Krystal Sexton

Primary breast cancer phenotypes associated with propensity for central nervous system metastases

There is anecdotal evidence that the incidence of central nervous system (CNS) metastases in breast cancer patients is increasing. It is unclear whether specific tumor biological properties or the use of systemic therapies influence this risk.

The rates of chemotherapy-induced amenorrhea in patients treated with adjuvant doxorubicin and cyclophosphamide followed by a taxane

Objective:Adjuvant chemotherapy in premenopausal women with breast cancer may induce amenorrhea, which can affect fertility, choice of hormonal therapy, and increase the risk of late toxicity. The incidence of chemotherapy-induced amenorrhea (CIA) resulting from doxorubicin and cyclophosphamide (AC) followed by a taxane (T) is poorly characterized. Methods:We retrospectively surveyed women who were premenopausal and less than age 50 at initiation of chemotherapy to determine the rates of CIA in women receiving AC followed by T compared with AC alone. Results:One hundred ninety-one eligible women completed the survey. The rate of CIA in women who received AC followed by T was 64% (95% confidence interval [CI] = 55–72%) compared with 55% (95% CI = 43–66%) in AC alone. As expected, CIA rates were higher in older than younger women (82% vs. 55%, P = 0.004). Multivariate logistic regression analysis revealed that age >40 was associated with a 4.6-fold increased risk of CIA (95% CI = 1.7–12.1, P = 0.002). It also revealed that receiving T after AC was associated with an odds ratio of 1.9 for CIA as compared with receiving AC alone (95% CI = 1.0–3.5, P = 0.05). Despite ≥6 months of amenorrhea, many women ≤40 resumed menses (40%). CIA was more likely to be irreversible in those >40. The addition of taxanes did not alter the rate of reversibility for the group as a whole (P = 0.36). Conclusions:Older age and the addition of taxane to AC increased the risk of CIA and the amenorrhea was more likely to be irreversible for women >40. Women ≤40 often resume menstruation even after 6 months of amenorrhea, and the addition of T does not play a role. Subsequent resumption of menstrual function must be considered when initiating appropriate hormonal therapy.

Identification of Novel Kinase Targets for the Treatment of Estrogen Receptor–Negative Breast Cancer

Purpose: Previous gene expression profiling studies of breast cancer have focused on the entire genome to identify genes differentially expressed between estrogen receptor (ER) α–positive and ER-α–negative cancers. Experimental Design: Here, we used gene expression microarray profiling to identify a distinct kinase gene expression profile that identifies ER-negative breast tumors and subsets ER-negative breast tumors into four distinct subtypes. Results: Based on the types of kinases expressed in these clusters, we identify a cell cycle regulatory subset, a S6 kinase pathway cluster, an immunomodulatory kinase–expressing cluster, and a mitogen-activated protein kinase pathway cluster. Furthermore, we show that this specific kinase profile is validated using independent sets of human tumors and is also seen in a panel of breast cancer cell lines. Kinase expression knockdown studies show that many of these kinases are essential for the growth of ER-negative, but not ER-positive, breast cancer cell lines. Finally, survival analysis of patients with breast cancer shows that the S6 kinase pathway signature subtype of ER-negative cancers confers an extremely poor prognosis, whereas patients whose tumors express high levels of immunomodulatory kinases have a significantly better prognosis. Conclusions: This study identifies a list of kinases that are prognostic and may serve as druggable targets for the treatment of ER-negative breast cancer. (Clin Cancer Res 2009;15(20):6327–40)

Effect of Lapatinib on the Development of Estrogen Receptor–Negative Mammary Tumors in Mice

