Structural and Molecular Mechanisms of Cytokine-Mediated Endocrine Resistance in Human Breast Cancer Cells.

Human breast cancers that exhibit high proportions of immune cells and elevated levels of pro-inflammatory cytokines predict poor prognosis. Here, we demonstrate that treatment of human MCF-7 breast cancer cells with pro-inflammatory cytokines results in ERα-dependent activation of gene expression and proliferation, in the absence of ligand or presence of 4OH-tamoxifen (TOT). Cytokine activation of ERα and endocrine resistance is dependent on phosphorylation of ERα at S305 in the hinge domain. Phosphorylation of S305 by IKKß establishes an ERα cistrome that substantially overlaps with the estradiol (E2)-dependent ERα cistrome. Structural analyses suggest that S305-P forms a charge-linked bridge with the C-terminal F domain of ERα that enables inter-domain communication and constitutive activity from the N-terminal coactivator-binding site, revealing the structural basis of endocrine resistance. ERα therefore functions as a transcriptional effector of cytokine-induced IKKß signaling, suggesting a mechanism through which the tumor microenvironment controls tumor progression and endocrine resistance.

Leukaemia-related protein 16 is highly expressed in oestrogen-dependent endometrial carcinoma and potentially promotes Ishikawa human endometrial cancer cells growth - a histopathological study.

Leukaemia-related protein 16 (LRP16) has been found to be highly expressed in various tumours and to be related to poor prognosis. However, the role of LRP16 in endometrial carcinoma remains to be explored. We aimed to investigate the prognosis and role of LRP16 in endometrial carcinoma. Overall, 160 endometrial carcinoma (EC) tissues and 60 benign samples were collected. The expression of LRP16 protein in EC tissues was significantly increased compared with that in normal endometrial tissues, and high LRP16 expression was related to poor patient prognosis. Reduced LRP16 expression markedly inhibited cancer cell growth. The proliferation rates in the prophylactic bilateral salpingectomy (PBS) group and the shNon group were 0.727 ±0.015 and 0.743 ±0.009, respectively, while the proliferation rate in the shLRP16 group was only 0.373 ±0.012. The migration experiment showed that the number of cells passing through the basement membrane in the shLRP16 group was 34.2 ±5.1, which was significantly different to the shNon (161.6 ±7.8) and PBS groups (138.0 ±7.2). The results of the invasion experiment showed that the number of cells was 39.2 ±6.2 in the shLRP16 group, 146.7 ±8.2 in the shNon group, and 141.2 ±8.1 in the PBS group ( p < 0.05). Leukaemia-related protein 16 is highly expressed in oestrogen-dependent EC and may promote cancer cell growth.

Impact of FKBP52 on cell proliferation and hormone-dependent cancers.

FK506 binding protein 52 (FKBP52) (gene name FKBP4) is a 52 kDa protein that belongs to the FKBP family; it binds to the immunosuppressant FK506 and has proline isomerase activity. In addition to its FK domain-containing peptidylprolyl isomerase activity, FKBP52 also acts as a cochaperone through the tetratricopeptide repeat domain that mediates binding to heat shock protein 90. Previous studies have reported that FKBP52 is associated with hormone-dependent, stress-related, and neurodegenerative diseases, revealing its diverse functions. In particular, the effects of FKBP52 on cancer have attracted considerable attention. FKBP52 promotes the growth of hormone-dependent cancers by activating steroid hormone receptors. Recent studies have shown that the expression of FKBP52 is increased not only in steroid hormone-dependent cancer cells but also in colorectal, lung, and liver cancers, revealing its diverse functions that contribute to cancer growth. This review summarizes reports related to hormone-dependent cancer and cell proliferation in terms of the structure of FKBP52 and its function on interacting molecules.

Estrogen Receptors-Mediated Apoptosis in Hormone-Dependent Cancers.

It is known that estrogen stimulates growth and inhibits apoptosis through estrogen receptor(ER)-mediated mechanisms in many cancer cell types. Interestingly, there is strong evidence that estrogens can also induce apoptosis, activating different ER isoforms in cancer cells. It has been observed that E2/ERα complex activates multiple pathways involved in both cell cycle progression and apoptotic cascade prevention, while E2/ERß complex in many cases directs the cells to apoptosis. However, the exact mechanism of estrogen-induced tumor regression is not completely known. Nevertheless, ERs expression levels of specific splice variants and their cellular localization differentially affect outcome of estrogen-dependent tumors. The goal of this review is to provide a general overview of current knowledge on ERs-mediated apoptosis that occurs in main hormone dependent-cancers. Understanding the molecular mechanisms underlying the induction of ER-mediated cell death will be useful for the development of specific ligands capable of triggering apoptosis to counteract estrogen-dependent tumor growth.

