# Canine Cushing's syndrome

Canine Cushing's syndrome - named after Harvey Cushing, who first described disease in humans - or canine hyperadrenocorticism (from ancient Greek ὑπέρ hyper, and Latin cortex adrenalis adrenal cortex) is a common hormonal disease of the domestic dog (Canis lupus familiaris). It is characterized by high blood levels of glucocorticoids, especially cortisol produced in the adrenal gland (hypercortisolism). The cause is usually tumors of the pituitary gland, which release an excess of the hormone ACTH, which controls the adrenal gland, and thus trigger an overproduction of cortisol. Cushing's syndrome occurs even more frequently due to the administration of synthetic glucocorticoids such as prednisolone or dexamethasone. These are frequently used in veterinary medicine to treat allergys, autoimmune diseases or inflammations and can trigger the same clinical picture with long-term use (iatrogenic Cushing's syndrome).
Typical symptoms of the disease are increased drinking, increased urine output, increased appetite, pot belly, thin skin, muscle wasting and susceptibility to infections. Various laboratory tests can be used to confirm the diagnosis, but a definitive diagnosis is not always possible. To treat iatrogenic Cushing's syndrome, the synthetic glucocorticoid is gradually discontinued. In other forms of Cushing's disease, the drug Trilostane is usually used, which suppresses the formation of cortisol via enzyme inhibition. Surgical removal of the tumors causing the disease is also possible, but is performed much less frequently due to the frequent complications.

## Basics

The adrenal gland is a small, paired organ of the endocrine system. It lies in front of the respective kidney and is about 20 × 5 mm in size in dogs. In the outer layer, the adrenal cortex, glucocorticoids such as cortisol and corticosterone are produced by zona faciolata cells arranged like strands. Cortisol production is controlled via the hypothalamic-pituitary-adrenal axis. When required, the hypothalamus corticotropin-releasing hormone (CRH), which stimulates the production of adrenocorticotropin in the pituitary gland (ACTH) in the pituitary gland. High levels of cortisol inhibit the formation and release of the higher-level control hormones CRH and ACTH (feedback inhibition). Cortisol can be reversibly converted into the inactive form cortisone by oxidation or hydrogenation at the double bond between the fourth and fifth carbon atom by liver enzymes and thus irreversibly inactivated.
Cortisol has numerous effects in the body. It binds to specific receptors in the cell nucleus and stimulates the formation of new glucose (gluconeogenesis) in the liver, inhibits protein biosynthesis in muscle and connective tissue, increases protein breakdown (proteolysis), increases fat breakdown (lipolysis), suppresses the immune response and inhibits wound healing.

## Pathogenesis and occurrence

In principle, five causes can be distinguished:
* Treatment with synthetic glucocorticoids (iatrogenic Cushing's syndrome)
* Spontanes Cushing-Syndrom
** tumors of the pituitary gland (secondary hyperadrenocorticism)
** Tumors of the adrenal gland (primary hyperadrenocorticism)
** Ectopic ACTH syndrome
** Feeding-dependent Cushing's syndrome
The most common cause of Cushing's syndrome is treatment with synthetic glucocorticoids (iatrogenic, i.e. caused by the veterinarian). Prednisolone or dexamethasone are often used to treat allergys, autoimmune diseases or inflammations. The longer glucocorticoids are used, the more frequently clinical pictures such as Cushing's syndrome or diabetes occur as side effects, as these active substances lead to an inhibition of the hypothalamic-pituitary-adrenal axis.
Spontaneous Cushing's syndrome is caused in 80 to 85 % of cases by cell proliferation of the ACTH-producing cells of the anterior pituitary, usually a tumor of this part of the organ. The cells produce an increased amount of ACTH, which stimulates the cells of the adrenal cortex to produce an excessive amount of cortisol. In dogs, benign tumors (adenomas) predominate and almost always only affect the ACTH-producing cells of the pituitary gland. In around 15 to 20 % of cases, tumors (adenomas and carcinomas) of the adrenal cortex itself are responsible for the increased cortisol production. The other two forms are very rare. Ectopic ACTH syndrome is caused by ACTH-producing tumors outside the pituitary gland, usually malignant tumors of the diffuse neuroendocrine system, whose cells produce an increased amount of this hormone. Feeding-dependent Cushing's syndrome occurs in animals that develop defective receptors in the adrenal cortex, especially for the glucose-dependent insulinotropic peptide. In these animals, there is a pathological increase in cortisol production after each food intake.
A British study from 2022 determined an incidence of 0.06% for spontaneous Cushing's syndrome. The disease occurs mainly in older animals (> 7 years). Most commonly affected are Bichon Frisé, Border Terrier and Dwarf Schnauzer. Yorkshire Terriers, Jack Russell Terriers and Dachshunds are also affected more frequently. For other dog breeds such as Fox Terrier, Cavalier King Charles Spaniel, Boxer, Shih Tzu, Poodle, Irish Setter and Basset Hound, a breed-related accumulation has been described, although this information is based on relatively small random samples. Dog breeds such as Golden Retriever, Labrador Retriever, Border Collie and Cocker Spaniel, on the other hand, appear to suffer from Cushing's syndrome much less frequently. The increased incidence described in the literature in spayed/neutered female dogs described in the literature could not be confirmed by the British study. Whether there is a connection between sex hormones and the disease is also disputed among experts and is currently considered unlikely.
The first description of the disease in humans was made in 1912 by the US neurosurgeon Harvey Cushing. Two years earlier, Cushing had already presented an individual case report. In dogs, Cushing's syndrome was first described in 1939 by Verstraete and Thoonen.

