NOVEMBER 1973

The American Journal of Medicine

VOLUME 55

NUMBER 5

EDITORIAL

Parathyroid Hormone: Coming of Age in

Clinical Medicine

CLAUDE D. ARNAUD, M.D Clinicians have been intensely interested in disorders of the
Rochester, Minnesota parathyroid glands since Fuller Albright and his colleagues [l]
demonstrated that these disorders were common, were chal-
lenging diagnostic exercises and often were curable. The first
impetus to both clinical investigation and improved diagnosis
came with the development of reliable, precise technics for
measuring calcium [2] and phosphorus [3] in biologic fluids.
These are still keystones, and presently available automated
analytical technics permit the measurement of these com-
pounds in small quantities of serum in multiphasic screening
programs. As a result, the detection of abnormalities of calci-
um and phosphorus metabolism has increased dramatically
[4,5]. This has resulted in better patient care and an approach
toward preventive medicine in this area of endocrinology not
previously thought possible, but also it has given rise to con-
sternation. The physician is now being faced much more fre-
From the Mineral Research Laboratory, De- quently with the tricky differential diagnoses related to hyper-
partment of Endocrine Research, Mayo Clin- calcemia and hypocalcemia but has had little more than rela-
ic, Rochester, Minnesota 55901. This study tively ancient indirect tests of parathyroid function to help him.
was supported in part by grants from the
In 1963, when Berson and co-workers [S] first reported a
NIH (AM 12302, 69-2166 and Ca 11911AA)
and the Mayo Foundation. Manuscript re- workable radioimmunoassay fqr human parathyroid hormone
ceived October 12,1973. (PTH).in serum, using a cross-reacting bovine PTH antiserum,

November 1973 The American Journal of Medicine Volume 55 577

PARATHYROID HORMONE-ARNAUD

it was thought that widespread clinical application PTH (l-84) both in peripheral organs and in the
of the assay would rapidly follow. This did not parathyroid gland; and (3) the C fragments have
happen. Aside from the special technical difficul- long survival times (hours), and both PTH (l-84)
ties involved in performing the PTH radioimmu- and the N fragments have short survival times
noassay [7], a new set of problems slowly emerged, (minutes).
and these have yet to be completely solved. The practical consequences of these facts in
First, and probably least important, only nonhu- the routine application of PTH radioimmunoassays
man species of PTH (bovine and porcine) were to clinical’ diagnostic situations will be reported in
available for the large-scale immunization pro- detail in the upcoming Third F. Raymond Keating,
grams needed to produce high-affinity anti-PTH Jr., Memorial Symposium entitled Parathyroid
serums, and cross reaction of these antiserums Hormone, Calcitonin, and Vitamin D: Clinical Con-
with human PTH was at best incomplete. This siderations to be published in the May 1974 issue
problem may now be solved because the amino of The American Journal of Medicine. In essence,
acid sequence of the amino-terminal region of it appears that the previously reported discrepan-
human PTH has been deduced [8], and synthetic cies can be satisfactorily explained on the basis of
preparations of the human peptide [9] should be- the use, by different laboratories, of PTH assays
come available soon for use as an antigen. that have different specificities for the different
Second, and probably most important, it was immunoreactive species of PTH in serum or plas-
demonstrated by Berson and Yalow [lo] that the ma. Assays using antiserums that react primarily
PTH circulating in hyperparathyroid man is with the C terminal region of the PTH molecule
immunoheterogeneous, that is, more than one im- measure C fragments (long survival time) as well
munoreactive form of PTH is present in serum. as native PTH (l-84) (short survival time), where-
Although it was not appreciated at the time, the as assays using antiserums that react primarily
impact of serum PTH immunoheterogeneity on the with the N terminal region of the PTH molecule
practical application of radioimmunoassay to clini- measure N fragments (short survival time) and
cal diagnostic problems began to be felt when, in native PTH (l-84) (short survival time).
the period 1968 to 1971, discrepancy and dis- Direct comparison of assays with N and C
cordance characterized radioimmunoassay results specificities in studies of serum iPTH during
reported by various laboratories. Overlap of serum, steady-state conditions and induced hypercalce-
immunoreactive parathyroid hormone (IPTH) mia show the following:
values between normal subjects and patients
(1) C-specific assays give higher values for
with proved hyperparathyroidism varied from
serum iPTH than do N-specific assays because,
0 to 50 per cent. One laboratory [l11 reported
during the steady state, C fragments accumulate
that induced hypercalcemia failed to sup-
in the circulation as a consequence of their long
press serum iPTH values in patients with parathy-
survival time.
roid adenomas, supporting the concept of autono-
my; another laboratory [12] reported that, in simi- (2) Serum iPTH values obtained with C-specific
lar patients, serum iPTH values not only were de- assays reflect the state of chronic hyperparathy-
creased by calcium infusions but also were sharp- roidism much better than do those with N-specific
ly increased by induced hypocalcemia. assays (that is, there is less overlap of serum
Although these results appeared irreconcilable, iPTH values between normal and hyperparathyroid
reasonable explanations were forthcoming. A subjects).
number of laboratories [13-201 systematically
(3) N-specific assays, because of the short sur-
began to identify the various circulating forms of
vival times of both N fragments and PTH (l-84),
PTH and to characterize their metabolic handling,
can more easily demonstrate rapid changes in
using gel filtration of serum and sequence-specific
PTH secretion (such as suppression with induced
immunoassays as tools. This work has been re-
hypercalcemia) than can C-specific assays and
cently reviewed in detail [21]. Summarized (Fig-
probably can better demonstrate increases in
ure l), the data so far published indicate that (1)
iPTH concentrations in serum obtained from veins
hyperparathyroid plasma contains the “native”
draining parathyroid adenomas. This is because
9,500 molecular weight 84-amino acid “glandular”
N-specific assays primarily “see” the PTH (l-84)
form of the hormone (PTH l-84) and multiple,
secreted by parathyroid tumors and are “blind” to
amino-terminal (N) (potentially biologically active)
the C fragments that also are present in parathy-
and carboxyl-terminal (C) (biologically inactive)
roid venous drainage.
fragments of PTH (l-84); (2) the C and N frag-
ments are probably derived from degradation of On the surface, the complexities of these inter-

