A thesis submitted to the Faculty of Health Sciences, University of the Witwatersrand, Johannesburg, in fulfillment of the requirements for the degree of Doctor of Philosophy July, 2015 / Introduction: Anastrozole is clinically effective in ovulation induction, but it has not been well researched. The aim of this study was to determine the optimal dose for anastrozole as a superovulator and ascertain its effects on implantation in Wistar rats; also to determine its effects on uterine morphology during early pregnancy using light microscopy and scanning electron microscopy (SEM). [Abbreviated Abstract. Open document to view full version] / AC2016

CoordenaÃÃo de AperfeiÃoamento de Pessoal de NÃvel Superior / Conselho Nacional de Desenvolvimento CientÃfico e TecnolÃgico / FundaÃÃo Cearense de Apoio ao Desenvolvimento Cientifico e TecnolÃgico / A deficiÃncia de estrÃgeno tem mostrado aumentar a remodelaÃÃo Ãssea, podendo afetar algumas doenÃas Ãsseas como a periodontite. A biossÃntese desse hormÃnio à catalisada pela enzima aromatase e sua inibiÃÃo à importante para terapia do cÃncer de mama. Os pacientes que usam inibidores da aromatase como o Anastrozol (ANA) mostram maior nÃmero de fraturas e menor densidade Ãssea. Considerando, entÃo, o papel do estrÃgeno na resposta inflamatÃria e no metabolismo Ãsseo, alÃm dos efeitos adversos do ANA e da periodontite ser caracterizada por um processo inflamatÃrio que resulta em perda Ãssea alveolar, o objetivo desse estudo foi de investigar se o ANA afeta a periodontite em ratas ovariectomizadas. Para isto, o ANA (0,02, 0,1 e 0,5 mg/kg-v.o.) foi avaliado em um modelo de periodontite induzida por ligadura em ratos durante 11 dias. A periodontite foi analisada atravÃs de macroscopia, de histologia e da atividade de mieloperoxidase (MPO). As dosagens sÃricas de estrÃgeno, o leucograma, a variaÃÃo de massa corpÃrea e as funÃÃes hepÃtica, renal e esplÃnica tambÃm foram consideradas. A ovariectomia reduziu os nÃveis sÃricos de estrÃgeno apÃs 14 dias, mas nÃo exacerbou a periodontite ou aumentou a atividade de mieloperoxidase (MPO) quando comparada aos animais falso ovariectomizados (F-OVX). Embora o ANA nÃo tenha aumentado a perda Ãssea alveolar (POA), a administraÃÃo deste fÃrmaco por 11 dias aumentou a atividade de MPO. A ovariectomia combinada ou nÃo ao ANA nÃo promoveu nenhuma mudanÃa nas funÃÃes hepÃtica, renal ou esplÃnica, mas causou uma leucocitose promovida por cÃlulas mononucleares no 11 dia. Os animais ovariectomizados que receberam salina (SAL) tambÃm mostraram maior ganho de massa corpÃrea e a curva de peso do ANA foi similar à curva do SAL. Em conclusÃo, embora a reduÃÃo de estrÃgeno por 25 dias e a administraÃÃo de ANA por 11 dias nÃo tenha aumentado a POA, o ANA aumentou a atividade de MPO. / Estrogen deficiency was shown to increase bone remodeling, may affect some bone diseases as periodontitis. Biosynthesis of this hormone is catalyzed by aromatase enzyme and her inhibition is important for breast cancer therapy. The patients who use aromatase inhibitors as Anastrozole (ANA) show greater number of fractures and lower bone density. Since estrogen role in the inflammatory response and the bone metabolism, besides those adverse effects of ANA and periodontitis is characterized by an inflammatory process that results in alveolar bone loss, the aim of this study was to investigate whether ANA affects the periodontitis in ovariectomized rats. For this, ANA (0.02, 0.1 and 0.5 mg/kg-v.o.) was evaluated in ligature-induced periodontits for 11 days in ovariectomized rats. Periodontitis was analyzed through macroscopy, histology and by myeloperoxidase (MPO) activity. Serum dosage of estrogen, leukogram, corporal mass variation and liver, renal and spleen functions were also analyzed. The ovariectomy reduced the serum levels of estrogen after 14 days, but did not worsen the periodontitis nor increased MPO activty when compared to sham ovariectomized (S-OVX) animals. Although, ANA did not increase alveolar bone loss (ABL), administration of this drug for 11 days increased the MPO activity. The ovariectomy combined or not with ANA no promoved any changes in liver, renal or spleen functions, but caused leukocytosis promoved by mononuclear cells at 11th day. Ovaricetomized animals that received saline (SAL) also showed more gain of corporal mass and ANA weight curve were similar to SAL curve. In conclusion, although the estrogen reduction for 25 days and the administration of anastrozole for 11 days did not increase the ABL, the ANA increased the MPO activity.

