Norethisterone enanthate as an injectable contraceptive in puerperal and non-puerperal women

CONTRACEPTION

NORETHISTERONE ENANTHATE AS AN INXXYAELE CONTPACEPTIVE IN PUERPERAL AND NON-PUERPERAL WOMEN G.B.Melis,F.Strigini,F.Fruzzetti,A.M.Paoletti, E.Rainer,B.Diisterberg,P.Fioretti Dept.Obstetrics

and Gynecology

- University

of Pisa Italy Clinical Research Service - Schering S.p.A. - Segrate Milan0 - Italy Dept.Biodynamik - Schering AG - West Berlin - Germany Federal Republic ABSTRACT Norethisterone enanthate (NET-EN) was intramuscularly administered to 5 puerperal women and 20 non-puerperal women for a total of 366 months.Contraceptive effectiveness and side effects of the drug were evaluated.Basal levels of LH,FSH,prolactin (PRL),estradiol 17 (E2) and progesterone (P) were measured in blood B samples collected from 5 non-puerperal women,while LH,FSH,PRL and norethisterone (NET) plasma levels were evaluated in puerperal women-NET was also assayed in plasma from breastfed newborns.No woman became pregnant-side effects consisted of only menstrual abnormalities.Ovulation (P plasma levels higher than 2000 pg/ml) was achieved in 3 patients during the first month of NET-EN treatment but luteal function appeared to be insufficient.In puerperal women,NET plasma levels showed a course similar to the one observed outside puerperium.Lactation was not inhibited, and NET transfer to newborn through milk was negligible, since NET was undetectable in newborn plasma when maximal levels were measured in the mother.It was concluded that NET-EN is an effective contraceptive drug,deprived of major side effects,and particularly useful in women affected by metabolic diseases or during puerperium. This work has been partially supported by C.N.R. (Centro Nazionale Ricerche)-Rome,Italy : project "Biology of the reproduction" and grant n.79.01891.04 Address for reprints : Dr.G.B.Melis - Clinica Ostetrica e Ginecologica - Universita degli Studi di Pisa - Via Roma 35 - 56100 Pisa - Italy.

Accepted

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December

1981 VOL. 23 NO. 1

26, 1980

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INTRODUCTION So far it is widely accepted that oral contraceptives (OC) or intrauterine devices (IUD) are the most utilized methods for female contraception,but they cannot be useu in various pathophysiological conditions.Particularly OC cannot be assumed by women with hematological (sickle cell anemia),cardiovascular or metabolic (diabetes mellitus, liver diseases) alterations,or at risk for these pathologies (l).On the other hand,IUD may be dangerous in women with malformative or inflammatory genital diseases, or suffering from fibromyomas.Barrier methods could be an alternative to OC or IUD although neither are as effective nor always accepted nor correctly used. Particular problems about contraception are represented by puerperium.It is well known that ovulation may occur during lactation,which cannot always be considered as a period of absolute refractoriness to exogenous as well as endogenous stimuli (2,3).OC represent an adjunctive risk of thromboembolic disease (4),interference with milk secretion (5) and may induce estrogenic modifications of breast-fed newborns (6). Depot gestagens may be advisable in some of these cases, since they seem to be deprived of the negative metabolic effects linked to the estrogenic component of OC (7,8), while they are seemingly not contraindicated in women affected by genital diseases or even during puerperium. Therefore,the aim of this paper was to evaluate clinical and endocrinological effects of the depot gestagen norethisterone enanthate (NET-EN) as contraceptive in a group of selected women. MATERIALS AND METHODS Twenty women with relative or absolute contraindications for OC or IUD and 5 puerperal women were submitted to the study.Among puerperal women,one subject (N.S.) was affected by diabetes mellitus and required insulin therapy (60 UI/ die);in the same patient lactation was suppressed by means of methergoline (4 mg 3 times daily for 10 days).The others continued breast-feeding for the first month and thereafter decided to interrupt it. NET-EN (200 mg in oil/vial) was intramuscularly injected on the 5th day of menstrual cycle to non-puerperal women and between the 2nd and 5th post-partum day to puerperae. The stated dosage of the drug were always administered every 60 days,as suggested by W.H.O. report (9).After physical evaluation and history,every subject was required to observe either characteristics of subsequent menstrual cycles or side effects of treatment.Laboratory examination, including serum glucose,creatinine,urea,bilirubin,SGOT, prothrombin index,cholesterol and SGTP,blood picture, urinalysis,was performed before treatment and every 6 months thereafter. Blood samples were withdrawn from 5 non-puerperal women before treatment and 7 and 14 days after injection of

