnancy

AND “SPECIFIC”

and

tumors. Am J Obstet Gynecol 66. Weintraub bon

YM:

to human

atomammotropin:

Application

to

munoassay.

Endrocrinol

Metab

J Clin

radioim33:432,

67

Wetntraub

BD:

motropin

(HCS)

Applicatron

to

Fishman

chemical

WH:

SE, Porter

kin’s disease.

Evidence

LL.

production

isoenqme

for

Fishman

of an alkaline with

phosphatase

cancer.

N

Engl

KD:

placental-type

A

J

radiotm-

alkaline

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Res 32:.2-l 13, I972

‘3,.

73. Edynak

tn human

A fetal

antigen

P: Foetal

“antrgens”

Old

associated

Vrana

in can-

with

M,

human

et al: neo-

DL,

tt) Sci

JM,

Higgins HP, Starnes WK.

mole and human

tropin.

Metabolism

Lung

thyroid

l9:735.

MA,

carcinoma

Eilber

FR.

Malmgren

RA:

R. Schaaf

assoctated

of three placental gonadotropin,

placental

286:

I 169.

and 1972

tn

(tic(;) patrents

Doppman

N

JL. et

Preoperative

and drfferentiation

hyperplasia.

BD phus-

197 I

PM,

hyperparathyrotdism.

localization

alkaline

(HCS)

with cancer. Clin Res 19497,

tumor

alkalrne

Weintraub

gonadotropin

somatomammotropin

al: Primary

Human chorionic

Med (in press)

secretion of placental chorrontc

et

ectopi<

proteins:

79. Sussman H. Rosen SW. phatase (PAP).

M,

wrth

human

and

Arch Intern

rn

thyro-

1970

production

phosphatase.

stimulator chorionic

Claypool

chorionic

adenoma

plasia. N tngl J Med 2X6:1 178, 1972 Morton

An

pa-

cancer test. lsr J Med

80. Powell D, Shimkin LJ,

of cancer attempt

hydatidiform

and

137, 1972 EM,

in serum

between

Discordant

reversion

1971

women:

somatomammotroprn

P: Antigenic

72. Alexander

protein

pregnant

S: Hodg-

M: Presence of serologr-

Differences

al:

WH:

cancer. Ann Rev Med 22:85, 1971 cer. Nature

and

7X. Charles

B, Bagshawe

Cancer

71. Gold

Ann

lY69

Jacoby

munoassay

bio-

phosphatase

MJ,

Hellman

6: 708, 1970

1969

Krant

distinct

77. Hershman and

to alkaline

in patients

Med 2X1:757,

phatasr.

Biochrm

M,

for a tumor-associated

N Engl J Med 285:471.

develop a diagnostic

39:83, 1970

analysis: The Regan isoenzymr.

69. Stolbach

tally tients

chromatography:

Immunologic

NY Acad Sci 166:745, Ectopic

and

somatomam-

radioimmunoassay.

approaches

isorn/yme

chorionic

by affinity

Biophys Res Commun 68.

Concentration

of human

Mahgnant

Res 14:25. 1971

75. Order antigen.

cancer:

and soft tissue sarcomas.

76. Tal C. Halperin

puritication

74.

in human

skeletal

Prog Exp Tumor

som-

1971

70

factors

melanomas,

Fractiona-

chorionic

1127

MARKERS

Immune

trophoblastx

102: 115, 1968

BD, Kadesky

of antibodies

TUMOR

Engl

between J

Mrd