578 November 1973 The American Journal of Medicine Volume 55

 PARATHYROID HOHMONF ARNAUD

C
ASSAYS
___------

N
ASSAYS

PARATHYROID PERIPHERAL
CIRCULATION
Figure 1. Simplified schema of current concept of the metabolic alterations
in PTH (l-84) that result in immunoheterogeneity. Proparathyroid hormone
(proPTH) [27-291 (mol wt > 11,000) is converted to PTH (l-84) (mol wt =
9,500) by specific enzymatic cleavage. It is not presently known if proPTH is
released from the gland into the blood. PTH (l-84) is the major secreted
species of PTH, but there is evidence that the gland contains a calcium-regu-
lated enzyme that can cleave PTH (l-84) to COOH-terminal and NH2-termi-
nal (C and NJ fragments [27]. These hagments also might be released into
the circulation, but there is no direct evidence for this. Once secreted, PTH
(l-84) is converted peripherally into C and N fragments. PTH (l-84) and N
fragments have a fast turnover rate, and their extracellular pool sizes are
probably small compared with those of C fragments, which have a slow turn-
over rate. N-specific radioimmunoassays of serum iPTH give low values and
C-specific radioimmunoassays give high values because of the differences in
these pool sizes. This scheme is based on data obtained from the study of
serum from hyperparathyroid patients. Data on normal serum are not avail-
able. See text for further information on practical consequences of these phe-
nomena in the application of PTH radioimmunoassay in clinical endocrinolo-

relationships might tempt the casual observer to with one assay (Hawker) but inappropriately low
turn his head and join those who, out of initial dis- with another assay (Arnaud). One set of serum
enchantment caused by the reported discrepan- iPTH values (Arnaud) reflected the apparent true
cies, tend to “put down” the clinical usefulness of biologic state of the patient (hypoparathyroidism)
PTH radioimmunoassay. It is clear, however, that whereas the other set (Hawker) reflected the ap-
important new knowledge has been born of dis- parent parathyroid secretory state but not the bio-
crepancy and, far from invalidating the radioim- logic state of the patient.
munoassay of PTH as a clinical diagnostic and These workers have logically concluded that
investigative tool, the unraveling of these complex the PTH secreted by the patient’s parathyroid
molecular interrelationships should prove the great glands was not biologically effective, and they
intrinsic potential of the procedure. suggested that the problem is due to the failure of
The discrepant results of the assays of serum peripheral conversion of a secreted, precursor
iPTH in the patient so well studied and reported by form of PTH to an active form of PTH. By the
Nusynowitz and Klein in this issue of The Ameri- same reasoning, an alternative explanation would
can Journal of Medicine may turn out to be as im- be that there is defective intraglandular conver-
portant as the discrepancies that originally stimu- sion of a precursor form of PTH with release of
lated a systematic investigation of the immunohet- only the biologically inactive precursor into the
erogeneity of plasma PTH. In this case, the serum circulation. On the other hand, the data might be
iPTH values in a hypocalcemic, hyperphospha- explained on the basis of the rather heretical no-
temic patient, whose renal response to exogenous tion that PTH (l-84) itself is not biologically active
PTH was not impaired, were appropriately high and that it requires specific cleavage in peripheral

