Endocrine System

Contents

23.1 Hypothalamic-Pituitary Dysfunction . . . 365

23.1.1 Incidence and Etiology . . . 365

23.2 Growth Hormone Deficiency . . . 366

23.2.1 Treatment . . . 367

23.2.1.1 Investigation . . . 367

23.2.1.2 Growth Hormone Replacement Therapy . . . 367

23.2.2 Prognosis . . . 367

23.3 Hypothalamic-Pituitary-Gonadal Axis . . . 367

23.3.1 Gonadotrophin Deficiency . . . 367

23.3.2 Early or Precocious Puberty . . . 368

23.3.2.1 Treatment . . . 368

23.3.2.2 Prognosis . . . 368

23.4 Thyroid Disorders . . . 369

23.4.1 Treatment . . . 369

23.5 Hypothalamic-Pituitary-Adrenal Axis . . . 369

23.6 Other Pituitary Hormones . . . 370

23.6.1 Fertility . . . 370

23.6.1.1 Testicular Failure . . . 370

23.6.1.2 Ovarian Failure . . . 370

26.6.2 Treatment . . . 370

23.6.2.1 Assessment of Gonadal Function . . . 370

23.6.2.2 Treatment for Gonadal Dysfunction . . . 370

26.6.3 Prognosis . . . 371

References . . . 373

Sklar (1999) suggests that approximately 40% of can- cer patients present with at least one endocrine ab- normality at follow-up. Endocrine disturbances are attributed to location of disease, the type and dosage of chemotherapy, and the amount and schedule of ra- diotherapy (Cohen, 2003).

23.1 Hypothalamic-Pituitary Dysfunction

Table 23.1 summarises the physiology of hypothala- mic-pituitary function.

23.1.1 Incidence and Etiology

The hypothalamic-pituitary axis is central to the con- trol of the endocrine system (Brougham et al., 2002).

Pituitary hormone deficiencies can occur as a result of

▬ Radiation therapy for central nervous system, or- bital, facial, or nasopharyngeal tumours

▬ Radiation therapy for acute lymphoblastic leuk- aemia (ALL)

▬ Total body irradiation pre-bone marrow (stem cell) transplant (Cohen, 2003)

The resultant hormone deficiency depends on sever- al factors, as shown in Table 23.2.

Endocrine System

Deborah Tomlinson · Ethel McNeill

23.2 Growth Hormone Deficiency

Growth hormone (GH) deficiency is the most com- mon endocrine abnormality following cranial radia- tion. The main presentation of GH deficiency in chil- dren is short stature; however, the causes for short stature are not dependent on GH alone. Other treat- ments that inhibit growth include spinal irradiation, glucocorticoids, and chemotherapeutic agents, as well as altered nutrition/inadequate caloric intake.

Incidence rates of the deficiency vary because of these confounding factors.

GH deficiency has also been associated with re- duced lean body mass and increased fat mass, meta- bolic abnormalities (including glucose intolerance), a reduction in bone mineral density, and consequent impaired quality of life.

Growth can be divided into three stages:

1. Infancy – from birth to 2 years, growth is influ- enced by adequate nutrition and is at least partly independent from GH

2. Childhood – GH is the dominant factor and along with adequate nutrition will influence growth

Table 23.1. Hypothalamic releasing hormones and the pituitary trophic hormones (Drury, 1990)

Hypothalamic hormones Pituitary hormones Peripheral hormones

Gonadotrophin-releasing hormone Luteinising hormone (LH) Oestrogens/androgens

(GnRH, LHRH) Follicle stimulating hormone (FSH)

Prolactin inhibiting factor (PIF) Prolactin –

Growth hormone-releasing factor (GHRH) Growth hormone (GH) Somatomedin C (insulin-like

Somatostatin (GHRIH) growth factor) and others

Thyrotropin-releasing hormone (TRH) Thyroid-stimulating hormone (TSH) Thyroxine (T

), triiodothyronine (T

) Both are present as free

and bound forms Corticotrophin-releasing factor (CRF) Adrenocorticotrophic hormone Cortisol

Vasopressin, antidiuretic hormone (ADH) – –

Oxytocin – –

Table 23.2. Factors related to pituitary hormone deficiencies in paediatric oncology (Cohen, 2003, Brougham et al., 2002)

Dependent factor Effect on hypothalamic-pituitary dysfunction

Radiation dose received Directly related. Growth hormone is the most radiosensitive, caused by doses as low as 18 Gy. Increasing radiation doses cause deficiencies in gonadotrophin, corticotrophin, and thyrotropin secretion

Fractionation schedule Inversely related, i.e. therapy administered over more fractions will reduce the degree of dysfunction

Interval since completion of therapy Dysfunction becomes progressively more severe with time since radiation treatment, possibly due to delayed effects of radiotherapy on the axis or to pituitary dysfunction secondary to earlier damage to the hypothalamus.

