Patient-derived response estimates from zero-passage organoids of luminal breast cancer.

Background: Primary luminal breast cancer cells lose their identity rapidly in standard tissue culture, which is problematic for testing hormone interventions and molecular pathways specific to the luminal subtype. Breast cancer organoids are thought to retain tumor characteristics better, but long-term viability of luminal-subtype cases is a persistent challenge. Our goal was to adapt short-term organoids of luminal breast cancer for parallel testing of genetic and pharmacologic perturbations. Methods: We freshly isolated patient-derived cells from luminal tumor scrapes, miniaturized the organoid format into 5 µl replicates for increased throughput, and set an endpoint of 14 days to minimize drift. Therapeutic hormone targeting was mimicked in these "zero-passage" organoids by withdrawing ß-estradiol and adding 4-hydroxytamoxifen. We also examined sulforaphane as an electrophilic stress and commercial neutraceutical with reported anti-cancer properties. Downstream mechanisms were tested genetically by lentiviral transduction of two complementary sgRNAs and Cas9 stabilization for the first week of organoid culture. Transcriptional changes were measured by RT-qPCR or RNA sequencing, and organoid phenotypes were quantified by serial brightfield imaging, digital image segmentation, and regression modeling of cellular doubling times. Results: We achieved >50% success in initiating luminal breast cancer organoids from tumor scrapes and maintaining them to the 14-day zero-passage endpoint. Success was mostly independent of clinical parameters, supporting general applicability of the approach. Abundance of ESR1 and PGR in zero-passage organoids consistently remained within the range of patient variability at the endpoint. However, responsiveness to hormone withdrawal and blockade was highly variable among luminal breast cancer cases tested. Combining sulforaphane with knockout of NQO1 (a phase II antioxidant response gene and downstream effector of sulforaphane) also yielded a breadth of organoid growth phenotypes, including growth inhibition with sulforaphane, growth promotion with NQO1 knockout, and growth antagonism when combined. Conclusions: Zero-passage organoids are a rapid and scalable way to interrogate properties of luminal breast cancer cells from patient-derived material. This includes testing drug mechanisms of action in different clinical cohorts. A future goal is to relate inter-patient variability of zero-passage organoids to long-term outcomes.

Yolk Sac Differentiation in Endometrial Carcinoma: Incidence and Clinicopathologic Features of Somatically Derived Yolk Sac Tumors Versus Carcinomas With Nonspecific Immunoexpression of Yolk Sac Markers.

Endometrial somatically derived yolk sac tumors are characterized by yolk sac morphology with AFP, SALL-4, and/or Glypican-3 immunoexpression. Yolk sac marker expression, however, is not limited to tumors with overt yolk sac histology. Three hundred consecutive endometrial malignancies were assessed for immunomarkers of yolk sac differentiation. Of these, 9% expressed ≥1 yolk sac marker, including 29% of high-grade tumors. Only 3 (1%) met morphologic criteria for yolk sac differentiation; these were originally diagnosed as serous, high-grade NOS, and dedifferentiated carcinoma. Two were MMR-intact and comprised exclusively of yolk sac elements, while the dedifferentiated case was MMR deficient and had a background low-grade endometrioid carcinoma; this case also showed BRG1 loss. All 3 were INI1 intact. Nonspecific yolk sac marker expression was seen in 14 carcinosarcomas, 4 endometrioid, 2 serous, 1 clear cell, 1 dedifferentiated, 1 mixed serous/clear cell, and 1 mesonephric-like carcinoma. INI1 was intact in all cases; one showed BRG1 loss. Twenty were MMR-intact, and 4 were MMR deficient. All MMR-deficient cases with yolk sac marker expression, both with and without true yolk sac morphology, had no evidence of residual disease on follow-up, whereas 82% of MMR-intact cases developed recurrent/metastatic disease. In summary, endometrial somatically derived yolk sac tumors were rare but under-recognized. While AFP immunostaining was specific for this diagnosis, Glypican-3 and SALL-4 expression was seen in a variety of other high-grade carcinomas. INI1 loss was not associated with yolk sac morphology or immunomarker expression in the endometrium, and BRG1 loss was rare. All patients with MMR-deficient carcinomas with yolk sac immunoexpression +/- morphology were disease-free on follow-up, whereas the majority of MMR-intact cancers showed aggressive disease.

