PLASMA-PROLACTIN IN HUMAN BREAST CANCER

433 PLASMA-PROLACTIN IN HUMAN BREAST CANCER

by centrifugation in another laboratory false-positives may assay, fetal red blood-cells

are

removed

occur.

We have evaluated as a screening procedure the radioimmunoassay of A.F P. in maternal serum and confirm the earlier observations of Seppala and Ruoslahti.9,10 High maternal serum-A.F.P. levels do not occur consistently with open spina bifida or anencephaly even when the level in amniotic fluid obtained simultaneously is high. The onlyunequivocally raised serum value in our series occurred before the spontaneous abortion of a macerated anencephalic fetns. Seppälä and Ruoslahti11 have already noted raised values in association with fetoplacental dysfunction and impending intrauterine death. The kinetics of A.F.P. metabolism during pregnancy have been discussed previously.12 Amnioticfluid A.F.P. reaches a peak at about 14 weeks gestation, falling to below 1 µg. per ml. by the beginning of the third trimester and is then undetectable by immunodiffusion using the Behringewerke plate. In contrast the peak in maternal serum is at about 32 week’s gestation, and even higher values occur in certain types of disorders in the fetoplacental unit. It is not entirely clear from what source maternal A.F.P. is derived, but the occurrence of maternal peak values at a time when the total amount of A.F.P. in the fetus is at its highest suggests a fetal origin of maternal A.F.P. Unless more A.F.P. than normal is produced by fetuses with spina-bifida/anencephaly it seems likely that previous reports’ and our case no. 2 in which high maternal A.F.P. levels occurred in anencephaly are due to fetoplacental dysfunction rather than neural-tube malformations per se. Since most babies with spina bifida and anencephaly are born to mothers who have no relevant history to alert the obstetrician, a simple and safe screening procedure is urgently required. Although amniotic-fluid A.F.P. assay seems to he reliable, amniocentesis could not be used in more than a minority of pregnancies, and our results do not encourage us to believe that widespread use of A.F.P. radioimrnunoassay of maternal sera in randomly selected early pregnancies would detect many affected but viable fetuses. However, it might be argued that since raised maternal serum-A.F.p. levels are almost invariably associated with severely disturbed pregnancies little harm would be done if these pregnancies were terminated even though they are not related to anencephaly or spina bifida. The cumulative efficiency of all available methods, including A.F.P. assay, and the continuing search for new methods will allow the detection with increasing efficiency of neural-tube malformations early in pregnancy. We thank our many obstetric colleagues for their cooperation, Dr Norman Nevin (case 9 and ref. 4) and Dr A. Milford Ward (cases 10 and 11) for specimens of liquor and serum. Mrs Claire Pownall, Mrs E. Quinn, Miss Jane Fennell, Mrs R. Harris, and Miss A. Gallagher for their assistance with the collection of the dati and the preparation of the paper. One of us (K. M L.) is especially grateful for the support of the National Fund for Research into

Crippling

Diseases.

Requests for reprints should

be addressed to R. H.

M. DE JONG-BAKKER F. J. CLETON

H. G. KWA E. ENGELSMAN

and Internal Medicine, The Netherlands Cancer Institute, Amsterdam, The Netherlands

Departments of Biology

Plasma-prolactin was assayed in 115 patients with breast cancer and 115 matched controls. Mean plasma-prolactin levels were 6.0 ± 3.7 ng. per ml. and 5.9 ± 2.9 ng. per ml., respectively. Plasma-prolactin levels were normally Summary

distributed in both groups. Plasma levels in 64 members of nine families with a high frequency of breast

irregularly distributed, with a mean level of 10.4 ± 8.1 ng. per ml. Statistical prolactin evaluation demonstrated that breast-cancer patients and controls may be regarded as one population but that the prolactin levels in the high-risk group represent a different population (P < 0.0004). cancer

were

Introduction THERE is

evidence that mammary be induced in mice from strains with a low frequency of breast cancer by exposing the animals to high levels of prolactin for a long time.1,2 tumours

