Archivio Italiano di Urologia e Andrologia 2017; 89, 112 ORIGINAL PAPER Daidzein plus isolase associated with zinc improves clinical symptoms and quality of life in patients with LUTS due to benign prostatic hyperplasia: Results from a phase I-II study Daniele Tiscione 1, Luca Gallelli 2, Irene Tamanini 1, Lorenzo Giuseppe Luciani 1, Paolo Verze 3, Alessandro Palmieri 3, Vincenzo Mirone 3, Riccardo Bartoletti 4, Gianni Malossini 1, Tommaso Cai 1 1 Department of Urology, Santa Chiara Regional Hospital, Trento, Italy; 2 Department of Health Science, School of Medicine, University of Catanzaro, Catanzaro, Italy; 3 Department of Urology, University of Naples, Federico II, Naples, Italy; 4 Department of Urology, University of Pisa, Pisa, Italy. Objective: In the last years there is a grow- ing interest in nutraceutical substances that seems able to improve clinical symptoms in patients with lower urinary tract symptoms (LUTS) due to benign prostatic hyperplasia (BPH). In this paper, we evaluated both efficacy and safety of a combination of daidzein with isolase and zinc in patients with LUTS due to BPH. Materials and methods: In a phase I-II study clinical trial we enrolled patients with clinical and instrumental diagnosis of LUTS associated to BPH that received a six-month treatment with a combination of daidzein with isolase and zinc (1 tablet/day). Clinical, laboratory and instrumental analyses were carried out at the time of admission (T0) and 6 months after the ending of the treatment (T1). The Italian version of International Prostatic Symptom Score (IPSS), International Index of Erectile Function (IIEF-5) and Quality of Well-Being (QoL) questionnaires were used. The development of adverse drug reactions (ADRs) and drug interactions (DDIs) were recorded using the Naranjo scale and drug interaction proba- bility scale. Student’s t test and Anova test were used for sta- tistical analysis, and the threshold of statistical significance was set at P < 0.05. Results: We enrolled 71 patients, 62 (87.3%) completed the follow-up and we documented a significant differences between T0 and T1 in terms of IPSS [21.5 ± 1.2 vs 16.2 ± 1.5; (-4.8); p < 0.001], Cmax [9.7 ± 3.7vs 15.3 ± 2.5; (+5.6); p < 0.001] and QoL [0.56 ± 0.15 vs 0.84 ± 0.19; (+0.28); p < 0.001]. In contrast, no significant difference were recorded in terms of IIEF-5 [p = 0.50] and PSA [p = 0.67]. Finally, we did not record any significant ADRs or DDIs during the study. Conclusions: In this study, we documented that a combination of daidzein with isolase and zinc, reduces the clinical symp- toms of LUTS and improves the quality of life in patients with BPH, without the development of ADRs or DDIs. KEY WORDS: BPH; Zinc; LUTS; Equol; Isoflavones; Medical treatment. Submitted 20 January 2017; Accepted 18 February 2017 Summary No conflict of interest declared. vant lower urinary tract symptoms (LUTS) that can reduce the quality of life (QoL) leading to serious clini- cal manifestations, e.g. acute urinary retention, urinary incontinence, recurrent urinary tract infection, obstruc- tive uropathy, up to surgery (2). To date, two main cat- egories of drugs are prescribed to treat patients with symptomatic BPH: 1) α1-adrenoreceptor antagonists that improve the dynamic component of urination (acti- vation of bladder smooth muscles) and relieve the symp- toms (3); 2) 5-ARIs that block the conversion of testos- terone to dihydrotestosterone, and deprive the prostatic tissue of trophic androgenic influence (3). The guide- lines for BPH/LUTS recommend a combination therapy with α-blockers and 5ARI in patients with moderate-to- severe LUTS and enlarged prostates (3). Unfortunately, these treatments may be related with the development of ADRs that can induce the discontinuation of the current therapy with an impaired of both clinical symptoms and quality of life (4). Even if the current therapeutic strate- gies for the management of patients affected by LUTS associated to BPH are able to both control the progres- sion of the disease and relief the symptoms, their use is related with the development of adverse drug reactions (ADRs) particularly on sexual function (5). In a recent meta-analysis Liu and co-workers, evaluating the effects of 5α-Reductase inhibitors (5ARIs) on sexual function in men with BPH, documented that both dutasteride and finasteride compared with placebo were associated with the development of sexual dysfunction (6). The develop- ment of ADRs represents a common problem during drug administration and represents the most common cause of low adherence