Genomic profiles and clinical outcomes in primary versus secondary metastatic hormone-sensitive prostate cancer.

BACKGROUND: Clinical outcomes may differ among patients presenting with primary (de novo) metastatic hormone-sensitive prostate cancer (mHSPC) versus secondary (metachronous) mHSPC occurring after local therapy. It is unknown what molecular features distinguish these potentially distinct presentations. METHODS: A single-center retrospective study of mHSPC patients classified as primary mHSPC (n = 121) or secondary mHSPC (n = 106). A targeted set of genes was analyzed: BRCA2, PTEN, RB1, TP53, SPOP, CDK12, any two out of PTEN/RB1/TP53 alterations, and homologous recombination deficiency mutations. TP53 mutations were categorized as loss-of-function (LOF) versus dominant-negative (DN). The impacts of genetic features on progression-free survival (PFS) and overall survival (OS) were assessed using univariate and multivariate Cox proportional hazards regression. RESULTS: Median PFS was 15 and 30 months for men with primary and secondary mHSPC, respectively (hazard ratio: 0.57, 95% confidence interval: 0.41-0.78; p < .01). OS did not show a significant difference between groups. There were more men with Gleason 8-10 disease in the primary versus secondary mHSPC groups (83% vs. 68%; p < .01). In univariate and multivariate analyses, TP53 DN mutations showed a statistically significant association with OS for the entire mHSPC population. Conversely, SPOP mutations were associated with improved OS. Additionally, TP53 mutations (DN and LOF) were associated with worse OS for secondary mHSPC. A combination of PTEN/RB1/TP53 mutations was associated with worse OS and PFS for secondary mHSPC, while no genomic alteration affected outcomes for primary mHSPC. CONCLUSIONS: TP53 DN mutations, but not all TP53 alterations, were the strongest predictor of negative outcomes in men with mHSPC, while SPOP mutations were associated with improved outcomes. In subgroup analyses, specific alterations were prognostic of outcomes in secondary, but not primary, mHSPC.

Molecular pathogenesis of breast cancer: impact of miR-99a-5p and miR-99a-3p regulation on oncogenic genes.

Our recent research has revealed that passenger strands of certain microRNAs (miRNAs) function as tumor-suppressive miRNAs in cancer cells, e.g., miR-101-5p, miR-143-5p, miR-144-5p, miR-145-3p, and miR-150-3p. Thus, they are important in cancer pathogenesis. Analysis of the miRNA expression signature of breast cancer (BrCa) showed that the expression levels of two miRNAs derived from pre-miR-99a (miR-99a-5p and miR-99a-3p) were suppressed in cancerous tissues. The aim of this study was to identify oncogenic genes controlled by pre-miR-99a that are closely involved in the molecular pathogenesis of BrCa. A total of 113 genes were identified as targets of pre-miR-99a regulation (19 genes modulated by miR-99a-5p, and 95 genes regulated by miR-99a-3p) in BrCa cells. Notably, FAM64A was targeted by both of the miRNAs. Among these targets, high expression of 16 genes (C5orf22, YOD1, SLBP, F11R, C12orf49, SRPK1, ZNF250, ZNF695, CDK1, DNMT3B, TRIM25, MCM4, CDKN3, PRPS, FAM64A, and DESI2) significantly predicted reduced survival of BrCa patients based upon The Cancer Genome Atlas (TCGA) database. In this study, we focused on FAM64A and investigated the relationship between FAM64A expression and molecular pathogenesis of BrCa subtypes. The upregulation of FAM64A was confirmed in BrCa clinical specimens. Importantly, the expression of FAM64A significantly differed between patients with Luminal-A and Luminal-B subtypes. Our data strongly suggest that the aberrant expression of FAM64A is involved in the malignant transformation of BrCa. Our miRNA-based approaches (identification of tumor-suppressive miRNAs and their controlled targets) will provide novel information regarding the molecular pathogenesis of BrCa.