experimental

can

In D.M.B.A. (7,12-dimethylbenz[a] anthracene)-treated rats, there was a correlation between plasmaprolactin levels and the genetically determined susceptibility to breast cancer.3 We have studied plasma-

high-risk group-i.e., members of famihigh frequency of breast cancer. Their plasma-prolactin levels were compared with those of a group of breast-cancer patients and matched conprolactin in

lies witli

a

a

trols. Patients and Methods From 1970 to 1972, 115 patients treated for breast canstudied after mastectomy. In some of these, there was evidence of recurrence. Full clinical records and detailed data on tumour histology were available. A normal age, race, and parity matched control was selected for each patient. The median age was fifty-one years in the patients and fifty-four years in the controls Information was obtained on the use of drugs (antihypertensives, tranquillisers, and contraceptives) that might influence plasmaprolactin levels. Members of nine families in which more than2 first-degree relatives had breast cancer were investigated. The positive history of breast cancer was checked in all cases by review of clinical records or of tumour hiscer were

DR

1. 2. 3.

4. 5. 6.

7. 8. 9 10. 11. 12.

HARRIS

AND

OTHERS :

REFERENCES

Brock, D. J. H., Sutclifle, R. G. Lancet, 1972, ii, 197. Lorber, J., Stewart, C. R., Milford Ward, A. ibid. 1973, i, 1187. Seller, M. J., Campbell, S., Coltart, T. M., Singer, J. D. ibid. 1973, ii, 73. Nevin, N. C., Nesbitt, S., Thompson, W. ibid. 1973, i, 1383. Allan, L. D., Ferguson-Smith, M. A., Donald, I., Sweet, E., Gibson, A. A. M. ibid. 1973, ii, 522. Brock, D. J. H., Bolton, A. E., Monaghan, J. M., ibid. 1973, ii, 923. Ruoslahti, E., Seppälä, M. Int. J. Cancer, 1971, 8, 374. Laurence, K. M , Turnhull, A. C., Harris, R., Jennison, R. F., Ruoslahti, E., Seppala, M. Lancet, 1973, ii, 860. Seppälä, M , Ruoslahti, E ibid. 1973, i, 155. Seppälä, M., Ruoslahti, E. ibid. 1973, i, 375. Seppälä, M., Ruoslahti, E. Am. J. Obstet. Gynec. 1973, 115, 48. Seppälä, M., Ruoslahti, E. J. perinat. Med. 1973, 1, 107.

434

tology. Pedigrees were constructed, which included at least three generations. The 64 family members studied included 17 men. The median age was forty-six years. The patients and their controls came to the outpatient department to give blood-samples, between 9 and 11A.M., after a normal breakfast. Blood from members of families was collected at home, at the same time of day. Plasma samples were stored at -20°C. Radioimmunoassay was based on highly purified human prolactin given by Dr U. J. Lewis (iodinated with ’2I and used as a reference preparation) and an antiserum induced in rats, as described elsewhere.4 All samples were assayed undiluted and in dilutions 1/2, 1/4, 1/8, and 1/16. Results were expressed as ng. per ml.; 1 ng. being equal to 8 standard milliampoules of M.R.C. 71/222.

00

32 8 20 4 12 16 24 28 PLASMA- PROLACTIN LEVELS (ng.perml.)