This body of research investigates the effects of sex and estrogen on higher brain functions, in general, and on working memory, in particular. A female advantage for object-location and verbal working memory has been reported, and estrogen supplementation facilitates performance of the same. Less is known about whether the female advantage is due to the verbalizability of the stimuli, and whether estrogen-suppression adversely affects performance on working memory and other neuropsychological domains sensitive to estrogen. Study 1 examined sex-related differences in young adults on an object-location working memory measure that varied in verbalizability of stimuli and task presentation (i.e., manual or computer); as expected, females performed better than males regardless of the verbalizability of the stimuli or task presentation. Study 2 examined sex-related differences in young adults on the n-back working memory task across verbal, spatial, and object conditions. Contrary to the hypotheses, there was no sex effect for the verbal version of the n-back task, and males actually performed better than females on the object version; as expected, males performed better than females on the spatial version. Study 3 investigated the effect of estrogen suppression in middle-aged and older adult females undergoing treatment for estrogenic breast cancer using the experimental working memory measures from Studies 1 and 2, and a comprehensive neuropsychological battery that included measures considered to be either sensitive (e.g., letter fluency) or insensitive (e.g., spatial ability) to the female advantage and the effects of estrogen. The estrogen suppression group performed more poorly than healthy age-matched controls on certain estrogen-sensitive measures (i.e., speeded visuomotor attention, speeded manual dexterity, and letter fluency), but unexpectedly, the groups did not differ on any of the memory measures presumed to be estrogen-sensitive. This body of research suggests that although certain working memory measures are sensitive to sex effects, the direction depends on the domain, and that estrogen suppression does not impact working memory in postmenopausal women but does adversely impact speeded performance measures.

This body of research investigates the effects of sex and estrogen on higher brain functions, in general, and on working memory, in particular. A female advantage for object-location and verbal working memory has been reported, and estrogen supplementation facilitates performance of the same. Less is known about whether the female advantage is due to the verbalizability of the stimuli, and whether estrogen-suppression adversely affects performance on working memory and other neuropsychological domains sensitive to estrogen. Study 1 examined sex-related differences in young adults on an object-location working memory measure that varied in verbalizability of stimuli and task presentation (i.e., manual or computer); as expected, females performed better than males regardless of the verbalizability of the stimuli or task presentation. Study 2 examined sex-related differences in young adults on the n-back working memory task across verbal, spatial, and object conditions. Contrary to the hypotheses, there was no sex effect for the verbal version of the n-back task, and males actually performed better than females on the object version; as expected, males performed better than females on the spatial version. Study 3 investigated the effect of estrogen suppression in middle-aged and older adult females undergoing treatment for estrogenic breast cancer using the experimental working memory measures from Studies 1 and 2, and a comprehensive neuropsychological battery that included measures considered to be either sensitive (e.g., letter fluency) or insensitive (e.g., spatial ability) to the female advantage and the effects of estrogen. The estrogen suppression group performed more poorly than healthy age-matched controls on certain estrogen-sensitive measures (i.e., speeded visuomotor attention, speeded manual dexterity, and letter fluency), but unexpectedly, the groups did not differ on any of the memory measures presumed to be estrogen-sensitive. This body of research suggests that although certain working memory measures are sensitive to sex effects, the direction depends on the domain, and that estrogen suppression does not impact working memory in postmenopausal women but does adversely impact speeded performance measures.