Genetic Contribution of Endometriosis to the Risk of Developing Hormone-Related Cancers.

Endometriosis is a common gynecological disorder that has been associated with endometrial, breast and epithelial ovarian cancers in epidemiological studies. Since complex diseases are a result of multiple environmental and genetic factors, we hypothesized that the biological mechanism underlying their comorbidity might be explained, at least in part, by shared genetics. To assess their potential genetic relationship, we performed a two-sample mendelian randomization (2SMR) analysis on results from public genome-wide association studies (GWAS). This analysis confirmed previously reported genetic pleiotropy between endometriosis and endometrial cancer. We present robust evidence supporting a causal genetic association between endometriosis and ovarian cancer, particularly with the clear cell and endometrioid subtypes. Our study also identified genetic variants that could explain those associations, opening the door to further functional experiments. Overall, this work demonstrates the value of genomic analyses to support epidemiological data, and to identify targets of relevance in multiple disorders.

Long-term outcome and prognostic value of Ki67 after perioperative endocrine therapy in postmenopausal women with hormone-sensitive early breast cancer (POETIC): an open-label, multicentre, parallel-group, randomised, phase 3 trial.

BACKGROUND: Preoperative and perioperative aromatase inhibitor (POAI) therapy has the potential to improve outcomes in women with operable oestrogen receptor-positive primary breast cancer. It has also been suggested that tumour Ki67 values after 2 weeks (Ki672W) of POAI predicts individual patient outcome better than baseline Ki67 (Ki67B). The POETIC trial aimed to test these two hypotheses. METHODS: POETIC was an open-label, multicentre, parallel-group, randomised, phase 3 trial (done in 130 UK hospitals) in which postmenopausal women aged at least 50 years with WHO performance status 0-1 and hormone receptor-positive, operable breast cancer were randomly assigned (2:1) to POAI (letrozole 2·5 mg per day orally or anastrozole 1 mg per day orally) for 14 days before and following surgery or no POAI (control). Adjuvant treatment was given as per UK standard local practice. Randomisation was done centrally by computer-generated permuted block method (variable block size of six or nine) and was stratified by hospital. Treatment allocation was not masked. The primary endpoint was time to recurrence. A key second objective explored association between Ki67 (dichotomised at 10%) and disease outcomes. The primary analysis for clinical endpoints was by modified intention to treat (excluding patients who withdrew consent). For Ki67 biomarker association and endpoint analysis, the evaluable population included all randomly assigned patients who had paired Ki67 values available. This study is registered with ClinicalTrials.gov, NCT02338310; the European Clinical Trials database, EudraCT2007-003877-21; and the ISRCTN registry, ISRCTN63882543. Recruitment is complete and long-term follow-up is ongoing. FINDINGS: Between Oct 13, 2008, and April 16, 2014, 4480 women were recruited and randomly assigned to POAI (n=2976) or control (n=1504). On Feb 6, 2018, median follow-up was 62·9 months (IQR 58·1-74·1). 434 (10%) of 4480 women had a breast cancer recurrence (280 [9%] POAI; 154 [10%] control), hazard ratio 0·92 (95% CI 0·75-1·12); p=0·40 with the proportion free from breast cancer recurrence at 5 years of 91·0% (95% CI 89·9-92·0) for patients in the POAI group and 90·4% (88·7-91·9) in the control group. Within the POAI-treated HER2-negative subpopulation, 5-year recurrence risk in women with low Ki67B and Ki672W (low-low) was 4·3% (95% CI 2·9-6·3), 8·4% (6·8-10·5) with high Ki67B and low Ki672W (high-low) and 21·5% (17·1-27·0) with high Ki67B and Ki672W (high-high). Within the POAI-treated HER2-positive subpopulation, 5-year recurrence risk in the low-low group was 10·1% (95% CI 3·2-31·3), 7·7% (3·4-17·5) in the high-low group, and 15·7% (10·1-24·4) in the high-high group. The most commonly reported grade 3 adverse events were hot flushes (20 [1%] of 2801 patients in the POAI group vs six [<1%] of 1400 in the control group) and musculoskeletal pain (29 [1%] vs 13 [1%]). No treatment-related deaths were reported. INTERPRETATION: POAI has not been shown to improve treatment outcome, but can be used without detriment to help select appropriate adjuvant therapy based on tumour Ki67. Most patients with low Ki67B or low POAI-induced Ki672W do well with adjuvant standard endocrine therapy (giving consideration to clinical-pathological factors), whereas those whose POAI-induced Ki672W remains high might benefit from further adjuvant treatment or trials of new therapies. FUNDING: Cancer Research UK.