## Clinical picture

The most common symptoms of Cushing's syndrome are increased drinking (polydipsia), increased urine output (polyuria), greatly increased appetite with binge eating (polyphagia), pendulous belly, thinning of the skin (skin atrophy) with hair loss and skin calcifications (calcinosis cutis) as well as muscle atrophy and increased susceptibility to infections. The increased urine output is presumably mediated by the cortisol-induced inhibition of adiuretin secretion and the increase in glomerular filtration rate and the resulting fluid loss is compensated for by increased drinking. The binge eating results from the direct stimulation of the hunger center in the hypothalamus, and the release of leptin may also be inhibited. The pendulous abdomen is caused by atrophy of the abdominal muscles, an increase in fat in the abdomen, enlargement of the liver (hepatomegaly) and the often very full urinary bladder. Mild forms of Cushing's syndrome often only manifest themselves in skin changes with susceptibility to infection in the form of skin fungal disease, purulent skin inflammation or generalized hair follicle mite disease (demodicosis). (demodicosis).
* Sagging belly and hair loss
* Atrophie
* Calcification of the skin (calcinosis cutis)
* Generalized demodicosis
Other signs of the disease include fatigue, panting and high blood pressure. As cortisol interacts with numerous other components of the hormonal system, diabetes, diurethritis, secondary underactive thyroid, secondary overactive parathyroid and reproductive disorders can occur as concomitant diseases. In addition to kidney disease, these are also the most important Differential diagnoses.

## Diagnostic

When fully developed, the clinical picture is already strongly indicative of the disease. However, the diagnosis must be confirmed by laboratory tests. However, there is no test that can reliably prove or rule out Cushing's syndrome, so there are always false positives or false negatives laboratory findings. The laboratory results must therefore always be considered in the context of the clinical picture and several tests may need to be combined.

### Laboratory findings

The blood count very often shows a stress leukogram due to the influence of cortisol on the blood cells. A stress leukogram is characterized by an increase in the number of white blood cells (leukocytosis), especially neutrophil granulocytes (neutrophilia), as well as the decrease in lymphocytes (lymphopenia) and eosinophils (eosinopenia). In addition, the number of red blood cells is usually moderately increased (erythrocytosis). The influence of high cortisol levels leads to activation of the liver enzymes, storage of glycogen and thus to enlargement of the liver cells and the liver as a whole ("steroid liver"). The blood chemistry examination is therefore dominated by elevated liver values, especially alkaline phosphatase, which is significantly elevated in 85 to 90 % of affected dogs. Cholesterol and blood lipids are also often elevated.
In the urine examination, the greatly reduced relative density is particularly noticeable. As cortisol also favors bladder infections, the typical changes (pyuria, bacteriuria) can also occur secondarily.

