Treatment of Hormone-Producing Adrenocortical Cancer With o,p’DDD and Streptozocin
BARBRO ERIKSSON, MD,* KJELL ÖBERG, MD,* THORE CURSTEDT, MD,t ANDERS HEMMINGSSON, MD,# PROFESSOR HENRY JOHANSSON,§ ERIK LINDH, MD,* PER-GUNNAR LINDGREN, MD,# KARL-ÅKE THUOMAS, MD, ERIK WILANDER, MD,| AND GÖRAN ÅKERSTRÖM, MD§
Three patients with advanced adrenocortical carcinoma were treated with a combination of intermittent streptozocin and continuous o,p’DDD. Two patients were treated preoperatively and the primary tumors, initially considered as inoperable, could be resected after 19 and 5.5 months, respectively. In the patient with the longer treatment (35 months), lung and lymph node metastases disappeared and she has no evidence of recurrent disease 6.5 years after start of therapy. One patient was followed by magnetic resonance imaging (MRI) and urinary steroid secretion. The MRI gave a good visualization of the tumor. Measurements of relaxation times showed a significant decrease in T1 values. The urinary steroid profile showed an increased secretion of 38-hydroxy-5-ene steroids and tetrahydro-11-deoxy-cortisol. Treatment with streptozocin and o,p’DDD initially increased 16-oxygenation of dehydroepiandrosterone and androst- 5-en-36,178-diol, followed by a decrease in the secretion of all urinary steroids. The third patient received postoperative treatment with no effect on metastatic disease in the lungs, she died 9 months after start of treatment. The therapeutic approach with the combination regimen of streptozocin and o,p’DDD pre- treatment plus aggressive surgery has to be further evaluated, as well as MRI and urinary steroid profile as methods to monitor the effect of therapy.
Cancer 59:1398-1403, 1987.
C ARCINOMA OF THE ADRENAL CORTEX is a rare tumor in man, affecting about two per million popula- tion.1-3 In most cases, the diagnosis is made at a late stage when the tumor is inoperable or has metastasized.4 In approximately 50% of the reported cases, endocrine man- ifestations, such as Cushing’s syndrome, hyperaldoster- onism, virilization, or feminization have been present.5 The prognosis has been considered extremely poor be- cause of the unfavorable results with conventional surgical treatment, radiotherapy, and chemotherapy.4 An oral chemotherapeutic agent with adrenolytic effects, or- tho,para’DDD (o,p’DDD, mitotane), has been used in this disease since 1960. Totally, there have been eight complete responses to this drug described in the world literature.6 Streptozocin, an alkylating agent employed in gastroin- testinal endocrine tumors, has been reported to concen- trate into mice adrenal cortex.7 We thus have explored the usefulness of this drug in the treatment of adrenocor-
tical cancer. Three patients with advanced tumors were treated with a combination of intermittent streptozocin and continuous o,p’DDD.
Methods
Peripheral blood and urinary samples were collected before and regularly during treatment in addition to dif- ferent hormones; also, routine hematology, liver enzymes, serum creatinine, and urinary albumin were checked.
The levels of serum cortisol, 11-desoxycortisol, estra- diol, testosterone, dehydroepiandrosterone sulphate (S- DHAS) and urinary cortisol and aldosterone were mea- sured by commercial radioimmunoassays. Total urinary 17-oxosteroid levels were determined according to the method of Vestergaard.8
Isolation and separation of individual urinary steroids were performed as described previously9 with the excep- tion that the steroid conjugates were not separated before enzymatic hydrolysis and solvolysis. The steroids were analyzed as 0-methyloxime-trimethylsilyl ether derivatives by gas chromatography using a fused silica capillary col- umn coated with OV-1. Mass response factors were de- termined for all major naturally occurring steroids. For other components the response factor of 1.0 was used. The identity of the compounds constituting the gas chro- matographic peaks were confirmed by gas chromatogra-
From the Departments of *Internal Medicine, Radiology, §Surgery, and | Pathology, University Hospital, Uppsala, and +Department of Clinical Chemistry, Karolinska Hospital, Stockholm, Sweden.