Evidence indicates that the hypothalamus is more sensitive to radiotherapy than the pituitary

Age at treatment Younger children are more sensitive to radiation-induced damage

of the hypothalamic-pituitary axis compared with older children and adults.

3. Puberty – the sex hormones along with GH and nutrition influence growth until epiphyseal fusion is complete and growth is finished

Consequently, children treated for cancer at an early age are less likely to achieve their expected stature unless they receive GH replacement.

23.2.1 Treatment 23.2.1.1 Investigation

Statural growth should be monitored for 6 months before the fusion of growth plates. If there is any de- cline in growth velocity, formal testing of GH reserve should be performed (Oberfield and Sklar, 2002). Be- fore establishing any hormone treatment, the follow- ing assessment should be made:

1. Accurate measurement of supine length (if under 2 years), standing height, sitting height, and weight 2. Measurement of parents

3. Calculation of decimal age

4. Plotting of growth measurements on appropriate chart

5. Calculation of mid-parental height and target range

6. Calculation of height velocity 7. Pubertal staging

8. Bone age

A GH stimulation test is performed once there is a suspicion of GH deficiency. Numerous tests can be performed, including clonidine, arginine, and insulin tolerance tests. The gold standard test is the insulin tolerance test, which is a safe and reliable test provid- ed that a strict procedure is followed (Galloway et al., 2002). Pituitary function tests used to diagnose GH deficiency rely on these pharmacological provoca- tion tests. However, although peak responses to these tests may remain normal, the physiologic secretion of pituitary hormones is often impaired (neurosecreto- ry dysfunction). The assessment of physiological se- cretion is difficult, which emphasises the importance of continued long-term follow-up.

The insulin test should only be performed in pae- diatric endocrine units and in children over 5 years of age.

23.2.1.2 Growth Hormone Replacement Therapy

Once it is confirmed that the child is GH-deficient and is 2 years post-cancer treatment, replacement with recombinant GH therapy can be advocated.

Somatropin (synthetic human growth hormone) is given in the form of daily subcutaneous injections.

The recommended dose is 5 mg/m

/week.

GH replacement therapy is normally discontinued once the patient’s final height has been achieved.

However, consideration needs to be given to the con- tinuation of GH replacement beyond this time in or- der to avoid the other adverse consequences associat- ed with GH deficiency mentioned previously (Glee- son et al., 2002). Therefore, replacement therapy may continue, at a reduced dose, into adulthood (Broug- ham et al., 2002).

23.2.2 Prognosis

Growth hormone is potentially mitogenic, and there have been numerous clinical studies investigating any evidence of recurrence of disease in the children treated with GH. There has been no evidence found to support that there is a significant risk. It is there- fore thought that GH treatment benefits most GH-de- ficient children. Oberfield and Sklar (2002) observe the importance of tracking adults who have survived childhood cancers and who are now starting to re- ceive GH as part of the newly suggested lower-dose GH replacement regimens for adults.

23.3 Hypothalamic-Pituitary-Gonadal Axis 23.3.1 Gonadotrophin Deficiency

Higher doses of cranial irradiation (>35 Gy) can damage the hypothalamic-pituitary axis, with a result in gonadotrophin secretion deficiency (Cohen, 2003).

Patients have demonstrated deficiencies in the secre- tion of

▬ Follicle stimulating hormone (FSH)

▬ Luteinising hormone (LH)

Clinically, gonadotrophin deficiency can exhibit a broad spectrum of severity, from subclinical abnor- malities, which are detectable by gonadotrophin re- leasing hormone (GnRH) testing, to a significant re- duction in circulating sex hormone levels and de- layed puberty (Brougham et al., 2002). The etiology of hypogonadism following cranial irradiation is often hypothalamic GnRH deficiency. Exogenous GnRH replacement therapy may possibly restore go- nadal function and fertility.

