Luteinizing Hormone-releasing Hormone Receptor Expression Research Articles

Immunohistochemical Expression of LHRH Receptor in Different Compartments of Female Genital Tract in Patients With Endometrial Cancer.

Luteinizing hormone-releasing hormone receptor (LHRHR) expression has been reported in various cancers, including endometrial neoplasms. Thus, LHRHR provides a potential point for therapeutic approach using LHRH analogs as carrier molecules for chemotherapeutic agents in this cancer population. However, clinical data did not prove any potential benefits for patients. We decided to assess LHRHR expression in patients with endometrial cancer to explain possible lack of efficacy in previous clinical reports. LHRHR expression was assessed immunohistochemically in different anatomic and histogenetic compartments of female genital tract of patients with endometrial cancer. The study sample consisted of paraffin tissue blocks obtained from patients who has undergone primary surgery owing to endometrial cancer. Strong LHRHR expression was found in endometrial cancer, fallopian tube, and concurrent atypical hyperplasia. Interestingly, LHRHR expression showed significant differences depending on the respective compartment of the ovary analyzed. Level of LHRHR expression in patients with primary advanced and unresectable disease, particularly in certain ovarian compartments may be substantially lower, which may influence the use of new targeted therapy regimens. The studies on secondary Müllerian system compartment and its hormonal receptor status may be crucial to understand mechanisms of lack of efficacy of LHRH hybrid molecules anti-cancer treatment.

Experimental therapy of doxorubicin resistant human uveal melanoma with targeted cytotoxic luteinizing hormone-releasing hormone analog (AN-152)

BackgroundCytotoxic analogs of LHRH (luteinizing hormone-releasing hormone) can be successfully used for the treatment of hormone-dependent cancers such as prostatic, ovarian, endometrial, but our knowledge about their effect on hormone-independent cancers such as human uveal melanoma (UM) is limited. Previously, we have demonstrated that 46% of UM express full-length LHRH receptors. This finding has led us to further examine the mechanism of action of LHRH receptor based targeted therapies in this malignancy. AimsIn the present study we investigated the cellular uptake of doxorubicin (DOX) and cytotoxic LHRH analog AN-152 (AEZS-108, zoptarelin doxorubicin) on human UM cell lines (OCM3) and its DOX resistant form OCM3DOX320 by confocal laser scanning microscopy. The LHRH receptor expression was characterized by RT-PCR and immunocytochemistry. ResultsWe were able to establish a new, stable and DOX resistant human UM cell line OCM3DOX320. Our results demonstrated the expression of splice variants and isoforms of receptor for LHRH in OCM3 UM cell line and its doxorubicin resistant form OCM3DOX320.It has been revealed by MTT assay that AN-152 inhibited cell proliferation in a dose dependent manner in OCM3DOX320 cells. Furthermore, receptor-mediated uptake of AN-152 was demonstrated using confocal laser scanning microscopy in both cell line. ConclusionsOur results suggest that the antiproliferative effect of AN-152 can be detected even if only LHRH receptor isoforms are expressed. Our study also demonstrates the LHRH receptor-mediated uptake of AN-152 in DOX resistant OCM3DOX320 cells. Our experiments provide new insights into a potential targeted therapy of UM and give further details about the accumulation of AN-152 in hormone-independent DOX-resistant cells expressing splice variants of the LHRH receptors.

A phase II trial of zoptarelin doxorubicin in castration- and taxane-resistant prostate cancer.

210 Background: Luteinizing hormone-releasing hormone receptors (LHRH-R) are expressed in ~86% of prostate cancers, with sustained expression despite prolonged exposure to LHRH agonists. Limited expression of LHRH-R on normal cells suggests LHRH-R may be a therapeutic target for prostate cancer. Zoptarelin doxorubicin is a cytotoxic hybrid molecule linking doxorubicin to an LHRH analog that selectively targets doxorubicin to cancer cells expressing LHRH-R. Methods: This Phase 2 trial evaluated the clinical benefit of zoptarelin doxorubicin for metastatic castration-resistant prostate cancer (mCRPC) following androgen deprivation therapy and ≥1 taxane-based regimen. The primary endpoint was clinical benefit (CB) defined as progression-free survival (PFS) per RECIST v1.1 at 12 weeks with no dose-limiting toxicities (DLT) or other toxicities resulting in treatment cessation. Secondary endpoints were time to overall progression, response rate, pain response, overall survival (OS), and PSA response. Results: Of 25 patients enrolled, 20 had ≥1 measurable lesion at baseline with PFS ≥12 weeks achieved in 56% and PSA response of PR (4%) and SD (84%). Median PFS and OS were 3.8 and 6.0 months, respectively. Reduced pain was reported by 59%. Treatment was well tolerated, with no cardiotoxicity and neutropenia (all grades) the most common hematologic adverse event. Conclusions: This phase 2 trial demonstrated clinical benefit in 56% of patients with no DLT or other toxicity requiring treatment termination. These results suggest a role for zoptarelin doxorubicin for mCRPC following progression on second- and third-line hormonal therapy. Clinical trial information: NCT01240629. [Table: see text]