Supported by the Swedish Medical Research Committee (project No. 6676), Swedish Cancer Research Fund (1925-B85-02XA), and Pharmacia Research Fund.
Address for reprints: Barbro Eriksson, MD, Department of Internal Medicine, University Hospital, S-751 85 Uppsala, Sweden.
Accepted for publication November 14, 1986.
phy-mass spectrometry using a Finnigan 1020 (Finnigan Mat, San Jose, California) instrument with an electron energy of 40 eV and equipped with a 25-m OV-1 fused silica capillary column. After a splitness injection with the oven at room temperature, the temperature was rap- idly increased to 190℃ whereupon it was programmed to 260°℃ at a rate of 2°℃/minute.
Histopathologic examination included routine fixations and the Grimelius silver nitrate stain.10
Angiography, computerized tomography (CT), and ul- trasound were used to assess changes in tumor size. In Case 2, treatment response was followed by magnetic res- onance imaging (MRI) operated at 0.35 Tesla (Magnetom, Siemens AG, Berlin). Spin echo images were obtained at repetition times (TR) of 500 and 1500 ms and echo delay times (TE) of 35, 70, and 120 ms. The slice thickness was 10 mm with 10-mm slice intervals. The matrix size was 256 × 256. Calculations of the relaxation times were made by the standard soft ware program (Numaris, Siemens AG, Erlangen, version B1).
Ultrasonically guided tumor biopsy specimens, using a 1.2-mm specially made cutting needle, were concomi- tantly taken for histopathologic investigations.11
o,p’DDD (mitotane) was administered orally. The doses are described in each individual case. Streptozocin was given intravenously with an induction course of 1 g/d for 5 days. The first patient started streptozocin after 2 weeks of o,p’DDD and received 5-day courses every month with a few interruptions (in all, five courses), whereafter she received monthly injections of 1.5 to 2.0 g. In the second and third patients, streptozocin was initiated at the same time as o,p’DDD. After one induction course, they got monthly single injections of 1.5 to 2.0 g. Cortisone re- placement therapy was initiated concomitantly with the o,p’DDD treatment in all three patients (dexamethasone 2 mg per day in Cases 1 and 2, cortisone acetate 50 mg per day in Case 3). In Case 1, mineral corticoid replace- ment therapy was started after 6 months. In the third case, mineral corticoid replacement was given initially but had to be discontinued after 1 month because of hy- pertension.
Case Reports
Case 1
A 35-year-old woman presented with hypertension and ele- vated hemoglobin and platelet levels in January 1979. She had a 3-year history of increasing pain in the upper right part of the abdomen. Acne had appeared 3 years earlier and she had noticed a tendency of blushing and swelling of her face and neck, bruises, proximal muscle weakness, and hirsutism 1 year before admis- sion.
Biochemical testing demonstrated markedly elevated urinary cortisol secretion, 2300 nmol/d (reference level, <300 nmol/d) (Fig. 1). Plasma cortisol levels were increased and without cir- cadian rhythm. Urinary aldosterone also was elevated, 225 nmol/
2000
U-Cortisol(nmol/d)
1000
.
S mo
10
15
20
25
30
34 37 42 48 55
62
STZ + + +
+
+
+
.
+
+
+
+
op’DOD
op
op
d (reference level, <38 nmol/d). Serum estradiol, testosterone and 17-urinary oxosteroids all were normal. Radiologic exam- ination (CT scan) revealed an abdominal tumor mass, measuring 19 × 13 cm (Fig. 2A). Lymph node and lung metastases (Fig. 3A) were present. The patient was referred for surgery in January 1980, but at operation the tumor was found to be unresectable.
No biopsy was performed because of the high vascularity of the tumor.