### Specific diagnostics

The gold standard for spontaneous Cushing's syndrome is the low-dose dexamethasone suppression test (LDDS), but it is not suitable for detecting iatrogenic hyperadrenocorticism. For the test, a blood sample is taken in the morning and the dog is then injected with a low dose of dexamethasone. Four and eight hours later, blood is taken again and cortisol is determined in all three samples. In healthy dogs, the dexamethasone would have the effect of feedback inhibition (see above) would inhibit ACTH and thus cortisol secretion. If the cortisol level is above 39 nmol/l (1.4 µg/dl) after 8 hours, this is a strong indication of Cushing's syndrome; if it is below 28 nmol/l (1 µg/dl), it is unlikely. The 4-hour value is used to differentiate between primary and secondary hyperadrenocorticism. In secondary hyperadrenocorticism, i.e. tumors of the pituitary gland, there is a reduction in ACTH secretion and thus a drop in cortisol secretion in around 60 % of animals in the first few hours; this suppression does not occur in animals with an adrenal tumor. The Sensitivity of the LDDS is 97 %, the Specificity is 70 %. The high-dose dexamethasone suppression test (HDDS) can also be used to differentiate between primary and secondary hyperadrenocorticism. For this test, five times the amount of dexamethasone is administered compared to the LDDS. In the case of primary hyperadrenocorticism, cortisol formation is hardly suppressed with this test either. There are occasional cases in which the clinical findings suggest Cushing's syndrome but the suppression tests show normal findings. These are referred to as atypical hyperadrenocorticism.
In the ACTH stimulation test, the blood level of cortisol is determined, ACTH is injected and the cortisol level in the blood is measured again after one hour. This test is of limited suitability for the diagnosis of spontaneous Cushing's syndrome, but it is the only test that can be used to detect iatrogenic Cushing's syndrome. Stimulated cortisol levels below 138 nmol/l are indicative of adrenocortical insufficiency or iatrogenic Cushing's syndrome, levels between 138 and 497 nmol/l are physiological, levels between 497 and 662 nmol/l are suspicious and levels above this are highly suspicious of spontaneous Cushing's syndrome. For atypical Cushing's cases, it is also suggested not to determine the cortisol itself, but its precursor molecules in the synthesis pathway (sex hormones such as 17α-hydroxyprogesterone, progesterone, estradiol and androstenedione).
The determination of cortisol in the blood does not provide any information about the disease. Dogs with Cushing's syndrome have statistically higher blood cortisol levels than healthy dogs, but as cortisol is released irregularly (pulsatile), stressful situations lead to increased cortisol levels and there are considerable individual differences, individual determinations are not suitable for diagnosis. The determination in a 24-hour urine sample is somewhat more meaningful, as this compensates for diurnal fluctuations. Ideally, urine from two consecutive days is collected and cortisol is considered in relation to creatinine (cortisol-creatinine quotient). The sensitivity is almost 100%, meaning that all dogs with Cushing's syndrome are detected, but the specificity is low (21%), there are also many other causes for an increased cortisol-creatinine quotient. Even the size of the dog has an influence, as creatinine excretion is dependent on muscle mass. The daily fluctuations in cortisol production can also be compensated for by taking blood samples every hour for eight hours; dogs with Cushing's syndrome generally have higher cortisol concentrations than healthy dogs.
Determining the endogenous (body's own) ACTH concentration in blood plasma is another test procedure. In primary Cushing's syndrome, ACTH is usually low (< 4.4 pmol/l) as a result of feedback inhibition; in secondary Cushing's syndrome, it is usually high (> 8.8 pmol/l). However, this test does not provide clear results in around 20 % of cases. In addition, this test is more expensive and logistically more complex, as the plasma must be transported to the laboratory frozen.