A genetic mosaic mouse model illuminates the pre-malignant progression of basal-like breast cancer.

Basal-like breast cancer (BLBC) is highly aggressive, and often characterized by BRCA1 and p53 deficiency. Although conventional mouse models enabled the investigation of BLBC at malignant stages, its initiation and pre-malignant progression remain understudied. Here, we leveraged a mouse genetic system known as mosaic analysis with double markers (MADM) to study BLBC initiation by generating rare GFP+Brca1, p53-deficient mammary cells alongside RFP+ wild-type sibling cells. After confirming the close resemblance of mammary tumors arising in this model to human BLBC at both transcriptomic and genomic levels, we focused our studies on the pre-malignant progression of BLBC. Initiated GFP+ mutant cells showed a stepwise pre-malignant progression trajectory from focal expansion to hyper-alveolarization and then to micro-invasion. Furthermore, despite morphological similarities to alveoli, hyper-alveolarized structures actually originate from ductal cells based on twin-spot analysis of GFP-RFP sibling cells. Finally, luminal-to-basal transition occurred exclusively in cells that have progressed to micro-invasive lesions. Our MADM model provides excellent spatiotemporal resolution to illuminate the pre-malignant progression of BLBC, and should enable future studies on early detection and prevention for this cancer.

The POWER-PAK Score Characterizes Tumor Response to 3 Months of Preoperative Endocrine Therapy.

BACKGROUND: The Pre-Operative Window of Endocrine Therapy to Inform Radiation Therapy Decisions (POWER, NCT04272801) trial aims to determine whether 3 months of preoperative endocrine therapy (pre-ET) informs adjuvant radiation therapy decisions among older women with early stage, ER-positive breast cancer. We propose the POWER Pathologic Assessment and Ki-67 (POWER-PAK) scoring system to characterize the histologic effects of pre-ET. METHODS: Histologic evaluation was performed on core biopsy and lumpectomy specimens from 37 POWER trial participants who completed pre-ET and surgery. The POWER-PAK score consists of tumor regression, decrease in Ki-67 expression, and ER expression, each ranging from 0 to 2. Scores were aggregated to create the POWER-PAK score with a range from 0 to 6. Participants with no residual tumor were labelled 6-NRT. RESULTS: ER expression did not decrease after pre-ET. Ki-67 decreased from 13% in biopsy specimens to 5% in the lumpectomy specimens (p < 0.001). Cellularity decreased from 40% to 23% (p < 0.001). There was heterogeneity of POWER-PAK scores ranging from 2 to 6-NRT: score of 2, n = 2 (5.4%); 4, n = 8 (21.6%); 5, n = 4 (10.8%); 6, n = 16 (43.2%); and 6-NRT, n = 7 (18.9%). Participants with a score ≥ 5 were more likely to have smaller tumors after pre-ET compared with those with a score < 5 (p = 0.04). CONCLUSIONS: The tumor responses following treatment with pre-ET are heterogenous. We propose that the POWER-PAK scoring system can be used to quantify response to pre-ET. Future studies will explore the use of POWER-PAK to support informed decision-making for adjuvant therapy options for older women with early stage breast cancer.

Dichotomous ovarian cancer-initiating potential of Pax8+ cells revealed by a mouse genetic mosaic model.

Different cellular compartments within a tissue present distinct cancer-initiating capacities. Current approaches to dissect such heterogeneity require cell-type-specific genetic tools based on a well-understood lineage hierarchy, which are lacking for many tissues. Here, we circumvented this hurdle and revealed the dichotomous capacity of fallopian tube Pax8+ cells in initiating ovarian cancer, utilizing a mouse genetic system that stochastically generates rare GFP-labeled mutant cells. Through clonal analysis and spatial profiling, we determined that only clones founded by rare, stem/progenitor-like Pax8+ cells can expand on acquiring oncogenic mutations whereas vast majority of clones stall immediately. Furthermore, expanded mutant clones undergo further attrition: many turn quiescent shortly after the initial expansion, whereas others sustain proliferation and manifest a bias toward Pax8+ fate, underlying early pathogenesis. Our study showcases the power of genetic mosaic system-based clonal analysis for revealing cellular heterogeneity of cancer-initiating capacity in tissues with limited prior knowledge of lineage hierarchy.