23.3.2 Early or Precocious Puberty

In contrast, lower doses of cranial irradiation can cause premature activation of the hypothalamic-pi- tuitary-gonadal axis. This can lead to early or preco- cious puberty. True central precocious puberty oc- curs before the age of 8 years in girls and before 9 years in boys. Early puberty is defined as onset be- tween ages 8 and 10 in girls (with the development of pubic hair) and between 9 and 11 in boys (using tes- ticular enlargement). The etiology of radiation-in- duced early puberty is thought to be through the dis- inhibition of cortical influences on the hypothalamus (Brougham et al., 2002). The incidence of radiation- induced early puberty is dependent on dose. The dose threshold is then gender-specific; low doses of cranial radiation, 18–24 Gy (used in ALL treatment), may cause early puberty predominately in girls, but higher doses of 25–47 Gy (used to treat brain tu- mours) affects both genders equally (Cohen, 2002).

Additionally, a patient may enter puberty early but subsequently develop gonadotrophin deficiency, sug- gesting the differential effects of radiotherapy with time (Brougham et al., 2002).

Entering puberty early causes a premature growth spurt that is followed by early epiphyseal fusion with a consequential reduction in final height. Puberty is usually of normal duration, but early puberty is often combined with GH deficiency, which further reduces the final height potential.

23.3.2.1 Treatment

These patients also require clinical assessment with regard to their growth and puberty. This assessment includes

▬ Pubertal status

▬ Auxological assessment, including standing and sitting heights

▬ Biochemical assessment of growth hormone and gonadotrophin secretion

▬ Radiological assessment of bone age

It is advantageous to suppress puberty by using a GnRH analogue, e.g. Zoladex LA 10.8 mg every 10–12 weeks. In this situation, combined treatment of GH with GnRH analogue will improve the height poten- tial. To achieve improved auxological outcome, Glee- son et al. (2003) recommend

1. The use of standardised GH schedules and better dosing regimens

2. A reduction in the time interval between finishing radiotherapy and starting GH replacement 3. The use of GnRH analogue in addition to GH re-

placement in carefully selected patients 23.3.2.2 Prognosis

Despite treatment, final height obtained is generally lower than the target height.Also, growth is often dis- proportionate due to a reduction in sitting height.

This is due to disruption to spinal growth caused by

▬ Early onset of puberty and GH deficiency (spinal growth is significant in the latter part of the pu- bertal growth spurt)

▬ Spinal irradiation

23.4 Thyroid Disorders

Thyroid disorders can occur following treatment for childhood cancer due to

▬ Disruption of the hypothalamic-pituitary-thyroid axis

▬ Direct damage to the thyroid gland

The thyroid gland is radiosensitive and can be affect- ed by craniospinal radiation and total body irradia- tion, as well as treatments with iodine radiolabeled monoclonal antibodies, such as I

-MIBG for neu- roblastoma.

Disorders manifest as

▬ Dysfunction of the thyroid gland – Hypothy- roidism is the most common abnormality follow- ing direct thyroid damage. Radiation-induced thy- roid stimulating hormone (TSH) deficiency is dose-dependent and uncommon with doses less than 40 Gy in children. Hyperthyroidism is less common but can occur, particularly when patients receive higher doses.

▬ Thyroid nodules – Gleeson et al. (2002) report a varying incidence of thyroid nodules, 2–65%, in children and adolescents treated for Hodgkin’s disease.

▬ Slight risk of secondary thyroid function – Papil- lary carcinoma is the most common thyroid cancer that develops secondary to irradiation (Brougham et al., 2002).

Direct damage to the thyroid is usually secondary to radiotherapy in which the radiation field includes the neck. Chemotherapy has been reported to potentiate this damage; however, van Santen and colleagues (2003) have concluded that chemotherapy does not contribute to damage on the thyroid axis.

23.4.1 Treatment

Biochemical diagnosis of central hypothyroidism re- lies on measurement of TSH and thyroid hormone as free T

. Primary hypothyroidism results in an elevat- ed TSH and a low free T

; compensated hypothy- roidism is indicated by an elevated TSH and a normal

free T

. However, TSH and free T

may both fall with- in normal range, but more detailed investigation of TSH dynamics, including thyrotrophin secretion, can suggest significant central hypothyroidism. This detail may be important when deciding thresholds for treatment with thyroxine supplements and may be particularly important in paediatric patients be- cause reduced thyroid function may affect growth (Brougham et al., 2002).