Cytotoxic treatment with o,p’DDD was initiated in February 1980, with 1 g twice per day, increased to 2 g twice per day after 1 month. Cortisol secretion decreased from 2300 to 1640 nmol/ d after 2 weeks of o,p’DDD. Streptozocin was introduced in March 1980. Cortisol secretion then was normalized within 3 months (Fig. 1) and a similar pattern could be seen in the urinary aldosterone secretion, which was normalized within 2 months. Radiologically, the lymph node and lung metastases (Fig. 3B) had disappeared at a 4-month check up. At 5 months, the patient had an episode of fever and ultrasound scan demonstrated a necrotic area in the tumor. After 6 months, she developed an Addisonian crisis with severe hypotension. Dexamethasone was increased and later replaced by cortisone acetate 50 mg per day, and mineral corticoid replacement with fluodrocortisone acetate 0.15 mg daily was commenced. One month later, cytotoxic treatment could be continued, and in May 1981 she had received a total of 27.5 g of streptozocin. Repeated CT scans had shown a decreasing tumor mass and in August 1981 the maximum diameter of the tumor was less than 10 cm (Fig. 2B). At 19 months, a surgical resection of a fibrotic and necrotic adrenal tumor was performed. Histopathologic examination showed no viable cancer cells. Postoperatively, streptozocin and o,p’DDD treatment was reinitiated. The maintenance dose of o,p’DDD was kept relatively low, 1.0 to 1.5 g, until it was discontinued at 30 months (11 months postoperative because of side effects, nausea, and vomiting). The patient received 1.0 to 1.5 g of strep- tozocin every third month until December 1982; the total dose had then reached 33.5 g. Since then, she has received no che- motherapy.
After 35 months of therapy CT showed a suspect tumor in the left adrenal gland and another laparotomy was performed. This operation, however, displayed a totally atrophic adrenal
gland, which was excised and the “tumor” suspected at CT probably corresponded to her atrophic spleen. Microscopic ex- aminations of the atrophic adrenal gland showed total cortical atrophy and capsular fibrosis. At 4.5 years after start of therapy, the patient was operated upon because of cholelithiasis and at that time no recurrent abdominal tumor could be seen. She has now been followed for more than 6.5 years and biochemically and radiologically there is no evidence of any recurrency. Her cortisol levels have varied (as can be seen in Fig. 1) with the substitution dose, which she still requires, since both her adrenal glands have been removed. During treatment liver enzymes in- creased, especially alkaline phosphatase and alanine amino- transferase, and these elevations were persistent until treatment was stopped. Urinary albumin increased after 3 months of treat- ment and at 7 months it was 145 mg/d (reference level, <20 mg/d). This elevation has then persisted. Aside from nausea, vomiting, and hypotension, pain in the bladder region bothered the patient intermittently and repeatedly during treatment. No hematuria was seen. Irregular bleeding and colpitis called for estrogen substitution, which was started after 30 months.
Case 2
A 31-year-old woman with a 4-year history of intermittent episodes of pain in her upper right abdomen with concomitant fever and night sweat. No clinical signs of endocrine dysfunction were present.
Biochemically, a slight excess in urinary cortisol excretion was evident, 290 nmol/d (reference level, <200 nmol/d). Fur- thermore, S-DHAS was elevated, 21 umol/l (reference level, <10 umol/d (Fig. 4). Urinary steroid profile measurements revealed an excessive secretion of cortisol precursors (36-hydroxy-5-ene steroids and tetrahydro-11-deoxycortisol) (Table 1, first column). Angiography performed in January 1984 demonstrated a tumor
in the right adrenal gland. The CT scan showed a lobulated, partly calcified tumor. On MRI examination, a tumor measuring 102 × 156 mm was demonstrated (Figs. 5A and 5B). Calculations of relaxation times showed an elevation of T1 values, as an av- erage 1500 ± 350 SD ms (normal 700 ms), whereas T2 values were in the normal range, i.e., 70 ± 10 SD ms. Ultrasound guided biopsy was performed and the diagnosis of an adrenocortical tumor was confirmed.