Tamoxifen and aromatase inhibitors are treatment options for women with breast cancer and evidence on the risk of stroke is important in choosing between these two options. A systematic review of two randomized controlled trials and their nine related trial reports showed different methods for adverse event reporting and inconsistent estimates of stroke risk. In an observational cohort study of 5443 Ontario women, aged 66 years or older with early stage breast cancer, 86 ischemic stroke events (1.6%) occurred during follow-up of 5 years. There was no statistically significant difference in the risk of stroke between the hormone therapy groups [adjusted HR for tamoxifen compared to AI 1.330 (0.810, 2.179)]. Results were similar across cardiovascular disease risk groups and were robust to different follow up periods and analytic methods. This study suggests that there is no significant difference in stroke between these treatment options.

Tamoxifen and aromatase inhibitors are treatment options for women with breast cancer and evidence on the risk of stroke is important in choosing between these two options. A systematic review of two randomized controlled trials and their nine related trial reports showed different methods for adverse event reporting and inconsistent estimates of stroke risk. In an observational cohort study of 5443 Ontario women, aged 66 years or older with early stage breast cancer, 86 ischemic stroke events (1.6%) occurred during follow-up of 5 years. There was no statistically significant difference in the risk of stroke between the hormone therapy groups [adjusted HR for tamoxifen compared to AI 1.330 (0.810, 2.179)]. Results were similar across cardiovascular disease risk groups and were robust to different follow up periods and analytic methods. This study suggests that there is no significant difference in stroke between these treatment options.

Made available in DSpace on 2015-07-22T20:50:08Z (GMT). No. of bitstreams: 0 Previous issue date: 2009-11-25. Added 1 bitstream(s) on 2015-08-11T03:25:59Z : No. of bitstreams: 1 Publico-11912.pdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) / Introdução: A endocrinoterapia é uma das principais responsáveis pela redução de mortalidade do câncer de mama. Biomarcadores preditivos de resposta celular precoce vêm sendo estudados com intuito de prever precocemente a hormonioresistência. Freqüentes deleções e mutações têm sido descritas no gene p16 em diversos tipos de tumores, mas pouco se sabe sobre seu papel no câncer de mama e seu comportamento após endocrinoterapia neoadjuvante com tamoxifeno e anastrozol. Objetivos: Estudar a expressão do p16 e dos receptores de estrogênio e progesterona (RE e RP) em pacientes na pós - menopausa com carcinoma de mama RE e/ ou RP (+) após curto período (26 dias) de tratamento com tamoxifeno, anastrozol e placebo. Métodos: Estudo prospectivo randomizado duplo-cego realizado com 58 pacientes dos Hospitais Pérola Byington e São Paulo da UNIFESP (São Paulo - Brasil) com carcinoma ductal infiltrativo de mama, nos estádios II e III, que no período pré-operatório foram subdivididas em três grupos: P (placebo, N=25), T (tamoxifeno 20mg/dia, N=15) e A (anastrozol 1mg/dia, N=18). A biópsia foi realizada no momento do diagnóstico e após a cirurgia definitiva (26º dia) e os tumores foram isolados por micro-arranjos teciduais. O estudo imunoistoquímico foi realizado com anticorpos para p16 (Dako- OA315), RE (Neomarkers-M7047), RP (Dako- M3569). Realizou-se o estudo semiquantitativo utilizando-se os critérios de Allred e o estudo estatístico pelo teste paramétrico de Anova. Resultados: A positividade do p16 variou de 22 para 17%, respectivamente pré e pós tratamento com anastrozol; de 8 para 4% no grupo placebo e não houve variação, com 7% de positividade, no grupo que recebeu tamoxifeno. A comparação entre grupos e tempos não apresentou relação significativa para o p16 (p=0,17). Não foi encontrada correlação entre a positividade do p16 e o status hormonal (RE e RP). Conclusão: Não houve diferença estatística significante entre os três grupos estudados. Futuros estudos com métodos moleculares poderão esclarecer dúvidas suscitadas a respeito do tempo de exposição à droga necessária para interferir nos biomarcadores