LHRH receptor expression in sarcomas of bone and soft tissue.

Luteinizing hormone releasing hormone (LHRH) is a neurohormone, secreted by the hypothalamus, which regulates the secretion of gonadotropins, luteinizing hormone (LH) and follicle stimulating hormone (FSH) from the pituitary. LHRH acts by binding to receptors located in the pituitary gland. These receptors (LHRH receptors) have also been found in the cytoplasm of many tumor cells that involve both the reproductive and non-reproductive organs. These receptors have been demonstrated in prostate and breast cancers, endometrial carcinomas, renal cell carcinoma, lymphoma, carcinoma of liver, pancreas and skin. So far, the expression of LHRH receptors on sarcomas (i.e. malignant tumors of mesenchymal origin) has not been studied, except for endometrial sarcomas. It has also been demonstrated that both LHRH agonists and antagonists can down-regulate these receptors and thus inhibit these tumor cells. Another major therapeutic implication is that these receptors can be targeted specifically by peptides conjugated to anti-cancer drugs. The purpose of this study was to determine if LHRH receptors are expressed in primary and/or metastatic sarcomas of human origin. We looked at LHRH receptor expression in 38 consecutive sarcoma specimens, using immunohistochemistry. The specimens were either from office biopsy or from resected tumor; these were confirmed as sarcomas by histopathological examination. The receptor staining characteristics and the staining intensity were also documented. The pattern of staining was classified either as "focal or diffuse staining of the cytoplasm" and the intensity of staining was graded on a scale from 1+ to 4+. Positive receptor staining was seen in 25 of the 38 (66%) specimens. Twelve of the specimens stained diffusely and 13 had focally positive staining. Three tumors had 1+ staining, 10 had 2+ staining, six had 3+ staining, and six tumors had 4+ staining. The tumors included undifferentiated pleomorphic sarcoma, synovial sarcoma, osteosarcoma, myofibroblastic sarcoma, myxofibrosarcoma, liposarcoma, dermatofibrosarcoma protuberans, metastatic chondrosarcoma and chordoma. Sarcomas express LHRH receptors with a varying incidence and degree. Our study suggests that those sarcomas that are LHRH receptor positive could potentially be treated with targeted chemotherapy.

Novel LHRH-receptor-targeted cytolytic peptide, EP-100: first-in-human phase I study in patients with advanced LHRH-receptor-expressing solid tumors

PurposeTo conduct a phase I study determining the safety, pharmacokinetics and preliminary efficacy of EP-100, a novel anticancer drug consisting of natural luteinizing-hormone-releasing hormone (LHRH) ligand linked to a cationic membrane-disrupting peptide.MethodsPatients with advanced, solid tumors, positive for LHRH receptor by immunohistochemistry (IHC), received EP-100 weekly or twice weekly for 3 of 4 weeks in a 28 day cycle. A modified Fibonacci 3 + 3 dose-escalation schema was used. Initial cohorts received EP-100 once weekly (cohorts 1–7, 0.6–7.8 mg/m2, n = 21). Later cohorts received doses twice weekly (cohorts 7–11, 7.8–40 mg/m2, n = 16).ResultsLHRH-receptor expression was confirmed by IHC in 52 of 89 consented patients; 37 patients received at least 1 dose. Cohorts receiving doses of 5.2 mg/m2 and above achieved therapeutic levels from in vitro studies Clearance was rapid (mean half-life 7.1 ± 3.8 to 15.9 ± 3.6 min). The maximum-tolerated dose was not reached at the highest dose evaluated (40 mg/m2 twice weekly). Grade 2 increase in alanine aminotransferase/serum aspartate aminotransferase in one patient resolved, did not recur upon re-treatment, and was not observed in other patients. The only drug-related adverse event was transient infusion-related dermatologic reactions (10 patients). No complete or partial tumor responses were observed; seven patients had stable disease of 16 weeks.ConclusionsEP-100 was well tolerated in patients with advanced, LHRH-receptor-expressing solid tumors. The recommended phase 2 dose is 40 mg/m2 twice weekly for 3 of 4 weeks per cycle.Electronic supplementary materialThe online version of this article (doi:10.1007/s00280-014-2424-x) contains supplementary material, which is available to authorized users.

