Trilostane, Prednisone, and ACTH Stimulation Testing in Dogs with Cushing's Disease

My problem patient is Buddy, an 8-year old, male neutered Labrador Retriever weighing 42 kg. Buddy first developed diabetes, which was difficult to control because of insulin resistance. He was later found to have concurrent pituitary-dependent Cushing's disease, confirmed by both ACTH simulation and low-dose dexamethasone suppression testing).

Buddy has been doing well on insulin and trilostane (Vetoryl) for the past 5 months. We started him on a Vetoryl dose of 60 mg, given twice daily with food (2.9 mg/kg/day). When we repeated the ACTH stimulation test after a month, we had a low-normal serum cortisol value (1.4 µg/dl) with absolutely no increase in the post-ACTH cortisol value. This blunted response worried us, so we reduced his Vetoryl dose to 30 mg once a day in the morning (1.4 mg/kg/day).

Recently, Buddy's appetite began to wane, so we added 5-mg of prednisone to his treatment regime. Almost immediately after adding the glucocorticoid treatment, his appetite perked up. 

Buddy has just started showing signs of polyuria and polydipsia, so we repeated his ACTH stimulation test once again. Results of this testing showed a basal serum cortisol value of 2.3 µg/dL, but the post-ACTH cortisol value was only 2.0 µg/dL. 

My question is this: Could the low-dose prednisone treatment that we are administering be suppressing this dog's adrenal response to the ACTH injection and providing us with inaccurate information? With the polyuria and polydipsia, I would have expected high cortisol levels, not low cortisol values.

My Response:

Effects of prednisone supplementation on ACTH stimulation test results
In a dog with pituitary-dependent Cushing's disease, we generally are dealing with an ACTH–secreting pituitary adenoma, which is resistant to negative-feedback suppression with glucocorticoids. This is the basis for using the low- and high-dose dexamethasone suppression tests, which we use to diagnose and differentiate the causes of Cushing's syndrome in these dogs (1).

Therefore, adding 5-mg of prednisone (0.12 mg/kg/day) to this large breed dog's treatment regime would not suppress pituitary ACTH secretion or change your ACTH stimulation test results. If the dog was normal, that small dose of glucocorticoid could possibly be enough to suppress or blunt pituitary ACTH secretion, but it is much too small of a dose to change ACTH secretion in dogs with Cushing's syndrome.

It is possible, however, that you are measuring some of the prednisone in the assay for cortisol, especially if you are giving the prednisone within 24 hours of the ACTH stimulation test. Prednisone and prednisolone will both cross-react in the cortisol assay to falsely increase the measured serum cortisol values. If this is true, basal and post-ACTH stimulated cortisol values in this dog could be very low, and the dose of the trilostane may need to be decreased even further or possibly discontinued.

Adjusting the trilostane dose when ACTH-stimulated cortisol values fall too low
Even even if the prednisone isn't having any effect on cortisol measurements, a post-ACTH stimulated cortisol value of 2.0 µg/dL is too low for me (1,2). I'd recommend that you decrease the trilostane (Vetoryl) dose down to 30 mg once a day, and repeat the ACTH stimulation test again in 2-4 weeks. If the post-ACTH serum cortisol value remains less than 2-2.5 µg/dl on the lowered Vetoryl dosage, I'd even stop the drug completely for 2 weeks to see if the cortisol concentrations will go back up (generally to above 10 µg/dL), as an untreated dog with Cushing's disease should do within a few hours to days.

In many dogs that I treat with Vetoryl, the dose can be decreased over time. In some of these dogs, the dose can even be permanently stopped, and their cortisol secretion remains "normal" and never goes up high enough again to cause signs of Cushing's disease (3,4). The aldosterone secretion is not affected in these dogs, and they never develop any serum electrolyte changes associated with hypoadrenocorticism. This "cure" presumably is the result of mild adrenal necrosis, but we don't know for certain. In any case, when it does happen it certainly is not a bad thing, and the owners are generally thrilled!

Permanent adrenal necrosis, however, can result in life-threatening adrenal crisis (undetectably cortisol values with hyperkalemia and hyponatremia) if the dog is not properly monitored and the dose of the drug lowered or stopped as needed.

What is the cause of dog's polyuria and polydipsia?
Obviously, based on the information we have, it's impossible for me to determine the cause of this dog's polyuria and polydispsia. However, the list of possible differentials in this dog is quite long and includes poor diabetic control or urinary tract infection. A serum chemistry panel and a glucose curve is certainly recommended.

In addition, since both diabetic and Cushingoid dogs are predisposed to having urinary tract infections, a complete urinalysis with culture must be done (6,7). Even without clinical signs of an urinary tract infection, I'd recommend performing a urine culture twice a year since these infections are commonly subclinical and these urinary tract infections are so common in these dogs.

Bottom Line

Again, permanent adrenal necrosis can result in a life-threatening adrenal crisis in dogs treated with trilostane if not properly monitored. For this reason, we never want to "cover up" trilostane-induced lowering of the circulating cortisol values by adding in a glucocorticoid supplement, as you did in this dog. We want to lower the dose of the trilostane instead.

References:

1. Melián C, M. Pérez-Alenza, D, Peterson ME. Hyperadrenocorticism in dogs, In: Ettinger SJ (ed): Textbook of Veterinary Internal Medicine: Diseases of the Dog and Cat (Seventh Edition). Philadelphia, Saunders Elsevier, 2010;1816-1840.
2. Ramsey IK. Trilostane in dogs. Vet Clin North Am Small Anim Pract 2010;40:269-283. 
3. Chapman PS, Kelly DF, Archer J, et al. Adrenal necrosis in a dog receiving trilostane for the treatment of hyperadrenocorticism. J Small Anim Pract 2004;45:307-310. 
4. Ramsey IK, Richardson J, Lenard Z, et al. Persistent isolated hypocortisolism following brief treatment with trilostane. Aust Vet J 2008;86:491-495. 
5. Reusch CE, Sieber-Ruckstuhl N, Wenger M, et al. Histological evaluation of the adrenal glands of seven dogs with hyperadrenocorticism treated with trilostane. Vet Rec 2007;160:219-224. 
6. Nichols R. Complications and concurrent disease associated with canine hyperadrenocorticism. Vet Clin North Am Small Anim Pract 1997;27:309-320.  
7. Forrester SD, Troy GC, Dalton MN, et al. Retrospective evaluation of urinary tract infection in 42 dogs with hyperadrenocorticism or diabetes mellitus or both. J Vet Intern Med 1999;13:557-560.  

Protocol for ACTH Stimulation Testing to Monitor Dogs on Trilostane Therapy: Timing, Feeding, and ACTH Preparation All Matter

I have a question about the use of ACTH stimulation testing in dogs we are monitoring on Vetoryl (trilostane) therapy.  When we recheck these dogs, when should the ACTH stimulation test be started after the morning dose of the drug is administered? 

In other words, how many hours should we wait after giving the morning medication and starting the ACTH stimulation test?  I've read anywhere from 3 hours up to 6 hours?

We normally fast our dogs overnight before any cortisol testing. Should the dog be fed or fasted the morning of the ACTH stimulation test?

My Response:

Most of the initial studies recommended that the ACTH stimulation test be done 4-6 hours after dose administration times, but more recent investigation indicate that the drug generally has a peak effect earlier (1). I like to have my dog owners make the appointment at 2-4 hours because this interval ensures that we are testing at close to the peak effects of the drug (2).

Four important points here:
First, trilostane should always be given together with food. It has been shown that feeding immediately after the administration of the drug significantly increases its absorption (1,2).

Second, It's very important to be consistent in timing when using ACTH stimulation test to make dosage adjustments. We don't want to start one test at 10 AM, then do the next follow-up ACTH stimulation test at 2 PM and compare the results. Because of the relatively short-acting effects of trilostane, the results of ACTH stimulation testing varies considerably with the time of testing relative to dosing.

Third, when we state that the ACTH stimulation test be done 2-4 hours after the morning dose, that's when the basal serum cortisol is collected and the ACTH injected.

Fourth, the form of ACTH injected makes a difference. If synthetic ACTH (cosyntropin or Cortrosyn) is used, the post-ACTH cortisol sample is usually collected at 1 hour after ACTH administration. If the ACTH preparation used is a gel, the post-cortisol sample is usually collected at 2 hours.

• If we use ACTH get and start the test at 4-6 hours after the morning dose, we have a good chance of missing the time of the peak action of trilostane.  Remember that after administration of trilostane, peak concentrations are seen within 1.5 hours and decrease to baseline values in about 18 hours (less in some dogs). 
• In addition, many of the ACTH gel products are formulated by compounding pharmacies, and these may not always be reliable (3).
• For these reasons, I prefer using synthetic cosyntropin over ACTH gel when doing an ACTH stimulation test to monitor trilostane therapy.

Bottom Line:  

1. When monitoring dogs on Vetoryl (trilostane) for Cushing's disease, start your ACTH stimulation test 2 to 4 hours after the morning dose of the drug has been administered.
2. The dog should never be fasted on the morning of the test. Trilostane is a fat-soluble drug and must be given with food to help increase its absorption. 
3. Ideally, use synthetic cosyntropin to do the ACTH stimulation test to ensure consistent and reliable results.