e proteínas. / Background: Frequent deletions or mutations of the INK4 gene, which encodes the cyclin-dependent kinase 4 inhibitor p16INK4a, have been documented in various human cancers, but little is known about the role of this tumor suppressor gene in primary breast cancer and there is a lack in the literature about its expression behavior in neoadjuvant endocrinetherapy with tamoxifen or anastrozole. Objective: Analysis of p16INK4a expression in patients with invasive ductal carcinomas (IDC) prior to tamoxifen and anastrozole neoadjuvant treatment and possible correlation between predictive and prognostical factors – estrogen receptor (ER), progesterone receptor (PgR). Methods: We examined p16INK4a mRNA expression and its relationship with short period (26 days) neoadjuvant endocrine therapy with tamoxifen and anastrozole in 58 primary breast cancers with palpable ER-positive IDC. They were double-blind randomized in three neoadjuvant treatment groups for 21 days: Anastrozole 1mg/day (n= 17), Placebo (n=25) and Tamoxifen 20mg/day (n=15). Biomarkers status (ER, PgR and p16) were obtained by comparing single immunohistochemical evaluation of pre and post-surgery samples using Allred’s method. Statistical analyses were performed using the SPSS software for Windows. Results: Variation in p16 was 22% to 17% in anastrozole group, 8% to 4% in placebo group and there was no variation in tamoxifen group, standing in 7%. There was no significant statistical diference in p16INK4a expression among the three groups (p=0.17). Variation in p16 was 12% to 9% (p<0.05) when considering the three groups together. There was a significant decrease of p16 expression in pre and post surgery results. There was no significant statistical correlation between p16 expression and hormonal status (RE and RP). Conclusions: There was no significant statistical diference in p16INK4a expression among the three groups. There was a significant statistical decrease in p16INK4a expression when compared pre and post surgery values. These findings could indicate that expression of p16 and variation in pre and post surgery samples are associated with hormone responsiveness and mechanisms of resistance. There were no significant statistical correlation between p16 expression and hormonal status (RE and RP). Further studies are necessary to understand their functional interrelationships and whether high p16INK4a expression may be associated with a lack of hormone responsiveness in breast cancer. / TEDE / BV UNIFESP: Teses e dissertações

<p> This thesis focuses on an examination of cortical and trabecular bone density and geometry at the radius and tibia in postmenopausal women, primarily women with history of breast carcinoma, while also assessing musculoskeletal changes in postmenopausal breast cancer patients after treatment with the Aromatase Inhibitor, Anastrozole. The first sub-study is an investigation of the reproducibility of the pQCT measurement parameters at the radius and tibia in healthy pre-and postmenopausal women. Results indicated that the reproducibility was good at the radius and even better at the tibia for all parameters measured. The second study is an appraisal of the level of osteoporosis knowledge in a cohort of postmenopausal women. The participants were assessed via the Facts on Osteoporosis Quiz, a well validated questionnaire, and the data revealed significantly lower test scores among the breast cancer subjects in comparison with healthy postmenopausal women. In the remaining group of studies, pQCT technology was utilized to describe trabecular and cortical bone at the radius and tibia in postmenopausal women and women with a history of breast carcinoma whom had been prescribed Anastrozole. The following measurement sites were significantly lower in the breast cancer subjects: TOT_DEN and TOT_CNT at the 4% radius; CRT_DEN, TOT_CNT, and CRT_CNT at the 20% radius; TOT_DEN at the 4% tibia; and CRT_DEN at the 38% tibia. With respect to time on Anastrozole, TOT_CNT at the 4% radius (r=-0.36); TOT_CNT (r=-0.33), CRT_CNT (r=-0.34) and CRT_DEN (r=-0.44) at the 20% radius; and CRT_DEN (r=-0.39) and CRT_CNT (r=-0.27) at the 38% tibia were significantly negatively correlated with days on Anastrozole. Furthermore, after