Surgically, the tumor was considered inoperable, because of the size and suspect invasive growth into the right liver lobe angiographically. Cytotoxic treatment was initiated in February 1985 with continuous o,p’DDD and a 5-day course of strepto- zocin. The o,p’DDD dose was increased up to 4 g/d, but then had to be reduced to a maintenance dose of 2 g/d because of intolerable nausea and vomiting. After the induction, strepto- zocin was repeated with a single monthly injection of 1.5 to 2.0 g. Biochemically, cortisol secretion and S-DHAS normalized after 1 month (Fig. 4). The decrease of urinary dehydroepian- drosterone (DHA) (mostly as sulphate) and androst-5-ene- 36,176-diol was initially accompanied by a concomitant increase of 16-oxygenation of these steroids, resulting in secretion of 16x- hydroxy-DHA,androst-5-ene-38,178-diol-16-one and androst-5- ene-36,16a,176-triol. Further treatment decreased all 36-hy- droxy-5-ene steroids and after 3 to 4 months only trace amounts of steroids could be detected (Table 1).
Repeated MRI examinations showed decreasing T1 values, at 5 months as an average 800 ± 80 SD ms, whereas T2 values had increased in certain areas up to 120 ± 20 SD ms. After 5 months of therapy (10 g of streptozocin), the tumor measured 89 × 92 mm (Figs. 5C and 5D).
A hemorrhagic cystitis was noted after the second course of streptozocin, and after 5 months elevated liver enzymes (alanine aminotransferase 5.59 ukat/l and aspartate aminotransferase 2.95 ukat/1; reference level, <0.6 ukat/l) were seen and the treatment discontinued. The patient was referred to surgery, which was
20
S-DHAS (umol/l)
10
f mo
2 mo
3 mo
4 mo
5 mo
6 mo
7 mg
STZ It
+
1 g x 5
2 g
1,5 g
1,5 g
1,5 g
op
o.p’DDD
performed at 5.5 months. The entire tumor mass was resected and histopathologic examination showed necrotic and fibrotic tissue with a few viable tumor cells. Postoperatively, there were infectious complications, but she recovered and cytotoxic treat- ment was restarted. Treatment was stopped 5 months after sur- gery because of elevated liver enzymes. We suspect an autoim- mune reaction within the liver. She has now been followed for 20 months without signs of recurrent disease.
Case 3
A 37-year-old woman presented with tiredness, weight loss, and dyspnea. Hypertension and hypokalemia was noted a few months later in December 1980. No clinical endocrine signs could be seen.
| Peak no. in Fig. 7 | Steroid | Before treatment | Duration of treatment (umol/24 h) | Controls* | |||
|---|---|---|---|---|---|---|---|
| 1 mo | 2 mo | 3 mo | 4 mo | ||||
| 1 | Androsterone | 24 | 2.9 | Tr | 0.6 | 1.7 | 8.5± 3.6 |
| 2 | Etiocholanolone | 18 | 9.3 | 2.8 | 2.4 | 0.6 | 7.8 ±3.2 |
| 3 | Dehydroepiandrosterone | 103 | 11 | Tr | ND | ND | 1.3±1.5 |
| 4 | Androst-5-ene-33,178-diol | 17 | 10 | 0.8 | ND | ND | <1.0 |
| 5 | 113-Hydroxyandrosterone | 6.2 | 2.3 | 1.5 | 0.6 | ND | 4.2 ± 1.6 |
| 17a-Hydroxydehydroepiandrosterone | |||||||
| 6 | 16a-Hydroxydehydroepiandrosterone | 18 | 58 | 11 | Tr | 1.3 | <2.0 |
| 7 | Pregnanetriol | 9.2 | 2.7 | 0.4 | 0.3 | 0.2 | 2.0± 1.1 |
| 8 | Androst-5-ene-38,173-diol-16-one | Tr | 10 | 2.8 | Tr | ND | ND |
| 9 | Androst-5-ene-36,16a,173-triol | Tr | 44 | 11 | 1.9 | 1.5 | <1.0 |
| 10 | Tetrahydro-11-deoxycortisol | 15 | 13 | 7.3 | 5.0 | ND | ND |
| 11 | Pregn-5-ene-38,17a,20a-triol | 85 | 30 | 7.0 | 1.4 | 0.5 | <1.0 |