References:

1. Melián CM, Pérez-Alenza D, Peterson ME. Hyperadrenocorticism in dogs In: Ettinger SJ, Feldman EC, eds. Textbook of Veterinary Internal Medicine: Diseases of the Dog and Cat  (Seventh Edition) Philadelphia, Saunders Elsevier, pp 1816-1840, 2010.
2. Ramsey IK. Trilostane in dogs. Vet Clin North Am Small Anim Pract 2010;40:269-283. 
3.  Kemppainen RJ, Behrend EN, Busch KA. Use of compounded adrenocorticotropic hormone (ACTH) for adrenal function testing in dogs. J Am Anim Hosp Assoc 2005;41:368-372. 

Top Endocrine Publications of 2012: Canine and Feline Reproductive Endocrinology

Fertility axis. GnRH, LH and FSH 
stimulate secretion of testosterone, 
progesterone, estrogen, and inhibin. 
Controlled via negative feedback effect
on hypothalamus and pituitary gland.

In my 9th compilation of the canine and feline endocrine publications of 2012, I’m moving on to endocrine disorders of the canine and feline gonads and mammary gland.

Listed below are 23 research papers written in 2012 that deal with a variety of issues of clinical importance in reproductive endocrinology in dogs and cats.

These range from the changes in the pituitary-gonadal axis in dogs with disorders of sexual development (1) to the effects of gonadotropin-releasing hormone (GnRH) in dogs and cats (3,7,18,19); from studies on the ovarian remnant syndrome (4,10,14,18, 22) to a review of the common lesions of the male reproductive tract (5); and from studies of the adrenal sex steroids (6,13) associated with atypical hyperadrenocortisim to studies of the progesterone and prolactin receptors in the mammary gland of cats and dogs (9,12).

Other studies include a review of the reproductive causes of hypocalcemia in dogs and cats (2) to the  reproductive effects of hypothyroidism (15); and finally, from the effects of age and cryptorchidism on plasma levels of insulin-like peptide and testosterone in male dogs (16) to studies investigating the interaction of circulating reproductive hormones on the canine prostate gland (17,23).

References:

1. Buijtels JJ, de Gier J, Kooistra HS, et al. Disorders of sexual development and associated changes in the pituitary-gonadal axis in dogs. Theriogenology 2012;78:1618-1626. 
2. Davidson AP. Reproductive causes of hypocalcemia. Top Companion Anim Med 2012;27:165-166. 
3. de Gier J, Buijtels JJ, Albers-Wolthers CH, et al. Effects of gonadotropin-releasing hormone administration on the pituitary-gonadal axis in male and female dogs before and after gonadectomy. Theriogenology 2012;77:967-978. 
4. Demirel MA, Acar DB. Ovarian remnant syndrome and uterine stump pyometra in three queens. J Feline Med Surg 2012;14:913-918. 
5. Foster RA. Common lesions in the male reproductive tract of cats and dogs. Vet Clin North Am Small Anim Pract 2012;42:527-545, vii. 
6. Ginel PJ, Sileo MT, Blanco B, et al. Evaluation of serum concentrations of cortisol and sex hormones of adrenal gland origin after stimulation with two synthetic ACTH preparations in clinically normal dogs. Am J Vet Res 2012;73:237-241. 
7. Gobello C. Effects of GnRH antagonists vs agonists in domestic carnivores, a review. Reprod Domest Anim 2012;47 Suppl 6:373-376. 
8. Gomes Poppl A, Costa Valle S, Hilario Diaz Gonzalez F, et al. Estrus cycle effect on muscle tyrosine kinase activity in bitches. Vet Res Commun 2012;36:81-84. 
9. Gracanin A, de Gier J, Zegers K, et al. Progesterone receptor isoforms in the mammary gland of cats and dogs. Reprod Domest Anim 2012;47 Suppl 6:313-317. 
10. Gunzel-Apel AR, Buschhaus J, Urhausen C, et al. Clinical signs, diagnostic approach and therapy for the so-called ovarian remnant syndrome in the bitch. Tierarztl Prax Ausg K Kleintiere Heimtiere 2012;40:35-42. 
11. Kim H, Choi H, Choi J. A giant parovarian cyst in a dog with a granulosa cell tumor. J Vet Med Sci 2012;74:385-389. 
12. Michel E, Feldmann SK, Kowalewski MP, et al. Expression of prolactin receptors in normal canine mammary tissue, canine mammary adenomas and mammary adenocarcinomas. BMC Vet Res 2012;8:72. 
13. Monroe WE, Panciera DL, Zimmerman KL. Concentrations of noncortisol adrenal steroids in response to ACTH in dogs with adrenal-dependent hyperadrenocorticism, pituitary-dependent hyperadrenocorticism, and nonadrenal illness. J Vet Intern Med 2012;26:945-952. 
14. Oliveira KS, Silva MAM, Brun MV, et al. Ovarian remnant syndrome in small animals. Semina: Ciências Agrárias, Londrina 2012;33:363-380.
15. Panciera DL, Purswell BJ, Kolster KA, et al. Reproductive effects of prolonged experimentally induced hypothyroidism in bitches. J Vet Intern Med 2012;26:326-333. 
16. Pathirana IN, Yamasaki H, Kawate N, et al. Plasma insulin-like peptide 3 and testosterone concentrations in male dogs: changes with age and effects of cryptorchidism. Theriogenology 2012;77:550-557. 
17. Ponglowhapan S, Church DB, Khalid M. Expression of luteinizing hormone and follicle-stimulating hormone receptor in the dog prostate. Theriogenology 2012;78:777-783. 
18. Rohlertz M, Strom Holst B, Axner E. Comparison of the GnRH-stimulation test and a semiquantitative quick test for LH to diagnose presence of ovaries in the female domestic cat. Theriogenology 2012;78:1901-1906. 
19. Struthers RS. Gonadotropin-releasing hormone targeting for gonadotroph ablation: an approach to non-surgical sterilization. Reprod Domest Anim 2012;47 Suppl 4:233-238. 
20. Switonski M, Payan-Carreira R, Bartz M, et al. Hypospadias in a male (78,XY; SRY-positive) dog and sex reversal female (78,XX; SRY-negative) dogs: clinical, histological and genetic studies. Sex Dev 2012;6:128-134. 
21. Tejerizo G, Domenech A, Illera JC, et al. Altered plasma concentrations of sex hormones in cats infected by feline immunodeficiency virus or feline leukemia virus. Domest Anim Endocrinol 2012;42:113-120. 
22. Urhausen C, Buschhaus J, Wolf K, et al. Identification of ovarian remnant tissue in the bitch after incomplete ovariectomy or ovariohysterectomy. Proceedings of the 7th International Symposium on Canine and Feline Reproduction, 2012. 
23. Wolf K, Kayacelebi H, Urhausen C, et al. Testicular steroids, prolactin, relaxin and prostate gland markers in peripheral blood and seminal plasma of normal dogs and dogs with prostatic hyperplasia. Reprod Domest Anim 2012;47 Suppl 6:243-246. 

Trilostane Treatment of Cats with Cushing's Disease

Moe is a 13-year old, DSH male neutered cat who was diagnosed recently with pituitary-dependent hyperadrenocorticism (Cushing's disease), based on the results of a dexamethasone screening test, as well as an abdominal ultrasound examination.

He has been diabetic for about 9 months, well regulated on 5 units of glargine insulin BID and a strict low-carbohydrate diet (< 10% of calories as carbs). However, he never regained his youthful attitude and healthy hair coat after the diabetes was regulated, which eventually led to this diagnosis of Cushing's disease. His main clinical signs include very thin skin, mild truncal hair thinning, lethargy, and a grumpy mood.

With the dexamethasone suppression test, the morning baseline cortisol concentration was a bit high 6.1 µg/dl (reference range, 1-3.5 µg/dl), with no suppression 4 or 8 hours after the administration of the dexamethasone (0.1 mg/kg, IV). On ultrasound, he had marked bilateral adrenomegaly with no evidence of adrenal neoplasia.

Based on these results, we started him on trilostane (Vetoryl) at a dose of 30 mg once daily. Over the last 2 weeks on that dose, the owner noticed that the cat will get up and walk around a bit more often, but his wandering is more aimless, almost like he is restless. He has also developed a habit of meowing or howling loudly at odd times.

We just did an ACTH stimulation test to evaluate the trilostane dosage, using a cosyntropin (Cortrosyn) dose of 125 µg, IV.

• Baseline cortisol: 1.5 µg/dl
• 1 hour post-ACTH: 2.1 µg/dl
• 2 hour post ACTH: 1.7 µg/dl

These cortisol results look pretty good to me, although clinically, Moe does not seem much improved. Do we just need to give him more time? What range of cortisol value should I be aiming for, and when should I retest?  I am planning on another ACTH stimulation test in 2 more weeks.

Do you have very much experience with trilostane in cats, and seen these side effects?

My Response:
Starting dose: Trilostane treatment has been reported only in a few cats with hyperadrenocorticism (1-5), but it can be a valuable medical treatment for cats with this disorder. Based on both reported studies and my personal experience, the recommended starting dose in cats with hyperadrenocorticism is 20–30 mg/cat orally per day, administered once daily or divided between feeding times. The daily trilostane dose frequently needs to be adjusted in cats, based upon resolution of clinical signs, serum biochemistry results, and repeat ACTH response testing.

Recheck evaluation times: Cats on trilostane treatment should be evaluated at 2 weeks, 1 month, 2–3 months and every 1–3 months thereafter (3). At each recheck (scheduled at approximately 3–4 hours after the morning trilostane dose), the owner should be questioned and the cat examined. Blood is then collected for routine clinicopathological analyses, and an ACTH response test is performed.