two years of Anastrozole treatment in a small cohort of breast cancer subjects, there was a significant decrease in CRT_DEN (p=0.025) at the 20% diaphyseal radius and also at the 38% diaphyseal tibia (p=0.051). Together, the sub-studies that comprise this thesis demonstrate that there are noteworthy deficiencies in osteoporosis knowledge among postmenopausal women, particularly those with a history of breast carcinoma, and yet, these are the same women that have an increased need to understand the preventative and treatment options regarding this disease as they demonstrate reduced bone density at all measurement sites. It also appears that time on Anastrozole primarily affects cortical bone density in these women. In summary, this thesis provides novel details regarding cortical bone in breast cancer subjects and emphasizes the need for a normative database of bone quality parameters at different skeletal sites in order to gain a better understanding of the utility of each skeletal site with regard to fracture risk prediction. </p> / Thesis / Doctor of Philosophy (PhD)

Background: In the ‘Arimidex’, Tamoxifen Alone or in Combination (ATAC) trial, the aromatase inhibitor (AI) anastrozole had a ignificantly better efficacy and safety profile than tamoxifen as initial adjuvant therapy for hormone receptor-positive (HR+) early breast cancer (EBC) in postmenopausal patients. To compare the combined long-term clinical and economic benefits, we carried out a cost-effectiveness analysis (CEA) of anastrozole versus tamoxifen based on the data of the 100- month analysis of the ATAC trial from the perspective of the German public health insurance. Patients and Methods: A Markov model with a 25-year time horizon was developed using the 100-month analysis of the ATAC trial as well as data obtained from published literature and expert opinion. Results: Adjuvant treatment of EBC with anastrozole achieved an additional 0.32 quality-adjusted life-years (QALYs) gained per patient compared with tamoxifen, at an additional cost of D 6819 per patient. Thus, the incremental cost effectiveness of anastrozole versus tamoxifen at 25 years was D 21,069 ($ 30,717) per QALY gained. Conclusions: This is the first CEA of an AI that is based on extended follow-up data, taking into account the carryover effect of anastrozole, which maintains the efficacy benefits beyond therapy completion after 5 years. Adjuvant treatment with anastrozole for postmenopausal women with HR+ EBC is a cost-effective alternative to tamoxifen. / Hintergrund: Bei der adjuvanten Therapie von postmenopausalen Patientinnen mit Hormonrezeptor-positivem (HR+) Mammakarzinom belegen die ATAC-100-Monatsdaten (ATAC-Studie: ‘Arimidex’, Tamoxifen Alone or in Combination) einen signifikanten Vorteil von Anastrozol gegenüber Tamoxifen in Bezug auf Rezidivrisiko und Verträglichkeit. Es wurde eine Kosten-Nutzwert-Analyse von Anastrozol im Vergleich zu Tamoxifen aus der Sicht des deutschen Gesundheitssystems durchgeführt. Material und Methoden: Als Berechnungsbasis wurde ein Markov- Modell zur Abschätzung der Kosteneffektivität entwickelt. Der Modellierungszeitraum umfasste 25 Jahre. Die Daten wurden anhand der ATAC-100-Monatsdaten, vorliegender Literatur und durch ein interdisziplinäres Expertenteam ermittelt. Ergebnisse: Eine adjuvante Therapie mit Anastrozol erzielte 0,32 quality-adjusted life-years (QALYs) pro Patientin mehr, verglichen mit einer adjuvanten Tamoxifentherapie. Die zusätzlichen Kosten der Therapie mit Anastrozol lagen bei 6819 D pro Patientin. Im Vergleich mit Tamoxifen erzielte Anastrozol einen ICER (Incremental Cost-Effectiveness Ratio) von 21 069 D (30 717 $)/QALY über den gesamten Modellierungszeitraum. Schlussfolgerung: Diese Kosten- Nutzwert-Analyse eines Aromatasehemmers basiert erstmals auf einer Datenanalyse, die auch das Follow-Up und den sogenannten Carryover- Effekt nach einer abgeschlossenen 5-Jahres-Therapie beinhaltet. Anastrozol ist auch nach dieser Analyse aus der Sicht des deutschen Gesundheitssystems eine kosteneffektive Therapieoption für postmenopausale Patientinnen mit einem HR+ frühen Mammakarzinom. / Dieser Beitrag ist mit Zustimmung des Rechteinhabers aufgrund einer (DFG-geförderten) Allianz- bzw. Nationallizenz frei zugänglich.

