A unifying biology of sex steroid-induced apoptosis in prostate and breast cancers

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   Figure 1

   A schematic representation of the androgen and estrogen deprivation therapy in prostate cancer and pre- and postmenopausal women with breast cancer. (A) The hypothalamic–pituitary–gonadal and adrenal axis in prostate cancer with their therapeutic targets. The hypothalamus produces gonadotropin-releasing hormone (GnRH), which stimulates the adenohypophysis of the pituitary to produce adrenocorticotropic hormone (ACTH). This in turn, stimulates the adrenal gland cortex to produce androgens: dehydroepiandrosterone sulfate (DHEA-S) predominately, DHEA and androstenedione (AD) into the circulation. These androgens (A), alongside testosterone (T) from the testes, are converted in the prostate to their potent form, dihydrotestosterone (DHT). Dihydrotestosterone stimulates the growth of prostate cancer cells and exerts a negative feedback loop onwards to the hypothalamus and pituitary. Both, GnRH agonists/antagonists suppress LH production and cause a subsequent decline in serum testosterone to castrate levels. However, GnRH agonists (with chronic use) lead to the downregulation of GnRH receptors, whereas, GnRH antagonists usually cause an immediate blockade to the receptor. At the adrenal level, abiraterone inhibits adrenal androgen de novo steroidogenesis. At the prostate level, androgen receptor (AR) inhibitors are used and they have different mechanisms of action. For example, enzalutamide competitively inhibits the AR binding to DHT, inhibits nuclear translocation, and DNA and cofactor binding. Whereas, Bicalutamide is a highly selective, competitive and silent antagonist to the AR, which was also found to accelerate AR degradation. (B) The hypothalamic–pituitary–gonadal axis in premenopausal women with breast cancer and their therapeutic targets. The hypothalamus produces gonadotropin-releasing hormone (GnRH), which stimulates the adenohypophysis of the pituitary to produce luteinizing hormone (LH) and follicle-stimulating hormone (FSH). This in turn, stimulates the granulosa cells in the ovarian follicles to produce estrogen. However, FSH in particular stimulates the granulosa cells to produce inhibin, which suppresses FSH in a feedback loop and activin, a peripherally produced hormone that stimulates GnRH cells. Estrogen stimulates the growth of breast cancer cells, and exerts a negative feedback loop onwards to the hypothalamus and pituitary. Ovarian suppression can be achieved with LHRH superagonists such as goserelin, which is an analogue of LHRH, and a GnRH or LHRH agonist. Goserelin initiates a flare of LH production and ultimately leads to receptor downregulation. Antiestrogens can be estrogen receptor (ER) competitive blockers such as the Selective ER Modulators (SERMs, i.e. tamoxifen), or pure antiestrogens or what is known as a Selective ER Downregulators (SERDs, i.e. fulvestrant). Third-generation aromatase inhibitors (i.e. anastrozole, letrozole, exemestane) selectively block the aromatase enzyme system at the breast cancer level and therefore suppress estrogen synthesis. (C) The hypothalamic–pituitary–gonadal axis in postmenopausal women with breast cancer and their therapeutic targets. The differences from premenopausal women is that the ovarian follicles are depleted, therefore there is no active production of estrogen and progesterone. This leads to a dramatic increase in GnRH, an increase in FSH serum level relatively to that of LH through the feedback loops. Ovarian suppression is not used as a treatment option.
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   Figure 2