Goal of therapy: Although the ideal target range for the post-ACTH cortisol concentration for cats receiving trilostane has yet to be determined, a post-ACTH cortisol concentration of 1.5-5.0 µg/dl (50–150 nmol/L) should be targeted. In cats with persistent clinical signs and serum cortisol values higher than this range, the dose of trilostane is increased to 30–60 mg/cat per day, administered once daily or divided at the time of feeding. Additional dosage adjustments are made as required, based upon subsequent recheck examinations and ACTH response testing. In some cats, daily doses as high as 90–120 mg have been required to control clinical signs and lower ACTH-stimulated cortisol concentrations into the ideal range.

It's certainly too early to expect to see resolution of the clinical signs. This generally takes at least a month or two before we can expect to see improvement. Does he have diabetic neuropathy? Many of those cats hurt and they can be grumpy due to that.

Follow-up: 4-week update:
At Moe's 4-week recheck (on 30 mg trilostane once daily), the abnormal crying behavior has lessened, although is not gone completely. He has no obvious signs of diabetic neuropathy. The diabetic control is improving and the owner has been able to decrease his insulin from 5 units BID to 2 units BID. However, the cat still has very thin skin and his hair is not growing back yet.

Results of the ACTH stimulation test, done 4 hours after the morning dose:

• Baseline cortisol: 2.5 µg/dl
• 1 hour post-ACTH: 5.6 µg/dl
• 2 hour post ACTH: 4.2 µg/dl

So, looks like so far, so good. I am relieved to finally see some clinical improvement. The current trilostane dose will be continued.

Follow-up: 8-week update
Moe is doing great on the trilostane. He has more energy, and is acting more like his old self. His fur is finally starting to grow back, and his skin is losing that tissue-paper look.

Moe's insulin requirements have continued to drop, and he has been off insulin all together for the last week. He is still eating canned, low-carb food. The owner did a glucose curve at home this week, and his blood sugars ranged from 94 to 130 mg/dl!

Follow-up: 12-week update:
Moe has been on the same dose of trilostane (30 mg once daily) for 12 weeks now, and the owner reports great improvement at home. His attitude is greatly improved, and he is more active and alert. The fur is growing back (mostly), and his skin quality/thickness has improved.

He has been off insulin for the last month. The owner continues to spot check his blood glucose 2-3 times weekly, and she is getting numbers between 80 and 120 mg/dl.

Results of the ACTH stimulation test, done 4 hours after the morning dose:

• Baseline cortisol: 0.6 µg/dl
• 1 hour post-ACTH: 1.3 µg/dl
• 2 hour post ACTH: 1.9 µg/dl

As you can see, these cortisol values are a little lower than his previous tests, but they still look acceptable to me.

My Follow-up Response:
These cortisol values are lower than we'd like.  I'd try decreasing the dose to 20 mg per day (10 mg divided may be best).

Follow-up: 5-month update:
Based on your recommendations, we did decrease the trilostane to 10 mg BID. Within 1 week, however, the owner found that his blood sugar levels, which had previously been in the low-to-mid 100's without insulin, were creeping up above 200 mg/dl and even 300 mg/dl. 

Because of the relapse in his diabetes, we decided to switch him back to the original trilostane dose of 30 mg once daily. Do you agree?

My Follow-up Response:
I wouldn't expect the pancreas to ever really become normal again in Cushing's cats like Moe, in which the diabetes goes into remission after we control the disorder with trilostane.

I'd keep him on this higher dose of trilostane for now and start low dose insulin therapy (e.g., 1-2 U glargine, divided BID).

Follow-up: 7-month update
Moe is doing great again back on the higher dose of the Vetoryl. His blood sugars are all in the normal range, with many readings even below 100 mg/dl, so we are backing him off to 1/2 unit BID and will watch closely.  

Final update:
Since my last post, Moe's diabetes did go back into remission, and his owner says that ever since starting the Vetoryl back in March, Moe did seem to feel much better, so at least we were able to improve his quality of life for a little while.

Last week, however, Moe developed neurologic symptoms (circling) and was admitted to the local teaching hospital. An MRI was performed, and a large pituitary macroadenoma was identified. Because of the poor prognosis, Moe was euthanized.

My Response:
Feline hyperadrenocorticism is a serious disease with a guarded to grave prognosis (3-5). Without treatment, most cats will succumb to complications of the disease within a few weeks to months of diagnosis. The use of trilostane has allowed us to help improve the quality of life for at least some these cats. But some of these cats, like dogs with pituitary-dependent disease, will die of complications related to a pituitary macrotumor (3,4).

More work certainly needs to be done to continue to find better ways to diagnose and treat feline Cushing's syndrome.

References:

1. Boag AK, Neiger R, Church DB. Trilostane treatment of bilateral adrenal enlargement and excessive sex steroid hormone production in a cat. J Small Anim Pract 2004;45:263–266.
2. Neiger R, Witt AL, Noble A, et al. Trilostane therapy for treatment of pituitary-dependent hyperadrenocorticism in 5 cats. J Vet Intern Med 2004;18:160-164.
3. Peterson ME. Feline hyperadrenocorticism In: Mooney CT, Peterson ME, eds. BSAVA Manual of Canine and Feline Endocrinology. Fourth ed. Quedgeley, Gloucester: British Small Animal Veterinary Association. 2012;199-203.
4. Cross E, Moreland R, Wallack S. Feline pituitary-dependent hyperadrenocorticism and insulin resistance due to a plurihormonal adenoma. Top Companion Anim Med 2012;27:8-20. 
5. Mellett Keith AM, Bruyette D, Stanley S. Trilostane therapy for treatment of spontaneous hyperadrenocorticism in cats: 15 cases (2004-2012). J Vet Intern Med 2013 (in press).

Performing an ACTH Stimulation Test in Dogs Treated with Fludrocortisone (Florinef)

Fludrocortisone acetate (Florinef), a drug used as mineralocorticoid replacement in dogs with primary hypoadrenocorticism (Addison's disease) 

Thanks for your last post on "Confirming the Diagnosis of Addison's Disease after Treatment of an Adrenal Crisis." I found it to very helpful in managing these dogs in my practice. However, after reading it, I have 2 questions:

1. Are you at all concerned about the glucorticoid effect of fludrocortisone (Florinef) and its effect on the hypothalamic-pituitary-adrenal (HPA) axis?
2. Will fludrocortisone cross-react in the cortisol assay to falsely elevate the measured cortisol result? I've read that the cross reaction is limited to 6% or less — is that correct?
3. How long would I have to stop this drug before I do an ACTH stimulation test in a dog suspected of having Addison's disease?

My Response:

All of your questions are good ones. When we consider mineralocorticoid replacement for dogs with Addison's disease, we have two choices — fludrocortisone acetate (Florinef), administered orally on a daily basis, or desoxycorticosterone pivalate (Percorten-V) generally administered by injection every 3-5 weeks (1-3).

Fludrocortisone acetate is a synthetic adrenocortical steroid that has potent mineralocorticoid and glucocorticoid activities. Compared to cortisol, this drug has 125-times the mineralocorticoid activity and has 10-times the glucocorticoid activity. In this regard, fludrocortisone is very different than desoxycorticosterone pivalate, which is a pure mineralocorticoid agent and possesses no glucocorticoid activity.

The potent glucocorticoid activity of fludrocortisone explains why some dogs will develop polyuria and other signs of iatrogenic Cushing's syndrome, especially when on high daily doses of the drug. It also is the reason why many dogs treated with fludrocortisone do not require additional daily prednisone or prednisolone supplementation.

Because of its glucocorticoid activity, fludrocortisone will indeed cross-react in the cortisol assay to a some extent, as you noted. But even more importantly, it can suppress the HPA axis, especially when given in large doses (4,5), and may lead to a lowering of the basal cortisol concentration and blunting of the cortisol response to ACTH stimulation.

Ideally, we would do the ACTH stimulation test early in the course of the disease, before the dog has been treated for more than a few days with fludrocortisone but especially before the drug dosage has been increased to high daily levels. In these dogs, I simply withhold the drug for 24 hours prior to doing the ACTH stimulation test. That time interval will allow most of the drug to be eliminated so we don't have to worry about measuring the fludrocortisone in the cortisol assay.

If a dog has been on long-term and high-dose fludrocortisone therapy, it is always a good idea to switch to Percorten-V therapy for their mineralocorticoid supplementation for a month or two prior to ACTH stimulation testing. Because Percorten doesn't have any glucocorticoid activity, this might allow the HPA to recover if it has been chronically suppressed by the high-dose fludrocortisone therapy.

References:

1. Kintzer PP, Peterson ME. Treatment and long-term follow-up of 205 dogs with hypoadrenocorticism. J Vet Intern Med 1997;11:43-49. 
2. Church DB. Canine hypoadrenocorticism In: Mooney CT, Peterson ME, eds. BSAVA Manual of Canine and Feline Endocrinology. Fourth ed. Quedgeley, Gloucester: British Small Animal Veterinary Association, 2012;156-166.
3. Kintzer PP, Peterson ME. Canine hypoadrenocorticism In: Bonagura JD, Twedt DC, eds. Kirk's Current Veterinary Therapy, Volume XV. Philadelphia: Saunders Elsevier, 2013;in press.
4. Otte C, Jahn H, Yassouridis A, et al. The mineralocorticoid receptor agonist, fludrocortisone, inhibits pituitary-adrenal activity in humans after pre-treatment with metyrapone. Life sciences 2003;73:1835-1845. 
5. Karamouzis I, Berardelli R, Marinazzo E, et al. The acute effect of fludrocortisone on basal and hCRH-stimulated hypothalamic-pituitary-adrenal (HPA) axis in humans. Pituitary 2013;16:378-385. 