Fig. 3—Plasma-prolactin levels high frequency

Results

Levels of plasma-prolactin in breast-cancer patients (fig. 1) and in controls (fig. 2) were normally distributed. Mean plasma-prolactin levels in both groups (6.0 -±- 3.7 ng. per ml. and 5.9 --h 2.9 ng. per ml., respectively) were almost identical. The intake of drugs in patients and controls did not influence prolactin levels. Plasma-prolactin levels in 24 patients and 29 controls taking drugs ranged from 1.4 to 16.1 ng. per ml. (mean 6.5 ng. per ml.). Plasmaprolactin levels in members of breast-cancer families

44

in members of families with of breast cancer.

a

rather

irregularly distributed (fig. 3). The mean plasma-prolactin level for this group was 10.4 ± 8.05 ng. per ml., which is significantly higher than that m the breast-cancer patients and the controls. -Statistical evaluation (Wilcoxon’s ranking test) shows that breast-cancer patients and controls may he regarded as one population. The prolactin levels m the high-risk group represent a different population (p < 0.0004).

were

Discussion

Prolactin is widely regarded as a possible major factor in the carcinogenesis in the mammary gland.’ No

significant differences in plasma-prolactin were reported between breast-cancer patients and controls in other studies,3,6 this is confirmed in the present study. There seems to be no difference in prolactin levels in persons under or over fifty. Having near relatives with breast cancer is one of the recognised risk factors for breast cancer.-, Moreover, patients

PLASMA-PROLACTIN LEVELS

( ng. per ml.)

Fig. I-Plasma-prolactin

levels in

patients

with breast

cancer.

with breast cancer and their sisters have abnormal androgen excretion.,’ This may be relevant to onr finding significantly higher plasma-prolactin levels in members of breast-cancer families compared with breast-cancer patients and controls. It is remarkable that plasma samples from 5 women in one family were relatively low—i.e., between 1.7 and 6.7 ng. per ml. Prolactin levels in men tend to be lower than in women. There were 17 men among the 64 members of breast-cancer families; their mean prolactin level was 7 ng. per ml. These findings in men and in the women from one family described above emphasise the importance of the higher prolactin levels of the women in the other breast-cancer families. We thank Mr F. Verhofstad and Miss A. M. A. Sanders for technical assistance and Miss M. Heyster for her effort in collecting the blood-samples from the families.

Requests for reprints should be addressed to H. G. K., The Netherlands Cancer Institute, 108 Sarphatistraat, Amsterdam-C, Netherlands. REFERENCES 1 2. 3.

4

PLASMA- PROLACTIN

LEVELS

(ng. per mi.) Fig. 2—Plasma-prolactin

levels in controls.

Mubibock, O., Boot, L. M. Cancer Res. 1959, 19, 402. Kwa, H. G., Verhofstad, F. J. Endocrinology, 1967, 38, 81. Wilson, R. G, Buchan, R., Roberts, M. M., Forrest, A. P M. Boyns, A. R., Cole, E. N., Griffiths, K. Proc. R. Sec Med 1973, 66, 865. Kwa, H G., Engelsman, E, van der Gugten, A A. in H Prolactin (edited by J. I . Pasteels and C. Robyn); p. Amsterdam, 1973.

435

Hypothesis POSSIBLE ROLE OF A B-CELL MITOGEN IN HYPERGAMMAGLOBULINAEMIA IN MALARIA AND TRYPANOSOMIASIS B. M. GREENWOOD

Department of Medicine, Ahmadu Bello University, Zaria, Nigeria

especially Serum-immunoglobulins, IgM, increase considerably in patients with malaria and trypanosomiasis. This immunoglobulin contains not only specific anti-parasite antibodies but also antibodies reacting with other non-parasite antigenic determinants, including those of host tissue. It is suggested that the immunoglobulin response seen in malaria and trypanosomiasis is due in part to the production by malaria parasites and trypanosomes of a non-specific B-cell mitogen. Stimulation of a non-specific and disordered immunoglobulin response could help the paraSummary

site to survive. HYPERGAMMAGLOBULINAEMIA IN MALARIA AND TRYPANOSOMIASIS

Malaria and African trypanosome infections of lead to the rapid development of hypergammaglobulinaemia, especially of IgM.l,2 Invasion of the central nervous system by Trypanosoma gambiense leads to the production of large amounts of IgM within the nervous system,3 and a raised cerebrospinal fluid (C.S.F.) IgM is an important diagnostic feature of this disease.4 Serum IgG and IgM levels are increased in most people in many tropical areas.5-7 Black Americans have only slightly higher immunoglobulin levels than White Americans,8 suggesting that environmental factors are important in producing hypergammaglobulinaemia in Africans. Malaria is probably the most important of these environmental factors.9,lo The nature and function of the immunoglobulin produced in such large amounts in malaria and trypanosomiasis is uncertain. Some is specific antibody, but absorption experiments with malarialll and trypanosomah2 antigens suggest that specific antibody forms only a small fraction of the increased immunoglobulin concentrations seen in these two infections. man