ESR1 mutations are frequent in newly diagnosed metastatic and loco-regional recurrence of endocrine-treated breast cancer and carry worse prognosis.

BACKGROUND: Emerging mutations in the ESR1 gene that encodes for the estrogen receptor (ER) are associated with resistance to endocrine therapy. ESR1 mutations rarely exist in primary tumors (~ 1%) but are relatively common (10-50%) in metastatic, endocrine therapy-resistant cancers and are associated with a shorter progression-free survival. Little is known about the incidence and clinical implication of these mutations in early recurrence events, such as local recurrences or newly diagnosed metastatic disease. METHODS: We collected 130 archival tumor samples from 103 breast cancer patients treated with endocrine therapy prior to their local/metastatic recurrence. The cohort consisted of 41 patients having at least 1 sample from local/loco-regional recurrence and 62 patients with metastatic disease (of whom 41 newly diagnosed and 28 with advanced disease). The 5 most common ESR1 hotspot mutations (D538G, L536R, Y537S/N/C) were analyzed either by targeted sequencing or by droplet digital PCR. Progression-free survival (PFS), disease-free survival (DFS), and distant recurrence-free survival (DRFS) were statistically tested by Kaplan-Meier analysis. RESULTS: The prevalence of ESR1 mutations was 5/41 (12%) in newly diagnosed metastatic patients and 5/28 (18%) for advanced metastases, detected at allele frequency > 1%. All mutations in advanced metastases were detected in patients previously treated with both tamoxifen (TAM) and aromatase inhibitors (AI). However, in newly diagnosed metastatic patients, 4/5 mutations occurred in patients treated with TAM alone. PFS on AI treatment in metastatic patients was significantly shorter for ESR1 mutation carriers (p = 0.017). In the local recurrence cohort, ESR1 mutations were identified in 15/41 (36%) patients but only 4/41 (10%) were detected at allele frequency > 1%. Again, most mutations (3/4) were detected under TAM monotherapy. Notably, 1 patient developed ESR1 mutation while on neoadjuvant endocrine therapy. DFS and DRFS were significantly shorter (p = 0.04 and p = 0.017, respectively) in patients that had ESR1 mutations (> 1%) in their loco-regional recurrence tumor. CONCLUSIONS: Clinically relevant ESR1 mutations are prevalent in newly diagnosed metastatic and local recurrence of endocrine-treated breast cancer. Since local recurrences are amenable to curative therapy, these mutations may inform the selection of subsequent endocrine therapies.

TBCRC 002: a phase II, randomized, open-label trial of preoperative letrozole with or without bevacizumab in postmenopausal women with newly diagnosed stage 2/3 hormone receptor-positive and HER2-negative breast cancer.

BACKGROUND: In preclinical studies, the expression of vascular endothelial growth factor (VEGF) in hormone receptor-positive breast cancer is associated with estrogen-independent tumor growth and resistance to endocrine therapies. This study investigated whether the addition of bevacizumab, a monoclonal antibody against VEGF, to letrozole enhanced the antitumor activity of the letrozole in the preoperative setting. METHODS: Postmenopausal women with newly diagnosed stage 2 or 3 estrogen and/or progesterone receptor-positive, HER2-negative breast cancer were randomly assigned (2:1) between letrozole 2.5 mg PO daily plus bevacizumab 15 mg/kg IV every 3 weeks (Let/Bev) and letrozole 2.5 mg PO daily (Let) for 24 weeks prior to definitive surgery. Primary objective was within-arm pathologic complete remission (pCR) rate. Secondary objectives were safety, objective response, and downstaging rate. RESULTS: Seventy-five patients were randomized (Let/Bev n = 50, Let n = 25). Of the 45 patients evaluable for pathological response in the Let/Bev arm, 5 (11%; 95% CI, 3.7-24.1%) achieved pCR and 4 (9%; 95% CI, 2.5-21.2%) had microscopic residual disease; no pCRs or microscopic residual disease was seen in the Let arm (0%; 95% CI, 0-14.2%). The rates of downstaging were 44.4% (95% CI, 29.6-60.0%) and 37.5% (95% CI, 18.8-59.4%) in the Let/Bev and Let arms, respectively. Adverse events typically associated with letrozole (hot flashes, arthralgias, fatigue, myalgias) occurred in similar frequencies in the two arms. Hypertension, headache, and proteinuria were seen exclusively in the Let/Bev arm. The rates of grade 3 and 4 adverse events and discontinuation due to adverse events were 18% vs 8% and 16% vs none in the Let/Bev and Let arms, respectively. A small RNA-based classifier predictive of response to preoperative Let/Bev was developed and confirmed on an independent cohort. CONCLUSION: In the preoperative setting, the addition of bevacizumab to letrozole was associated with a pCR rate of 11%; no pCR was seen with letrozole alone. There was additive toxicity with the incorporation of bevacizumab. Responses to Let/Bev can be predicted from the levels of 5 small RNAs in a pretreatment biopsy. TRIAL REGISTRATION: This trial is registered with ClinicalTrials.gov (Identifier: NCT00161291), first posted on September 12, 2005, and is completed.