Emerging association between androgen deprivation therapy and male meningioma: significant expression of luteinizing hormone-releasing hormone receptor in male meningioma

There is emerging data suggesting a potential risk for meningioma growth stimulation in patients on luteinizing hormone-releasing hormone (LHRH) analogs for prostate cancer. We examined the expression of LHRH receptor (LHRH-R), progesterone receptor (PR) and Ki67 labeling index (LI) in specimens from male meningioma (MM) and female meningioma (FM) patients. A total of 24 MM and 24 FM paraffin blocks were retrieved from our institution between 1991 and 2008. Sections from the paraffin blocks were stained with mouse monoclonal antibodies against LHRH-R, PR and Ki67. All male patients had no previous history of prostate cancer (PCa) or previous history of hormone therapy. LHRH-R positivity was extensive in 92% of MM and 88% of FM samples, with both showing strong intensity (67% and 79%, respectively). PR was positive in 20 of 24 (83%) MM and 23 of 24 (96%) FM samples. MM is less likely to exhibit Ki67 LI >4% compared with FM. The majority of MM and FM samples were strongly positive for LHRH-R expression and PR expression. The emerging association of androgen deprivation therapy and meningioma growth should be recognized in urological practice. Caution should be taken when considering LHRH agonist administration for patients with PCa and concurrent meningioma or previous history of meningioma.

Inhibition of U-87 MG glioblastoma by AN-152 (AEZS-108), a targeted cytotoxic analog of luteinizing hormone-releasing hormone

Glioblastoma multiforme is the most frequent tumor of the central nervous system in adults and has a dismal clinical outcome, which necessitates the development of new therapeutic approaches. We investigated in vivo the action of the targeted cytotoxic analog of luteinizing hormone releasing hormone, AN-152 (AEZS-108) in nude mice (Ncr nu/nu strain) bearing xenotransplanted U-87 MG glioblastoma tumors. We evaluated in vitro the expression of LHRH receptors, proliferation, apoptosis and the release of oncogenic and tumor suppressor cytokines. Clinical and U-87 MG samples of glioblastoma tumors expressed LHRH receptors. Treatment of nude mice with AN-152, once a week at an intravenous dose of 413 nmol/20 g, for six weeks resulted in 76 % reduction in tumor growth. AN-152 nearly completely abolished tumor progression and elicited remarkable apoptosis in vitro. Genomic (RT-PCR) and proteomic (ELISA, Western blot) studies revealed that AN-152 activated apoptosis, as reflected by the changes in p53 and its regulators and substrates, inhibited cell growth, and elicited changes in intermediary filament pattern. AN-152 similarly reestablished contact regulation as demonstrated by expression of adhesion molecules and inhibited vascularization, as reflected by the transcription of angiogenic factors. Our findings suggest that targeted cytotoxic analog AN-152 (AEZS-108) should be considered for a treatment of glioblastomas.

Single Molecular Recognition Force Spectroscopy Study of a Luteinizing Hormone-Releasing Hormone Analogue as a Carcinoma Target Drug

The luteinizing hormone-releasing hormone- Pseudomonas aeruginosa exotoxin 40 (LHRH-PE40), is a candidate target drug associated with elevated LHRH receptor (LHRH-R) expression in malignant tumor tissue. The capability of LHRH-PE40 to recognize LHRH-Rs on a living cell membrane was studied with single molecular recognition force spectroscopy (SMFS) based on atomic force microscopy (AFM). The recognition force of LHRH-PE40/LHRH-R was compared with that of LHRH/LHRH-R by dynamic force spectroscopy. Meanwhile, cell growth inhibition assay and fluorescence imaging were presented as complementary characterization. The results show that LHRH moiety keeps its capability to recognize LHRH-R specifically, which implies that recombinant protein LHRH-PE40 can be a promising target drug.