to the treatment (7). In the last years, there is a grow interest for nutraceutical agents in several clinical manifestations for several reasons, e.g. low side-effect and costs, high level of adherence and a low rate of efficacy of standard treatments with subse- quent patient disappointment and drop-out (8-9). Several nutraceutical products for the management of BPH have been commercialised in Italy (10) and between these, a new nutraceutical is MIGAR® MG, an association between daidzein, isolase and zinc. The effects of each component have been well-reported in literature. DOI: 10.4081/aiua.2017.1.12 INTRODUCTION Benign prostatic hyperplasia (BPH) is one of the most common diseases affecting the elderly male and is the most common clinical condition among all urological outpatients (1). BPH is characterized by clinically rele- Tiscione_Stesura Seveso 04/04/17 09:05 Pagina 12 13Archivio Italiano di Urologia e Andrologia 2017; 89, 1 Nutraceuticals for the management of LUTS related to BPH Daidzein is a soy isoflavone, after oral intake it crosses the enterocytes, is reduced to S-(-)equol through the intermediate dihydrodaidzein, and then is converted by deoxygenation to yield the bioactive S-(-)equol by intes- tinal bacteria (11). Several bacteria are able to perform this reaction and one of these, Lactococcus garvieae, was found in some Italian cheeses and has been used to pro- duce the first natural S-(-) equol-containing nutraceuti- cal (12). The bioactivation of daidzein to S-(-)equol is related to gender, race and age and requires: specific equol-producing bacteria and an optimum intraluminal conditions in terms of redox potential for the above described reactions (13). Therefore, the aim of this study is to evaluate the efficacy and safety of MIGAR® MG Retard in patients affected by LUTS related to BPH. METHODS Study design We performed a non-sponsored phase I-II study in a sin- gle urological institution between January to June 2016. The study was conducted according in line with Good Clinical Practice guidelines and with the ethical principles of the Declaration of Helsinki. Before the beginning of the study, all participants signed the written informed con- sent. No placebo run-in period was performed. Experimental protocol Patients with clinical and instrumental diagnosis of mod- erated-severe LUTS related to BPH were recruited. We considered patients with moderate-severe LUTS and with an International Prostatic Symptom Score (IPSS) score > 8. At the time of admission (T0), the patients underwent self- administered baseline questionnaire [IPSS, International Index of Erectile Function (IIEF-5) and Quality of Well-Being (QoL)], urological examination with history interview, prostate ultrasound and uroflowmetry (Cmax) with evalua- tion of post-voided residual volume (PVR), in accordance with the procedure described in EAU guidelines (3). All patients who met the inclusion criteria were enrolled and received 1 tablet/day of a combination of daidzein with isolase and zinc (MIGAR® MG Retard). The follow-up was performed 6 months after the end of therapy (T1) when patients underwent IPSS, IIEF-5QoL questionnaires and urological examination with laboratory and instrumental examinations. Inclusion and exclusion criteria Patients male who were > 45 years, sexually active, with a maximal urinary flow rate (Cmax) < 15 mL/s and with a post-residual voided volume < 100 cc, with IPSS = 8 or greater, with Prostate Specific Antigen (PSA) < 4 ng/ml (or higher in presence of negative prostate biopsy) and IIEF-5 score > 21 were enclosed in this study. On the other hand, patients with diabetes, liver, and/or renal failure, cancer, prostatitis, active urinary tract infection, chronic retention or polycystic kidney disease were excluded from the study. Moreover, we also excluded patients with urethral stenosis or anatomical abnormality interfering with the evaluation of voiding function. Patients with a history of transurethral resections of the prostate, laser therapy, or thermo therapy or with allergy to one or more compounds of MIGAR® MG Retard were also excluded. Finally, were also excluded from the study the patients that received a treatment for LUTS/BPH in the last 6 months. Questionnaires and urological examinations The validated Italian versions of the IPSS (14), IIEF-5 (15) and QoL (16) were self-administered to each patient at the arrival to the urological institution. The QoL scale was selected because it has been successfully applied to acute illnesses, whereas other quality of life scales, including