Theodore C Friedman M D Ph D the

Theodore C. Friedman, M. D. , Ph. D. (the Wiz) Professor of Medicine-UCLA Chairman, Department of Internal Medicine Charles R. Drew University Dr. Friedman’s Endocrinology Clinic Familial Cushings: Could it Be Genetic? MAGIC Adult Convention Chicago, IL July 22, 2016

Cushing’s patients ask me • • Why did I get Cushing’s? Is it my genes (DNA)? Can it be passed on to my children? Should I be tested for genetic changes?

I have • 4 families that have more than one member with Cushing’s suggesting the importance of genetics. • So far their genetic testing has been negative (Professor Márta Korbonits). • I’m part of the International FIPA (Familial Isolated Pituitary Adenomas Consortium • I have several patients from the same location (central New Jersey) that suggest environment plays a role.

Genes vs Environment • Nature vs Nurture • Must be some reason one person gets Cushing’s and not the next person

What are genes? • Genes are the coded instructions that control the growth and development of our cells. • DNA (Deoxyribose Nucleic Acid) is the coding molecule that allows human cells to replicate and function. • Human DNA is organized into compact structures called chromosomes of which humans have 23 pairs. • In each pair, one chromosome comes from the mother, and one from the father.

What are genes? • Twenty-two pair of chromosomes are the autosomes and the 23 rd pair that determines your gender (XYmale and XX-female). • Genes come in pairs; one of each pair is inherited from each parent. • Some conditions tend to manifest when one copy of a pair of genes is altered. • These are called dominant disorders. • Other conditions need both genes to be affected in order to develop the condition. • These are called recessive disorders.

Dominant Disorders • Both men and women who have a one copy of an abnormal gene and one copy of the normal gene (a so called 'heterozygote' state) have a 50: 50 chance of passing the abnormal gene on to the next generation. • However, not all patients who carry the abnormal gene will develop the disease, this is called incomplete penetrance.

Dominant Disorders

Recessive Disorders • Both genes are needed to get the condition. • Patients with one gene are carriers and are unaffected, but can pass the genes to their children. • In some conditions, those with one gene are mildly affected.

Recessive Disorders

What kind of mutations are there? • Inheritable mutations (germline mutations) are present in an individual’s DNA from conception. These are inherited from the parents and can be passed on to their children. • Non-inheritable mutations (i. e. , sporadic mutations) occur after birth, most often in a single cell. These mutations are not inherited from the parents and cannot be passed on to their children. • These may be changed by someone’s environment (epigenetics)

Different types of Cushing’s Syndorme • Pituitary-Cushing’s Disease- overgrowth of one ACTHproducing cell in the pituitary, which multiplies abnormally to become a benign tumor that produces too much ACTH • Adrenal –Adrenal Adenoma- is caused by a single benign adrenal tumor (adenoma) on one adrenal gland. • Ectopic-Outside the pituitary-not yet found to be genetic • Adrenal enlargement (hyperplasia) (most often genetic)

Pituitary adenomas • Almost all pituitary tumours are benign, these are called adenomas. • This means that they are not cancerous, and do not spread. • Most of the time pituitary adenomas grow slowly and it takes years before they get diagnosed. • The vast majority of pituitary adenomas occur spontaneously which means that they are not inherited and don’t run in families. • However, a small group of patients with pituitary adenomas (about 1 in 20) also have family members with similar disease. • If no other hormone abnormality and family members have other pituitary tumors, the patients has Familial Isolated Pituitary Adenoma or FIPA. • Familial pituitary adenomas due to FIPA, MEN 1 or Carney

Pituitary adenomas • Families with pituitary adenomas, FIPA families, can be divided into two groups. • In about 80% of families the gene causing the disease is unknown. The pituitary tumor types occurring in these families are most commonly growth hormone-secreting adenomas (causing acromegaly or acromegalic gigantism), prolactin-secreting adenomas (prolactinomas) or nonfunctioning pituitary adenomas (NFPA), very rarely ACTH -secreting adenomas (causing Cushing's disease) or TSHsecreting adenomas. • The disease most often starts in adulthood, very rarely in childhood. • These families usually only have two or three patients known with the disease.