Receptor-targeted therapy of human experimental urinary bladder cancers with cytotoxic LH-RH analog AN-152 [AEZS- 108

Many bladder cancers progress to invasion with poor prognosis; new therapeutic methods are needed. We developed a cytotoxic LH-RH analog, AN-152 (AEZS-108) containing doxorubicin (DOX), for targeted therapy of cancers expressing LHRH receptors. We investigated the expression of LH-RH receptors in clinical bladder cancers and in HT-1376, J82, RT-4 and HT-1197 human bladder cancer lines. The effect of analog, AN-152, on growth of these tumor lines xenografted into nude mice was analyzed. Using molecular and functional assays, we also evaluated the differences between the effects of AN-152, and DOX alone. We demonstrated the expression of LH-RH receptors on 18 clinical bladder cancers by immunohistochemistry and on four human urinary bladder cancer lines HT-1376, J82, RT-4 and HT-1197 by Western blotting and binding assays. AN-152 powerfully inhibited growth of these bladder cancers in nude mice. AN-152 exerted greater effects than DOX and was less toxic. DOX activated strong multidrug resistance mechanisms in RT-4 and HT-1197 cancers, while AN-152 had no or less such effect. PCR assays and in vitro studies revealed differences in the action of AN-152 and DOX on the expression of genes involved in apoptosis. These results suggest that targeted cytotoxic LH-RH analog, AN-152 (AEZS- 108), should be examined for treatment of patients with LH-RH receptor positive invasive bladder cancers.

The in vivo antitumor activity of LHRH targeted methotrexate–human serum albumin nanoparticles in 4T1 tumor-bearing Balb/c mice

The use of targeted drug delivery systems is a growing trend in cancer treatment to decrease the adverse effect of anti-cancer drugs. In this study, we sought to conjugate methotrexate–human serum albumin nanoparticles (MTX–HSA NPs) with luteinizing-hormone releasing hormone (LHRH). The LHRH was intended to target LHRH receptors overexpressed on the several types of tumors. The expression of LHRH receptors on the 4T1 breast cancer cells was confirmed by FITC conjugated LHRH receptor antibody using fluorescence microscopy. Female Balb/c mice bearing 4T1 breast cancer tumor were treated with a single i.v. injection of free MTX, non-targeted MTX–HSA NPs and LHRH targeted MTX–HSA NPs. LHRH targeted MTX–HSA nanoparticles showed stronger anti-tumor activity in vivo. By 7 days after treatment, average tumor volume in the LHRH targeted MTX–HSA NPs treated group decreased to 8.67% of the initial tumor volume when the number of attached LHRH molecules on MTX–HSA NPs was the highest, while the average tumor volume in non-targeted MTX–HSA NPs treated mice grew rapidly and reached 250.7% of the initial tumor volume 7 days after the treatment. LHRH targeted MTX–HSA NPs could significantly extend the survival time of tumor bearing mice compared with the non-targeted MTX–HSA NPs and free MTX formulations.

Abstract 2829: Activity of EP-100 in Non-Hodgkin's Lymphoma - synergy in combination

Abstract EP-100 is a targeted anti-cancer peptide comprised of Luteinizing Hormone Releasing Hormone (LHRH) fused to a membrane-disrupting peptide (MDP). It is currently in Phase 1 clinical trial in tumors that over-express LHRH receptors. EP-100 kills cancer cells directly via membrane disruption. We tested a combination of EP-100 and doxorubicin in multi-drug resistant Non-Hodgkin's Lymphoma (NHL) cell lines and primary cells from refractory/relapsed NHL patients. Cells were cultured in the presence of EP-100 (0.00001-100 μM) or unconjugated MDP alone or in combination with doxorubicin (0.0000056 - 56.5 µM) and EP 100 at 0.5, 5, 50 and 500 nM. Cytotoxicity was determined by membrane integrity and cell viability assays. LHRH receptor expression was determined by flow cytometry. The effect of EP-100 on purified human recombinant p-glycoprotein (h-pgp) pump was measured by ATPase activity. The IC50 values [µM] for EP-100 alone were 0.52±0.13, 0.95±0.2, 2.8±0.5, 0.9±0.13 and values for unconjugated MDP were 59±1.5, 25.6±1.7, 6.1±0.8 after 5 h of incubation for Daudi, Raji, Toledo, Hut78 cells, respectively. EP-100 specifically killed NHL patient cells and unconjugated MDP was ineffective. The IC50 values for EP-100 were 1.2 ± 0.1µM for cells obtained from three Mantle Cell Lymphoma patients (N=3), 2.3± 0.1 µM for Diffuse Large B Cell Lymphoma patient (N=1), 1.7± 0.3 µM for Follicular Lymphoma patients (N=4), and 1.6 ± 0.1 µM for one Waldenstr[[Unable to Display Character: ő]]m Macroglobunemia patient. EP-100 or unconjugated MDP did not kill B-cells from normal subjects (N=2) after 5 hour incubation. LHRH receptors were over-expressed on cell lines and patient cells. Combination of EP-100 with doxorubicin resulted in synergistic responses after 72 hours of incubation and Combination Index was <1. EP-100 directly inhibited the h-pgp ATPase activity. These results show that specific killing of NHL cells by EP-100 is mediated via over-expression of LHRH receptors and the synergy between EP-100 and doxorubicin is due to direct inhibition of EP-100 on h-pgp pump. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2829. doi:1538-7445.AM2012-2829