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1'ET-EN.Other samples were obtained once between the 21st and 28th and between the 35th and 60th post-injection days. Circulating levels of gonadotropins (LH and FSH), prolactin (PRL),estradiol 17p (E2) and progesterone (P) were evaluated in all samples.During puerperium,blood samples were collected from every subject before treatment,daily for 7 days and on the 15th day after injection.Blood samples were also collected in the 4 breast-fed newborns on the 2nd day (3 subjects) or on the 5th day (1 subject) after injection to the mothers.Plasma norethisterone (NET) was measured in all samples while gonadotropins and PRL were also assayed in blood collected from puerperae. In all,366 months of treatment were observed. Three subjects (2 puerperae) were treated for 4 months,10 women (3 puerperae) for 12 months,9 for 18 months and the others reached 24 months (Table I). Table

I : Clinical

data

for women

Puerperae (5)

Months of treatment

4 12 18 24

2 3

treated

with

Non-puerperae (20)

NET-EN

Total (25)

1 7 9 3

3 10 9 3

2

3

Pregnancy Amenorrhea (more than

3 months)

'l

Prolonged bleeding (more than 15 days)

1

1

2

Irregular

2

9

11

3

3

bleeding

Other side effects (headache,nausea, depression)

LH,FSH and PRL plasma levels were measured by specific radioimmunoassay (RIA) methods using double antibody technique (Biodata kits).Within assay and between assay coefficients of variation were,respectively,lower than 5% and 10%. Unconjugated steroids were assayed by the previously described RIA (IO) after their extraction with dietiyl-ether. Charcoal-dextran was used to separate bound from free fraction.Plasma NET was also measured as previously described (11). The following reagents,purchased by Scherinc AG (West Berlin - FRG), have been used: the phosphate

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buffer saline (PBS) solution was 0.05 M sodium phosphate (pH 7.5) in 0.15 M sodium chloride, containing 0.01% merthiolate and 0.5% human serum albumineithe dextrancoated charcoal reagent was prepared as 0.5% suspension of Norit-A charcoal in a 03.05% solution ot dextran T-70 in PBS.Labelled NET(15,16H NET; specific activity 2109 GBq/mmol;30 pg/tube) was used as radioligand and rabbit anti-NET-llO( -BSA was used as antiserum. The standard curve ranged from 3 .9 to 1000 pq/tube.Plasma samples were extracted with diethyl-ether (1:5) and reconstituted with PBS; 0.1 ml of reconstituted PBS was assayed.The incubation time was 16 hours at O'C.Dextrancharcoal (0.5 ml/tube) was added to separate bound from free fraction-The coefficients of variation of this assay were respectively 5.4+0.4% (within assay) and 9.5+0.7% (between assays).The sensitivity was 50 pq/ml. RESULTS Clinical data During 366 months of treatment none of patients became pregnant.Only 2 patients complained of headache, while another suffered from nausea and depression during the first trimester of therapy;other general side effects were not observed.Also hematological parameters did not show significant modifications after NET-EN assumption.The values (expressed as mean + standard error, M+SE) observed before treatment and after-12 months of therapy were, respectively, 0.80+0.2 mq/ml and 0.79+0.3 mg/ml for serum glucose;0.92+0.05~mq% and 0.88+0.07 kq% for serum creatinine;27+5 mg% and 2822 mg% for serum urea;0.62+0.03 mg% and 0.65+0.02 mq% for serum bilirubin; 14.2452.0 U/ml and 15.12+2.3 U/ml for SGOT;8.16+3.4 U/ml and 9.2352.0 U/ml for SGTP;T1.3+1.2% and 92.2+1.4%-for prothrombin index; 180+13 mg% and 185+12 mq% for serum cholesterol. The-main side effect was then represented by alterations of menstrual cycle,which were observed in the majority of subjects (Table I). Four of the puerperal women resumed menses,respectively, 40,54,62 and 68 days post-partum,while one presented continous vaginal bleeding until she stopped treatment (4 months after delivery).One puerpera recorded amenorrhea after 4 regular menses. In the group of non-puerperal women,one had intermenstrunl bleeding during treatment,2 showed amenorrhea after the 2nd injection of NET-EN,while the others showed only minor disorders or altered rhythmicity of cycles.Eight patients did not show significant variations of their menstrual cycles in comparison to pre-treatment conditions.Owinq to these effects,therapy was interrupted in 2 patients,while 7 underwent other contraceptive methods just after one year of NET-EN treatment. Endocrine effects With reuard to hormonal warameters, significant modifications have not been observed either in puerperal or in non-puerperal women in comparison to untreated 2

