PITUITARY TUMORS

Anterior pituitary cell types express distinct hormone gene products. These include ACTH, GH, PRL, TSH, FSH and LH. Pituitary adenomas are distinguished by both excess proliferation of one of these differentiated cell types, as well as dysregulated specific hormone hypersecretion. Physiologically, the six pituitary trophic hormones regulate respective target hormone production and endocrine gland function in tightly controlled regulatory loops (Figure 1). Adenoma hypersecretion is usually a reflection of dysregulated hormone synthesis and or secretion.

Figure 1.

Hormone secretion from pituitary tumors, although excessive and associated with unique phenotypic features, often retains intact trophic control. For example, dopaminergic agents appropriately suppress PRL secretion by prolactinomas, and dexamethasone may suppress ACTH secretion in patients with pituitary Cushing disease.

From Melmed S 2003 Mechanisms for pituitary tumorigenesis: the plastic pituitary. J Clin Invest 112:1603-1618, with permission.

Each of these cell-specific adenomas are associated with a benign sellar mass, as well as a unique clinical syndrome associated with the hormone being hypersecreted (12). Thus, lactotroph adenomas result in hyperprolactinemia, with features of hypogonadanism accompanied by galactorrhea. Corticotroph adenomas hypersecrete ACTH leading to features of hypercortisolism associated with central obesity, hypertension, hyperglycemia, infections and psychologic disturbances. Somatotroph adenomas hypersecrete GH, leading to features of acromegaly including acral changes, arthritis, hypertension, headache, soft tissue swelling and hyperglycemia. Thyrotroph adenomas hypersecreting TSH are extremely rare, and may be associated with mild hyperthyroidisms and goiter. Tumors arising from gonadotroph cells rarely hypersecrete intact FSH or LH, and more commonly express either glycoprotein subunits, or no excess hormone. These adenomas are often discovered incidentally, and are usually associated with hypogonadism and pituitary failure, because of compressive effects of the expanding mass (Table 1).

Table 1.

Classification of pituitary adenomas

Cell tvDe Adenoma Type	Population Prevalence ftotal/lO5)	Tumor transcription factor expression	Unregulated differentiated gene expression	Clinical features
Lactotroph Sparsely or densely granulated	45–50	Pit-1	Prolactin	Hypogonadism and/or galactorhea
Gonadotroph	15–20	SF-l,GATA-2	FSH and/or LH and/or glycoprotein subunit. (depending on type, null or oncocytic)	Silent or pituitary failure Ovarian hyperstimulation (reproductive age women) Testicular enlargement (prepubertal)
Somatotroph	10	Pit-1
Sparsely granulated			GH	Acromegaly or gigantism
Densely granulated			GH	Acromegaly
Combined GH and			GHand	Acromegaly or
PRL cells			prolactin	gigantism
Mixed GH and			GHand	Hypogonadism and
PRL			prolactin GH and	acromegaly Acromegaly or
Mammos omatotroph			prolactin	gigantism
Acidophil Stem			Prolactin and	Hypogonadism and
Cell			GH	acromegaly
Silent			GH	Hypopituitarism
Corticotroph	5	T-Pit	ACTH
Cushing				Cushing Disease
Silent				Hypopituitarism
Nelson				Pituitary hyperplasia
Thyrotroph	<1	Pit-1	TSH	Hyperthyroidism
Plurihormonal	Unknown	All	GH, prolactin, ACTH, glycoprotein	Mixed

All tumor types exhibit features of a pituitary mass, visible on MRI. Glycoprotein refers to intact FSH or LH, or respective α or β glycoprotein subunits. Although null-cell tumors do not express hormone genes, they are classified as gonadotroph in origin.

Abbreviations: ACTH, adrenocorticotropic hormone; FSH, follicle-stimulating hormone; GATA2, endothelial transcription factor GATA-2; GH, growth hormone; LH, luteinizing hormone; PIT1, pituitary-specific positive transcription factor 1; STF1, steroidogenic factor 1; TBX19, T-box transcription factor TBX19 (also known as TPIT).

From Melmed S 2011 Pathogenesis of pituitary tumors. Nat Rev Endocrinol 7:257-266, with permission.