the Short Form-36 (SF-36) Health Survey, are more suitable in chronic cases (17). Higher scores on the QoL scale reflect a higher quality of life (16). The IIEF-5 questionnaire was used for exclude all patients with con- comitant erectile dysfunction and for evaluating the impact of treatment on sexual function at the end of the follow-up therapy. Composition and characterization of the extracts used MIGAR® MG Retard Each tablet contains Daidzein 20 mg, Genistein 20 mg, Isolase® 40 mg, Zinc 10 mg. Instrumental, laboratory and urological evaluation At the time of the enrolment (T0) and during the follow- up (T1), all patients underwent urological examinations with the prostate volume ultrasound evaluation. Ultrasound prostate evaluation was performed during the urological examination with a trans-abdominal approach. The trans-abdominal scan was performed with the patient's supine after applying ultrasonic gel to the supra- pubic regions (3). Moreover, an uroflowmetry (Cmax) with ultrasound evaluation of post-voided residual vol- ume was performed by using URODYN® 1000 System (Medtronic Functional Diagnostics A/S.) (3). All patients were asked to do the PSA dosage. In order to exclude uri- nary tract infection, both leukocyte esterase and nitrite levels were measured in urine samples through dipstick assay (Bayer Multistik Pro Reagent Strips) (3). Main outcome measure The main outcome measure was the improvement of quality of life at the end of the whole study period, in terms of changes in IPSS, IPSS-QOL, Cmax, PVR, and QoL from baseline to the evaluation point, that is, 6 months. As secondary endpoints we considered a change between T0 and T1 in the following parameters: PSA val- ues, prostate volume, progression of disease, and surgi- cal treatment. Clinical failure was defined as the persist- ence of symptoms after the treatment or the suspension of therapy for significant reported ADRs, in particular on sexual function (3). Finally, from the beginning of the study (T0) and up to its ending (T1), the development of ADRs or of drug-drug interactions (DDIs) were recorded in agreement with our previous paper (18) using the Naranjo scale and the Drug Interaction Probability Scale, in agreement with the common terminology criteria for adverse events (CTCAE) guidelines (19). Safety assess- ments included treatment-emergent adverse events (TEAEs) and serious AEs (SAEs). The study design is reported in Figure 1. Tiscione_Stesura Seveso 04/04/17 09:05 Pagina 13 Archivio Italiano di Urologia e Andrologia 2017; 89, 1 D. Tiscione, L. Gallelli, I. Tamanini, L.G. Luciani, P. Verze, A. Palmieri, V. Mirone, R. Bartoletti, G. Malossini1, T. Cai 14 Efficacy outcome The efficacy outcome was identified as a statistically sig- nificant difference between clinical data recorded at the time of admission (T0) and those recorded at the end of the study (T1). Drug adherence At the end of the study, the adherence to the treatment was calculated using a formula based on the number of tablets consigned at the time of admission and on the number of tablets returned unused at the end of the study. Statistical analysis The required sample size for the present study was cal- culated under the following conditions: Difference between T0 and T1, 4 ± 1 score points in the IPSS score; α error level, 0.05 two-sided; statistical power, 80%; and anticipated effect size, Cohen’s d = 0.5. The calculation yielded 45 individuals. At baseline, the independent sample 2- tailed t-test was used to compare vari- ables. For categorical parameters, chi- square test was applied. Changes from baseline to end of therapy were ana- lyzed using ranked one-way analysis of variance (ANOVA) with a term for treatment group. All data are expressed as mean ± standard devia- tion. The threshold of statistical sig- nificance was set at p < 0.05. All reported p-values are two-sided. All statistical analyses were performed by using SPSS 21.0 (IBM Corporation, Armonk, NY, USA), while G*Power (Institut für Experimentelle Psychologie, Heinrich Heine Universität, Dusseldorf, Germany) was used for power calculation. RESULTS During the study period, 71 patients were enrolled. Nine patients (12.7%) were lost during the follow-up and excluded from the analysis, while 62 patients (87.3%) completed the study (mean age 65.4 ± 5.6 years). Baseline characteristics History, clinical, laboratory, instrumental and question- naires data at the time of admission are reported in Table 1. Follow-up examination At T1 we recorded a significant difference in terms of IPSS in all patients [21.5 ± 1.2 vs 16.2 ± 1.5; (-4.8); p < 0.001], and Cmax [9.7 ± 3.7 vs 15.3 ± 2.5; (+5.6); p < 0.001], post-voiding residual volume [85 ± 20.6 vs 50 ± 18.5; (-35); p < 0.001], QoL [0.56 ± 0.15 vs 0.84 ± 0.19; (+0.28); p < 0.001]. We did not record any in terms of PSA evaluation [2.47 ± 5.6 vs 2.09 ± 4.3; p = 0.67], prostate volume [41.5 ± 5.8 vs 42.1 ± 7.5 p = 0.61] and IIEF-5 [22.1 ± 1.8 vs 22.3 ± 1.5 p = 0.50] between T0 and T1 (Table 2). Figure 1. The figure shows the study schedule. IPSS: International Prostatic Symptom Score; IIEF-5*: International Index of Erectile Function; QoL: Quality of Well-Being. V1: visit 1, at the enrolment; V2: visit 2 at the second follow-up (6 months). Table 2. Questionnaire results at the first follow-up visit (6 months). Table 1. Clinical, instrumental and laboratory patient’s data at the enrolment time. The table shows the mean change differences from baseline to 6 months relative to main outcome measures. IPSS†: International Prostatic Symptom Score; IIEF-5*: International Index of Erectile Function; QoL‡: Quality of Well-Being. The table shows all anamnestic, clinical and questionnaires data at enrolment. SD#: Standard Deviation; IPSS†: International Prostatic Symptom Score; IIEF-5*: International Index of Erectile Function; QoL‡: Quality of Well-Being Baseline values Follow-up values (SD* or %) (SD* or %) Efficacy outcomes IPSS† 21.5 ± 1.2 16.2 ± 1.5 Treatment difference -4.8 ± 1.1 (p < 0.001) IIEF-5* 22.1 ± 1.8 22.3 ± 1.5 Treatment difference 0.1 ± 0.2 (p = 0.50) QoL§ 0.56 ± 0.15 0.84 ± 0.19 Treatment difference 0.28 ± 0.02 (p < 0.001) Maximal urinary flow rate (ml/sec) 9.7 ± 3.7 15.3 ± 2.5 Treatment difference 5.6 ± 0.8 (p < 0.001) Post-voiding residual volume (ml) 85 ± 20.6 50 ± 18.5 Treatment difference -35 ± 1.9 (p < 0.001) Patients (n°) 62 Age (mean ± SD#) 65.4 ± 5.6 Marital status Married 56 (90.3) Unmarried 6 (9.7) Educational qualification Primary school 31 (50) High school 20 (32.2) University 11 (17.8) Sexually active (past month) 62 (100) Symptoms Score at baseline (mean ± SD#) IPSS† 21.5 ± 1.2 IIEF-5* 22.1 ± 1.8 QoL‡ 0.56 ± 0.15 Charlson Comorbidity Index 0-1 62 (100) ≥ 2 - Laboratory Serum totale PSA (ng/ml) 2.47 ± 5.6 Serum creatinine (ng/ml) 1.0 ± 0.6 Urine dipstick negative 62 (100) positive - Ultrasound and uroflowmetry data Prostate volume (ml) 41.5 ± 5.8 Post-voiding residual volume (ml) 85 ± 20.6 Maximal urinary flow rate (ml/sec) 9.7 ± 3.7 V1 Clinical, instrumental, laboratory evaluation, IPSS, IIEF-5, QoL and ENROLMENT Screening (up to 3 months) Treatment: 6 months V2 (6 months) Clinical, instrumental, laboratory evaluation, IPSS, IIEF-5, QoL MIGAR© MG RETARD 1 tablet q24h Tiscione_Stesura Seveso 04/04/17 09:05 Pagina 14 15Archivio Italiano di Urologia e Andrologia 2017; 89, 1 Nutraceuticals for the management of LUTS related to BPH ADRs During the study period, 1 patient (1.6%) developed a mild ADRs (nausea) that did not require drug discontin- uation or other treatments. Moreover, did not document the development of DDIs, while we recorded a 100% adherence to the treatment and a 100% compliance to the experimental protocol. DISCUSSION In this phase I-II clinical trial, we evaluated the efficacy and the safety of a new nutraceutical agent named MIGAR® MG containing daizein, isolase and zinc in the treatment of LUTS related to BPH. It has been docu- mented that in Italy, about 50% of treatment used in patients with BPH are phytotherapies, while in Germany and other European countries, these represent the first- line treatment for mild-to-moderate LUTS in patients with BPH. Isoflavones show a chemical structure similar to the estrogen which allows them to bind to both alpha and beta estrogen receptors, exerting estrogen-like effects and so also called as phytoestrogens (21). Whereas estro- gen binds to and transactivates both estrogen receptors, isoflavones preferentially bind to and transactivate estro- gen beta receptor exerciting tissue-selective effects (22) that is expressed in prostate epithelial cells and plays a role in cellular homeostasis inducing anti-proliferative, pro-differentiative (23), and pro-apoptotic effects (23- 24). In this concern, several clinical trials documented that isoflavones are able to suppress PSA expression in patients with prostate cancer cells. In a recent meta- analysis, Zhang et al., evaluating the effects of isoflavones on prostate cancer risk, analyzed 11.346 cases and 140.177 controls and documented that daidzein, genis- tein, and glycitein were associated with a reduction of prostate cancer risk with