| 12 | Tetrahydrocortisone | 11 | 1.7 | 2.4 | 2.8 | ND | 6.9 ±3.7 |
| 13 | Tetrahydrocortisol | 9.0 | 2.1 | 1.7 | 0.7 | ND | 4.3±2.0 |
| 14 | allo-tetrahydrocortisol | 4.2 | 0.5 | 0.2 | ND | ND | 2.9 ± 1.7 |
| 15 | a-Cortolone | 5.6 | 2.3 | 0.7 | 0.3 | ND | 2.4±1.0 |
| 16 | B-Cortolone + 3-cortol | 5.8 | 4.2 | Tr | Tr | ND | 3.2±0.9 |
| 17 | «-Cortol | 2.5 | 1.7 | Tr | Tr | ND | 1.0±0.5 |
* Mean ± SD from 16 women, 24-44 yr old.
Tr: trace; ND: not detected.
156 mm
192
mm
92 3R
€
E
89 mm
D
C
Biochemical testing revealed excessive urinary cortisol secre- tion, 910 nmol/d (reference level, <300 nmol/d). There also was markedly increased serum levels of 11-deoxycortisol, 102 nmol/ 1 (reference level, <30 nmol/l). 17-Oxosteroids were at the upper normal range. Angiography and CT performed in June 1981, demonstrating a left suprarenal tumor with a maximum diameter of 20 cm. Multiple lung metastases were present.
The patient was operated on in July 1981. The abdominal tumor, spleen, and left kidney were resected, but mesenterial lymph node metastases could not be excised. Histopathologic examination showed an adrenocortical cancer with an abun- dance of mitotic cells and tumor growth into the renal vein.
Cytotoxic treatment commenced 2 weeks after surgery with an induction course of streptozocin. o,p’DDD was increased up to 4 g/d within 10 days, but then rapidly had to be reduced to a maintenance dose of 1 g/d because of nausea and vomitting. The patient received four additional single injections of strep- tozocin. Her u-cortisol levels decreased slightly, but s-11-desox- ycortisol increased steadily as did the lung metastases. Strepto- zocin was discontinued after 4 months and other chemothera- peutic agents (cisplatin, 5-fluorouracil, and mitomycin C) were tried without any effect. The patient died 9 months after start of chemotherapy. Autopsy showed metastases in the lungs and the liver.
Discussion
Adrenocortical carcinomas are highly malignant tu- mors with a very poor prognosis. The mean survival for untreated cases is 2.9 months according to Macfarlane.5 Surgical treatment can be curative for small and localized tumors but most of the cases are encountered in advanced stages. Surgical resection, even when incomplete should be considered, especially in functional tumors to alleviate symptoms.12 Radiation therapy has been attempted post- operatively with a mean survival of 10.3 months in one
series.4 Approaches with nonspecific chemotherapy have been tried with mainly palliative effects. The only drug with some effect in adrenocortical carcinoma is o,p’DDD (mitotane).
In a series of 138 patients, Hutter and Kayhoe found no evidence of cure but a significant reduction in elevated steroid secretion in 69% with a mean duration of 4.8 months, and a significant reduction in tumor mass in 34% with a mean duration of 10 months.13 In 1973, Lubitz et al. described a 54% response rate, however, the duration of life from the start of the treatment only had a mean of 8.4 months.14 Didolkar et al.15 described in 1981 the re- sults with chemotherapy in 28 patients. Objective re- sponses were seen in two patients, both of which had re- ceived o,p’DDD (one complete response with a duration of 18 months). These earlier studies mainly included pa- tients in whom no effort had been made to resect all pos- sible carcinoma.