AIP • In about 20% of families a gene has been identified causing the disease, called Aryl hydrocarbon receptor Interacting Protein, or AIP. • Patients with AIP mutations most often have acromegaly or occasionally prolactinoma, very rarely other types of adenomas. • The majority of the patients who carry a mutation in the AIP gene and develop a pituitary adenoma, become diagnosed before the age of 30 years. • Interestingly, two third of the patients with AIP mutation positive pituitary adenoma are males. • 15 -20% of childhood onset acromegaly patients, with no apparent family history, carry an AIP mutation. • Two of my families with Cushing’s disease were tested for AIP and did not have it.

AIP • If you carry one abnormal copy of the AIP gene, you do not necessarily develop the disease. • Only a third of those who inherit such a genetic change go on to develop a pituitary tumor (penetrance is only around 30%). • If you as a parent have been identified to have the gene, then your future children would have a 50: 50 chance of inheriting the genetic changes in the family.

Multiple Endocrine Neoplasia type 1 (MEN 1) • MEN 1 is an inherited disorder that causes tumors in various endocrine glands. • MEN 1 is sometimes called multiple endocrine adenomatosis or Wermer’s syndrome. • MEN 1 is rare, occurring in about one in 30, 000 people. • MEN 1 is autosomal dominant. • 3 Ps-parathyroid (95%), pancreas (80%), pituitary (40%). • In MEN 1 patients, all four parathyroid glands tend to be overactive, causing hyperparathyroidism. • This leads to hypercalcemia-reasonable to measure Ca if suspect MEN 1.

Multiple Endocrine Neoplasia type 1 (MEN 1) • Pancreatic tumors involve the islet cells, giving rise to gastrinomas or insulinomas. • Duodenal tumors can also produce gastrinomas • Very acidic stomach-high gastrin levels (don’t measure gastrin when on PPI). • Severe ulcers in the stomach and small intestine • Diarrhea • Insulinomas-hypoglycemia

Multiple Endocrine Neoplasia type 1 (MEN 1) • Pituitary tumors (25%) • Prolactinomas most common • High prolactin levels can cause excessive production of breast milk, irregular periods or interfere with fertility in women or with libido and fertility in men. • Other pituitary tumor types in MEN 1 can be nonfunctioning pituitary adenoma or growth hormonesecreting adenoma. • Cushing’s disease is very rare in MEN 1 • MEN 2 (parathyroid, medullary carcinoma of thyroid and pheochromocytoma) does not involve the pituitary.

Carney complex • Carney complex is a hereditary condition associated with spotty skin pigmentation, myxomas (benign or noncancerous connective tissue tumors), and benign or cancerous tumors of the endocrine glands such as the adrenal, thyroid and pituitary gland. • Symptoms of Carney complex typically develop when a person is in his or her early 20 s. • Skin pigmentation and heart myxomas or other heart problems are usually the first signs of the condition. • The spotty skin pigmentation is found on lips, inner and outer corners of the eyes, the conjunctiva (membrane lining) of the eye, and around the genital area.

Carney complex • Other common features of Carney complex are Cushing’s syndrome and multiple thyroid nodules or growth hormone -secreting tumors. • The Cushing’s syndrome from Carney complex is usually adrenal and will be discussed there, but rarely can be pituitary. • Although people with Carney complex have an increased risk of cancer, most tumors are benign.

Testicular orphan receptor 4 (TR 4) • Testicular orphan receptor 4 (TR 4) (Dr. Tony Heaney) was found in higher quantities than normal in human Cushing’s disease tumors. • TR 4 was shown to stimulate ACTH secretion and tumor growth in cells in test tubes. • Additional studies are needed to determine if TR 4 can explain the development of pituitary tumors and if drugs could be targeted to decrease its levels.

Pituitary transforming gene (PTTG) • Pituitary transforming gene (PTTG) is a protein that regulates important signals that stimulate cell multiplication. • A drug called R-roscovitin, which targets PTTG, was able to decrease ACTH, cortisol production, and tumor growth in animal models. • Further studies are needed to determine the role of PTTG in patients with Cushing’s Disease.

USP 8 gene (deubiquitinase gene) • In 2014, Reinke and many others reported that 4 out of 10 (40%) ACTH-secreting tumors studied showed somatic (not inheritable) mutations in the USP 8 gene. • These mutations were shown to increase the activity of a receptor for an important growth factor called epidermal growth factor (EGF) which stimulates the secretion of ACTH by the tumors. • In 2015 Ma and others in China found similar mutations in the USP 8 gene in 67 of 108 (62%) ACTH secreting pituitary tumors. • The USP 8 mutations were found mostly in smaller tumors compared to larger tumors.