Confirming the Diagnosis of Addison's Disease after Treatment of an Adrenal Crisis

I have a question about using ACTH stimulation testing to confirm primary hypoadrenocorticism (Addison's disease) in dogs. My patient is a 9-year old female spayed Standard Poodle (weighing 25 kg) that was diagnosed with an acute Addisonian crisis at an emergency hospital. At the time of clinical presentation, she was very weak and had collapsed, and had been anorexic for 2 days; physical exam revealed severe dehydration with a normal heart rate (100 bpm).

Routine laboratory workup showed hypoglycemia (glucose, 46 mg/dl), mild azotemia (serum creatinine 1.8 mg/dl), and moderate to severe hyperkalemia (potassium, 7.5 mEq/L) with a normal sodium level (142.4 mg/dl).

At presentation, the owner declined an ACTH stimulation test due to the expense, so the emergency clinic only did a baseline cortisol, which was low at < 0.8 µg/dl. The dog was treated with IV fluids and a shock dose of dexamethasone. She had a good clinical response was discharged after 2 days. On the day of release, she was treated with Percorten-V (50 mg, IM) and started on oral prednisone (2.5 mg BID for 3 days, then 2.5 mg SID).

Today, the owner called me and asked me about doing an ACTH stimulation test to make sure we have the right diagnosis.

My many questions are listed below:

1. Is it always better to do the ACTH stimulation test prior to any glucocorticoid treatment? 
2. Can we do an ACTH stimulation test during the initial adrenal crisis? 
3. If I stop the prednisone and do an ACTH stimulation test now, will I be able to interpret the results?
4. How long does she need to be off of the prednisone before doing the ACTH stimulation test?
5. The owners are concerned that the dog could develop another adrenal crisis if she doesn't get the prednisone every day. How concerned do we have to be worried about that issue?

My Response:

To make a definitive diagnosis of hypoadrenocorticism (Addison's disease), we need to prove that the dog has inadequate adrenal reserve. As you know, this is done by performing an ACTH stimulation test, which is considered to be the gold standard diagnostic test for hypoadrenocorticism in both man and dogs (1-3).

Ideally, the ACTH stimulation test would be done prior to administration of any glucocorticoid agent. The reason for this is two-fold.

Reason 1— Many glucocorticoids (e.g., cortisone, hydrocortisone, prednisolone and prednisone) will cross­-react in the cortisol assay to falsely elevate the measured cortisol value. Therefore, to accurately measure a dog's circulating cortisol concentrations, the ACTH response test should either be performed before these glucocorticoids are administered or after stopping them for at least 24 hours to allow their elimination from the circulation (2,3). The only glucocorticoid that does not cross-react in most cortisol assays is dexamethasone (see below).

Reason 2— Secondly, if high doses of any glucocorticoid preparation are administered, the circulating glucocorticoid will feedback to the hypothalamus and pituitary gland to shut off the secretion of corticotropin-releasing hormone and ACTH, respectively (see Figure 1). This is certainly true for dexamethasone as well, since it is a very potent glucocorticoid.

In the normal dog, this will lead to an acute decrease in circulating ACTH concentrations, and within a few hours to days, lead to mild atrophy of the adrenal cortex. Remember that without circulating ACTH, the basal cortisol value will fall and the response to ACTH stimulation will also become "blunted," even if that dog is not suffering from Addison's disease (2-4).

Figure 1

ACTH stimulation testing during initial treatment of adrenal crisis with dexamethasone
The ACTH stimulation test can be performed at time of initial glucocorticoid treatment for acute adrenal crisis, but only if dexamethasone is used for glucocorticoid replacement. Again, since dexamethasone does not interfere with the cortisol assay, it will not lead to falsely high measured cortisol values.

As noted above (under Reason 2), however, dexamethasone is a very potent glucocorticoid and will act via negative feedback on the HPA axis to inhibit endogenous cortisol production. However, this effect usually takes at least 4–6 hours to be expressed. Consequently, any artifactual lowering of post­-ACTH cortisol concentrations can be avoided by ensuring that the ACTH response test is completed within 2–3 hours of dexamethasone administration (3).

Overall, it is always best to delay glucocorticoid therapy until the ACTH response test is completed. This is rarely problematic, provided that concurrent intravenous fluid therapy is instituted.

Protocol for ACTH stimulation testing in suspect Addison's dogs that have been treated with oral prednisone or another glucocorticoid 
Once a day or more has passed and the dog has received large doses of IV and/or oral glucocorticoids, interpretation of ACTH stimulation test results becomes more problematic. Remember that administration of even moderate doses of a glucocorticoid can result in suppression of the HPA axis and lead to mild to moderate atrophy of the adrenal cortex. Even in a dog without Addison's disease, this can result in a low basal cortisol level with a "blunted" or suppressed response to ACTH stimulation.

If large doses of potent IV glucocorticoids, such as dexamethasone, have been administered for adrenal crisis, it may be best to postpone testing for a week or more to allow the HPA axis to partially recover. If oral prednisone or prednisolone is being administered, the dosage should be tapered down to the low maintenance range (0.1 mg/kg/day) for a few days. The prednisone or prednisolone should then be withheld for at least 24 hours before ACTH stimulation testing to ensure that the glucocorticoid is not detected in the cortisol assay.

As long as the dog has received mineralocorticoid treatment and has normal serum electrolytes, withholding all glucocorticoids for 1 day will not result in adrenal crisis.

Interpreting the results of ACTH stimulation testing in dogs treated with oral prednisone or another glucocorticoid 
In normal dogs administration of a supraphysiologic dose of ACTH produces a rise in serum cortisol to values usually above 10 µg/dl (275 nmol/L). In contrast, dogs with hypoadrenocorticism have an absent or blunted response to ACTH administration. Basal and post-ACTH serum cortisol concentrations are less than 1 µg/dl (27 nmol/L) in over 75% of dogs and less than 2 µg/dl (55 nmol/L) in virtually all dogs with primary hypoadrenocorticism (1-3). Similarly, the great majority of dogs with secondary hypoadrenocorticism also have ACTH-stimulated cortisol concentrations of less than 2 µg/dl.

So what kind of ACTH stimulation response can we expect to find in a dog without Addison's disease that is being treated with chronic glucocorticoids supplementation?  If we are lucky, the dog may show a completely normal cortisol response, thereby making it clear that Addison's disease is ruled out.

However, many of these dogs will show low basal cortisol concentrations or have a slightly blunted response to ACTH stimulation. Even in the these dogs, however, their post-ACTH stimulated cortisol values tend to be higher than 3-4 µg/dl (>80-110 nmol/L). Because we know that dogs with naturally-occurring hypoadrenocorticism will have a much lower adrenocortical reserve than this, and we also know that exogenous glucocorticoids can suppress the HPA axis to inhibit endogenous cortisol production, this type of cortisol response also helps exclude Addison's disease.

References:

1. Peterson ME, Kintzer PP, Kass PH. Pretreatment clinical and laboratory findings in dogs with hypoadrenocorticism: 225 cases (1979-1993). J Am Vet Med Assoc 1996;208:85-91. 
2. Church DB. Canine hypoadrenocorticism In: Mooney CT, Peterson ME, eds. BSAVA Manual of Canine and Feline Endocrinology. Fourth ed. Quedgeley, Gloucester: British Small Animal Veterinary Association, 2012;156-166.
3. Kintzer PP, Peterson ME. Canine hypoadrenocorticism In: Bonagura JD, Twedt DC, eds. Kirk's Current Veterinary Therapy, Volume XV. Philadelphia: Saunders Elsevier, 2013;in press.
4. Kemppainen RJ, Sartin JL, Peterson ME. Effects of single intravenously administered doses of dexamethasone on response to the adrenocorticotropic hormone stimulation test in dogs. Am J Vet Res 1989;50:1914-1917. 

Low-Dose ACTH Stimulation Testing in Cats

Cortisol and Aldosterone Response to Various Doses of Cosyntropin in Healthy Cats

Amy E. DeClue, Linda G. Martin, Ellen N. Behrend, Leah A. Cohn, David I. Dismukes, and Hollie P. Lee

Journal of the American Veterinary Medical Association 2011;238:176-182. 

Adrenocorticotropic hormone (ACTH) stimulation testing is commonly used to evaluate adrenocortical function in both dogs and cats (1-3). In cats, ACTH stimulation testing has been used primarily to test adrenocortical production of cortisol (4-6), but has also been used to evaluate the adrenal sex steroid (progestins and androgens) response (7-10).

Although the primary regulators of aldosterone secretion are the renin-angiotensin system and extracellular potassium concentration (11,12), ACTH acts as an additional stimulant (13). In accord with that fact, administration of exogenous ACTH to cats has been reported to cause a reliable increase in aldosterone secretion. Therefore, ACTH stimulation testing can also be used for evaluation of adrenal mineralocorticoid function (14).

Traditionally, a standard dose of cosyntropin (125 μg/cat, IV) has been recommended for ACTH stimulation testing (5). A previous study (15) documented that lower doses of cosyntropin will stimulate maximal cortisol secretion in cats. However, that study used per-cat dosing as opposed to per-body-weight dosing and did not evaluate the serum aldosterone response to ACTH stimulation.