In 1982, Schteingardt et al.4 reported a mean survival of 46.6 months in 17 patients who had been subjected to aggressive surgery plus o,p’DDD treatment. In his series, the longest survival, 74 months, was found in patients who received adjuvant o,p’DDD before any evidence of metastatic spread of the disease. In the literature there have been a total of eight complete responses to o,p’DDD (one biopsy-proven lasting for 4 years) reported by Boven et al.6 in a review article in 1984.
The observation that streptozocin, an alkylating agent used in gastrointestinal endocrine tumors, was taken up in the adrenal cortex of mice7 encouraged us to try a com- bined regimen of streptozocin and o,p’DDD. There was a significant reduction in tumor size in two cases. In the first patient with the longer treatment, the primary tumor decreased by more than 50% and the lung and lymph
node metastases disappeared. Furthermore, in the excised tumor, there were no viable cancer cells.
In the second case, the adrenal tumor was reduced by slightly less than 50% after 5 months of treatment ac- cording to measurements of the two maximum perpen- dicular diameters on the MRI images. Serial biopsy spec- imens showed increasing necrosis and fibrosis and deple- tion of tumor cells. The decrease in T1 values could reflect decreased tumor cell growth rate. The increase in T2 val- ues in certain areas of the tumor might indicate tumor necrosis. Adrenocortical carcinomas often excrete exces- sive amounts of 36-hydroxy-5-ene steroids,16-18 tetrahy- dro-11-deoxycortisol,16,17 or other steroids, 19,20 which was confirmed in the second patient. Treatment of this patient with streptozocin and o,p’DDD initially increased the 16- oxygenation (Table 1), resulting in steroids seen in new- born infants with a fetal adrenal cortex.21 The normal- ization of S-DHAS after 1 month probably reflects an increased 16-oxygenation and not a decreased synthesis of steroids in the tumor. However, prolonged treatment decreased the total amount of all steroids and after 10 months the patient only excreted trace amounts. The uri- nary steroid profile is normal 20 months after start of treatment.
The first patient has now been followed for more than 6.5 years with no evidence of recurrent disease. Patient 2 recovered from surgery and was reinitiated on cytotoxic treatment which was stopped after 5 months because of liver toxicity. There is no signs of recurrent disease 20 months after start of treatment. In both cases, the com- bined treatment of streptozocin plus o,p’DDD showed objective tumor responses and enabled surgery later on.
In the third patient who received postoperative treat- ment, this combination regimen was unsuccessful and she died from metastatic disease in the lungs and the liver. Perhaps this might indicate that cytotoxic treatment before surgery is more advantageous.
Concerning side effects, the usually reported side effects of streptozocin, i.e., vomiting and renal toxicity occurred, but both were temporary and did not require withdrawal of the drug. As expected, o,p’DDD causes adrenocortical insufficiency, which is easily managed by replacement therapy. Both liver toxicity and hemorrhagic cystitis were noticed. By combining two cytotoxic drugs the required doses can be reduced and thereby the toxicity of the in- dividual drugs.
Because of the small number of patients and the shorter follow-up time in the second patient, one might be cau- tious in the interpretations of these results. We found a clear objective response in two patients and none in the third. However, we report these results in order to stim- ulate further investigations of this therapeutic approach
with the combination regimen of streptozocin and o,p’DDD followed by aggressive surgery in this rare dis- ease, which has earlier been described as incurable. The MRI and urinary steroid profile measurements may be adjuncts in the evaluation of the therapy responses.