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subjects. On the day of the first injection (the 5th day after the beginning of menstrual cycle),4 non-puerperal women showed mean (M+SE) baseline levels of PRL (12.7+2.4 ng/ml),FSH (12.6~2.1 mU/ml),and LH (11.0+1.9 mU/ml)within the normal range for early follicular phase (Table II).After treatment hormone plasma levels were not significantly modified (PRL:12.9+2.3 ng/ml;FSH 9.2+0.7 mU/ml;LH 12.8+4 mu/ml). On the contrary, one patient showed LH and FS?? peak (respectively, 24 mu/ml and 100 mu/ml) on the day of the first NET-EN injection;in the same sample E2 concentration was 165 pg/ml,and P peak (3500 pg/ml) was observed on the P peak was also observed in another 7th day after injection. 2 patients (respectively 32C0 and 3800 pg/ml) on the 14th post-injection day during the first month of treatment. In the same cycles E2 plasma levels consistent with initial follicular maturation (more than 100 pa/ml) were detected. In all other samples E2 concentrations were similar to those measured in normal subjects during the early follicular phase (32.8+4.7 pg/ml on the day of first injection and 29.4+4.6-pg/ml during treatment). Also in puerperal women,hormonal pattern was not different in comparison to the values usually detected during puerperium in our laboratory (Fig.l).FSH was barely detectable (c3.1 mU/ml),while LH concentrations were at first very high (>I00 mU/ml),because of the cross-reaction between LH and hCG.Since hCG is progressively eliminated; on the 15th day post-partum,LH concentrations were 7.1+1.5 mU/ml.PRL plasma concentrations reached very high levels in every patient‘except N.S.,who was taking methergoline. Norethisterone concentrations ____ In puerperal women, NET plasma levels,which were obviously undetectable before treatment,reached the highest values (5.920.8 ng/ml) between 4 and 7 days after drug injection (Table III,Fig.2).Subsequently plasma levels decreased up to the 15th day (4.79+0.8 ng/ml) (Table III,Fig.2).In one subject plasma samples were obtained also 29 and 45 days after injection-In these samples NET concentrations were, respectively,0.40 ng/ml and 0.24 ng/ml. In the 4 breast-fed newborns,NET levels were undetectable (CO.05 ng/ml) in plasma samples collected between 2 and 5 days after injection to the mothers. DISCUSSION Our data confirm the effectiveness of NET-EN as a depot contraceptive and its possible utilization in different pathophysiological conditions. The main negative effect consisted of menstrual abnormalities, inducing treatment discontinuation in 2 out of 25 patients.In comparison with the most widely used medroxyprogesterone acetate,NET-EN seems to induce lower incidence of spotting and amenorrhea, while the two drugs are comparable with regard to other menstrual abnormalities (12).Present results seem to confirm (7,8) that NET-EN can be useful also when

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Before

12.6+2.1 -

(mU/ml)

(mu/ml)

FSH

LH

11.0+1.9 _

12.7f2.4 _

(ng/ml)

PRL

treatment

I

I -

10.4+3.9 -

7.3+0.9 -

11.0+2.6 _

7

14

after

10.6+1.1 -

7.3+2.0 -

15.9+3.6 -

11.5+2.5 -

1

21-28

injection

8.9+0.9 -

12.7+2.5 -

Days

35-60

(mean + standard error) of prolactin (PRL) and gonadotropins Table II : Plasma levels (LH and FSH) in 5 non-puerperal subjects treated with NET-EN (200 mg i.m.).All values were consistent with follicular phase of normal menstrual cycle.