The purpose of the study reported here by DeClue et al (16) were to determine the lowest dose of cosyntropin (Cortrosyn) on a per-body-weight basis that would produce maximal cortisol and aldosterone secretion in cats.  A secondary purpose was to determine the ideal timing of blood sample collection for cortisol and aldosterone concentrations after ACTH injection in these healthy cats.

Objective—To determine the lowest dose of cosyntropin on a per body weight basis that would produce maximal cortisol and aldosterone secretion and the ideal timing of blood sample collection after ACTH stimulation in healthy cats.

Design—Randomized crossover trial.

Animals—7 adult sexually intact male purpose-bred cats.

Procedures—Each cat received saline (0.9% NaCl) solution (control) and 5 doses (125 μg/cat and 10, 5, 2.5, and 1 μg/kg) of cosyntropin IV with a 2-week washout period between treatments. Blood samples were obtained before (baseline) and at 15, 30, 45, 60, 75, and 90 minutes after administration of saline solution or cosyntropin.

Results—Serum cortisol and aldosterone concentration increased significantly, compared with baseline values, after administration of all cosyntropin doses. Lower doses of cosyntropin resulted in an adrenocortical response equivalent to the traditional dose of 125 μg/ cat.

The lowest doses of cosyntropin that stimulated a maximal cortisol and aldosterone response were 5 and 2.5 μg/kg, respectively. Lower doses of cosyntropin resulted in a shorter interval between IV administration of cosyntropin and peak serum cortisol and aldosterone concentrations.

Conclusions and Clinical Relevance—Low-dose ACTH stimulation testing with IV administration of cosyntropin at 5 μg/kg followed by blood sample collection at 60 to 75 minutes resulted in concurrent peak serum cortisol and aldosterone concentrations that were equivalent to those achieved following administration of cosyntropin at 125 μg/cat, the standard dose currently used.

My Bottom Line:

This study confirms our earlier work that low doses of ACTH (e.g., 5 μg/kg body weight of cosyntropin) will maximally stimulate cortisol secretion in cats (15). In the cats of this study, as in our previous study, lower doses of cosyntropin resulted in an adrenocortical response that was equivalent to the traditional dose of 125 μg/cat (15, 16). The efficacy of of this low-dose ACTH stimulation testing protocol is also well documented in dogs (17-19), and has become a widely used testing dosage in clinical practice for evaluation of both hyper- and hypoadrenocorticism.

Based on the results of these cat studies, the following test protocol can be recommended:

1. Collect blood sample for basal cortisol (± aldosterone or sex steroids).
2. Calculate the cosyntropin (Cortrosyn) dosage (5 μg/kg of cat's body weight). To draw up this amount, it's best to reconstitute and dilute the Cortrosyn powder and store the remaining ACTH product (20).
3. Administer the cosyntropin dose to the cat by the IV route.
4. Collect a post-ACTH blood sample at 60-75 minutes after cosyntropin injection.

It is extremely important to point out that cosyntropin must be administered by the IV route in cats, especially when this low-dose protocol is used for ACTH stimulation testing. When given intramuscularly (IM) to cats, cosyntropin is not well absorbed and will not produce a maximal adrenocortical response (21). In cats, ACTH given by the IV route induces a greater and more prolonged adrenocortical stimu­lation than intramuscular administration.

This difference in the cortisol response between IV and IM administration is in contrast to the situation in dogs, in which IV or IM low-dose ACTH stimulation protocols produce similar adrenocortical responses (22). This difference between cats and dogs should not surprise us— we all know that cats are not just small dogs, especially when it comes down to endocrinology!

References:

1. Behrend EN, Kemppainen RJ. Diagnosis of canine hyperadrenocorticism. Vet Clin North Am Small Anim Pract 2001;31:985-1003.
2. Church DB. Canine hypoadrenocorticism In: Mooney CT, Peterson ME, eds. BSAVA Manual of Canine and Feline Endocrinology. Fourth ed. Quedgeley, Gloucester: British Small Animal Veterinary Association, 2012;156-166.
3. Herrtage ME, Ramsey IK. Canine hyperadrenocorticism In: Mooney CT, Peterson ME, eds. BSAVA Manual of Canine and Feline Endocrinology. Quedgeley, Gloucester: British Small Animal Veterinary Association, 2012;167-189.
4. Peterson ME, Greco DS, Orth DN. Primary hypoadrenocorticism in ten cats. J Vet Intern Med 1989;3:55-58. 
5. Duesberg C, Peterson ME. Adrenal disorders in cats. Vet Clin North Am Small Anim Pract 1997;27:321-347. 
6. Peterson ME. Feline hyperadrenocorticism In: Mooney CT, Peterson ME, eds. BSAVA Manual of Canine and Feline Endocrinology. Fourth ed. Quedgeley, Gloucester: British Small Animal Veterinary Association, 2012;199-203.
7. DeClue AE, Breshears LA, Pardo ID, et al. Hyperaldosteronism and hyperprogesteronism in a cat with an adrenal cortical carcinoma. J Vet Intern Med 2005;19:355-358. 
8. Briscoe K, Barrs VR, Foster DF, et al. Hyperaldosteronism and hyperprogesteronism in a cat. J Feline Med Surg 2009;11:758-762. 
9. Quante S, Sieber-Ruckstuhl N, Wilhelm S, et al. Hyperprogesteronism due to bilateral adrenal carcinomas in a cat with diabetes mellitus. Schweiz Arch Tierheilkd 2009;151:437-442. 
10. Meler EN, Scott-Moncrieff JC, Peter AT, et al. Cyclic estrous-like behavior in a spayed cat associated with excessive sex-hormone production by an adrenocortical carcinoma. J Feline Med Surg 2011;13:473-478. 
11. Gogerly RL, Coghlan JP, Morgenroth P, et al. A compartmental model of acute stimulation of aldosterone secretion in vivo by potassium and ANG II. Am J Physiol 1993;265:E190-196. 
12. Pratt JH. Role of angiotensin II in potassium-mediated stimulation of aldosterone secretion in the dog. J Clin Invest 1982;70:667-672. 
13. Crabbe J, Reddy WJ, Ross EJ, et al. The stimulation of aldosterone secretion by adrenocorticotropic hormone (ACTH). J Clin Endocrinol Metab 1959;19:1185-1191. 
14. Zimmer C, Horauf A, Reusch C. Ultrasonographic examination of the adrenal gland and evaluation of the hypophyseal-adrenal axis in 20 cats. J Small Anim Pract 2000;41:156-160. 
15. Peterson ME, Kemppainen RJ. Dose-response relation between plasma concentrations of corticotropin and cortisol after administration of incremental doses of cosyntropin for corticotropin stimulation testing in cats. Am J Vet Res 1993;54:300-304. 
16. DeClue AE, Martin LG, Behrend EN, et al. Cortisol and aldosterone response to various doses of cosyntropin in healthy cats. J Am Vet Med Assoc 2011;238:176-182. 
17. Frank LA, Oliver JW. Comparison of serum cortisol concentrations in clinically normal dogs after administration of freshly reconstituted versus reconstituted and stored frozen cosyntropin. J Am Vet Med Assoc 1998;212:1569-1571. 
18. Kerl ME, Peterson ME, Wallace MS, et al. Evaluation of a low-dose synthetic adrenocorticotropic hormone stimulation test in clinically normal dogs and dogs with naturally developing hyperadrenocorticism. J Am Vet Med Assoc 1999;214:1497-1501. 
19. Lathan P, Moore GE, Zambon S, et al. Use of a low-dose ACTH stimulation test for diagnosis of hypoadrenocorticism in dogs. J Vet Intern Med 2008;22:1070-1073. 
20. Peterson ME. How to extend your supply of cortrosyn and lower the cost of ACTH stimulation testing. Insights into Veterinary Endocrinology, Blog post, March 22, 2011.
21. Peterson ME, Kemppainen RJ. Comparison of intravenous and intramuscular routes of administering cosyntropin for corticotropin stimulation testing in cats. Am J Vet Res 1992;53:1392-1395. 
22. Behrend EN, Kemppainen RJ, Bruyette DS, et al. Intramuscular administration of a low dose of ACTH for ACTH stimulation testing in dogs. J Am Vet Med Assoc 2006;229:528-530. 

How Does the New Generic Cosyntropin Compare to Cortrosyn for ACTH Stimulation Testing?

PAPER REVIEW

Comparison of IV and IM Formulations of Synthetic ACTH

for ACTH Stimulation Tests in Healthy Dogs

by T.A. Cohen, and E.C. Feldman

Journal of Veterinary Internal Medicine 2012;26:412-414.

Background

Corticotropin (adrenocorticotropic hormone or ACTH) stimulation testing is widely accepted as the gold standard for confirming a diagnosis of hypoadrenocorticism in dogs and cats. The test is also commonly used as as a screening test for hyperadrenocorticism (Cushing's syndrome). Finally, the ACTH stimulation test is widely accepted as the best means of monitoring medical therapy for hyperadrenocorticism with either mitotane or trilostane (1).

Natural ACTH is a single-chain 39-amino acid peptide hormone synthesized in the corticotrophs of the anterior lobe (pars distalis) of the pituitary gland. Although the amino acid sequence of ACTH varies among species, the first 24 amino acids are identical among all species studied (2). Canine ACTH differs from human ACTH by only one amino acid residue, at position 37, although the amino terminal end of the ACTH molecule (amino acids 1-18) is responsible for its biologic activity.