### Imaging diagnostics

To differentiate between primary and secondary hyperadrenocorticism, ultrasound examination (sonography) of the adrenal gland. This examination technique is now widely available in veterinary medicine and can detect adrenal tumors relatively reliably. In general, the adrenal gland is also enlarged in primary Cushing's patients, as the increased hormone production leads to an enlargement of the organ. A transverse diameter of more than 6 mm is suspicious for hyperadrenocorticism, the sensitivity is 75 %, the specificity 94 %. Ultrasound can also be used to detect the typical "steroid liver", in which the liver tissue is diffusely echoic. However, differentiation from other diseases with this sonographic finding is only possible by means of a liver biopsy and subsequent histological examination.
Pituitary tumors, which are much more common as a cause of disease, could be detected with magnetic resonance imaging or computed tomography, but this is only available in a few veterinary facilities and can only be performed under anesthesia. In addition, these methods can only detect tumors of a certain size. However, they are absolutely essential for planning surgical tumor removal.
* X-ray image of calcinosis cutis in the back area of a dog
* Ultrasound image of an enlarged left adrenal gland in pituitary Cushing's syndrome
* Ultrasound image of a tumorously altered adrenal gland in a dwarf spitz
* X-ray of a carcinoma of the right adrenal gland (left in the picture) of a dog, which is recognizable by calcification at the level of the last rib.
* Ultrasound image of a liver altered in the course of Cushing's syndrome in a dog: steroid hepatopathy

### Consensus paper on diagnostics

In 2013, leading experts in the field of canine and feline endocrinology from the American College of Veterinary Internal Medicine (ACVIM) agreed on standardized criteria for the diagnosis of canine Cushing's syndrome. In this consensus paper (consensus statement), it is recommended that laboratory diagnostics should not be initiated if there is no clinical suspicion of the disease. If only a few symptoms are present, then it is either increased drinking and urination or hair loss and skin changes that should raise suspicion of this disease. A poor response to insulin diabetes mellitus or persistent hypertension would also be indications for a specific Cushing's diagnosis, whereas mere changes in blood count and blood chemistry would not. None of the tests is preferred, except in cases of suspected iatrogenic Cushing's syndrome, in which case only the ACTH stimulation test is useful. If a test shows a negative result despite clinical suspicion of spontaneous Cushing's syndrome, alternative tests should be performed. If the results are questionable, the tests should be repeated after 3 to 6 months. If other serious illnesses are present, testing is not very useful, as these can falsify the results of the Cushing's tests. Imaging diagnostics alone are not suitable for diagnosis. If atypical Cushing's syndrome is suspected, in which the involvement of sex hormones has been postulated by some scientists, their determination is nevertheless not considered very useful, as the specificity is very low.

## Treatment

The most common form, iatrogenic hyperadrenocorticism, is the easiest to treat, namely by phasing out glucocorticoid therapy. Abrupt discontinuation would lead to hypoadrenocorticism (Addison's disease), as the adrenal glands need some time to produce sufficient glucocorticoids again after prolonged suppression of hormone production. Spontaneous Cushing's syndrome is usually treated conservative by administering medication. Since tumors are often the trigger, this does not eliminate the cause of the disease, but it does restore a reasonable quality of life. Surgical or radiotherapy treatment of tumours is associated with some risks, is not suitable for every patient and can only be carried out in specialized facilities.