Levels of different subtypes of tumour-infiltrating lymphocytes correlate with each other, with matched circulating lymphocytes, and with survival in breast cancer.

PURPOSE: Breast cancer tumour-infiltrating lymphocytes associate with clinico-pathological factors, including survival, although the literature includes many conflicting findings. Our aim was to assess these associations for key lymphocyte subtypes and in different tumour compartments, to determine whether these provide differential correlations and could, therefore, explain published inconsistencies. Uniquely, we also examine whether infiltrating levels merely reflect systemic lymphocyte levels or whether local factors are predominant in recruitment. METHODS: Immunohistochemistry was used to detect tumour-infiltrating CD20+ (B), CD4+ (helper T), CD8+ (cytotoxic T) and FoxP3+ (regulatory T) cells in breast cancers from 62 patients, with quantification in tumour stroma, tumour cell nests, and tumour margins. Levels were analysed with respect to clinico-pathological characteristics and matched circulating levels (determined by flow-cytometry). RESULTS: CD4+ lymphocytes were the most prevalent subtype in tumour stroma and at tumour edge and CD8+ lymphocytes were most prevalent in tumour nests; FoxP3+ lymphocytes were rarest in all compartments. High grade or hormone receptor negative tumours generally had significantly increased lymphocytes, especially in tumour stroma. Only intra-tumoural levels of CD8+ lymphocytes correlated significantly with matched circulating levels (p < 0.03), suggesting that recruitment is mainly unrelated to systemic activity. High levels of stromal CD4+ and CD20+ cells associated with improved survival in hormone receptor negative cases (p < 0.04), while tumour nest CD8+ and FoxP3+ cells associated with poor survival in hormone receptor positives (p < 0.005). CONCLUSIONS: Lymphocyte subtype and location define differential impacts on tumour biology, therefore, roles of tumour-infiltrating lymphocytes will only be unravelled through thorough analyses that take this into account.

Use of MarginProbe as an adjunct to standard operating procedure does not significantly reduce re-excision rates in breast conserving surgery.

PURPOSE: A positive margin after breast conserving surgery has consistently been shown to be a significant predictor for ipsilateral breast tumor recurrence. Currently, there is no standard for intraoperative margin assessment during lumpectomy, and up to 20% of cases result in positive margins. MarginProbe is a device that provides real-time evaluation of lumpectomy margins during surgery. The aim of this study was to evaluate the impact of MarginProbe as an adjunct to standard operating procedure (SOP). METHODS: Patients diagnosed with breast cancer scheduled for breast conserving surgery were consented for intraoperative use of MarginProbe. Shaved margins were excised based on margin assessment using the surgeon's SOP which included specimen radiography and gross pathologic examination, and feedback from the device. The primary endpoint was re-excision rate. Secondary endpoints included sensitivity, specificity, false-positive and negative rates. RESULTS: Of the 60 breast cancers, initial histologically close/positive margins were identified in 18 patients (30%). The re-excision rate in the overall cohort was 6.6%, compared to a historical re-excision rate of 8.6% (p < 0.01). Based on 360 measurement sites, MarginProbe demonstrated a sensitivity of 67% and specificity of 60%, with a positive predictive value of 16%, and of negative predictive value of 94%, which was similar to the accuracy of SOP. CONCLUSIONS: MarginProbe performs equally as well as specimen radiography and gross pathologic examination. In this setting where the baseline re-excision rate was low, the use of MarginProbe as an adjunct to SOP resulted in a small 2% absolute reduction in re-excision rate.