A genetic mosaic mouse model illuminates the pre-malignant progression of basal-like breast cancer.

Basal-like breast cancer is an aggressive breast cancer subtype, often characterized by a deficiency in BRCA1 function and concomitant loss of p53 . While conventional mouse models enable the investigation of its malignant stages, one that reveals its initiation and pre-malignant progression is lacking. Here, we leveraged a mouse genetic system known as M osaic A nalysis with D ouble M arkers (MADM) to generate rare GFP-labeled Brca1 , p53 -deficient cells alongside RFP+ wildtype sibling cells in the mammary gland. The mosaicism resembles the sporadic initiation of human cancer and enables spatially resolved analysis of mutant cells in comparison to paired wildtype sibling cells. Mammary tumors arising in the model show transcriptomic and genomic characteristics similar to human basal-like breast cancer. Analysis of GFP+ mutant cells at interval time points before malignancy revealed a stepwise progression of lesions from focal expansion to hyper-alveolarization and then to micro-invasion. These stereotyped morphologies indicate the pre-malignant stage irrespective of the time point at which it is observed. Paired analysis of GFP-RFP siblings during focal expansion suggested that hyper-alveolarized structures originate from ductal rather than alveolar cells, despite their morphological similarities to alveoli. Evidence for luminal-to-basal transition at the pre-malignant stages was restricted to cells that had escaped hyper-alveoli and progressed to micro-invasive lesions. Our MADM-based mouse model presents a useful tool for studying the pre-malignancy of basal-like breast cancer. Summary statement: A mouse model recapitulates the process of human basal-like breast tumorigenesis initiated from sporadic Brca1, p53 -deficient cells, empowering spatially-resolved analysis of mutant cells during pre-malignant progression.

Breast Cancer Antiestrogen Resistance 3 (BCAR3) promotes tumor growth and progression in triple-negative breast cancer.

Triple-Negative Breast Cancers (TNBCs) constitute roughly 10-20% of breast cancers and are associated with poor clinical outcomes. Previous work from our laboratory and others has determined that the cytoplasmic adaptor protein Breast Cancer Antiestrogen Resistance 3 (BCAR3) is an important promoter of cell motility and invasion of breast cancer cells. In this study, we use both in vivo and in vitro approaches to extend our understanding of BCAR3 function in TNBC. We show that BCAR3 is upregulated in ductal carcinoma in situ (DCIS) and invasive carcinomas compared to normal mammary tissue, and that survival of TNBC patients whose tumors contained elevated BCAR3 mRNA is reduced relative to individuals whose tumors had less BCAR3 mRNA. Using mouse orthotopic tumor models, we further show that BCAR3 is required for efficient TNBC tumor growth. Analysis of publicly available RNA expression databases revealed that MET receptor signaling is strongly correlated with BCAR3 mRNA expression. A functional role for BCAR3-MET coupling is supported by data showing that both proteins participate in a single pathway to control proliferation and migration of TNBC cells. Interestingly, the mechanism through which this functional interaction operates appears to differ in different genetic backgrounds of TNBC, stemming in one case from potential differences in the strength of downstream signaling by the MET receptor and in another from BCAR3-dependent activation of an autocrine loop involving the production of HGF mRNA. Together, these data open the possibility for new approaches to personalized therapy for individuals with TNBCs.

Pan-Cancer Drivers Are Recurrent Transcriptional Regulatory Heterogeneities in Early-Stage Luminal Breast Cancer.