an odds ratio: 0.85, 0.87 and 0.89, respectively (25). In agreement, in an experimental model performed in human prostate cancer cell lines Lu et al., recorded that s-equol has significant anti-prostate cancer activities probably related to the activation of the transcription factor Forkhead box O3 via an Akt-specific pathway and inhibitory effects on MDM2 expression (26). Moreover, in an experimental animal model Bae et al. reported that genistein and equol are strong inhibitors of testosterone 5α-reductase enzyme, suggesting that isoflavones could be used in the management of patients with LUTS (27). In fact, Wong et al. documented in eld- erly men that the dietary intake of genistein, glycitein or daidzein was related with a significant decrease of LUTS (28). In our study, we documented that oral intake of MIGAR® MG reduces LUTS with an improvement of uroflowmetry parameters, IPSS score and QoL. Probably these improvements could be related to a potentiating of daidzein induced by isolases that seems to be able to improve the action time of equol reducing its catabolism and increasing its clinical efficacy. On the other hand the presence of zinc in this studied formulation could plays synergic effects with daidzein and isolases. In fact, a long period clinical study (4-year) documented that chronic prostate inflammation is related to both severity and pro- gression of BPH and LUTS and zinc has antioxidant and anti-inflammatory activity and changes in prostate zinc concentrations play a role in the improvement of BPH (29). Take together these published data confirm our results documenting the efficacy of this new nutraceuti- cal formulation in patients with LUTS. However, nutraceuticals are dietary supplements, and it is not nec- essary to demonstrate data on efficacy or safety before marketing therefore could be possible the development of ADRs or DDIs during their use. In the present study, using the Naranjo probability scale and the Drug Interaction Probability Scale, we failed to report the development of severe ADRs or DDIs during the treat- ment with this compound. In particular, we recorded the development of a probable mild ADR in 1 patient only but this manifestation did not induce the discontinuation of the treatment, documenting the safety of this nutraceutical. Moreover, must be underlined that enrolled patients did not develop sexual dysfunction, and the optimal sexual activity should be considered as one principal aim in the management of patients with LUTS associated to BPH (30). Previously, we documented that a better sexual quality of life is correlated with a higher overall quality of life regardless of the urinary function in patients affected by LUTS due to BPH10. Finally, in our study we documented a complete adherence (100%) to the treatment and a complete compliance (100%) to the experimental protocol and it may be related to the improvement of QoL. The present study has two limita- tions: first, the lacks of placebo arm, because in Italy eth- ically patients must receive a treatment during a sponta- neous study, as ours; second, we evaluated the safety and efficacy of this new compound for a short time that did not allow evaluating its effects for a long time. CONCLUSIONS In this study, we demonstrated that combination of daidzein plus isolase and zinc (MIGAR® MG Retard) reduces the symptoms and improves the quality of life in patients affected by LUTS due to BPH, without the devel- opment of ADRs. ACKNOWLEDGEMENTS We are grateful to Professor John Denton (Department of Modern Philology, University of Florence) for manuscript language revision. 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Antioxidant and anti-inflammatory effects of zinc. Zinc-dependent NF-αB signaling. Inflammopharmacology 2017; 25:11-24 30. Messina R, Mirone V. Benign Prostatic Hyperplasia - An eco- nomic assessment of fixed combination therapy based on a literature review. Arch Ital Urol Androl. 2015; 87:185-9. Correspondence Daniele Tiscione, MD Irene Tamanini, MD Lorenzo Giuseppe Luciani, MD Gianni Malossini, MD Tommaso Cai, MD ktommy@libero.it Department of Urology, Santa Chiara Regional Hospital Largo Medaglie d'Oro 9 - Trento, Italy Luca Gallelli, MD Department of Health Science, School of Medicine, University of Catanzaro, Catanzaro, Italy Paolo Verze, MD Alessandro Palmieri, MD Vincenzo Mirone, MD Department of Urology, University of Naples, Federico II, Naples, Italy Riccardo Bartoletti, MD Department of Urology, University of Pisa, Pisa, Italy Tiscione_Stesura Seveso 04/04/17 09:05 Pagina 16