   A schematic representation of the signal transduction pathways in ER-positive breast cancer cells and prostate cancer cells. (A) At the adrenal level, adrenal androgen de novo steroidogenesis occurs. Cholesterol is produced and converted to Pregnenolone with the aid of CYP11A1 enzyme. Pregnenolone is converted to dehydroepiandrosterone (DHEA) with the aid of CYP17A1. Finally, DHEA is converted to androstenedione (AD) with the aid of 3-β hydroxysteroid dehydrogenase enzyme. Then, AD is converted to testosterone via 17-β hydroxysteroid dehydrogenase. At the adipose tissue level, Both androstenedione and testosterone are converted with the aid of the aromatase enzyme system to estrone (predominant in postmenopausal women), and estradiol (predominant in premenopausal women), sequentially. Estrogen normally binds to the ER in the cytoplasm, the estrogen:ER complex translocates to the nucleus, gets phosphorylated, and binds to estrogen responsive elements (EREs) with the recruitment of coactivators. This creates a transcription complex (TC). This in turn, will initiate a cascade of protein synthesis and subsequent tumor proliferation through the activation of estrogen-sensitive genes. Whereas, SERMs:ER follows a similar pattern but recruits corepressors and inhibits protein synthesis; causing tumor regression. For SERDs, they bind to the ER causing an alien conformation. This leads to the destruction of the ER through the ubiquitin proteasome system; subsequently tumor regression. (B) At the adrenal level, adrenal androgen de novo steroidogenesis occurs. Cholesterol is produced and converted to Pregnenolone with the aid of CYP11A1 enzyme. Pregnenolone is converted to dehydroepiandrosterone with the aid of CYP17A1. Finally, DHEA is converted to DHEA-S with the aid of following enzymes: steryl-sulfatase (STS) and bile salt sulfotransferase. At the prostae level, DHEA-S in Leydig cells is converted back to DHEA via STS and then DHEA is converted to AD via enzyme 3β-HSD. Then, AD is converted to testosterone via enzyme AKR1C3, and finally to DHT via steroid 5α-reductase. Dihydrotestosterone normally binds to the AR in the cytoplasm, the DHT:ER complex translocates to the nucleus, gets phosphorylated, binds to androgen responsive elements (AREs) with the recruitment of coactivators. This creates a transcription complex (TC). This in turn, will initiate a cascade of protein synthesis and subsequent tumor proliferation through the activation of androgen-sensitive genes. Whereas, AR inhibitors:AR complex follows a similar pattern but recruits corepressors and inhibits protein synthesis; causing tumor regression. For SARDs, they bind to the AR causing the degradation of the receptor; subsequently tumor regression. Androgen receptor inhibitors vary in their mechanisms of action. For example, enzalutamide competitively inhibits the AR binding to DHT, inhibits nuclear translocation of AR, and DNA and cofactor binding. Whereas, bicalutamide is a highly selective, competitive and silent antagonist to the AR, which was also found to accelerate AR degradation. Abiraterone inhibits CYP17A1 and subsequently adrenal androgen de novo steroidogenesis. Dutasteride is a 5α-reductase inhibitor that blocks testosterone conversion into DHT.
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   Figure 3

   A schematic representation of the treatment paradigms used clinically for breast and prostate cancers. (A) Early-stage prostate cancer (PC) is usually approached with active surveillance, local treatments such as: surgery and radiation therapy. Hormone therapy can be given for early-stage PC men if they were at high-risk, or if they cannot undergo surgery or radiation therapy. The newer treatments for early-stage PC are: Intensity-Modulated Radiation Therapy, Proton beam therapy, and Cryosurgery. If early-stage PC progresses to metastatic PC (MPC) or what is known as castration-sensitive PC (CSPC), it will be treated with androgen deprivation therapy (ADT) using GnRH agonists, or complete androgen blockade (CAB) using a GnRH agonist plus flutamide for example, or secondary hormone therapy (SHT) using abiraterone, or enzalutamide as examples. If CSPC progresses to castration-resistant PC (CRPC), it will be treated with ADT or SHT. About 60% of PC is diagnosed in men >65, with 97% in men age >=50. The median age at the time of diagnosis in the U.S. is about 66. (B) Early-stage BC can be treated with local treatments such as: surgery and radiotherapy or systemic treatments such as: hormone therapy. What sets early-stage BC treatment apart from prostate cancer is adjuvant therapy with tamoxifen or AIs for 5–10 years. If early-stage BC progresses to metastatic BC (MBC), one therapeutic option is fulvestrant. Breast cancer rates increase after age 40 and are highest in women >70. The median age of diagnosis of BC for women in the U.S. is 62.
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   Figure 4

   Molecular modeling of the wild-type and mutant ER and AR bound with agonists and antagonists. (A) wtAR:DHT LBD complex (PDB ID: 3L3X); (B) the best docking pose of the wtAR:bicatulamide complex (PDB ID: 3RLJ), obtained via flexible docking (the experimental structure used for docking was selected based on the 3D similarity between bicatulamide and the available ligands co-crystalized with AR WT, thus the experimental structure 3RLJ was selected due high similarity between the native ligand, S-22 and bicatulamide). The major interactions are shown in dashed lines and colored as follows: hydrophobic interactions in lavender, pi-pi interactions in purple, water-mediated H-bonds are shown in blue, and classical H-bonds are depicted in green.; (C) T741L AR mutant:bicatulamide LDB complex (PDB ID: 1Z95), helix 12 is colored in green and the major interactions are shown in dashed lines and colored as follow: hydrophobic interactions in lavender, pi-pi interactions in purple, water-mediated H-bonds are shown in blue, and classical H-bonds are depicted in green; (D) wtER:E2 LBD complex (PDB ID: 1GWR); (E) wtER:endoxifen LBD complex; (F) Superposition of E2 D538G mutant with ERα D358G apo LBD structures (helix 12 is shown in red for apo conformation and pink in the E2 bound mutant structure). The major interactions are shown in dashed lines and colored as follow: hydrophobic interactions in lavender, pi-pi interactions in purple, water-mediated H-bonds are shown in blue, and classical H-bonds are depicted in green.
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   Figure 5