Lapatinib, a selective orally available inhibitor of epidermal growth factor receptor (EGFR) and ErbB2 receptor tyrosine kinases, is a promising agent for the treatment of breast cancer. We examined the effect of lapatinib on the development of mammary tumors in MMTV-erbB2 transgenic mice, which express wild-type ErbB2 under the control of the mouse mammary tumor virus promoter and spontaneously develop estrogen receptor (ER)-negative and ErbB2-positive mammary tumors by 14 months of age. Mice were treated from age 3 months to age 15 months with vehicle (n = 17) or lapatinib (30 or 75 mg/kg body weight; n = 16 mice per group) by oral gavage twice daily (6 d/wk). All statistical tests were two-sided. By 328 days after the start of treatment, all 17 (100%) of the vehicle-treated mice vs five (31%) of the 16 mice treated with high-dose lapatinib developed mammary tumors (P < .001). Among MMTV-erbB2 mice treated for 5 months (n = 20 mice per group), those treated with lapatinib had fewer premalignant lesions and noninvasive cancers in their mammary glands than those treated with vehicle (P = .02). Lapatinib also effectively blocked epidermal growth factor-induced signaling through the EGFR and ErbB2 receptors, suppressed cyclin D1 and epiregulin mRNA expression, and stimulated p27 mRNA expression in human mammary epithelial cells and in mammary epithelial cells from mice treated for 5 months with high-dose lapatinib. Thus, cyclin D1, epiregulin, and p27 may represent useful biomarkers of lapatinib response in patients. These data suggest that lapatinib is a promising agent for the prevention of ER-negative breast cancer.

Cancer stem cell markers are enriched in normal tissue adjacent to triple negative breast cancer and inversely correlated with DNA repair deficiency

IntroductionWe hypothesized that cells present in normal tissue that bear cancer stem cell markers may represent a cancer cell of origin or a microenvironment primed for tumor development, and that their presence may correlate with the clinically defined subtypes of breast cancer that show increased tumorigenicity and stem cell features.MethodsNormal tissues sampled at least 5 cm from primary tumors (normal adjacent tissue) were obtained from 61 chemotherapy-naive patients with breast cancer treated with mastectomy. Samples were stained simultaneously with immunofluorescence for CD44/CD49f/CD133/2 stem cell markers. We assessed the association between CD44+CD49f+CD133/2+ staining in normal adjacent tissue and breast cancer receptor subtype (defined by the expression of the estrogen (ER), progesterone (PR), or human epidermal growth factor-2 (Her2) receptors). We also examined the correlation between CD44+CD49f+CD133/2+ immunofluorescence and each of two previously published gene signatures, one derived from stem-cell enriched tissue and one from BRCA mutated tissue expected to have defective DNA repair.ResultsPatients with triple negative breast cancer (ER–/PR–/HER2–) expressed CD44+CD49f+CD133/2+ in 9 of 9 normal adjacent tissue samples compared with 7 of 52 ER+ and/or Her2+ tumors (P < 0.001). Further, expression of CD44+CD49f+CD133/2+ by normal adjacent tissue correlated positively with a stem cell-derived tumorigenic signature (P <0.001) and inversely with a defective DNA-repair signature (P <0.001).ConclusionNormal cells bearing cancer stem cell markers are associated with the triple negative receptor subtype of breast cancer. This study suggests stem cell staining and gene expression signatures from normal breast tissues represent novel tissue-based risk biomarkers for triple negative breast cancer. Validation of these results in additional studies of normal tissue from cancer-free women could lay the foundation for future targeted triple negative breast cancer prevention strategies.