### Drug treatment

The only active substance approved for the treatment of canine Cushing's syndrome is trilostane. Trilostane causes the formation of cortisol via reversible inhibition of 3β-hydroxysteroid dehydrogenase. Prior to the introduction of Trilostane in 2001 by Dechra Pharmaceuticals, Mitotane was also used, which selectively induces death of the glucocorticoid-producing cells in the adrenal cortex. Treatment must be monitored in any case, because if the dose is too high, adrenal hypofunction (Addison's disease) can be triggered. This manifests itself primarily in fatigue, loss of appetite, weakness, reduced performance, weight loss and vomiting. However, the symptoms quickly disappear again when the trilostane is discontinued or the dose is reduced. A study from 2022 showed that a lower starting dose of 0.2-1.1 mg/kg twice daily than the 3-6 mg/kg once daily recommended by the manufacturer slightly improved the survival time. Age, calcinosis cutis, a body condition score <3 and an elevated platelet count (thrombocythemia) have a negative effect on survival time.
The ACTH stimulation test was long regarded as the gold standard for monitoring and dose adjustment of treatment with Trilostane. The stimulation value should be between 6 and 18 µg cortisol/dl if the drug is well adjusted. As ACTH was no longer produced for some time, this test was no longer possible and alternatives to trilostane monitoring were sought. The serum cortisol level before Trilostane administration (prepill), the ACTH plasma concentration, the serum haptoglobin concentration, the alanine aminotransferase (ALT), the Γ-glutamyltransferase (γGT) and alkaline phosphatase, the specific density of urine and the cortisol creatinine quotient in urine are tested as possible markers. Haptoglobin is best suited for therapy monitoring; in a study from 2021, 90% of well-controlled dogs and two-thirds of under-treated dogs could be detected with it.
In addition to the blood chemistry check, it is important for the dog owner to monitor the dog's condition. The manufacturer of Trilostan has developed two questionnaires with the Royal Veterinary College London for this purpose. One is used to record drinking behavior and urine output, appetite, behavior and activity, while the second is used to assess quality of life.
A new therapeutic approach in the experimental stage is the administration of ATR-101, a selective inhibitor of the enzyme ACAT1 and thus of ACTH production, which has also shown promising results in dogs.

### Surgical treatment

Since in the majority of cases spontaneous hyperadrenocorticism is caused by tumors, surgical removal of the triggering tumor initially appears to be the most obvious treatment option. Several studies are available for unilateral adrenal tumors. These show that some tumors were inoperable, there can be numerous complications leading to euthanasia, and in some cases the increased cortisol secretion persisted or recurred after some time. Surgical treatment is therefore only the treatment of choice for unilateral adrenal tumors without metastasiss and good differentiation from the surrounding tissue. The removal of a tumor of the pituitary gland is a very difficult operation with numerous risks. The mortality rate is between 8 and 19 %. However, since sufficient hormone-producing pituitary cells often remain in the body when a pituitary tumor is removed, this is an effective therapeutic option.

### Radiotherapy

The effectiveness of radiation of pituitary tumors has so far only been investigated in small numbers of cases. A study on 45 dogs determined an average survival time of 311 days. Clinical improvement occurred in 82% of the animals, 22% of the dogs showed acute side effects. In 69% of the animals, neurological deficits increased in the late phase.

## Cushing's syndrome in humans and other mammals

As the effects of cortisol are largely the same in all mammals, the clinical picture and laboratory findings are also very similar in all of them. There are also hardly any differences in diagnostics. In humans, pituitary hyperadrenocorticism is sometimes conceptually differentiated from adrenal Cushing's disease as Cushing's syndrome. However, with 1 to 3 new cases per 1 million inhabitants (for comparison: dog 1-2 per 1000 dogs per year), spontaneous hyperadrenocorticism is very rare. It mainly affects women over the age of 40. As in dogs, the pituitary form dominates with 70 to 80 % of cases. A special feature in humans is an alcohol-induced form ("pseudo-Cushing's syndrome") and "biochemical hypercortisolism" in depressions and those taking the contraceptive pill. In humans, tumor-related hyperadrenocorticism is usually treated surgically; if this is not possible, radiotherapy or drug treatment is used. For the latter, ACTH secretion can be blocked by the somatostatin analog pasireotide. Since 2020, osilodrostat, an inhibitor of steroid 11β-hydroxylase (CYP11B1) and thus cortisol synthesis has been approved.
In veterinary medicine, Cushing's syndrome plays a certain role in horses and cats in addition to dogs. Here, too, secondary, i.e. pituitary hyperadrenocorticism dominates, even if these animal species are affected much less frequently than dogs. In horses (Equine Cushing's syndrome), Cushing's syndrome is caused by increased production of ACTH or other proopiomelanocortin derivatives. Since their formation is under the control of dopaminergic nerve cells, is used to treat the dopamine receptor agonist in horses. pergolide is used. In cats (Feline Cushing's syndrome), the disease is almost always associated with diabetes mellitus, which does not respond to insulin treatment (insulin resistance). In this animal species, in contrast to dogs, tumors of the growth hormone-producing cells in the pituitary gland are significantly more common than those of the ACTH-producing cells. Zur Therapie wird wie beim Hund Trilostan eingesetzt.