br Breastfeeding also suppresses ovulation reduces gonadotropin levels and

Breastfeeding also suppresses ovulation, reduces gonadotropin levels, and alters the hormonal milieu [14]; thus, it may reduce EOC risk. Most case-control studies report a protective association with ever breastfeeding [7,15,16]. Some also report an increasing Ferrostatin-1 in risk associated with increasing cumulative duration [7,15,16]. In con-trast, the handful of cohort studies examining breastfeeding found a weak, statistically non-significant reduction in risk [15], except for the Nurses' Health Study, which reported a statistically significant protec-tive effect only after 18 cumulative months of breastfeeding [17].
Beyond the association with ever breastfeeding and breastfeeding duration, there has been little exploration of factors that may influence the breastfeeding-EOC association. Questions remain about the effects of age at breastfeeding, time since breastfeeding, and influence of birth order among offspring breastfed. We used data from a large, population-based case-control study to examine these questions.
2. Material and methods
Details of the Hormones and Ovarian cancer PrEdiction (HOPE) Study are described elsewhere [18]. Cases were women diagnosed with incident, histologically-confirmed epithelial ovarian, peritoneal, or fallopian tube cancer from February 2003 to November 2008 in the contiguous regions of western Pennsylvania, eastern Ohio, and south-western New York. Eligible participants were at least 25 years old, resid-ing within the catchment region, and within 9 months of diagnosis at the time of interview. Women were identified through a network of hospital and physician practices using pathology records, physician practice records, and hospital cancer registries. Among 2878 potentially eligible cases, 1608 were excluded due to ineligibility (time since diag-nosis more than 9 months, residence outside catchment region, prior di-agnosis of ovarian cancer, inability to speak English, deceased). Of the 1270 remaining eligible cases, 902 (71%) consented to study participation.
Controls were identified through random-digit dialing and were frequency-matched to cases by 5-year age groups and 3-digit telephone 
Institutional Review Board approval for the study was obtained from hospitals in which cases were identified and from the University of Pittsburgh. All participants provided written, informed consent.
2.2. Data collection and exposure assessment
Trained interviewers conducted a standardized 2-hour in-person in-terview to obtain detailed information on reproductive, medical, and demographic data from birth until a reference date. To aid recall, a life events calendar with milestones, such as marriages, births, and deaths, was used [19]. Each pregnancy was denoted on the calendar by coloring the month of pregnancy initiation until pregnancy end. Breastfeeding episodes were similarly noted on the calendar. For each pregnancy, a woman was asked the outcome (live birth, still birth, miscarriage, abor-tion). For each live birth, she was asked detailed information, including “Did you ever breastfeed this baby?” If she responded “yes”, the total number of months breastfed was elicited by asking “For how many months did you nurse?”
The reference date was calculated as 9 months prior to diagnosis (cases) or interview (controls) to ensure that exposures occurred before ovarian cancer diagnosis in cases and within a similar time frame for controls.
Pathology data were extracted from pathology reports by two inde-pendent readers. Differences were reviewed by study staff to assign final pathology data.
2.3. Statistical analyses
Analyses were limited to the 1572 controls and 689 cases who re-ported at least one live birth. A woman was classified as having ever breastfed if she responded positively to the question “Did you breastfeed this baby?” for any live birth she reported. Total duration of breastfeeding was calculated by summing the number of months breastfed across all breastfeeding episodes. Age at first breastfeeding episode was calculated using the month and year of the pregnancy end corresponding to the first breastfeeding episode and the month and year of a subject's birth. Age at last breastfeeding episode was sim-ilarly calculated using the month and year of the pregnancy end corre-sponding to the last breastfeeding episode. Time since the first breastfeeding episode was obtained by calculating the time elapsed since the end of the first breastfeeding episode until the reference date. Time since the last breastfeeding episode was similarly obtained using the time elapsed since the end of the last breastfeeding episode.