REFERENCES
1. Ferber B, Hardy VH, Gerhardt PR, Solomon M. Cancer in New York State, Exclusive of New York City. Bureau of Cancer Control, New York State Department of Health. Albany, 1962.
2. Griswold MH, Wilder CS, Cutler SJ, Pollak ES. Cancer in Con- necticut 1935-1951. Connecticut State Department of Health. Hartford, 1955.
3. Clemmesen J. Statistical studies in the aetiology of malignant neo- plasms: II. Munksgaard, Copenhagen, 1965. Danish Cancer Registry under the National Anti-Cancer League.
4. Schteingardt DE, Motazedi A, Noonan RA et al. Treatment of adrenal carcinomas. Arch Surg 1982; 117:1142.
5. Macfarlane DA. Cancer of the adrenal cortex: The natural history, prognosis and treatment in a study of 55 cases. Ann R Coll Surg Engl 1958; 23:155.
6. Boven E, Vermorken J, van Sloten H, Pinedo H. Complete response of metastasized adrenal cortical carcinoma with o,p’DDD. Cancer 1984; 53:26-29.
7. Tjälve H, Wilander E, Johansson EB. Distribution of labelled streptozocin in mice: Uptake and retention in pancreatic islets. J En- docrinol 1976; 69:455-456.
8. Vestergaard P. Rapid micro-modification of Zimmermann/Callow procedure for determination of 17-ketosteroids in urine. Acta Endocrinol 1951; 8:193-214.
9. Cronholm T, Curstedt T, Schmidt DN, Sjövall J. Steroid profiles in urine and plasma of alcoholics during withdrawal. Alcohol 1985; 2: 677-682.
10. Grimelius L, Wilander E. Silver stains in the study of endocrine cells of the gut and pancreas. Invest Cell Pathol 1980; 3:3-12.
11. Lindgren PG. Ultrasonically guided punctures: A modified tech- nique. Acta Radiol Diagn 1982; 23:653-656.
12. Thompsson NW. Adrenocortical carcinomas. In: Thompsson NW, Vinik AI, eds. Endocrine Surgery Update. New York: Grune and Stratton, 1983.
13. Hutter AM, Kayhoe DE. Adrenal cortical carcinoma: Results of treatment with o,p’DDD in 138 patients. Am J Med 1966; 41:581.
14. Lubitz JA, Freeman L, Okun R. Mitotane use in inoperable ad- renal cortical carcinoma. JAMA 1973; 223:1109.
15. Didolkar MS, Bescher A, Elias G, Moore RH. Natural history of adrenal cortical carcinoma: A clinicopathologic study of 42 patients. Cancer 1984; 47:2153-2161.
16. Drafta D, Franchi F, Luisi M, Stroe E. Urinary, plasma and adrenal tissue steroids in a male patient with Cushing’s syndrome due to adrenal carcinoma, Endocrinology 1978; 16:41-49.
17. Minowada S, Kinoshita K, Hara M, Isurugi K, Uchikawa T, Ni- ijima T. Measurement of urinary steroid profile in patients with adrenal tumor as a screening method for carcinoma. Endocrinol Jpn 1985; 32: 29-37.
18. Honour JW, Price DA, Taylor NF, Marsden HB, Grant DB. Ste- roid biochemistry of virilising adrenal tumours in childhood. Eur J Pediatr 1984; 142:165-169.
19. Watanobe H, Kannari K, Kimura K, Takebe K. Adrenocortical carcinoma with Cushing’s syndrome presenting unusual urinary 17-ke- tosteroid fractionation. J Endocrinol Invest 1985; 8:249-252.
20. Shackleton CHL, Taylor NF, Honour JW. An Atlas of Gas Chro- matographic Profiles of Neutral Urinary Steroids in Health and Disease. Delft, Netherlands: Packard-Becker BV, 1980.
21. Shackleton CHL, Gustafsson J-ÅA, Mitchell FL. Steroids in new- born and infants. Acta Endocrinol 1973; 74:157-167.