November 1973 The American Journal of Medicine Volume 55 579

PARATHYROID HORMONE-ARNAUD

organs before it can exert its biologic effects In the present dilemma (Nusynowitz and Klein’s
[22,23]. Although Nusynowitz and Klein attempted patient), in which two assays with apparently the
to test this possibility by administering PTH (l-84) same specificities gave different results, I would
to the patient, they found that the purified PTH predict that ultimately it will be shown that certain
they used was ineffective both in the patient and antiserums that ostensibly are C-specific may rec-
in a control subject. A normal renal response was ognize a structural peculiarity of a unique, biologi-
obtained with Lilly Para-Thor-Monee in the patient. cally active, circulating N fragment of PTH that
However, this does not exclude the possibility that many antiserums (either N-or C-specific) recog-
the patient had a defective peripheral conversion nize poorly or not at all. Unfortunately, at the
mechanism for PTH (l-84) because Lilly Para- present time, our immunologic technics and re-
Thor-Mone is a crude, hot hydrochloric acid extract agents are probably not sophisticated enough to
of bovine parathyroid glands and is composed permit investigation of this problem.
largely of fragments of PTH (l-84) [24]. These latter comments are, clearly speculative
From an immunochemical point of view, sever- and comprise the current thinking and flow of re-
al observations made in my laboratory on the rel- search activity in my laboratory. Of greater impor-
ative specificities of the Arnaud and Hawker anti- tance is that much work can be accomplished
serums used in the study of this patient have been now in the partial characterization of potentially
disquieting. Neither of these antiserums recog- “abnormal” forms of circulating PTH. Methods
nizes either synthetic bovine or human amino-ter- have been recently developed [25] that allow for
minal PTH (l-34) except at extremely high the identification of molecular species of circulat-
and nonphysiologic concentrations. By exclusion, ing PTH by scaled-up molecular seiving technics,
therefore, they are both primarily directed at the C even in plasma samples containing very low con-
terminal region of the PTH molecule and should centrations of iPTH. Initial efforts with this ap-
recognize both PTH (l-84) and C fragments in proach have shown that the original observation
serum. Yet, assays of serum iPTH using these by Riggs and his co-workers [26] that, for a given
two antiserums have given different results in serum calcium value, serum iPTH concentration
studies of this patient as well as in other compara- was relatively lower in ectopic hyperparathyroid-
tive studies of serum from normal and hyperpar- ism (nonparathyroid cancer) than in primary hy-
athyroid subjects. perparathyroidism, is due to an alteration in the
These observations, along with other as yet un- ratio of immunoreactive forms of circulating PTH
published immunochemical studies of highly puri- in the two syndromes [25]. Until studies such as
fied human PTH (l-84) and synthetic fragments of this are carried out in patients similar to the one
human PTH using sequence-specific antihuman described by Nusynowitz and Klein, the case they
PTH antiserums, suggest the real possibility that have described will, at the very least, have
the schema of serum PTH immunoheterogeneity caused us to reexamine our current concepts of
described earlier in this editorial (Figure 1) may the interpretation of the results of sequence-spe-
not be the whole story. If so, I believe that new cific radioimmunoassays and, more important, will
information may be developed in regard to the im- alert us to the existence of another “accident of
munologic recognition of the secondary and terti- nature” that has the potential of giving us a great
ary structures of the circulating molecular species deal of information about parathyroid physiology
of PTH. and pathophysiology.

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 PARATHYnOlD HORMONE--ARNAUD

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