1066 EFFECTIVE TREATMENT OF URINARY BLADDER CANCERS BY TARGETED CYTOTOXIC LHRH ANALOG AEZS-108 (AN-152) A PRECLINICAL REPORT

You have accessJournal of UrologyBladder Cancer: Basic Research III1 Apr 20121066 EFFECTIVE TREATMENT OF URINARY BLADDER CANCERS BY TARGETED CYTOTOXIC LHRH ANALOG AEZS-108 (AN-152) A PRECLINICAL REPORT Ferenc Rick, Luca Szalontay, Gustavo Fernandez-Castro, Norman Block, Gunhild Keller, Karoly Szepeshazi, and Andrew Schally Ferenc RickFerenc Rick Miami, FL More articles by this author , Luca SzalontayLuca Szalontay Miami, FL More articles by this author , Gustavo Fernandez-CastroGustavo Fernandez-Castro Miami, FL More articles by this author , Norman BlockNorman Block Miami, FL More articles by this author , Gunhild KellerGunhild Keller Hamburg, Germany More articles by this author , Karoly SzepeshaziKaroly Szepeshazi Miami, FL More articles by this author , and Andrew SchallyAndrew Schally Miami, FL More articles by this author View All Author Informationhttps://doi.org/10.1016/j.juro.2012.02.1172AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookTwitterLinked InEmail INTRODUCTION AND OBJECTIVES Urinary bladder cancer is the fifth most common new cancer diagnosis and among the most expensive cancers to treat in the United States. The management of muscle-invasive tumors presents a clinical challenge because of the toxicity and limitations in durability, and efficacy of current therapeutic modalities. Novel therapeutic strategies for this disease are of paramount importance. Targeted cytotoxic luteinizing hormone-releasing hormone (LHRH or GnRH) analogs powerfully inhibit growth of various experimental cancers; they are more powerful and better tolerated than the cytotoxic radical, doxorubicin (DOX) alone. We evaluated, therefore, the effects of cytotoxic LHRH analog AN-152 on four human bladder cancers xenografted into nude mice. METHODS We investigated the effects of AN-152 on growth of HT-1376, J82, RT-4 and HT-1197 tumors xenografted into nude mice. Surgical specimens of 18 human primary urothelial bladder carcinomas were immunohistochemically stained for LHRH receptor, also. RESULTS In all four in vivo experiments, final tumor volumes (62%, 84%, 85%, and 89% inhibition, respectively; p<0.05) and tumor weights were significantly lower in the groups receiving AN-152; AN-152 had greater effect and was less toxic than DOX. Treatment with DOX alone resulted in significant reduction of tumor volume only in HT-1197 tumors. Positive staining for LHRH receptors was observed in all examined surgical specimens and enhanced staining of the plasma membrane as well as cytoplasmatic staining was detected in these malignant cells and in positive control (human pituitary). CONCLUSIONS We demonstrated the effectiveness of the potent targeted cytotoxic LHRH analog, AN-152, in inhibiting the growth of experimental models of bladder cancer in vivo. The expression of LHRH receptors was detected in all surgical specimens of human primary urothelial bladder carcinomas. Our findings warrant further development of AN-152 for clinical therapy of bladder cancer. © 2012 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 187Issue 4SApril 2012Page: e432-e433 Advertisement Copyright & Permissions© 2012 by American Urological Association Education and Research, Inc.MetricsAuthor Information Ferenc Rick Miami, FL More articles by this author Luca Szalontay Miami, FL More articles by this author Gustavo Fernandez-Castro Miami, FL More articles by this author Norman Block Miami, FL More articles by this author Gunhild Keller Hamburg, Germany More articles by this author Karoly Szepeshazi Miami, FL More articles by this author Andrew Schally Miami, FL More articles by this author Expand All Advertisement Advertisement PDF downloadLoading ...