All these adenomas, regardless of cell type, are invariably benign. Although often locally aggressive and invasive, they very rarely progress to true malignancy with documented extracranial metastases (1). Mechanisms underlying the constraints buffering pituitary adenomas against malignant transformation include development of DNA damage and premature proliferative arrest, i.e. cell senescence. Senescent pituitary cells are growth-constrained by CDK inhibitors including p21 for somatotroph tumors. p27 for corticotroph tumors and p15/p165 for non-functioning adenomas (3). These CDK inhibitors lead to cell cycle arrest, while maintaining differentiated hormone secretion and preventing the malignant transformation of respective adenoma cell types.

Another pathogenetic feature of these tumors is their monoclonality (7). As pituitary tumors appear to arise from a single cell, it would be unlikely that broad, generalized signals such as hypothalamic hormone excess, or estrogen excess, lead to polyclonal hyperplasia with resultant hormone hypersecretion. In support of this postulate, activating mutations of hypothalamic hormone receptors in pituitary adenomas (GHRHR; CRHR; GnRHR; TRHR) have not been reported. Similarly, inactivating mutations of hypothalamic inhibitory factor receptors (SSTR1-5; D2R) have not been reported. Consistent with these observations, adenoma monoclonality is also suggested by the fact that surgical resection of small (<10 mm) discrete hormone-secreting adenomas (especially those expressing GH or ACTH) may result in long-term remission with sustained hormonal control. Interestingly, pituitary tissue surrounding the adenoma does not exhibit features of hyperplasia, further suggesting the discreet monocellular pathogenesis of these adenomas.

Multiple extracellular and intracellular signals determine pituitary cell proliferation and specific hormone synthesis and secretion (Figure 2). Mitogenic hormones (e.g. hypothalamic GHRH, CRH or GnRH) or growth factors (e.g. EGF) or steroids (e.g. estrogen) may induce pituitary cell proliferation as well as transcription of pituitary hormone genes. These mitogenic inputs are balanced by important constraining factors of pituitary cell proliferation and hormone synthesis including hypothalamic somatostatin and dopamine. Hormones and growth factors also drive the cell cycle mediated by CDK-cyclin complexes and Rb phosphorylation. Release of E2F by phosphorylated Rb acts to drive cell cycle progression. This complex process is disrupted in adenomas, leading to chromosomal instability and DNA damage (2). In fact, pituitary adenomas are remarkably aneuploidy despite their benign phenotypes. Importantly, dysregulated hormone gene transcription occurs pari passa with cell proliferative changes (11). It is apparent that disruption or activation of multiple sites within the hormone-secreting adenoma cell leads to the dual outcome of adenoma formation with subsequent proliferation, associated with a unique hormone hypersecretory phenotype.

Figure 2.

Pituitary adenoma signaling. Transcription of pituitary hormone genes and cell proliferation are induced by pituitary mitogenic factors including hypothalamic hormones and transcription factors, as well as peripheral hormones. Proliferative constraints include somastostatin and dopamine, as well as tumor suppressor genes. Cell cycle progression is mediated by CDK–cyclin complexes that phosphorylate Rb and cause it to release E2F, which drives cell proliferation. CDK inhibitors block kinase phosphorylation, thereby restraining cell cycle progression. Chromosomal instability, DNA damage and senescence may act to constrain malignant transformation of pituitary tumors.

From Melmed S 2009 Acromegaly pathogenesis and treatment. J Clin Invest 119:3189-3202, with permission.

Animal models

Although classic oncogene mutations are not encountered in sporadic tumors, several growth factor, hormone and cell cycle genes are reported to determine development of sporadic pituitary adenomas (13). Many have been tested in genetically modified mice which either exhibit gain of function or loss of function (Table 2). For example, heterozygote Rb^+/− mutant mice develop pituitary adenomas with high penetrance, usually arising from the intermediate lobe with overexpressed POMC (8). Although several reported transgenic knock-in, universal knockout, and conditional knockout mice may faithfully recapitulate human phenotypes with specific cell type adenoma development and hormone hypersecretion, few such models have yielded a viable therapeutic drug discovery.

Table 2.