   A schematic representation of the parallel cellular evolution of acquired hormone resistance to hormone deprivation in prostate and breast cancer cell models in vitro. (A) LNCaP cell line is an androgen-sensitive human prostate adenocarcinoma cell line. When LNCaP cells are cultured in an androgen depleted environment for 8–11 months in vitro, they become hypersensitive to androgen; and subsequently proliferate. With extended androgen depletion of 16–20 months, selection pressure occurs and LNCaP cells become vulnerable to androgens with death through apoptosis. Cells then exhibit the characteristic morphology of apoptosis with apoptotic membrane blebbing, followed by formation of membrane protrusions (apoptopodia, microtubule spikes, and beaded apoptopodia, beads-on-a-string appearance), ending with cellular fragmentation into apoptotic bodies. (B) MCF-7 cell line is an estrogen-sensitive human breast adenocarcinoma cell line. When MCF-7 cells are cultured in estrogen depleted environment for 6–12 months in vitro, they become hypersensitive to estrogen; and subsequently proliferate. With extended estrogen depletion of 12–18 months, selection pressure occurs and MCF-7 cells are now vulnerable to estrogens with death through apoptosis. Cells then exhibit the characteristic morphology of apoptosis.
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   Figure 6

   A schematic representation of the Study of Letrozole Extension (SOLE) trial. SOLE is a phase III randomized clinical trial of continuous vs intermittent letrozole in postmenopausal women who had received 4–6 years of adjuvant endocrine therapy for hormone receptor (HR)- positive, lymph node- positive, early-stage breast cancer (BC). The rationale of SOLE trial was to test if 3-month treatment-free intervals during extended adjuvant endocrine therapy, would improve disease-free survival (DFS). The underpinning of this hypothesis is based on the theory that letrozole withdrawal for 3 months would allow a degree of estrogenic stimulation toward residual resistant disease, and subsequently the residual disease would become susceptible to letrozole reintroduction. The primary endpoint was DFS (randomization until invasive local, regional, distant recurrence or contralateral BC; 2nd malignancy; death). Postmenopausal women with prior 4–6 years of adjuvant endocrine therapy, were randomized into two arms: first arm is control which is continuous letrozole of 2.5 mg/daily for 5 years, and the second arm is intermittent letrozole of 2.5 mg/daily for 9 months in the first 1–4 years and fully at year 5. The trial concluded no difference in DFS among the two arms but for the first time pre-planned medication non-adherence is not harmful. This can provide a treatment-side effects and financial relief to many patients.
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   Figure 7

   A schematic representation of the proposed design (alongside a proposed optimized version) for the preemptive salvage therapy. (A) Breast cancer patients who are ER- positive after surgery and at high risk of recurrence (this includes large primary tumors and positive lymph nodes at diagnosis), can harness the benefits of long-term estrogen deprivation, with a preemptive salvage therapy, aiming at clearing occult micrometastases. After 5 years of adjuvant antihormonal therapy with either tamoxifen or AIs, breast cancer cell populations undergo selection pressure. The new long-term estrogen-deprived (LTED) breast cancer cell populations are now vulnerable to a woman’s own estrogen through apoptosis (aka estrogen-independent). Whereas, they would normally grow with estrogen within 5 years past menopause (aka estrogen-dependent). The clinically observed response rate to low-dose estrogen therapy was 30% in metastatic breast cancer. Estrogen can act in synergy with other FDA approved breast cancer cell survival inhibitors or apoptosis promoters. This synergy can potentially increase the response rate above 30%. (B) Panel A can be optimized. Estrogen deprivation can be achieved with a new SERM that degrades the ER, preventing future drug resistance and receptor mutations. As one example, there is an orally active SERD, GW5638, which is metabolically hydroxylated to GW7604, in the same way, tamoxifen is metabolically activated to 4-hydroxytamoxifen. Unlike tamoxifen, GW7608 triggers the destruction of ER in BC cells, while retaining an estrogenic tickle at ER elsewhere (i.e. bones and serum lipids). Although GW7608 is a SERD for degrading the ER, it is also a SERM due to its agonistic and antagonistic mechanism of action at different tissue levels. A similar mechanistic SERM/SERD compound can improve estrogen deprivation (with an AI) by destroying the ER, while maintaining women’s health. In addition, estrogen in the proposed 3-month drug holiday can be replaced with selective human estrogen receptor partial agonist (ShERPA). These compounds mimic estrogen without causing significant uterine growth and were found to inhibit the growth of endocrine-independent tamoxifen-resistant breast cancer cell lines.

• © 2018 The authors