It is advisable that thyroid function be assessed at least twice a year following treatment. The greatest risk of hypothyroidism occurs during the first 5 years after treatment, but new cases continue to emerge more than 20 years after irradiation (Gleeson et al., 2002). If treatment is necessary, thyroxin is pre- scribed orally in a dose of 100 mcg/m

to fully sup- press and replace endogenous thyroid function.

23.5 Hypothalamic-Pituitary-Adrenal Axis

The hypothalamic-pituitary-adrenal axis appears to be relatively radio-resistant, and adrenal insufficien- cy due to radiotherapy or cranial tumour is rare.

Treatment doses of glucocorticoids may suppress the hypothalamic-pituitary-adrenal axis. Dexametha- sone at a dosage of 6 mg/m

/day for 28–42 days can mildly suppress the axis for up to 4–8 weeks (Cohen, 2002; Peterson et al., 2003). However, clinical manifes- tation of adrenal insufficiency in these cases is rare.

Deficiency in adrenocorticotrophic hormone (ACTH) can develop in patients treated with radia- tion doses in excess of 50 Gy and in patients treated for pituitary tumours (Brougham et al., 2002). Exces- sive tiredness in patients treated with cranial radia- tion warrants testing of the hypothalamic-pituitary- adrenal axis. Aldosterone is usually unaffected be- cause secretion depends more on angiotensin II than on ACTH.

Diagnosis is difficult, and the insulin tolerance test

is usually the assessment of choice, as with GH secre-

tion. Treatments of high-dose glucocorticoids should

be tapered. Hydrocortisone replacement may be nec-

essary, and increased doses are required during peri-

ods of illness and surgery (Brougham et al., 2002).

23.6 Other Pituitary Hormones

Prolactin secretion is inhibited by dopamine from the hypothalamus. High-dose cranial radiation, especially >50 Gy, can disrupt this inhibitory control and result in hyperprolactinemia (Cohen, 2002;

Brougham et al., 2002). However, this is unusual in children.

Antidiuretic hormone (ADH) deficiency causing central diabetes insipidus has not been attributed to cranial irradiation. However, central diabetes in- sipidus has been associated with intracranial tu- mours involving the pituitary gland following cranial surgery or secondary to leukemic infiltration. The de- ficiency in these cases usually persists despite treat- ment of the underlying malignancy (Brougham et al., 2002) and may require treatment with exogenous va- sopressin.

The syndrome of inappropriate ADH secretion (SIADH) with water retention and secondary hy- ponatremia has been associated with malignancy and its treatment. The associated malignancies are those more commonly seen in adults, including gas- trointestinal tumours, breast and prostatic cancers, and primary brain tumours. However, treatments used for adults and children that can cause SIADH include the vinca alkaloids, cisplatin, cyclophos- phamide, and melphalan. Other aspects of treatment associated with SIADH include intrathoracic infec- tion and artificial ventilation. Fortunately, SIADH is only a problem for the short term and is rarely a long- term complication (Brougham et al., 2002).

23.6.1 Fertility

Radiotherapy involving the pelvic area or systemic chemotherapy can cause direct damage to gonadal tissue (testes and ovaries). This damage can result in subfertility or infertility in both males and females.

23.6.1.1 Testicular Failure

The adult male testes has two functions:

▬ Steroidogenesis

▬ Spermatogenesis

Table 23.3 outlines the physiology of the testes and the effects of radiotherapy and chemotherapy on their function. The mechanism that makes the testes susceptible to the toxic effects of radiation and chemotherapy appears to involve the combination of destruction of the germ cell pool and, when the germ cells survive, inhibition of further differentiation (Thomson et al., 2002).

23.6.1.2 Ovarian Failure

Ovarian function is very complex. Table 23.4 outlines the main functions and the consequential effects of radiotherapy and chemotherapy.

Additionally, radiotherapy involving the uterus in childhood is associated with

▬ Increased incidence of nulliparity

▬ Spontaneous miscarriage

▬ Intrauterine growth retardation with decreased uterine volume and decreased elasticity

Obstetricians must be aware of potential problems.