Animal models of pituitary tumors include spontaneous tumors

Model type	Genetic modification	Type of tumor
TRANSGENIC
PyLT	Polyoma large T antigen overexpression	Corticotropinoma
POMC-SV40 large T antigen	SV40 large T antigen overexpression (POMC promoter)	Intermediate lobe
AVP-SV40 large T antigen	SV40 large T antigen overexpression (AVP promoter)	GH-producing adenomas
FSHβ-SV40 large T antigen	SV40 large T antigen overexpression (FSHβ promoter	Null cell adenomas
hGRHH	GHRH overexpression	Somatotrophs and mammosomatotrophs
PRL-TGFα	TGFα overexpression	Prolactinomas
PRL-ptd-FGFR4	Overexpression of N-terminally truncated isoform of FGFR4	Prolactinomas
αGSU-PTTG1	PTTG1 overexpression	Gonadotroph hyperplasia
CMV-HMGA1	HMGA1 overexpression	Mixed growth hormone/prolactin adenomas
CMV-HMGA2	HMGA2 overexpression	Somatotrophs, lactotrophs and mammosomatotrophs
CONVENTIONAL KNOCK OUT
Men1 heterozygous	Men1 inactivation	Prolactinomas
Men1 heterozygous	Men1 inactivation	Prolactinomas, NFPA
Men1 heterozygous	Men1 inactivation	Prolactinomas, GH-producing adenomas
Men1 heterozygous	Men1 inactivation	Mammosomatotrophs (90%). ACTH-producing intermediate lobe (10%)
AIP heterozygous	AIP inactivation	GH-producing adenomas (90%), prolactinomas
Drd2 homozygous	Drd2 inactivation	Prolactinomas
Prl homozygous	Prl inactivation	Prolactinomas
Prlr homozygous	Prl receptor inactivation	Prolactinomas
Rb haterozygous	Rb inactivation	ACTH-producing intermediate lobe
P27Kip1 homozygous	p27Kip1 inactivation	Intermediate lobe
p18Ink4c homozygous	p18Ink4c inactivation	intermediate and anterior lobe
Compound p18Ink4c p27Kip1 homozygous	Double p18Ink4c and p27Kip1 inactivation	Intermediate and anterior lobe
Cdk4R24C/R24C mutant mice	Mutation rendering the Cdk4 protein insensitive to INK4 inhibitors	Anterior lobe
Compound Cdk4R24C/R24C p27Kip1 homozygous mutant mice	Mutation rendering the Cdk4 protein insensitive to INK4 inhibitors and and p27Kip1 inactivation	Poorly differentiated adenomas
CONDITIONAL KNOCK OUT
rGhrhr-Cre PRKAR1A	Pituitary-specific (somatotrophs, lactotrophs, and thyrotrophs) PRKAR1A inactivation	Somatotrophs, lactotrophs, and thyrotrophs
POMC-specific Rb heterozygous	Rb inactivation in POMC-expressing cells	Intermediate lobe
POMC-specific Rb homozygous	Rb inactivation in POMC-expressing cells	Intermediate lobe
POMC-specific Rb homozygous	Rb inactivation in POMC-expressing cells	Not reported
Ins-Cre Men1	Men1 homozygous inactivation in pituitary due to ectopic Cre expression	Prolactinomas
GFAP-Cre Bmi1	Bmi1 homozygous pituitary-specific inactivation	Intermediate lobe
Hesx1-Cre; Ctnnb1lox(ex3)+ Sox2-CreERT2;Ctnnb1lox(ex3)+	Pituitary-specific overexpression of constitutively active form of β-catenin	Adamantinomatous, craniopharyngioma

Animal models of pituitary tumors include spontaneous tumors, subcutaneous xenografted implantation, and genetically engineered mouse models may (transgenic and knockout mice). Adapted from Cano DA, Soto-Moreno A, Leal-Cerro A 2014 Front Oncol Aug 1;4:203, with permission.

Transgenic overexpression or disruption of several cell cycle-associated genes have been observed to uniquely lead to pituitary tumor formation in several animal models (Table 3). These experimental tumors exhibit varying degrees of penetrance, may be located in the anterior or intermediate lobes, and may or may not hypersecrete specific trophic hormones with resultant peripheral hormonal phenotypes. Two examples are presented of how further unraveling pituitary tumor signaling pathways in transgenic animal models may lead to translational discovery of novel subcellular therapeutic targets:

Table 3.