USP 8 gene • Other types of pituitary tumors were also studied and USP 8 mutations were not found in 150 non-ACTH secreting pituitary tumors. • Further work in cell culture with USP 8 mutated cells showed increased levels of the EFG receptor. • A specific blocker of the EFG receptor called gefitinib decreased ACTH secretion. • The authors concluded, Taken together, somatic gain-offunction USP 8 mutations are common and contribute to ACTH overproduction in Cushing’s disease. • More studies are needed to see if new drug therapy targeting EGF receptor overfunction could be used in Cushing’s disease.

Pituitary Genes • AIP, MEN 1, PTTG, TR 4, Carney and USP 8 all may contribute to familial Cushing’s (USP 8 -non-familial). • Currently only testing for AIP and MEN 1 is available and only under research settings. • Cushing’s still needs to be diagnosed biochemically.

Adrenal Tumors • In most cases, adrenal Cushing’s is caused by a single benign adrenal tumor (adenoma) on one adrenal gland. • Several different mutations have been identified in adrenal adenomas; however, they seemed to be important in only a very few cases. • Researchers found mutations of the PRKACA gene that increased the activity of protein kinase A, which is known to be important in the secretion of cortisol. • This mutation was found in 37% of adrenal adenomas from patients with Cushing’s syndrome. • This mutation was not found in normal adrenal tissue, nonfunctioning adrenal adenomas, tumors producing aldosterone, or those producing subclinical Cushing’s syndrome.

Bilateral Adrenal Hyperplasia • Some cases of bilateral adrenal hyperplasia have been reported to occur in families, indicating that inherited genetic defects are involved.

Primary Pigmented Nodular Adrenocortical Disease (PPNAD) • PPNAD is an adrenal disorder where very small nodules are found in both adrenals. • It can run in families and be associated with the Carney Complex. • Patients with Carney complex can also develop tumors in the heart and other endocrine tissues and have increased skin pigmentation. • Mutations of important genes PRKAR 1 A, PDE 11 A or PDE 8 B, which regulate signals to produce cortisol, have been identified in a large number of affected patients with PPNAD.

Bilateral Macronodular Adrenal Hyperplasia • Bilateral macronodular adrenal hyperplasia tissue can contain a mutation in a gene called ARMC 5. • Inactivation of ARMC 5 led to greater cell growth, perhaps leading to hyperplasia and also to changes in cortisol production.

Bilateral Macronodular Adrenal Hyperplasia • Bilateral macronodular adrenal hyperplasia tissue is enlarged adrenals with multiple large nodules. • Some mutations have been reported in bilateral macronodular adrenal hyperplasia tissue however, there is not one individual mutation that was shared among a large number of patients. • Some patients with bilateral macronodular adrenal hyperplasia show increased cortisol production in response to hormones other than ACTH , such as gastric inhibitory polypeptide (GIP), vasopressin, adrenalin, serotonin, and human chorionic gonadoptropin (h. CG), but the genetic basis of the hyperplasia is unknown.

Summary • In summary, the genetic basis for pituitary tumors responsible for Cushing’s disease is still unknown, but new target proteins that can regulate their growth are under study. • It is likely that there will be advances in genetic screening tools to detect affected family members at a much earlier stage than before. • While the work discussed is promising and the ultimate goal is to have highly effective pharmaceuticals, diagnosis and treatment of Cushing’s syndrome still needs to be done the traditional way.

What should a patient with Cushing’s syndrome be on the look out for on other family members? • Very unlikely that family members will get Cushing’s • If they do get it, it will probably will be the same type of Cushing’s • Same signs and symptoms as the patient. • 24 hr UFC, night time salivary cortisols, night time serum cortisol • Secondary 24 hr 17 OHS, 10 hr 10 pm -8 am UFC/Cr. • Imaging • Would not do genetic testing until 2 nd case of Cushing’s is confirmed.

If a second family member is diagnosed with Cushing’s. • Measure calcium and PTH to look for MEN 1. • If pituitary, consider contacting Dr. Korbanits at St. Bartholomew's Hospital in London info@fipapatients. org • If adrenal, consider contacting Dr. Stratakis at the NIH.

How do you contact Dr. Friedman’s office for an appointment or to get more information? • www. goodhormonehealth. com • Talk will posted in a few days • mail@goodhormonehealth. com • www. fipapatients. org • https: //csrf. net/category/doctors-articles/molecularmechanism-of-cushings/

Thanks • Magic Foundation for inviting me and doing great work! • Great job Dianne