26.6.2 Treatment

23.6.2.1 Assessment of Gonadal Function Table 23.5 summarises the patient history and inves- tigations necessary when assessing gonadal function in males and females.

23.6.2.2 Treatment for Gonadal Dysfunction

Oestrogen replacement in girls starts at a small dose

of ethinyl oestradiol 2 mcg, increasing in small incre-

ments over 2 years until full establishment of puber-

ty. A combined pill of oestrogen and progesterone

can then be given, e.g. Loestrin 30.

Boys will start with 100 mg Sustanon every 6 weeks increasing to every 4 weeks until puberty is es- tablished.

26.6.3 Prognosis

Concerns that the offspring of patients successfully treated for cancer might have an increased risk of congenital abnormality and childhood cancer have not been substantiated (Thomson et al., 2002).

Techniques in preserving fertility continue to be developed. Established options include cryop- reservation of spermatozoa and collection of mature oocytes with fertilisation and subsequent cryop- reservation of embryos in the female with a partner.

Options are limited in children. Experimental strate- gies are focusing on the harvesting and storage of go- nadal tissue, cryopreservation of immature sper-

matogenic cells or oocytes, gonadotrophin suppres- sion, and inhibition of follicle apoptosis (Thomson et al., 2002). Following cure, stored tissue could possibly be autotransplanted or stored tissue could be matured in vitro until sufficiently mature for fertili- sation with assisted reproductive techniques (Broug- ham et al., 2002). Human primordial follicles survive cryopreservation, and the return of ovarian hormon- al activity has been achieved after reimplantation;

however, no pregnancies have been reported (Thom- son et al., 2002). The potential developments raise many ethical and legal issues that must be addressed to ensure adequate regulation. Children with cancer must be ensured realistic, safe prospects for fertility in the future, and patients at risk of subfertility re- quire appropriate counselling as part of their routine care.

Table 23.3. Outline of the function of the testes and the associated effects of radiotherapy and chemotherapy (Drury,1990;

Cohen, 2002; Brougham et al., 2002)

Function Associated Target cells Action Effects of Effects of

hormones radiotherapy chemotherapy

Steroido- GnRH released Leydig’s cells Testosterone acts Leydig’s cells are more Similar to radiotherapy genesis from hypothala- of somatic systemically to resistant to damage. effects. Leydig’s cells

mus stimulates cells of testes produce male Damage occurs at are less sensitive and luteinising hor- to produce secondary sexual doses of 20 Gy in pre- may be unaffected.

mone (LH) testosterone characteristics, pubertal boys and up Higher doses may from the anabolism, and the to 30 Gy in sexually cause Leydig cell dys-

pituitary maintenance of mature males. Second- function

libido (Testosterone ary sexual characteris- feeds back on hypo- tics may develop thalamus/pituitary despite impaired sper- to inhibit GnRH matogenesis

secretion)

Spermato- GnRH released Sertoli’s cells Produces mature Damage depends on Extent of damage genesis from hypothala- in germinal sperm (and feedback field of treatment, depends on agent

mus stimulates epithelium hormone inhibin total dose, and frac- administered and dose follicle stimu- of the that appears to feed tionation schedule. received.

Gonado- lating hormone seminiferous back on pituitary to Doses as low as toxic chemotherapy (FSH) tubules decrease FSH secre- 0.1 Gy can affect sper- can cause oligosper- tion.) Testosterone matogenesis, with mia or azoospermia also acts locally to temporary azoospermia

aid spermatogenesis and oligospermia.

Doses over 4 Gy may have a permanent effect

a

Gonadotoxic agents include cisplatin, procarbazine, cytarabine, vinblastine and alkylating agents such as cyclophosphamide,

melphalan, chlorambucil

Table 23.4. Outline of the function of the ovaries and the associated effects of radiotherapy and chemotherapy (Drury, 1990;

Cohen, 2002; Brougham et al., 2002)

Function Associated Target cells Action Effects of Effects of

hormones radiotherapy chemotherapy

Oestrogen Pulses of LH stimulates Oestrogen initially, Ovaries may be in Prepubertal ovaries and pro- GnRH stimu- ovarian androgen then progesterone, the field of pelvic, appear to be more gesterone late release production cause uterine abdominal or spinal resistant to cytotoxic production of pituitary (mid-cycle LH endometrial proli- radiation. Degree of agents than post-

LH and FSH surge induces feration in prepara- impairment depends pubertal, possibly ovulation). tion for possible on dose, fractionation because they have

implantation; schedule, and age at more follicles.