Enhanced Anti-Tumoral Activity of Methotrexate-Human Serum Albumin Conjugated Nanoparticles by Targeting with Luteinizing Hormone-Releasing Hormone (LHRH) Peptide

Active targeting could increase the efficacy of anticancer drugs. Methotrexate-human serum albumin (MTX-HSA) conjugates, functionalized by luteinizing hormone-releasing hormone (LHRH) as targeting moieties, with the aim of specifically targeting the cancer cells, were prepared. Owing to the high expression of LHRH receptors in many cancer cells as compared to normal cells, LHRH was used as the targeting ligand in this study. LHRH was conjugated to MTX-HSA nanoparticles via a cross-linker. Three types of LHRH targeted nanoparticles with a mean particle size between 120–138 nm were prepared. The cytotoxicity of LHRH targeted and non-targeted nanoparticles were determined on the LHRH positive and negative cell lines. The internalization of the targeted and non-targeted nanoparticles in LHRH receptor positive and negative cells was investigated using flow cytometry analysis and fluorescence microscopy. The cytotoxicity of the LHRH targeted nanoparticles on the LHRH receptor positive cells were significantly more than non-targeted nanoparticles. LHRH targeted nanoparticles were also internalized by LHRH receptor positive cells significantly more than non-targeted nanoparticles. There were no significant differences between the uptake of targeted and non-targeted nanoparticles to the LHRH receptor negative cells. The active targeting procedure using LHRH targeted MTX-HSA nanoparticles could increase the anti-tumoral activity of MTX.

Effective targeted chemotherapy using AEZS-108 (AN-152) for LHRH receptor-positive pancreatic cancers

Pancreatic cancer is the fourth commonest cause of cancer-related mortality across the world. Because of the poor response to conventional chemotherapy, small molecules, radiation therapy and surgery, development of new targeted therapies is necessary. In the present study, we have analyzed expression of the luteinizing hormone releasing hormone (LHRH) receptor in specimens of human pancreatic cancers. Furthermore, we have investigated in vitro and in vivo whether the cytotoxic LHRH agonist AEZS-108 (AN-152) induces apoptosis in human pancreatic cancer cells that express LHRH receptors. LHRH receptor expression in tumor specimens of human pancreatic cancers was assessed using immunohistochemistry. Cell proliferation was analyzed using the alamar blue proliferation assay. Induction of apoptosis was analyzed using the TUNEL assay and quantified by measurement of loss of mitochondrial membrane potential. In vivo experiments were performed using nude mice bearing xenografted human pancreatic tumors. Thirteen of 40 human pancreatic adenocarcinomas (32.5%) expressed LHRH receptors. We were able to show that treatment of LHRH receptor-positive MiaPaCa-2 and Panc-1 human pancreatic cancer cells with AEZS-108 (AN-152) resulted in apoptotic cell death in vitro. The antitumor effects could be confirmed in nude mice. AEZS-108 (AN-152) inhibited the growth of xenotransplants of human pancreatic cancers in nude mice significantly, without any apparent side effects. The cytotoxic LHRH agonist AEZS-108 (AN-152) seems to be a suitable drug for treatment of LHRH receptor-positive human pancreatic cancers with little toxicity.

Luteinizing hormone-releasing hormone receptor targeted agents for prostate cancer

Introduction: Receptors for luteinizing hormone-releasing hormone (LHRH) are expressed on a variety of human cancer cells with relatively limited expression in normal tissues. The selective and persistent expression of these receptors in prostate cancer cells is the rationale for LHRH receptor targeted agents. With many agents in development and one entering early clinical study, it is important to be familiar with the concept of LHRH receptor targeting and the evidence supporting its use.Areas covered: This manuscript reviews the expression of LHRH receptors and the rationale for LHRH receptor targeted therapy in prostate cancer. Several different classes of agents targeting the LHRH receptor are discussed. The preclinical evidence supporting these agents is also reviewed and the clinical trial testing one of these agents is detailed.Expert opinion: LHRH receptor expression on prostate cancer cells has led to the rational design of many new compounds. The preclinical evidence is encouraging for these agents, which are in varying phases of development. AN-152 combines a modified LHRH agonist carrier with doxorubicin and is entering a Phase I–II clinical study.