Estrogen, progesterone, and human epidermal growth factor receptor 2 discordance between primary and metastatic breast cancer.

BACKGROUND: The estrogen receptor (ER), progesterone receptor (PR), and human epidermal growth factor receptor 2 (HER2) statuses are frequently discordant between the primary tumor and metastatic lesions in metastatic breast cancer. This can have important therapeutic implications. PATIENTS AND METHODS: In all, 541 patients with available receptor statuses from both primary tumor and metastatic lesion treated at Heidelberg and Tuebingen University Hospitals between 1982 and 2018 were included. RESULTS: Statistically significant discordance rates of 14% and 32% were found for ER and PR. HER2 status was statistically insignificantly discordant in 15% of patients. Gain in HER2 positivity was associated with an improved overall survival, whereas loss of HR positivity was associated with worse overall survival. Antiendocrine treatment differed in 20% of cases before and after biopsy and HER2-directed treatment in 14% of cases. CONCLUSIONS: Receptor statuses are discordant between primary tumor and metastasis in a considerable fraction of patients with metastatic breast cancer. Next to a highly presumed predictive value with respect to efficacy of endocrine and HER2-targeted therapy, discordance seems to provide prognostically relevant information. Where feasible, metastatic lesions should be biopsied in accordance with current guidelines.

G1T48, an oral selective estrogen receptor degrader, and the CDK4/6 inhibitor lerociclib inhibit tumor growth in animal models of endocrine-resistant breast cancer.

PURPOSE: The combination of targeting the CDK4/6 and estrogen receptor (ER) signaling pathways with palbociclib and fulvestrant is a proven therapeutic strategy for the treatment of ER-positive breast cancer. However, the poor physicochemical properties of fulvestrant require monthly intramuscular injections to patients, which limit the pharmacokinetic and pharmacodynamic activity of the compound. Therefore, an orally available compound that more rapidly reaches steady state may lead to a better clinical response in patients. Here, we report the identification of G1T48, a novel orally bioavailable, non-steroidal small molecule antagonist of ER. METHODS: The pharmacological effects and the antineoplastic mechanism of action of G1T48 on tumors was evaluated using human breast cancer cells (in vitro) and xenograft efficacy models (in vivo). RESULTS: G1T48 is a potent and efficacious inhibitor of estrogen-mediated transcription and proliferation in ER-positive breast cancer cells, similar to the pure antiestrogen fulvestrant. In addition, G1T48 can effectively suppress ER activity in multiple models of endocrine therapy resistance including those harboring ER mutations and growth factor activation. In vivo, G1T48 has robust antitumor activity in a model of estrogen-dependent breast cancer (MCF7) and significantly inhibited the growth of tamoxifen-resistant (TamR), long-term estrogen-deprived (LTED) and patient-derived xenograft tumors with an increased response being observed with the combination of G1T48 and the CDK4/6 inhibitor lerociclib. CONCLUSIONS: These data show that G1T48 has the potential to be an efficacious oral antineoplastic agent in ER-positive breast cancer.

Pretreatment elevated prognostic nutritional index predicts a favorable prognosis in patients with prostate cancer.

BACKGROUND: The prognostic nutritional index (PNI), an immunity and nutrition based prognostic score, was correlated with clinical outcomes in different tumors. However, the prognostic significance of PNI has not been investigated in hormone sensitive prostate cancer (PCa). The objective of this study was to determine the prognostic significance of PNI in hormone sensitive PCa. METHODS: Two hundred eighty PCa patients undergoing androgen deprivation therapy (ADT) as first line therapy at three centers were enrolled. The serum albumin levels and peripheral lymphocyte count were measured at the time of diagnosis. PNI was calculated as 10 * serum albumin (g/dL) + 0.005 * total lymphocyte count (per mm3). Patients were categorized in two groups using a cut-off point of 50.2 as calculated by the receiver-operating curve analysis. Univariate and multivariate cox regression analyses were performed to evaluate PNI as a favorable prognostic factor for progression-free survival (PFS), cancer-specific survival (CSS) and overall survival (OS). Prognostic accuracy was evaluated with the Harrell concordance index. RESULTS: Multivariate analyses identified PNI as an independent prognostic indicator with respect to PFS (hazard ratio (HR) = 0.521, p = 0.001), CSS (HR = 0.421, p = 0.002) and OS (HR = 0.429, p = 0.001). Patients with elevated PNI had better clinical outcomes. The addition of PNI to the final models improved predictive accuracy (c-index: 0.758, 0.830 and 0.782) for PFS, CSS and OS compared with the clinicopathological base models (c-index: 0.736, 0.801 and 0.752), which included Gleason score and incidence of metastasis. CONCLUSIONS: Elevated pretreatment PNI was a favorable prognostic indicator for PCa patients treated with ADT.