The heterogeneous composition of solid tumors is known to impact disease progression and response to therapy. Malignant cells coexist in different regulatory states that can be accessed transcriptomically by single-cell RNA sequencing, but these methods have many caveats related to sensitivity, noise, and sample handling. We revised a statistical fluctuation analysis called stochastic profiling to combine with 10-cell RNA sequencing, which was designed for laser-capture microdissection (LCM) and extended here for immuno-LCM. When applied to a cohort of late-onset, early-stage luminal breast cancers, the integrated approach identified thousands of candidate regulatory heterogeneities. Intersecting the candidates from different tumors yielded a relatively stable set of 710 recurrent heterogeneously expressed genes (RHEG), which were significantly variable in >50% of patients. RHEGs were not strongly confounded by dissociation artifacts, cell-cycle oscillations, or driving mutations for breast cancer. Rather, RHEGs were enriched for epithelial-to-mesenchymal transition genes and, unexpectedly, the latest pan-cancer assembly of driver genes across cancer types other than breast. These findings indicate that heterogeneous transcriptional regulation conceivably provides a faster, reversible mechanism for malignant cells to evaluate the effects of potential oncogenes or tumor suppressors on cancer hallmarks. SIGNIFICANCE: Profiling intratumor heterogeneity of luminal breast carcinoma cells identifies a recurrent set of genes, suggesting sporadic activation of pathways known to drive other types of cancer.See related articles by Schaff and colleagues, p. 1853 and Sutcliffe and colleagues, p. 1868.

ECTOPIC THYROID TISSUE MASQUERADING AS A FUNCTIONAL CAROTID BODY PARAGANGLIOMA.

OBJECTIVE: Ectopic thyroid tissue (ETT) is a rare entity resulting from thyroid gland dysembryogenesis. We present a case of ETT confirmed by histopathology that was misdiagnosed clinically as a carotid body tumor. METHODS: A 34-year-old female with a history of thyroidectomy for a goiter presented with 1 year of worsening tachycardia (heart rate ranging from 82 to 111 beats per minute), anxiety, hot flashes, and intolerance to heat. For further evaluation, we obtained imaging of her neck, including a thyroid ultrasound, a computed tomography (CT) scan, and an octreotide scan. We also performed laboratory studies including fractionated 24-hour urine meta-nephrines and thyroid function tests. RESULTS: Her thyroid ultrasound showed a mass at the right carotid bifurcation, which was confirmed on CT as well as on an octreotide scan. Her free thyroxine was 0.6 ng/dL (normal, 0.7 to 1.5 ng/dL) and her thyroid-stimulating hormone was 4.51 mIU/L (normal, 0.45 to 4.5 mIU/L). Her fractionated 24-hour total urine metanephrines were 1,502 mcg/24-hour (normal, 149 to 535 mcg/24-hour). She underwent resection of a vascular mass from the carotid bifurcation. Histologic examination revealed ETT with dilated follicles filled with colloid with no evidence of paraganglioma/carotid body tumor. CONCLUSION: The somatostatin receptor is typically present in paragangliomas; however, there are reports of octreotide uptake within thyroid goiters. It has been demonstrated that psychoactive medications can increase urine metanephrines. Given the patient's psychiatric history and that no other tumors were identified on imaging, it was felt that the elevated urine normetanephrine in this case was most likely due to psychoactive medication use. This case demonstrates the preoperative imaging findings and postoperative pathologic confirmation of an unusual presentation of ETT.

Sporadic activation of an oxidative stress-dependent NRF2-p53 signaling network in breast epithelial spheroids and premalignancies.

Breast and mammary epithelial cells experience different local environments during tissue development and tumorigenesis. Microenvironmental heterogeneity gives rise to distinct cell regulatory states whose identity and importance are just beginning to be appreciated. Cellular states diversify when clonal three-dimensional (3D) spheroids are cultured in basement membrane, and one such state is associated with stress tolerance and poor response to anticancer therapeutics. Here, we found that this state was jointly coordinated by the NRF2 and p53 pathways, which were costabilized by spontaneous oxidative stress within 3D cultures. Inhibition of NRF2 or p53 individually disrupted some of the transcripts defining the regulatory state but did not yield a notable phenotype in nontransformed breast epithelial cells. In contrast, combined perturbation prevented 3D growth in an oxidative stress-dependent manner. By integrating systems models of NRF2 and p53 signaling in a single oxidative stress network, we recapitulated these observations and made predictions about oxidative stress profiles during 3D growth. NRF2 and p53 signaling were similarly coordinated in normal breast epithelial tissue and hormone-negative ductal carcinoma in situ lesions but were uncoupled in triple-negative breast cancer (TNBC), a subtype in which p53 is usually mutated. Using the integrated model, we correlated the extent of this uncoupling in TNBC cell lines with the importance of NRF2 in the 3D growth of these cell lines and their predicted handling of oxidative stress. Our results point to an oxidative stress tolerance network that is important for single cells during glandular development and the early stages of breast cancer.