Abstract 1664: Maternal embryonic leucine aipper kinase (MELK) is a critical regulator of proliferation and is independently prognostic in estrogen receptor-negative breast cancer

Background: Molecularly targeted therapies such as tamoxifen, aromatase inhibitors, and trastuzumab are useful for the treatment of breast cancer. However, these targeted therapies do not treat all breast cancers, especially estrogen receptor alpha (ER)-negative, progesterone receptor (PR)-negative, HER2-negative (“triple-negative”) breast cancer. Our lab previously used gene expression profiling of human breast cancers to identify kinases overexpressed in ER-negative breast cancers. One of the kinases highly expressed in a poor prognosis group of triple negative breast cancers is maternal embryonic leucine-zipper kinase (MELK). The purpose of this study was to demonstrate that MELK expression is upregulated in ER-negative breast cancers, and to test the hypothesis that MELK is a critical regulator of ER-negative breast cancer cell growth. Methods: RNA from breast cancer cell lines and 60 human breast tumors was isolated for Q-PCR analysis of MELK. To determine if MELK regulates cell proliferation, ER-positive and ER-negative breast cancer cell lines were transfected with siRNA targeting MELK. Cells were replated 36 hours after transfection in 96-well plates at a density of 2,000/well. Cell number was measured by the MTS assay every day for 6 days. To determine if MELK levels are independently prognostic, we performed uni- and multivariate Cox regression analysis, including all available biological characteristics of the tumor in the model. Results: MELK mRNA levels were significantly higher (p Conclusions: MELK is highly expressed in ER-negative breast cancer, essential for ER-negative breast cancer cell growth, and is independently prognostic. These findings suggest that MELK is a promising target for the treatment of ER-negative breast cancer. Supported by a Susan G. Komen for the Cure Promise Grant. (KG081694). Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1664. doi:10.1158/1538-7445.AM2011-1664

Abstract CT127: Can potential curability be reached in high risk/metastatic breast cancer patients with a treatment optimization strategy that includes a novel GNT (gemcitabine, vinorelbine, docetaxel) in front line combination chemotherapy

Proceedings: AACR 106th Annual Meeting 2015; April 18-22, 2015; Philadelphia, PA Aiming at improving the therapeutic outcome of sequential taxane and anthracycline front line chemotherapy in breast cancer, G and N were incorporated into the taxane arm; GNT x 4 courses (G 1800-2500mg/m2, N 20-30mg/m2, T 100mg/m2, I.V. on day 1. GNT was administered sequentially with L-FAC x 4 courses; 72hr infusion of 400mg/m2/d 5-fluorouracil modulated by I.V. bolus 200mg/m2/d x 3 leucovorin. This was given concomitantly with 24hr d1 I.V. infusion 600mg/m2 cyclophosphamide, followed by 48hr I.V. infusion d2 + d3 60mg/m2 doxorubicin (ASCO 2006# 10741). Chemotherapy was given in a dose dense pattern with filgrastim support. As data became available, transtuzumab (for 1≤ year) was incorporated in HER2+ subset (15 patients) as of 11/2004, cis-platin (P) 70mg/m2/d1; GNTP in HER2+ and Triple negative (TN) subsets (20 patients) as of 5/2007 and bevacizumab 5mg/kg/wk I.V. in inflammatory and Stage IV HER2- subsets (6 patients) as of 10/2007. Hormone receptor positive (HOR+) subset (18 patients) received post chemotherapy hormone maintenance. Ten patients also received additional therapies. From 3/2001 to 2/2012, 52 patients (Median age 48) participated including the following stages: I; 3 (6%), II; 24 (46%), III; 17 (33%), IV; 8 (15%). The following subsets were represented; Triple negative 22 (42%), HER2+ 19 (37%), HER2-/HOR+ 11 (21%). Modified Blacks nuclear grade 3 was noted in 71% and Ki67 > 14% in 91%. 83% had breast conservation and 23 (44%) neoadjuvant chemotherapy; 87% achieved major pathologic response (14 PCR, 2 RCB1, 4 PET/CR/negative biopsy) at a median follow up of 75 months (m). Adjuvant chemotherapy follow up was longer at 116 m. At 81 m, 8/8 Stage IV patients achieved complete remission, though 1 patient (HER2+) had successfully treated intracranial relapse and another developed a second breast primary. At an overall follow up of 85 m (29-154), 4 relapses were observed; 2 HER+ (Stage III/without transtuzumab; Stage IV intracranial relapse only), 1 Inflammatory TN without PCR, 1 HOR+ late relapse > 10 year with short hormonal therapy. There were 3 deaths; 2 breast cancer related. Freedom from relapse for TN was 95% at 82 m, HER2+ 89% at 84 m, HER-/HOR+ 91% at 116 m. Dose dense limiting events included thrombocytopenia, muco-cutaneous reactions, and patient desired breaks. Other side effects noted were; decreased cardiac ejection fraction (2), decreased hearing (1), chronic peripheral neuropathy (2), AML (1), and MDS (1). Despite the limited study population number, this treatment approach resulted in high pathologic remission, complete remission in Stage IV and substantial freedom from relapse in high-risk and metastatic breast cancer subsets. This is highly suggestive that the natural history of aggressive breast cancer can be favorably influenced with this treatment approach. Citation Format: Khaled Jabboury, Lena Rogers, Krystal Sexton, Odelia Garcia. Can potential curability be reached in high risk/metastatic breast cancer patients with a treatment optimization strategy that includes a novel GNT (gemcitabine, vinorelbine, docetaxel) in front line combination chemotherapy. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr CT127. doi:10.1158/1538-7445.AM2015-CT127