CONTRACEPTION

Fig. 1: Gonadotropin (LH and FSH) and prolactin (PRL) plasma levels in 5 puerperal women treated with NET-EN (200 mg i-m.) between the 2nd and the 4th day post-partum. The values were similar to those usually measured during puerperium.Plasma PRL levels were low in N.S., who was _ treated with methergoline to inhibit lactation. contraception is required by women affected by metabolic disorders, since alterations of hematochemical parameters were not observed even after a long period of intake (until 2 years).On the other hand, elevations of PRL plasm? levels were not observed in non-puerperal women, in contrast with what happens in other animal species (13). Different hypotheses have been reported about the mechanism of action of NET-EN.In post-menopausal women, XET-EN treatment was shown to decrease gonadotropin plasma levels (14,15). Similar results have been obtained by Goebelsmann -& eta1 (16) in fertile women;they also suggested that NET was able to inhibit preovulatory LH ?>eak.All these effects have been only partially confirmed Ir?v other authors (14),who excluded an inhibitory effect of YET on basal gonadotropin levels in pre-menopausal women. Our data seem to confirm (17,18) that ovulation is not

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always inhibited by NET-EN treatment,since in 3 patients progesterone plasma levels,and in one patient also LH levels,were consistent with ovulation.In contrast with Fotherby's results (18),we have seen that ovulation also occurred during the first cycle of treatment and was followed by insufficient luteal function.All these data seem to suggest that NET-EN can act as a contraceptive steroid exerting antifertility effects at different levels. At the peripheral level it acts on cervical mucus, endometrium and fallopian tubes (17),while inhibition of ovulatory LH surge and of luteal function may sometimes occur depending on the levels of circulating NET. Table III : NET plasma levels (ng/ml) in 5 puerperal women injected with NET-EN between the 2nd and the 4th day postpartum-All values areexpressed as mean 2 standard error. Before treatment 1

2 Days

3

after

4

injection

5 6 7 15

(0.05 2.43+0.3 3.98+0.7 5.47+1.3 _ 6.53+1.3 _ 5.83+0.9 _ 5.91+0.8 _ 5.57+0.6 _ 4.79+0.8 -

As far as NET-EN administration to puerperal women is concerned,no different results were observed in comparison with non-puerperal subjects.The same kind of menstrual abnormalities and side effects were observed;not one became pregnant.The peripheral antifertility effect of NET-EN was important during puerperium as well as in nonpuerperal women,since variations of gonadotropins were not detected.Also NET plasma levels showed a pattern similar to that observed outside puerperium (16,19),which was characterized by an initial peak during the first week of treatment,followed by a relatively rapid decrease in the second week.According to Goebelsman (16),some discrepancies in absolute values of NET in plasma may be related to individual variations,rate of absorption,rate of cleavage from NET-EN to NET,metabolic clearance rate of NET and mass of body fat.In puerperal women,the high levels of sex hormone binding globulin may also modify NET plasma levels (21).

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days

from

injection

Fig. 2 : Plasma NET levels in 5 puerperal subjects injected with NET-EN between the 2nd and the 4th day postpartum.In maternal plasma,NET values showed a sharp peak in the first week after injection,followed by a decrease which was yet detectable on the 15th day post-injection.In breast-fed newborns,the steroid was undetectable even during maternal peak.

NET transfer from mother to newborn throughmilk appeared to be negligible.In fact,the steroid was undetectable ( < 0.05 ng/ml)in the plasma of breast-fed newborns when the highest plasma levels were detected in the mothers (2-5 days after NET-EN injection).Although it would have been more conclusive to measure NET in breast milk rather than in the newborns,our results are in agreement with previous data obtained by giving puerperal women 19-nor gestagens (21,22).It has been reported also that there is a positive effect of gestagens on lactation,in contrast with the inhibiting action of estrogens (5). Gestagens seem indeed to be able to increase milk production and newborn ponderal growth when administered to mothers (23). In our subjects milk production did not seem to be affected by the use of NET-EN,since every mother continued

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full breaseeeding until they decided to stop suckling. On the other hand,the physiologically high levels of PRL were not affected.All these factors suggest that NET-EN is also deprived of major side effects during puerperium for mothers and breast-fed newborns.However, the use of NET-EN in a greater number of puerperal women will be able to confirm its contraceptive effectiveness in these subjects. REFERENCES 1. Crosignani P.G. Follow-up during oral contraception.J. Endocrinol.Invest. 1:97-98 (1978). 2. Andreassen B.and Tyson J.E. Role of the hypothalamicpituitary-ovarian axis in puerperal infertility. J. Clin.Endocrinol.Metab. 42:1114-1121 (1976). 3.