Cosyntropin is a synthetic form of ACTH, which was created by isolating the first 24 amino acids from the 39-amino acid ACTH peptide. Since the introduction of cosyntropin to the veterinary literature 3 decades ago (3), this synthetic ACTH preparation (marketed as Cortrosyn, Amphastar Pharmaceuticals) has gained popularity and is currently recommended as the ACTH preparation of choice for use in dogs and cats (1).

Cortrosyn is available  as a sterile lyophilized powder in vials containing 0.25 mg of cosyntropin (4).  Once the lyophilized powder is reconstituted with 0.9% sodium chloride, the ACTH solution can be administered either by IV or IM injection with similar cortisol responses.

Recently, a 1-ml liquid form of generic cosyntropin that does not require reconstitution has become commercially available through Sandoz, Inc (5). This sterile solution contains 0.25 mg of cosyntropin.  The amino acid sequence of this generic cosyntropin product is identical to Cortrosyn, suggesting that the biologic activity of the generic product should be similar to that of the brand name Cortrosyn preparation. However, the manufacturer recommends that this generic liquid form of synthetic ACTH be given by the IV route only, and that any unused portion of the vial be discarded (5).

When considering use of one of these cosyntropin products versus the other, the question that can be raised is whether or not their biologic activities are comparable. If each testing agent causes maximal stimulation of cortisol from the canine and feline adrenal cortex, then these diagnostic testing products could be used interchangeably.  That is the question that was addressed in this study by Cohen and Feldman (6).

Objectives of study
To compare the biologic activity of the liquid and lyophilized forms of cosyntropin.

Methods
Eighteen privately owned healthy dogs were studied. Dogs were assigned to one of 2 groups of 9 dogs each. Group 1 dogs were tested with the lyophilized product first and the liquid solution 30 to 60 days later. The Group 2 dogs were tested with the liquid solution first and then tested with the lyophilized drug 30 to 60 days later.

For the ACTH stimulation tests, serum samples were collected before and 1 hour after IM administration of 0.25 mg reconstituted lyophilized product, or 1 hour after IV administration of 0.25 mg of liquid solution. Cortisol concentrations of all serum samples were measured by use of a commercial cortisol radioimmunoassay.

Results
Serum cortisol concentrations before and after ACTH stimulation did not differ significantly between groups (P = .57). In addition, no individual dog had as much as a 20% difference in serum cortisol concentrations after administration of either ACTH formulation.

Conclusions of study
Given the lack of significant differences of the ACTH stimulation test results, the lyophilized and liquid solution products can be used interchangeably.

My Bottom Line:

The results of this study suggest that injection of generic cosyntropin (liquid solution) stimulates adrenocortical secretion to an equivalent degree, as compared with Cortrosyn (lyophilized product). However, many questions about this generic cosyntropin product still need to be answered before it can be strongly recommended for use in dogs and cats.

1. First of all, only 18 clinically normal dogs were tested in this study (6). It would be very important to repeat the study in dogs with known adrenal disease to verify that both diagnostic testing preparations were truly equivalent for diagnosis of hypo- and hyperadrenocorticism.
2. Secondly, with Cortrosyn, it has been clearly shown that this product can be administered to dogs by either the intravenous or intramuscular routes with comparable results. Since IM administration can be more convenient in some cases, it would be of interest to investigate if the generic product could be given by that route or if it must be administered IV, as the insert states.
3. Thirdly, with Cortrosyn, it is common practice to dilute the product and administer lower doses (e.g., 5 μg/kg) of the cosyntropin to dogs (1, 7-9).  Since only a portion of the vial would be used in most dogs, the remaining cosyntropin is divided into aliquots and stored frozen for up to 6 months.  Can the generic product be diluted? And if so, will it maintain its potency when stored for months for later use? We simply don't know — the product insert states that "cosyntropin injection is intended as a single dose injection and contains no antimicrobial preservative; any unused portion should be discarded."
4. Finally, for reasons that are unclear to me, the current cost of a single vial of this generic preparation (≈$99 per vial) is about 30% more than the cost of an identical amount of Cortosyn.  Why would a generic product cost more than a time-honored brand-name product?

For all of the reasons listed above, I plan to continue using the lyophilized Cortosyn® preparation for  ACTH stimulation testing. 

References:

1. Melián C, M. Pérez-Alenza, D, Peterson ME. Hyperadrenocorticism in dogs, In: Ettinger SJ (ed): Textbook of Veterinary Internal Medicine: Diseases of the Dog and Cat (Seventh Edition). Philadelphia, Saunders Elsevier, 2010;1816-1840.
2. Eipper BA, Mains RE. Structure and biosynthesis of pro-adrenocorticotropin/endorphin and related peptides. Endocrine Reviews 1980;1:1-27. 
3. Feldman EC, Stabenfeldt GH, Farver TB, et al. Comparison of aqueous porcine ACTH with synthetic ACTH in adrenal stimulation tests of the female dog. American Journal of Veterinary Research 1982;43:522-524.
4. Cortrosyn Injection, package insert. Rancho Cucamonga, CA: Amphastar Pharmaceuticals, Inc.  
5. Cosyntropin Injection (Generic), package insert. Princeton, NJ: Sandoz Inc.
6. Cohen TA, Feldman EC. Comparison of IV and IM formulations of synthetic ACTH for ACTH stimulation tests in healthy dogs. Journal of Veterinary Internal Medicine 2012;26:412-414. 
7. Frank LA, Oliver JW. Comparison of serum cortisol concentrations in clinically normal dogs after administration of freshly reconstituted versus reconstituted and stored frozen cosyntropin. Journal of the American Veterinary Medical Association 1998;212:1569-1571. 
8. Kerl ME, Peterson ME, Wallace MS, et al. Evaluation of a low-dose synthetic adrenocorticotropic hormone stimulation test in clinically normal dogs and dogs with naturally developing hyperadrenocorticism. Journal of the American Veterinary Medical Association 1999;214:1497-1501. 
9. Martin LG, Behrend EN, Mealey KL, et al. Effect of low doses of cosyntropin on serum cortisol concentrations in clinically normal dogs. American Journal of Veterinary Research 2007;68:555-60. 

My related posts:

• What's the Best Protocol for ACTH Stimulation Testing in Dogs and Cats? http://endocrinevet.blogspot.com/2011/03/whats-best-protocol-for-acth.html
• How to Extend Your Supply of Cortrosyn and Lower the Cost of ACTH Stimulation Testing http://endocrinevet.blogspot.com/2011/03/how-to-extend-your-supply-of-cortrosyn.html
• How to Dilute and Store Cortrosyn for ACTH Stimulation Testing http://endocrinevet.blogspot.com/2012/03/how-to-dilute-and-store-cortrosyn-for.html

How to Dilute and Store Cortrosyn for ACTH Stimulation Testing

Can you please review how you recommend diluting Cortrosyn for low-dose ACTH stimulation testing? 

It's expensive and I don't want to give a whole vial to every dog, since most of my patients weigh les than 30 pounds.

My Response:

Yes, Cortrosyn is expensive. And once reconstituted, it has a limited shelf-life. However, by following the protocol outlined below, you can easily dilute, aliquot and store Cortrosyn after reconstitution for up to 6 months. This makes each ACTH stimulation test much less expensive, because each vial of Cortrosyn can be used to perform as many as five ACTH stimulation tests.

Cortrosyn is supplied in vials each containing 0.25 mg (250 μg) of synthetic ACTH (cosyntropin) in powder form. Because the dose of Cortrosyn used to perform an ACTH stimulation test is only 5 μg/kg, small to medium sized dogs require only a fraction of the ACTH contained in each vial.

How to Aliquot and Store Each Vial of Cortrosyn for Subsequent Use

1. Reconstitute the Cortrosyn power by adding exactly 2.5-ml of sterile saline solution to the vial. With this dilution, the resulting concentration of the Cortrosyn solution in the vial would be 100 μg/ml.
2. Once Cortrosyn is reconstituted, aspirate 50-μg doses (0.5 ml) into 5 plastic syringes. Or, if smaller ACTH doses are desired, aspirate 25-μg doses (0.25 ml) into 10 syringes. (For more accurate dosing, I use insulin or tuberculin syringes).
3. DO NOT store reconstituted Cortrosyn in glass containers or vials. The reason that the Cortrosyn needs to be stored in plastic is that ACTH will stick to glass, thereby lowering the available amount of Cortrosyn that would be injected at time of testing.
4. The syringes containing the reconstituted, diluted Cortrosyn should be labeled with the product, dose in each syringe, and the date the Cortrosyn was reconstituted.
5. Freeze each of the syringes at -20C. Avoid storing these syringes in a frost-free freezer, which must periodically warm up to defrost. Repeated freezing and thawing cycles would compromise the integrity of the Cortrosyn.
6. When frozen properly, aliquots can be stored for up to 6 months without loss of efficacy.
7. Alternatively, the Cortrosyn solution can be stored refrigerated (4C) where it has been shown to be bioactive and stable for at least 4 weeks.

Diluting the Cortrosyn and using a low-dose ACTH stimulation test protocol is very cost-effective. In this way, 1 vial of Cortrosyn can be used to test multiple patients without compromising the quality of the test results.

As an example, at the 5-µg/kg dose, 5 dogs weighing 10 kg (22 lbs) can be tested using a single vial, reducing the cost of the drug by 80%. By looking at the cost benefit, it is clear that the savings could be significant by using this testing protocol.

For more information on how to perform an ACTH stimulation test, see my post entitled, What's the Best Protocol for ACTH Stimulation Testing in Dogs and Cats?