Lutetium-177-PSMA-I&T as metastases directed therapy in oligometastatic hormone sensitive prostate cancer, a randomized controlled trial.

BACKGROUND: In recent years, there is increasing evidence showing a beneficial outcome (e.g. progression free survival; PFS) after metastases-directed therapy (MDT) with external beam radiotherapy (EBRT) or targeted surgery for oligometastatic hormone sensitive prostate cancer (oHSPC). However, many patients do not qualify for these treatments due to prior interventions or tumor location. Such oligometastatic patients could benefit from radioligand therapy (RLT) with 177Lu-PSMA; a novel tumor targeting therapy for end-stage metastatic castration-resistant prostate cancer (mCRPC). Especially because RLT could be more effective in low volume disease, such as the oligometastatic status, due to high uptake of radioligands in smaller lesions. To test the hypothesis that 177Lu-PSMA is an effective treatment in oHSPC to prolong PFS and postpone the need for androgen deprivation therapy (ADT), we initiated a multicenter randomized clinical trial. This is globally, the first prospective study using 177Lu-PSMA-I&T in a randomized multicenter setting. METHODS & DESIGN: This study compares 177Lu-PSMA-I&T MDT to the current standard of care (SOC); deferred ADT. Fifty-eight patients with oHSPC (≤5 metastases on PSMA PET) and high PSMA uptake (SUVmax > 15, partial volume corrected) on 18F-PSMA PET after prior surgery and/or EBRT and a PSA doubling time of < 6 months, will be randomized in a 1:1 ratio. The patients randomized to the interventional arm will be eligible for two cycles of 7.4GBq 177Lu-PSMA-I&T at a 6-week interval. After both cycles, patients are monitored every 3 weeks (including adverse events, QoL- and xerostomia questionnaires and laboratory testing) at the outpatient clinic. Twenty-four weeks after cycle two an end of study evaluation is planned together with another 18F-PSMA PET and (whole body) MRI. Patients in the SOC arm are eligible to receive 177Lu-PSMA-I&T after meeting the primary study objective, which is the fraction of patients who show disease progression during the study follow up. A second primary objective is the time to disease progression. Disease progression is defined as a 100% increase in PSA from baseline or clinical progression. DISCUSSION: This is the first prospective randomized clinical study assessing the therapeutic efficacy and toxicity of 177Lu-PSMA-I&T for patients with oHSPC. TRIAL REGISTRATION: Clinicaltrials.gov identifier: NCT04443062 .

Three-Dimensional Cell Cultures as an In Vitro Tool for Prostate Cancer Modeling and Drug Discovery.

In the last decade, three-dimensional (3D) cell culture technology has gained a lot of interest due to its ability to better recapitulate the in vivo organization and microenvironment of in vitro cultured cancer cells. In particular, 3D tumor models have demonstrated several different characteristics compared with traditional two-dimensional (2D) cultures and have provided an interesting link between the latter and animal experiments. Indeed, 3D cell cultures represent a useful platform for the identification of the biological features of cancer cells as well as for the screening of novel antitumor agents. The present review is aimed at summarizing the most common 3D cell culture methods and applications, with a focus on prostate cancer modeling and drug discovery.

Sequencing strategies in the new treatment landscape of prostate cancer.

Over the last 10 years, a number of new agents approved for the treatment of metastatic castration-resistant prostate cancer have led to a significant improvement in overall survival. The addition of new agents to androgen deprivation therapy has also allowed a paradigmatic change in the treatment of metastatic hormone-sensitive prostate cancer by improving overall survival in comparison with androgen deprivation therapy alone. Furthermore, recent data concerning the efficacy of three different androgen receptor-targeting agents in patients with nonmetastatic castration-resistant prostate cancer have opened up new scenarios for future patients' management. Defining the best sequencing strategies for men with prostate cancer is a currently unmet medical need, and choosing treatment is often challenging for clinicians because of the lack of direct comparisons of the available agents. The aim of this paper is to provide a comprehensive review of the literature concerning current sequencing strategies for prostate cancer patients.