Tyson J-r-.,in Prolactin and Human Reproduction (P.G. Press London Crosignani and C. Robyn,Editors),Academic New York San Francisco,1977,p.97-117.

4. Daniel D.G.,Campbell H. and Turnbull A.C. Puerperal thromboembolism and suppression of lactation. Lancet 2 :287-289 (1967). 5.

piller G.H. and Hugher L.R. Lactation and genital involution effect of a new low-dose oral contraceptive on breast-feeding mothers and their infants.Obstet. Gynecol. 35:44-50 (1970)

6. Curtis E.M. Oral-contraceptive feminization of a normal male infant.Report of a case.Obstet.Gynecol. 23:265 (1964). 7. Howard G.,Myatt C. and Helder M-G. The effects of intramuscular norethisterone enanthate used as a contraceptive on intravenous glucose tolerance and on blood coagulation factors VII and X. Br.J.Obstet. Gynaecol. 84:618-621 (1977). Dhall K.,Kumar M.,Rastogi G.K. and Devi P.K. Shortterm effects of norethisterone enanthate and medroxyprogesterone acetate on glucose,insulin,growth hormone and lipids. Fertil.Steril. 28:156-158 (1977). World Health Organization expanded programme of research ,development and research training in human reproduction.Task force on long-acting systemic agents for the regulation of fertility.Multinational comparative clinical evaluation of two long-acting injectable contraceptive steroids:norethisterone oenanthate and medroxyprogesterone acetate.1. Useeffectiveness.Contraception 15:513-533 (1977).

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10. Giannessi P.,Zucchelli G.C.,Simonini N.,Piro M.A.,Masini S. and Malvano R.Methodoloqical validation of human chorionic somatomammotrophin,estriol,estradiol and progesterone radioimmunoassay in preqnancy.J.Nucl. Med.All.Sci.22:71-78 (1978). 11.

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system radio-

12. World Health Organization special programme of research,development and research training in human reproduction.Task force on long-acting systemic agents for the regulation of fertility.Multinational comparative clinical evaluation of two long-acting injectable contraceptive, steroids:norethisterone oenanthate and medroxyproqesterone acetate.2-Bleeding patterns and side effects.Contraception 17:395-406 (1978). 13. Neumann.F.,Berswordt-Wallrabe R.,Elger W.,Grgf K.-J., Hasan S.H .,Mehrinq M.,Nishino Y. and Steinbeck H. Special problems in toxicity testing of long-acting depot con'craceptives.Acta Endocrinol. Supp1.185:315353 (1979).

14.

Franchimont P.,Cession G.,Ayalon D.,Mutsers A. and Legros J.J. Suppressive action of norethisterone enanthate and acetate on gonadotropin (FSH and LH) levels.Obstet.Gynecol. 36:93-99 (1970).

15.

Fotherby K.,Howard G.,Shdmanker K.,Elder M. and Bye P. G. Effect of norethisterone oenanthate on serum gonadotrophin levels. Contraception 16:591-604 (1977).

16.

GoebelsmannU,Stanczyk F.Z.,Brenner P.F.,GoebelsmannA E .,Gentzschein E.K.E. and Mishell D.R. Serum norethisterone (NET) concentrations following intramuscular NET enanthate injection.Effect upon serum LH,FSH,estradiol and proqesterone.Contraception 19: 283-312 (1979).

17. El Mahgoub S. and Karim M. The long-term Use Of injectable norethisterone enanthate as a contraceptive. Contraception 5:21-29 (1972). 18. Fotherby K.,Howard G.,Shrimanker K.,Elder M. and Bye G. Occurrence of ovulation in women receiving the injectable contraceptive norethisterone oenanthate. Contraception 18:535-542 (1978).

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22.

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