Q & A: Blunted Cortisol Response to ACTH in a Dog with Suspected Addison's Disease

My patient is a 6.5-year old male neutered Belgian Malinois who has a history of weight loss and intermittent vomiting of 3-weeks duration.

A CBC and serum chemistry panel were both unremarkable, with normal serum sodium and potassium values. Abdominal ultrasonography was performed, which revealed a splenic mass. A splenectomy and liver biopsy were performed, which revealed a splenic hematoma and hepatopathy, thought to be secondary to Rimadyl (1). The drug has subsequently been discontinued with no improvement.

Today we did an ACTH stimulation test to rule out Addison's disease, using a dose of 2.2 U ACTH gel administered intramuscularly. The baseline cortisol value was 2.2 μg/dl (reference range, 1-4 μg/dl) and the 2-hour post-ACTH cortisol was 3.6 μg/dl (reference range, 6-20 μg/dl).

Is this subnormal cortisol response consistent with mild Addison's disease?

My Response:

This ACTH stimulation test completely rules out spontaneous hypoadrenocorticism (Addison's disease). Most dogs with primary, secondary, or atypical hypoadrenocorticism will have a low basal cortisol concentration (< 1 μg/dl) and all will have a baseline cortisol value that is less than 2 μg/dl (2-4).

After ACTH stimulation, dogs with hypoadrenocorticism tend not to have any cortisol response to ACTH stimulation, and post-ACTH cortisol values should never be greater than 2.5-3.0 μg/dl (see Figure below)

In many dogs, the blunted cortisol response, as seen in this dog, is either due to prior administration of glucocorticoid sometime in the last few weeks. However, in this dog, the likely cause for the subnormal cortisol response is the compounded ACTH preparation that you used.

In a study performed a few years ago (5), normal dogs were tested with cosyntropin (Cortrosyn, 5 μg/kg, IV) and 4 different compounded forms of ACTH (all manufacturers recommend a dose of 2.2 U/kg, IM).  For all 5 of the ACTH stimulation tests, samples for cortisol analysis were collected before and 1 and 2 hours after ACTH injection.

After administration of the 4 forms of compounded ACTH, the time to reach peak cortisol concentrations varied, with some dogs showing the maximal cortisol value at 1 hour and others having peak cortisol concentrations at 2 hours.With 2 of the 4 products, serum cortisol concentration returned to baseline values by 2 hours in many of the dogs.

Thus, due to variability in duration of cortisol response after injection of the compounded ACTH products, the investigators recommend that we need to collect several post-ACTH samples for cortisol analysis after administration of a compounded ACTH preparation to ensure that we detect the peak cortisol response. At the minimum, we recommend collecting samples at 1 and 2 hours following injection.

Because of this variability in cortisol stimulation, I'm not a fan of using compounded ACTH preparations. I would rather use low-dose Cortrosyn (6) and collect only one post-ACTH blood sample at 1-hour and be assured that will produce the maximal cortisol response at that time.

References:

1. MacPhail CM, Lappin MR, Meyer DJ, et al. Hepatocellular toxicosis associated with administration of carprofen in 21 dogs. Journal of the American Veterinary Medical Association 1998; 212: 1895-1901.
2. Lifton SJ, King LG, Zerbe CA. Glucocorticoid deficient hypoadrenocorticism in dogs: 18 cases (1986-1995). Journal of the American Veterinary Medical Association 1996;15;209:2076-2081. 
3. Cook AK, Bond KG. Evaluation of the use of baseline cortisol concentration as a monitoring tool for dogs receiving trilostane as a treatment for hyperadrenocorticism. Journal of the American Veterinary Medical Association 2010;237:801-805.
4. Klein SC, Peterson ME. Canine hypoadrenocorticism: part II. Canadian Veterinary Journal 2010;51:179-184.
5. Kemppainen RJ, Behrend EN, Busch KA. Use of compounded adrenocorticotropic hormone (ACTH) for adrenal function testing in dogs. Journal of the American Animal Hospital Association 2005;41:368-372. 
6. Martin LG, Behrend EN, Mealey KL, et al.  Effect of low doses of cosyntropin on serum cortisol concentrations in clinically normal dogs. Journal of the American Veterinary Medical Association 2007;230:555-560. 

Q & A: Addison's Disease in a Siamese Cat

I would like to discuss a problem case of feline Addison's disease that I diagnosed a few months ago. The cat is a 6-year old female-spayed Siamese weighing 3 kg.

The cat presented for severe lethargy and depression, complete anorexia, and bloody diarrhea. On physical examination, she was 8-10% dehydrated and was severely depressed and weak. Her mucous membranes were slightly pale and tacky. Her only past history was a fungal infection causing hair loss on her face and neck, which has responded to itraconazole treatment with partial hair regrowth.

Results of a complete blood count revealed a hematocrit of 30%, and a white blood cell count of 14,300 with a normal differential. A serum chemistry panel showed a urea nitrogen of 22 mg/dl and a creatinine of 1.8 mg/dl. The serum concentrations of sodium (145 mEq/L), potassium (5.3 mEq/L), and chloride (107 mEq/L) were all within reference range limits. A basal serum cortisol value was slightly low at 0.8 μg/dl (normal, 1-4 μg/dl).

Based on the clinical presentations coupled with the low serum cortisol value, I made a diagnosis of hypoadrenocorticism and started the cat on prednisolone (5 mg, bid PO). There has been only a minimal response to treatment, and now the cat has developed ataxia. Repeat serum chemistry panel and survey radiographs were normal.

Why hasn't the cat responded better to my treatment?

My Response

In cats, as in other species, hypoadrenocorticism results from deficient adrenocortical secretion of glucocorticoids, either alone or concurrent with reduced secretion of mineralocorticoids. Hypoadrenocorticism can be a naturally occurring disease or can be iatrogenic and is extremely rare in cats (especially the naturally occurring disorder). The first cat with primary hypo­adrenocorticism was described approximately 30 years ago (1), and since then, fewer than 20 well-documented cases of naturally occurring adrenal insufficiency in cats have been reported (2-9).

It's highly unlikely that this cat has primary hypoadrenocorticism (Addison's disease) for the following reasons.

1. First of all, this is an extremely rare disorder of cats.
2. More importantly, all of the reported feline cases of Addison's disease have had serum electrolyte changes consistent with mineralocorticoid (aldosterone) deficiency (hyperkalemia, usually with hyponatremia and hypochloremia) (1-9). The fact that this cat has normal serum electrolytes alone tends to rule out the diagnosis of Addison's disease.
3. Thirdly, the serum cortisol of 1.1 μg/dl may be slightly low to low-normal, but cats with naturally occurring Addison's disease should have undetectable levels of circulating cortisol (1-10). So the fact that cortisol was detected alone goes against Addison's disease. However, I suspect that we may actually be measuring the prednisolone in the cortisol assay, since that steroid will cross react in the cortisol assay to produce a "falsely high" cortisol reading (10).
4. To diagnose Addison's disease, we would need to do an ACTH stimulation test. To do that, however, we would need to stop the prednisolone for at least a week (a month would be best) to allow the pituitary gland and adrenal glands to recover.

One common protocol for ACTH response testing in cats is to collect blood for determination of circulating cortisol concentration before and at 60 minutes after administration of 0.125 mg synthetic ACTH (cosyntropin; Cortrosyn) IV (10,11). It is important to administer ACTH intravenously, especially if the cat is dehydrated. In addition, findings in healthy cats indicate that ACTH given by the intravenous route induces a greater and more prolonged adrenocortical stimulation than does intramuscular administration (10,11).

It's highly likely that you have induced secondary hypoadrenocorticism in this cat.

• Remember that administration of any glucocorticoid to a cat will feed back to the cat's pituitary and suppress ACTH secretion. This can lead to secondary hypoadrenocorticism, with low circulating cortisol concentrations (10,12). In this scenario, however, aldosterone secretion would be be affected since pituitary ACTH has little stimulatory effect on aldosterone secretion. Therefore, normal serum electrolytes would be expected (10,12).
• In addition, it's quite possible that itraconazole that you have been giving is also blocking normal cortisol production, leading to iatrogenic hypoadrenocorticism. Antifungal agents, such as ketoconazole and itraconazole, interfere with the synthesis of steroid hormones and can block cortisol synthesis. Again, as long as you are giving the prednisolone, the cat is covered and should not show signs of hypoadrenocorticism.

Bottom Line: Overall, I do not know the cause of this cat's illness, and it's obvious that she is seriously ill. But I do not think this cat has naturally occurring hypoadrenocorticism — either primary (Addison's disease) or secondary (pituitary ACTH deficiency).