Targeting tumour re-wiring by triple blockade of mTORC1, epidermal growth factor, and oestrogen receptor signalling pathways in endocrine-resistant breast cancer.

BACKGROUND: Endocrine therapies are the mainstay of treatment for oestrogen receptor (ER)-positive (ER+) breast cancer (BC). However, resistance remains problematic largely due to enhanced cross-talk between ER and growth factor pathways, circumventing the need for steroid hormones. Previously, we reported the anti-proliferative effect of everolimus (RAD001-mTORC1 inhibitor) with endocrine therapy in resistance models; however, potential routes of escape from treatment via ERBB2/3 signalling were observed. We hypothesised that combined targeting of three cellular nodes (ER, ERBB, and mTORC1) may provide enhanced long-term clinical utility. METHODS: A panel of ER+ BC cell lines adapted to long-term oestrogen deprivation (LTED) and expressing ESR1 wt or ESR1 Y537S , modelling acquired resistance to an aromatase-inhibitor (AI), were treated in vitro with a combination of RAD001 and neratinib (pan-ERBB inhibitor) in the presence or absence of oestradiol (E2), tamoxifen (4-OHT), or fulvestrant (ICI182780). End points included proliferation, cell signalling, cell cycle, and effect on ER-mediated transactivation. An in-vivo model of AI resistance was treated with monotherapies and combinations to assess the efficacy in delaying tumour progression. RNA-seq analysis was performed to identify changes in global gene expression as a result of the indicated therapies. RESULTS: Here, we show RAD001 and neratinib (pan-ERBB inhibitor) caused a concentration-dependent decrease in proliferation, irrespective of the ESR1 mutation status. The combination of either agent with endocrine therapy further reduced proliferation but the maximum effect was observed with a triple combination of RAD001, neratinib, and endocrine therapy. In the absence of oestrogen, RAD001 caused a reduction in ER-mediated transcription in the majority of the cell lines, which associated with a decrease in recruitment of ER to an oestrogen-response element on the TFF1 promoter. Contrastingly, neratinib increased both ER-mediated transactivation and ER recruitment, an effect reduced by the addition of RAD001. In-vivo analysis of an LTED model showed the triple combination of RAD001, neratinib, and fulvestrant was most effective at reducing tumour volume. Gene set enrichment analysis revealed that the addition of neratinib negated the epidermal growth factor (EGF)/EGF receptor feedback loops associated with RAD001. CONCLUSIONS: Our data support the combination of therapies targeting ERBB2/3 and mTORC1 signalling, together with fulvestrant, in patients who relapse on endocrine therapy and retain a functional ER.

Abiraterone shows alternate activity in models of endocrine resistant and sensitive disease.

BACKGROUND: Resistance to endocrine therapy remains a major clinical problem in the treatment of oestrogen-receptor positive (ER+) breast cancer. Studies show androgen-receptor (AR) remains present in 80-90% of metastatic breast cancers providing support for blockade of AR-signalling. However, clinical studies with abiraterone, which blocks cytochrome P450 17A1 (CYP17A1) showed limited benefit. METHODS: In order to address this, we assessed the impact of abiraterone on cell-viability, cell-death, ER-mediated transactivation and recruitment to target promoters. together with ligand-binding assays in a panel of ER+ breast cancer cell lines that were either oestrogen-dependent, modelling endocrine-sensitive disease, or oestrogen-independent modelling relapse on an aromatase inhibitor. The latter, harboured wild-type (wt) or naturally occurring ESR1 mutations. RESULTS: Similar to oestrogen, abiraterone showed paradoxical impact on proliferation by stimulating cell growth or death, depending on whether the cells are hormone-dependent or have undergone prolonged oestrogen-deprivation, respectively. Abiraterone increased ER-turnover, induced ER-mediated transactivation and ER-degradation via the proteasome. CONCLUSIONS: Our study confirms the oestrogenic activity of abiraterone and highlights its differential impact on cells dependent on oestrogen for their proliferation vs. those that are ligand-independent and harbour wt or mutant ESR1. These properties could impact the clinical efficacy of abiraterone in breast cancer.