Pathologic Evaluation and Prognostic Implications of Nodal Micrometastases in Breast Cancer.

Using modern sentinel lymph node techniques, occult nodal metastases including micrometastases or isolated tumor cells are being increasingly discovered in up to 10% of early-stage breast cancers. Furthermore, the rate of nonsentinel lymph node involvement is approximately 10%. However, the impact of these findings on disease-free survival is low, particularly with regards to axillary recurrences. Current evidence suggests small-volume lymph node involvement in breast cancer patients is only one of several factors that should guide adjuvant therapy options. In otherwise favorable patients, adjuvant radiation and systemic therapy can help mitigate the risk of recurrence when omitting axillary lymph node dissection.

GATA3 Expression in Common Gynecologic Carcinomas: A Potential Pitfall.

GATA binding protein 3 (GATA3) immunohistochemistry is primarily used as a marker of breast and urothelial differentiation, particularly in metastatic settings. In the gynecologic tract it also serves a robust marker for mesonephric and trophoblastic tumors. However, expression has also been described in more common malignancies of gynecologic tract including ovarian, endometrial, and cervical carcinomas. Data on the distribution of GATA3 expression in gynecologic malignancies is somewhat limited, particularly across different histologic subtypes of ovarian, endometrial, and cervical carcinomas. To assess the rates of GATA3 expression among common gynecologic cancers of various histologic types, 100 ovarian carcinomas, 64 endometrial carcinomas/atypical hyperplasias, 16 cervical squamous cell carcinomas (SCCs), and 14 endocervical adenocarcinomas were evaluated by immunohistochemistry for GATA3 positivity. Eight percent of endometrial carcinomas expressed GATA3, including 2 serous carcinomas, 1 carcinosarcoma, and 1 case of atypical hyperplasia. Six percent of ovarian carcinomas were GATA3-positive including 2 clear cell carcinomas, 2 mucinous adenocarcinomas, and 2 high-grade serous carcinomas. Thirty-eight percent of cervical SCCs showed weak to moderate staining in up to 50% of tumor cells. All endocervical adenocarcinomas were entirely negative for GATA3. In summary, GATA3 shows focal weak to moderate expression in a subset of endometrial and ovarian carcinomas. In contrast, usual-type endocervical adenocarcinomas are typically negative for GATA3, which can be helpful in differentiating them from mesonephric proliferations or carcinomas. A larger proportion of cervical SCCs express GATA3, therefore caution should be exercised when using this stain in the setting of a lower genitourinary carcinomas.

Clinicopathologic characterization of breast carcinomas in patients with non-BRCA germline mutations: results from a single institution's high-risk population.

As multigene panel testing for hereditary cancer syndromes becomes commonplace, germline mutations in genes other than BRCA1/2 are increasingly identified in breast cancer patients. While histopathologic features of BRCA-mutated breast cancers have been well-characterized, less is known about non-BRCA-related hereditary cancers. We herein investigate the clinicopathologic characteristics of breast cancers in women with non-BRCA germline mutations. Out of 612 women who underwent germline testing, 16 (2.6%) women with 18 cancers had mutations in non-BRCA genes: ATM, CHEK2, PALB2, TP53, BMPR1A, BRIP1, MUTYH, and RAD50. An additional 2 cancers were identified in a woman with a diagnosis of Bloom syndrome (BLM mutation) who was not germline tested. Average age at diagnosis was 50 (range: 27-77), and 65% had no personal cancer history. The majority (79%) of tumors were grade 1 to 2; 35% were either lobular or ductal with lobular features. Stromal responses varied from absent to desmoplastic to sclerotic; 69% of cases had an in situ component. With the exception of a brisk lymphocytic response in BLM- and TP53-mutated cancers, lymphocytic infiltration was mild or absent. In summary, the majority of non-BRCA-related hereditary breast cancers represent the patient's sentinel malignancy. Lobular features were seen in a subset, and high-grade, immunogenic carcinomas were uncommon except in the setting of BLM and TP53 mutations. Overall, these findings demonstrate a range of involved genes in non-BRCA mutation carriers with breast cancer and histopathologic heterogeneity in the associated cancers, arguing against use of histomorphology to inform panel testing algorithms.