References

1. Johnessee JS, Peterson ME, Gilbertson SR: Primary hypoadrenocorticism in a cat. Journal of the American Veterinary Medical Association 1983;183:881-882.
2. Peterson ME, Greco DS, Orth DN: Primary hypoadrenocorticism in ten cats. Journal of Veterinary Internal Medicine 1989;3:55-58.
3. Berger SL, Reed JR. Traumatically induced hypoadrenocorticism in a cat. Journal of the American Animal Hospital Association1993; 29:337–339.
4. Ballmer-Rusca E. What is your diagnosis? Hypoadrenocorticism in a domestic cat. Schweizer Archiv für Tierheilkunde 1995;137:65–67.
5. Brain PH.Trauma-induced hypoadrenocorticism in a cat. Australian Veterinary Practitioner 1997;27:178–181.
6. Tasker S, MacKay AD, Sparkes AH. A case of feline primary hypoadrenocorticism. Journal of Feline Medicine and Surgery 1999;1:257–260,
7. Parnell NK, Powell LL, Hohenhaus AE, Patnaik AK, Peterson ME. Hypoadrenocorticism as the primary manifestation of lymphoma in two cats. Journal of the American Veterinary Medical Association 1999;214:1208–1211.
8. Stonehewer J, Tasker S. Hypoadrenocorticism in a cat. Journal of Small Animal Practice 2001;42:186–190.
9. Redden B. Feline hypoadrenocorticism. Compendium of Continuing Education for the Practicing Veterinarian 2005;27:697–706.
10. Peterson ME: Hypoadrenocorticism in cats. In: Mooney C.T., Peterson M.E. (eds), Manual of Canine and Feline Endocrinology (Fourth Ed), Quedgeley, Gloucester, British Small Animal Veterinary Association 2012; in press.
11. Peterson ME, Kemppainen RJ. Comparison of intravenous and intramuscular routes of administering cosyntropin for corticotropin stimulation testing in cats. American Journal of Veterinary Research 1992;531392-1395.
12. Middleton DJ, Watson AD, et al. Suppression of cortisol responses to exogenous adrenocorticotrophic hormone, and the occurrence of side effects attributable to glucocorticoid excess, in cats during therapy with megestrol acetate and prednisolone. Canadian Journal of Veterinary Research 1987;51:60–65.

Q & A: Prolonged Action of Percorten in a Dog with Addison's Disease

My patient is a 2-year old female/spayed Labrador Retriever who was initially examined 2 months ago because of anorexia and vomiting. Routine serum chemistry panel showed prerenal azotemia, hyperkalemia (potassium of 7.4 mEq/L; reference range, 3.5-5.7 mEq/L), hyponatremia (sodium, 127; reference range, 136-148 mEq/L) and a Na/K ratio of 18 (normal <27). An ACTH stimulation test revealed a low basal cortisol (0.2 μg/dl) with no response to ACTH stimulation (0.3 μg/dl). This lack of a cortisol response coupled with the classic serum electrolyte changes was considered diagnostic for primary hypoadrenocorticism (Addison's disease).

I started the dog on oral prednisone (0.15 mg/kg once daily) for glucocorticoid coverage. For mineralocorticoid supplementation, I administered a single dose of desoxycorticosterone pivalate (DOCP; Percorten-V, Novartis) at the recommended dose of 2.2 mg/kg intramuscularly.

The dog has done very well, with a complete clinical response. We have monitored the dog's serum chemistry panel at 15, 25, 35, and 51 days after the initial Percorten-V injection, but the dog's serum potassium value remains within the high-normal range (between 4.5-5.5).

Now on day 53, the dog is acting more lethargic and is not eating well. I just instructed the owner to administer the second Percorten-V injection; the owner is a nurse so they are able to give the injection at home.

I'm confused. Does this dog have Addison's disease or is my diagnosis incorrect? Can Addison's disease ever go into remission? Why is the serum potassium concentration still within the normal range?

My Response:

In answer to your first question, it is clear that this dog does indeed have primary hypoadrenocorticism (Addison's disease) (1-5). That diagnosis is based on the following data:

• Signalment (i.e., young adult female dog)
• History and clinical signs (anorexia, vomiting, lethargy)
• Routine laboratory findings (hyperkalemia, hyponatremia, prerenal azotemia)
• Low basal cortisol concentration
• Lack of a serum cortisol response to ACTH stimulation
• Complete response to glucocorticoid and mineralocorticoid treatment

In answer to your second question: No, dogs with true primary hypoadrenocorticism have destruction of all layers of the adrenal cortex and do not ever go into remission. These dogs require life-long replacement of the missing adrenocortical hormones (3-5).

In answer to your last question, I do not know for certain why an occasional dog will have a prolonged response to the Percorten-V injection, but it certainly does occur. Remember that circulating potassium concentrations are controlled by factors other than just aldosterone. That's one reason we sometimes see dogs with "atypical" Addison's dogs that have persistently normal serum potassium concentrations but no measurable aldosterone concentrations (6-9). Most of those dogs will eventually go on to develop hyperkalemia or hyponatremia, but in some dogs it may take months.

Dogs like this indicate that there are mechanisms which allow normal potassium and sodium balance to be maintained even without the presence of circulating aldosterone. This phenomenon is also recognized in human medicine, where it has been shown that up to 25% of human patients with primary hypoadrenocorticism may have normal serum potassium concentrations. However, all of these Addison's patients will have a high plasma renin to aldosterone ratio, which indicates a failing zona glomerulosa (10,11).

What's the mechanism for this maintenance of normal potassium concentrations in dogs or humans with Addison's disease? Well, that's not clear, but the following physiological mechanisms have been proposed (11-13):

• Hyperkalemia itself will increase potassium excretion; this may help maintain normal serum levels of potassium.
• A high renal tubular flow rate with increased distal delivery of potassium can increase the urinary excretion of potassium and again help maintain normokalemia.
• An increased sensitivity of the distal tubule to aldosterone will enhance the urinary excretion of potassium. Again, this would help maintain the whole body levels of potassium.
• Insulin may also act to compensate for aldosterone deficiency by promoting the transfer of potassium from the extracellular to intracellular space, thus maintaining normokalemia.

Based on my studies done 2 decades ago, it's clear that most dogs with Addison's disease can be maintained quite well with DOCP when given at much lower monthly doses than 2.2 mg/kg/injection (4,5).  In one study (4), the final median dose of DOCP needed in 33 dogs was 1.69 mg/kg/month, with a range of 0.75-3.4 mg/kg/month (Figure 1, below). 

Figure 1: Initial and final maintenance doses of DOCP (Percorten-V) administered to 33 dogs with Addison's disease. 
Data is displayed as "box plots," in the the whiskers represent the main body of data (i.e., the range). The box represents the interquartile range for the 25th to75th percentile (the middle half of the data). The horizontal bar through the box is the median value. Outlying data points are represented by open circles.

If this was my case, I would not waste any more time or money trying to determine the longest interval you can go between Percorten-V injections. Remember, it's not necessary for hyperkalemia to redevelop before the next injection; in fact, you don't want that to ever happen!  Rather, I recommend that you treat with low dose of Percorten-V once monthly. It's very difficult to "overdose" Percorten-V, so we don't have to worry about giving the injections too frequently, especially when we are giving a smaller monthly dose.

How low can you go with the monthly Percorten-V injections? I'd start by lowering the dose by about 10-20% and then check the serum electrolytes again in a month. If they remain normal, then I'd continue to decrease the dose by another 10-20%. Again, in my studies, over half the dogs did very well on a monthly dose of Percorten-V <1.6 mg/kg. Almost all dogs need a monthly dose of at least 1.0 mg/kg/month to maintain normal serum electrolytes on a long-term basis (Figure 1).

References

1. Peterson ME, Kintzer PP, Kass PH. Pretreatment clinical and laboratory findings in dogs with hypoadrenocorticism: 225 cases (1979-1993). Journal of the American Veterinary Medical Association 1996;208:85-91.
2. Klein SC, Peterson ME. Canine hypoadrenocorticism: part I. Canadian Veterinary Journal 2010;51:63-69.
3. Klein SC, Peterson ME. Canine hypoadrenocorticism: part II. Canadian Veterinary Journal 2010;5:179-184.
4. Kintzer PP, Peterson ME. Treatment and long-term follow-up of 205 dogs with hypoadrenocorticism. Journal of Veterinary Internal Medicine 1997; 11:43-49.
5. Melián C, Peterson ME. Diagnosis and treatment of naturally occurring hypoadrenocorticism in 42 dogs. Journal of Small Animal Practice 1996;37:268-275.
6. Bartges JW, Nielson DL. Reversible megaesophagus associated with atypical primary hypoadrenocorticism in a dog. Journal of the American Veterinary Medical Association 1992; 201: 889-891.
7. Feldman EC, Nelson RW. Hypoadrenocorticism (Addison's disease). In: Feldman EC, Nelson RW (Eds). Canine and Feline Endocrinology and Reproduction, 3rd Edition. WB Saunders. St. Louis, Missouri, 2004, pp 394-439.
8. Mueller C, Boretti FS, Wenger M, et al. Investigation on the aldosterone concentration before and after ACTH application in 44 dogs with hypoadrenocorticism. Kleintierpraxis 2007;52:216-224.
9. Baumstark ME, Mueller C, Boretti FS, et al.  Evaluation of aldosterone concentrations in dogs with Addison's disease (abstract). Proceeding of the 2011 ACVIM Forum.
10. Oelkers W, Diederich S, Baehr V. Diagnosis and therapy surveillance in Addison's disease: rapid adrenocorticotropin (ACTH) test and measurement of plasma ACTH, renin activity, and aldosterone. Journal of Clinical Endocrinology and Metabolism 1992;75:259-264.
11. Shiah CJ, Wu KD, Tsai DM, et al. Diagnostic value of plasma aldosterone/potassium ratio in hypoaldosteronism. Journal of the Formosan Medical Association 1995;94:248-524. 
12. Harvey TC. Addison's disease and the regulation of potassium: the role of insulin and aldosterone. Medical Hypotheses. 2007;69:1120-1126. 
13. Gagnon RF, Halperin ML. Possible mechanisms to explain the absence of hyperkalaemia in Addison's disease. Nephrology, Dialysis, Transplantation 2001;16:1280-1284.