Targeted chemotherapy for triple-negative breast cancers via LHRH receptor

Triple-negative breast cancer does not express estrogen and progesterone receptors and there is no overexpression/amplification of the HER2-neu gene. Therefore, this subtype of breast cancer lacks the benefits of specific therapies which target these receptors. About 60% of all human breast cancers express receptors for luteinizing hormone releasing hormone (LHRH, GnRH), which might be used as a target. The LHRH receptor can be used for targeted chemotherapy with cytotoxic luteinizing hormone releasing hormone agonists such as AEZS-108 (AN-152), in which doxorubicin is linked to [D-Lys6]LHRH. In the present study we have analyzed by in vitro and in vivo experiments whether the cytotoxic LHRH agonist AEZS-108 (AN-152) induces apoptosis in triple-negative human breast cancer cells that express LHRH receptors. LHRH receptor expression in tumor biopsy specimens of triple-negative breast cancers was tested using immunohistochemistry. Cell proliferation was analyzed using alamar blue proliferation assay. Induction of apoptosis was quantified by measurement of loss of mitochondrial membrane potential. In vivo experiments were performed using nude mice bearing xenografted human breast tumors.Thirty-one of 42 triple-negative breast cancers (73.8%) expressed LHRH receptors. We could show that treatment of triple-negative but LHRH-positive MDA-MB-231, HCC1806 and HCC1937 human breast cancer cells with AEZS-108 (AN-152) resulted in apoptotic cell death in vitro via activation of caspase-3. The antitumor effects were confirmed in nude mice. AEZS-108 (AN-152) inhibited the growth of xenotransplants of triple-negative human breast cancers in nude mice completely, without any apparent side effects. The cytotoxic LHRH agonist AEZS-108 (AN-152) seems to be a suitable drug for an efficacious therapy for triple-negative breast cancers with little toxicity.

Receptors for luteinizing hormone‐releasing hormone (LHRH) in benign prostatic hyperplasia (BPH) as potential molecular targets for therapy with LHRH antagonist cetrorelix

The majority of men will develop symptoms of benign prostatic hyperplasia (BPH) after 70 years of age. Various studies indicate that antagonists of LHRH, such as cetrorelix, exert direct inhibitory effects on BPH mediated by specific LHRH receptors. Our aim was to investigate the mRNA for LHRH and LHRH receptors and the expression of LHRH receptors in specimens of human BPH. The expression of mRNA for LHRH (n=35) and LHRH receptors (n=55) was investigated by RT-PCR in surgical specimens of BPH, using specific primers. The characteristics of binding sites for LHRH on 20 samples were determined by ligand competition assays. The LHRH receptor expression was also examined in 64 BPH specimens by immunohistochemistry. PCR products for LHRH were found in 18 of 35 (51%) BPH tissues and mRNA for LHRH receptors was detected in 39 of 55 (71%) BPH specimens. Eighteen of 20 (90%) samples showed a single class of high affinity binding sites for [D-Trp(6) ]LHRH with a mean K(d) of 4.04 nM and a mean B(max) of 527.6 fmol/mg membrane protein. LHRH antagonist cetrorelix showed high affinity binding to LHRH receptors in BPH. Positive immunohistochemical reaction for LHRH receptors was present in 42 of 64 (67%) BPH specimens. A high incidence of LHRH receptors in BPH supports the use of LHRH antagonists such as cetrorelix, for treatment of patients with lower urinary tract symptoms from BPH.

Expression of Receptors for Luteinizing Hormone-Releasing Hormone (LH-RH) in Prostate Cancers following Therapy with LH-RH Agonists