Effect of a tissue selective estrogen complex on breast cancer: Role of unique properties of conjugated equine estrogen.

The Women's Health Initiative studies reported that the menopausal hormone therapy (MHT) regimen containing conjugated equine estrogen (CEE) and medroxyprogesterone acetate increased, whereas CEE alone reduced breast cancer incidence. These observations suggest the possibility that CEE might exert unique actions on breast and also suggest the need to eliminate the progestogen from MHT regimens. A MHT regimen called a tissue selective estrogen complex (TSEC), containing CEE plus bazedoxifene (BZA), to avoid the need for a progestogen, was developed and FDA approved. Our study addressed two questions regarding this TSEC: (i) whether CEE exert effects on breast cancer which differ from those of estradiol (E2 ) and (ii) whether BZA antagonize the effects of E2 and CEE on breast cancer? Two rodent models (NMU and ACI) were used to compare the effect of CEE with E2 on mammary tumor formation, proliferation and apoptosis. In both the NMU and ACI models, E2 significantly increased tumor incidence and multiplicity whereas in striking contrast CEE did not, even though the estrogenic effects of CEE and E2 on uterine weight were identical. Mechanistically E2 blocked whereas CEE stimulated apoptosis (cleaved caspase-3) in ACI animals and only E2 stimulated proliferation (Ki67). BZA exerted highly potent anti-estrogenic effects on tumors by completely blocking palpable tumor formation. These data suggest that the CEE/BZA TSEC may be a safer, breast-antagonistic, MHT agent for women and might have potential to prevent breast cancer while relieving menopausal symptoms.

Tumor-Suppressor Inactivation of GDF11 Occurs by Precursor Sequestration in Triple-Negative Breast Cancer.

Triple-negative breast cancer (TNBC) is an aggressive and heterogeneous carcinoma in which various tumor-suppressor genes are lost by mutation, deletion, or silencing. Here we report a tumor-suppressive mode of action for growth-differentiation factor 11 (GDF11) and an unusual mechanism of its inactivation in TNBC. GDF11 promotes an epithelial, anti-invasive phenotype in 3D triple-negative cultures and intraductal xenografts by sustaining expression of E-cadherin and inhibitor of differentiation 2 (ID2). Surprisingly, clinical TNBCs retain the GDF11 locus and expression of the protein itself. GDF11 bioactivity is instead lost because of deficiencies in its convertase, proprotein convertase subtilisin/kexin type 5 (PCSK5), causing inactive GDF11 precursor to accumulate intracellularly. PCSK5 reconstitution mobilizes the latent TNBC reservoir of GDF11 in vitro and suppresses triple-negative mammary cancer metastasis to the lung of syngeneic hosts. Intracellular GDF11 retention adds to the concept of tumor-suppressor inactivation and reveals a cell-biological vulnerability for TNBCs lacking therapeutically actionable mutations.

Mismatch repair status and PD-L1 expression in clear cell carcinomas of the ovary and endometrium.

Clear cell carcinoma represents a distinct histologic type of müllerian carcinoma that is resistant to conventional chemotherapy. Expression of programmed cell death ligand (PD-L1) has been associated with immune evasion in numerous tumor types and may be used to identify patients who will benefit from targeted immunotherapy, particularly in the setting of mismatch repair defects. We evaluated PD-L1 expression in 23 ovarian clear cell carcinomas and 21 endometrial clear cell carcinomas, and correlated expression with mismatch repair status. Tumor PD-L1 staining was seen in 43% of ovarian tumors and 76% of endometrial tumors, including 71% of cases (67% of ovarian and 75% of endometrial) with mismatch repair defects. Extensive tumoral staining (>50%) was seen in only one case (an endometrial case with MSH6 loss). However, tumoral PD-L1 expression remained common in mismatch repair-intact tumors and mismatch repair status was not significantly correlated with PD-L1 expression. The increased incidence of PD-L1 positivity in tumor cells (P=0.04) in endometrial vs ovarian clear cell carcinomas suggests differences in the tumor microenvironment of these histologically and molecularly similar tumors that may inform treatment options. These results suggest that clear cell histology may be a useful susceptibility marker for immunotherapy targeting the PD-1/PD-L1 axis irrespective of mismatch repair status, particularly in endometrial carcinomas.