Selected cell cycle-associated genes associated with pathogenesis of pituitary adenomas

Gene	Function	Mode of activation/inactivation
CCNB2	Cyclin	Induced by HMGA
CCND1	Oncogene	Overexpression
CDKNIB	CDK inhibitor	Inactivating
HMGA2	Oncogene	Overexpression
FGFR4	Oncogene	Alternative transcription
PTTG	Securin	Overexpression
Rb	Tumor suppressor	Epigenetic silencing
CDKN2A	Cyclin-dependent kinase inhibitor	Epigenetic silencing

Select genes that contribute to the molecular pathogenesis of pituitary adenomas

Adapted from Melmed S 2009 Acromegaly pathogenesis and treatment. J Clin Invest 119:3189-3202, with permission.

TARGETING CDK SIGNALLING

PTTG, pituitary tumor transforming gene, isolated from rat pituitary tumor cells is the index mammalian securin, and drives pituitary tumor formation in murine and zebrafish models (15). The gene is also induced by estrogen in experimental prolactinomas (6) and is overexpressed in human pituitary tumor cells (16). In an attempt to recreate an ACTH-secreting pituitary adenoma, the zebrafish Pttg gene was linked to the POMC promoter to create a corticotroph targeted transgene. Transgenic zebrafish expressing pituitary corticotroph-directed zpttg develop phenotypic features which recapitulate human Cushing disease. These include corticotroph hyperplasia, hypercortisolism, hyperglycemia and fatty liver and cardiac effusions and hypertrophy (9). Furthermore, partial glucocorticoid resistance was also demonstrated. Using this animal model as a drug therapy, screening target, the transgenic zebrafish model was utilized for screening of small molecules exhibiting specific CDK inhibitory activity in the hyperplastic transgenic pituitary. Accordingly, R-Roscovitine, a selective CKD inhibitor, was shown to suppress tumorous pituitary POMC gene expression and ACTH secretion in both zebrafish and murine models of ACTH-secreting corticotroph cell adenomas. The drug also reversed phenotypic features in tumor-bearing mice. Suppressive effects of R-Roscovitine on ACTH secretion were also recapitulated in cell cultures derived from resected human pituitary tumor specimens (9). Based on these translational studies elucidating mechanisms for pituitary tumorigenesis, R-Roscovitine has now been approved to enter clinical trials to test for safety and efficacy in patients with ACTH-secreting pituitary adenomas and Cushing disease.

TARGETING Erb SIGNALLING

Mechanisms underlying prolactinoma pathogenesis are challenging to elucidate due to the unavailability of functional prolactin-secreting human pituitary tumor cell lines, as well as to the relative sparseness of available pituitary tumor tissue specimens. Both ErbB family receptors and ligands are expressed in normal anterior pituitary cells. Activation of this signaling pathway leads to induced PRL production by regulating PRL gene expression and lactotroph differentiation (14). To further elucidate mechanisms underlying prolactinoma tumorigenesis, transgenic mice expressing EGFR or HER2 driven by the tissue-specific rPRL enhancer/promoter to achieve pituitary over-expression were generated lactotroph-targeted murine transgenic hEGFR/HER2 expression induced murine prolactinomas secreting high PRL levels. When EGFR/HER2 activity was inhibited by treating mice with lapatinib, a dual tyrosine kinase inhibitor (TKI) for both EGFR and HER2, hyperprolactinemia and tumor MAPK and Akt pathways were attenuated (10).

Furthermore, in both ACTH-secreting murine AtT20 cells, as well as in primary human cell cultures derived from resected ACTH-secreting tumors were suppressed by TKI treatment. In ATCH-secreting AtT20 cells, overexpression of EGFR led to activation of MAPK-Erk signaling, which was inhibited by gefitinib, an EGFR TKI (5).

These observations provide a compelling rationale for the feasibility of employing ErbB targeted therapy for patients with prolactinomas and those with Cushing disease. Indeed, in a pilot clinical study, lapatanib was shown to attenuate PRL secretion in two patients harboring aggressive prolactinoma found to be tumors resistant to high dose dopamine agonist therapy (4).

SUMMARY

Pituitary tumors are commonly encountered intracranial neoplasms which are invariably benign. Classic oncogene mutations are not encountered in these tumors, and disrupted cell cycle control and growth factor signaling likely contribute to pathogenesis and natural history. They exhibit unique clinical features which are determined by the secreted hormone gene product.

Key Points.

• Pituitary tumors are commonly encountered intracranial neoplasms which are invariably benign

• Classic oncogene mutations are not encountered in these tumors, and disrupted cell cycle control and growth factor signaling likely contribute to pathogenesis and natural history

• Pituitary tumors exhibit unique clinical features which are determined by the secreted hormone gene product.