Abstract P3-10-01: Epidemiological risk factors and normal breast tissue markers in inflammatory breast cancer

Background: Inflammatory breast cancer (IBC) is a rare form of aggressive breast cancer with no existing identifiers for screening or prevention strategies. Women with triple-negative (TNBC, ER–, PR–, Her2–) non-inflammatory breast cancer are less likely to breastfeed, and we have shown in adjacent normal breast tissue that this tissue has more foci of stem cells compared to non-TNBC cancers. A disproportionately higher percentage of women with IBC have TNBC relative to women with non-IBC. We hypothesized that adjacent normal tissue in TNBC IBC vs. TN non-IBC may also display unique biological features based on epidemiologic characteristics. Methods: We examined epidemiologic factors by breast cancer receptor subtype in 144 patients diagnosed with IBC in 1991–2011 at MD Anderson Cancer Center. Breast cancer risk factors including parity and breastfeeding were compared between patients with TN and non-TN IBC with chi-square or Wilcoxon rank sum tests. Normal adjacent tissues were stained for stem cell markers CD44+CD49f+CD133/2+ and macrophage marker CD68. Results: The mean age at diagnosis was 52.3 years (range = 23–80) and 83% of patients were non-Hispanic white, 80% were overweight or obese (BMI >25), and 36% were TN IBC. Patients with TN IBC had significantly lower frequency of breastfeeding compared with non-TN IBC, 28% vs. 55%, (p = 0.01). No differences were found in the frequency of other breast cancer risk factors. All 8 IBC adjacent tissue samples showed a distinct spatial distribution of stem cell staining, not limited to the triple negative patients. Compared with 0/60 non-IBC cases, 0/8 triple negative non-IBC, (p = 0.001 Pearson chi-square). Given the high BMI among IBC patients, we further examined normal tissues for the presence of CD68+ cells distributed individually or as clusters exhibiting a “crown-like” pattern (multiple CD 68+ macrophages found around dead adipocytes), and found that 7 of the 8 IBC adjacent tissues were CD68+. Benign biopsies collected from 2 patients at 10 years before diagnosis displayed similar staining, including both stem cell and CD68 staining. Compared with 12/60 non-IBC adjacent tissues were positive for CD68, with 1/8 TN non-IBC, (p = 0.001 Pearson chi-square). Conclusion: We describe for the first time a stem-cell staining pattern unique to IBC present in all IBC tissues examined, including pre-cancer biopsies. Tissue samples from additional patients will be examined to further explore the relationship between stem cells and CD68 positivity with IBC subtypes. Citation Information: Cancer Res 2012;72(24 Suppl):Abstract nr P3-10-01.