Background: In the ‘Arimidex’, Tamoxifen Alone or in Combination (ATAC) trial, the aromatase inhibitor (AI) anastrozole had a ignificantly better efficacy and safety profile than tamoxifen as initial adjuvant therapy for hormone receptor-positive (HR+) early breast cancer (EBC) in postmenopausal patients. To compare the combined long-term clinical and economic benefits, we carried out a cost-effectiveness analysis (CEA) of anastrozole versus tamoxifen based on the data of the 100- month analysis of the ATAC trial from the perspective of the German public health insurance. Patients and Methods: A Markov model with a 25-year time horizon was developed using the 100-month analysis of the ATAC trial as well as data obtained from published literature and expert opinion. Results: Adjuvant treatment of EBC with anastrozole achieved an additional 0.32 quality-adjusted life-years (QALYs) gained per patient compared with tamoxifen, at an additional cost of D 6819 per patient. Thus, the incremental cost effectiveness of anastrozole versus tamoxifen at 25 years was D 21,069 ($ 30,717) per QALY gained. Conclusions: This is the first CEA of an AI that is based on extended follow-up data, taking into account the carryover effect of anastrozole, which maintains the efficacy benefits beyond therapy completion after 5 years. Adjuvant treatment with anastrozole for postmenopausal women with HR+ EBC is a cost-effective alternative to tamoxifen. / Hintergrund: Bei der adjuvanten Therapie von postmenopausalen Patientinnen mit Hormonrezeptor-positivem (HR+) Mammakarzinom belegen die ATAC-100-Monatsdaten (ATAC-Studie: ‘Arimidex’, Tamoxifen Alone or in Combination) einen signifikanten Vorteil von Anastrozol gegenüber Tamoxifen in Bezug auf Rezidivrisiko und Verträglichkeit. Es wurde eine Kosten-Nutzwert-Analyse von Anastrozol im Vergleich zu Tamoxifen aus der Sicht des deutschen Gesundheitssystems durchgeführt. Material und Methoden: Als Berechnungsbasis wurde ein Markov- Modell zur Abschätzung der Kosteneffektivität entwickelt. Der Modellierungszeitraum umfasste 25 Jahre. Die Daten wurden anhand der ATAC-100-Monatsdaten, vorliegender Literatur und durch ein interdisziplinäres Expertenteam ermittelt. Ergebnisse: Eine adjuvante Therapie mit Anastrozol erzielte 0,32 quality-adjusted life-years (QALYs) pro Patientin mehr, verglichen mit einer adjuvanten Tamoxifentherapie. Die zusätzlichen Kosten der Therapie mit Anastrozol lagen bei 6819 D pro Patientin. Im Vergleich mit Tamoxifen erzielte Anastrozol einen ICER (Incremental Cost-Effectiveness Ratio) von 21 069 D (30 717 $)/QALY über den gesamten Modellierungszeitraum. Schlussfolgerung: Diese Kosten- Nutzwert-Analyse eines Aromatasehemmers basiert erstmals auf einer Datenanalyse, die auch das Follow-Up und den sogenannten Carryover- Effekt nach einer abgeschlossenen 5-Jahres-Therapie beinhaltet. Anastrozol ist auch nach dieser Analyse aus der Sicht des deutschen Gesundheitssystems eine kosteneffektive Therapieoption für postmenopausale Patientinnen mit einem HR+ frühen Mammakarzinom. / Dieser Beitrag ist mit Zustimmung des Rechteinhabers aufgrund einer (DFG-geförderten) Allianz- bzw. Nationallizenz frei zugänglich.