In addition to their expression on pituitary cells, receptors for luteinizing hormone-releasing hormone (LH-RH) are found on most prostate cancer cells. These tumoral LH-RH receptors mediate the direct cytotoxic effects of LH-RH analogs and are potential therapeutic targets. Although pituitary LH-RH receptors are downregulated following prolonged exposure to LH-RH agonists, there is no evidence that tumoral receptors behave in a similar manner. To better characterize expression of tumoral LH-RH receptors, specimens of prostate cancer from various cohorts of patients were analyzed. Surgical specimens were obtained from untreated patients with prostate cancer and from patients with metastatic castration-resistant prostate cancer previously treated with bilateral orchiectomy. To address the possibility of receptor downregulation, two additional cohorts of patients who had been previously treated with LH-RH agonists were included. One group received neoadjuvant therapy prior to prostatectomy, and the other group was treated for metastatic disease with LH-RH agonists and, at progression, required palliative resection of the prostate. Lymph node metastases from previously untreated patients were subjected to similar analysis. Expression of LH-RH receptors was found in most specimens. The relative expression of LH-RH receptor mRNA in untreated patients was greater in patients whose tumor had received a Gleason score <8. LH-RH receptor expression persisted despite prolonged exposure to LH-RH agonists. These findings support the concept of targeting cytotoxic LH-RH analogs to prostatic LH-RH receptors, using these receptors to gain entry into cancer cells to deliver a hybridized cytotoxic moiety for the treatment of prostate cancer.

Mechanisms of inhibition of human benign prostatic hyperplasia in vitro by the luteinizing hormone‐releasing hormone antagonist cetrorelix

To assess the mechanism by which the luteinizing hormone-releasing hormone (LHRH) antagonist cetrorelix exerts its effects in men with benign prostatic hyperplasia (BPH), as it produces a long-lasting improvement in lower urinary tract symptoms that is only partly accounted for by the transient reduction in testosterone levels, and the beneficial results could be due to direct inhibitory effects of cetrorelix on the prostate exerted through prostatic LHRH receptors. Using the BPH-1 cell line we evaluated the effects of cetrorelix in vitro on the proliferation and the expression of receptors for LHRH, epidermal growth factor (EGF), α(1A) -adrenergic receptor, STAT-3 transcription factor and the response to growth factors insulin-like growth factor (IGF)-1 and -II and fibroblast growth factor (FGF)-2. There was expression of LHRH receptors in the human BPH-1 cell line. Cetrorelix had inhibitory effects on the proliferation rate of BPH-1 cells, also reflected by the decrease in the expression of the proliferating cell nuclear antigen (PCNA). Cetrorelix inhibited the stimulatory effect of the growth factors IGF-I and -II and FGF-2 on the proliferation of this line. Cetrorelix also downregulated the expression of the receptors for LHRH and EGF, as well as of α(1A) -adrenergic receptors, and inhibited the activation of the STAT3 transcription factor. The results show that in vitro cetrorelix can directly inhibit the proliferation rate of the human BPH-1 cell line by counteracting growth factors like IGF-I and -II and FGF-2, and downregulating the LHRH receptor and α-adrenergic receptors, as well as transcription factors.

Targeted Drug and Gene Delivery Systems for Lung Cancer Therapy

To evaluate the efficacy of a novel docetaxel derivative of deslorelin, a luteinizing hormone-releasing hormone (LHRH) agonist, and its combination in vivo with RGD peptide conjugated nanoparticles encapsulating an antiangiogenic, anti-vascular endothelial growth factor (VEGF) intraceptor (Flt23k; RGD-Flt23k-NP) in H1299 lung cancer cells and/or xenografts in athymic nude BALB/c mice. The in vitro and in vivo efficacy of the deslorelin-docetaxel conjugate was evaluated in H1299 cells and xenografts in athymic nude mice. Coadministration of deslorelin-docetaxel conjugate and RGD-Flt23k-NP was tested in vivo in mice. Tumor inhibition, apoptosis, and VEGF inhibition were estimated in each of the treatment groups. The conjugate enhanced in vitro docetaxel efficacy by 13-fold in H1299 cells compared with docetaxel at 24 hours, and this effect was inhibited following reduction of LHRH receptor expression by an antisense oligonucleotide. Combination of the conjugate with the RGD-Flt23k-NP in vivo resulted in an 82- and 15-fold tumor growth inhibition on day 39 following repeated weekly i.v. injections and a single intratumoral (i.t.) injection, respectively. These effects were significantly greater than individual targeted therapies or docetaxel alone. Similarly, apoptotic indices for the combination therapy were 14% and 10% in the i.v. and i.t. groups, respectively, and higher than the individual therapies. Combination therapy groups exhibited greater VEGF inhibition in both the i.v. and i.t. groups. Docetaxel efficacy was enhanced by LHRH receptor-targeted deslorelin conjugate and further improved by combination with targeted antiangiogenic nanoparticle gene therapy. Combination of novel targeted therapeutic approaches described here provides an attractive alternative to the current treatment options for lung cancer therapy.