FSH stimulates if implantation time of treatment. Ovarian function is follicular develop- does not occur, often retained or ment and aroma- corpus luteum The human oocyte is recovered with tase activity regresses, progeste- highly sensitive to standard doses of (Follicle differen- rone secretion radiation, with 50 % chemotherapy.

tiates into corpus falls, and menstrua- death of oocytes at High doses of alkylat- luteum and tion occurs. a dose of 4 Gy. The ing agents or secretes progeste- (Pregnancy causes younger the girl at myeloablative rone and oestra- human chorionic the time of radio- therapy for bone diol during 2nd gonadotrophin therapy, the larger marrow transplant are half of cycle; production from the oocyte pool and likely to result in aromatase is an corpus luteum hence the later onset permanent ovarian enzyme required until 10–12 weeks. of premature meno- failure

to convert ovarian Placenta then pause. Premature androgens to makes sufficient menopause can oestrogen). oestrogen and occur in women

progesterone). >40 years of age fol-

FSH stimulates lowing treatment

inhibin from Oestrogens induce dose of 6 Gy. (Early ovarian stromal secondary sexual menopause has impli- cells (feedback characteristics cations for fertility, of inhibin inhibits but also other medical

FSH release) complications includ-

ing osteoporosis).

Doses >20 Gy can cause complete ovarian failure.

Table 23.5. Assessment of gonadal function

Females – Males –

ovarian function testicular function

Pubertal staging Pubertal staging Menstrual history Testicular volume using

Prader orchidometer

Basal LH and FSH Basal LH, FSH, and inhibin B

Basal oestradiol Basal testosterone

Pelvic ultrasound Semen analysis

References

Brougham MFH, Kelnar CJH,Wallace WHB (2002) The late en- docrine effects of childhood cancer. Pediatric Rehabilita- tion 5(4): 191–201

Cohen LE (2003) Endocrine late effects of cancer treatment.

Current Opinion in Pediatrics 15: 3–9.

Drury PL (1990) Endocrinology. In Kumar PJ, Clark ML Clini- cal Medicine, 2nd edn. Balliere Tindall, London

Galloway PJ, McNeill E, Paterson WF, Donaldson (2002) Safety of the insulin tolerance test. Archives of Disease in Child- hood 87(4): 354–356

Gleeson HK, Darzy K, Shalet SM (2002) Late endocrine, meta- bolic and skeletal sequelae following treatment of child- hood cancer. Best Practice and Research Clinical En- docrinology and Metabolism 16(2): 335–348

Gleeson HK, Stoeter R, Ogilvy-Stuart AL, Gattamaneni HR, Brennan BM, Shalet SM (2003) Improvements in final height over 25 years in growth hormone (GH)-deficient childhood survivors of brain tumors receiving GH replace- ment. Journal of Clinical Endocrinology and Metabolism 88(8): 3682–3689

Oberfield SE, Sklar CA (2002) Endocrine sequelae in survivors of childhood cancer. Adolescent Medicine 13(1): 161–169 Petersen KB, Muller J, Rasmussen M, Schmiegelow K (2003)

Impaired adrenal function after glucocorticoid therapy in children with acute lymphoblastic leukemia. Medical Pedi- atric Oncology 41(2): 110–114

Sklar CA (1999) Overview of the effects of cancer therapies: the nature, scale and breadth of the problem. Acta Paediatrica (supplement) 88: 1–4

Thomson AB, Critchley HOD, Kelnar CJH, Wallace WHB (2002) Late reproductive sequelae following treatment of childhood cancer and options for fertility preservation.

Best Practice and Research Clinical Endocrinology and Me- tabolism 16(2):311–334

Van Santen HM, Vulsma T, Dijkgraaf MG, Blumer RM, Heinen

R, Jaspers Mw, Geenen MM, Offringa MO, de Vijlder JJ, van

den Bos C (2003) No damaging effect of chemotherapy in

addition to radiotherapy on the thyroid axis in young adult

survivors of childhood cancer. Journal of Clinical Endo-

crinology Metabolism 88(8): 3657–3663