ABERRANT ANTIBODY RESPONSE IN MALARIA AND TRYPANOSOMIASIS

The immunoglobulin produced panosomiasis commonly contains

5.

in malaria and tryantibodies reacting

der Gugten, A. A., Verstraeten, A. A. in Methods in Cancer Research (edited by H. Busch); vol X, p. 161. New York,

van

1974. 6.

Boyns, A. R., Cole, E N., Griffiths, K., Buchan, R., Roberts, M. M , Wilson, R. G , Forrest, A. P. M. Eur. J. Cancer 1973, 9, 99. 7. Anderson, D. E. Cancer Bull. 1973, 25, 23. 8. Bulbrook, R. D. Proc. R. Soc. Med. 1972, 65, 646.

with non-parasite antigenic determinants, including those of host tissues. Heterophile agglutinins are found in patients with African trypanosomiasis13,14 and occur in high titre in Nigerians living in areas of high malaria endemicity.15,16 Antibodies reacting with the Wassermann antigen develop in patients with malaria.17 IgM rheumatoid factors appear rapidly in the serum of animals experimentally infected with malaria and trypanosomiasis18 and are found in patients with African trypanosomiasis.19 Malaria probably plays an important role in the production of the rheumatoid factors commonly found in apparently healthy residents of many tropical countries .20 Malaria leads to the formation of a speckled form of antinuclear factor,21 and a high frequency of organ-specific autoantibodies was found in Rwandans moving from an area of low malaria endemicity to one of high endemicity.22 Autoantibodies also develop in trypanosome infections.23 POSSIBLE ROLE OF A B-CELL MITOGEN IN

MALARIA AND TRYPANOSOMIASIS

There is thus massive production of imunoglobulin in malaria and trypanosomiasis, the immunoglobulin containing specific antibody and also antibodies reacting with a wide range of non-parasite antigenic determinants. The presence of free, light chains in the c.s.F. and urine of patients with sleeping-sickness24,25 suggests that in this infection immunoglobulin synthesis is occurring in a disorganised way. I suggest that the disorder of immunoglobulin function seen in malaria and trypanosomiasis can best be explained by the assumption- that malaria parasites and trypanosomes produce a B-cell mitogen which has a non-specific stimulatory effect on immunoglobulin synthesis. This possibility has been raised by other workers.26,27 Production of a B-cell mitogen by these parasites could explain not only the high IgM levels seen in these infections but also the occurrence of anomalous antibodies, for it has recently been shown 28 that endotoxin, a B-cell mitogen, can stimulate the synthesis of antibody to sheep and horse red blood-cells by spleen cells, although these antigens were not present in the culture system. Production of a B-cell mitogen would be expected to lead to an increase in lymphocyte D.N.A. synthesis. It is therefore of interest that two recent studies have shown a high uptake of [3H]thymidine by lymphocytes from children with acute malaria.29,30 The tropical splenomegaly syndrome responds to treatment with antimalarials,31 suggesting that malaria plays an important part in its pathogenesis. Abnormal production of IgM is probably the basic abnormality in this condition and might be due to an abnormal and prolonged response to a B-cell mitogen produced by malaria parasites. The nature of the postulated mitogen is a matter for speculation. An endotoxin-like substance is a possible candidate since, in addition to its mitogenic effect, such a compound might be playing a part in the production of the intravascular coagulation, complement activation, kinin activation, and stimulation