The Association of Mammographic Density and Molecular Breast Cancer Subtype.

Background: Mammographic density (MD) is associated with increased breast cancer risk, yet limited data exist on an association between MD and breast cancer molecular subtypes.Methods: Women ages 18 years and older with breast cancer and available mammograms between 2003 and 2012 were enrolled in a larger study on MD. MD was classified by the Breast Imaging Reporting and Data System (BI-RADS) classification and by volumetric breast percent density (Volpara Solutions). Subtype was assigned by hormone receptor status, tumor grade, and mitotic score (MS). Subtypes included: Luminal-A (ER/PR+ and grade = 1; ER/PR+ and grade = 2 and MS = 1; ER+/PR- and grade = 1; n = 233); Luminal-B (ER+ and grade = 3 or MS = 3; ER+/PR- and grade = 2; ER/PR+ and grade = 2 and MS = 2; n = 79); Her-2-neu+ (H2P; n = 59); triple-negative (ER/PR-, Her-2-; n = 86). Precancer factors including age, race, body mass index (kg/m2), family history of breast cancer, and history of lobular carcinoma in situ were analyzed.Results: A total of 604 patients had invasive cancer; 457 had sufficient information for analysis. Women with H2P tumors were younger (P = 0.011) and had the highest volumetric percent density (P = 0.002) among subgroups. Multinomial logistic regression (LA = reference) demonstrated that although quantitative MD does not significantly differentiate between all subtypes (P = 0.123), the association between MD and H2P tumors is significant (OR = 1.06; confidence interval, 1.01-1.12). This association was not seen using BI-RADS classification in bivariable analysis but was statistically significant (P = 0.047) when controlling for other precancer factors.Conclusions: Increased MD is more strongly associated with H2P tumors when compared with LA.Impact: Delineating risk factors specific to breast cancer subtype may promote development of individualized risk prediction models and screening strategies. Cancer Epidemiol Biomarkers Prev; 26(10); 1487-92. ©2017 AACR.

Mesothelium expression of vascular cell adhesion molecule-1 (VCAM-1) is associated with an unfavorable prognosis in epithelial ovarian cancer (EOC).

BACKGROUND: Mesothelium vascular cell adhesion molecule-1 (VCAM-1) expression in the metastatic epithelial ovarian cancer (EOC) microenvironment is induced by tumor and mediates tumor cell invasion. VCAM-1 imaging suggests expression during treatment is an indicator of platinum resistance. Here, we assess the potential prognostic significance of mesothelium VCAM-1 expression and prospectively evaluate whether soluble VCAM-1 (sVCAM-1) is a surrogate for mesothelium expression. METHODS: A retrospective review of EOC patients was performed to evaluate outcomes with mesothelium VCAM-1 expression determined by immunohistochemistry of peritoneum or omentum specimens. A prospective cohort of EOC patients was identified and followed through primary treatment. Serum for sVCAM-1 evaluation, which was performed via enzyme-linked immunosorbent assay, was collected before surgery or neoadjuvant chemotherapy and at each treatment cycle. Peritoneal specimens were obtained during debulking to assess mesothelial VCAM-1 expression. RESULTS: A retrospective review identified 54 advanced-stage EOC patients. Patients expressing mesothelium VCAM-1 had shortened overall survival (44 vs 79 months, P = 0.035) and progression-free survival (18 vs 67 months, P = 0.010); the median time to platinum resistance was 36 months for VCAM-1-expressing patients and not yet determined for the VCAM-1-negative group. In our prospective observational cohort, 18 EOC patients completed primary treatment; 3 were negative for mesothelium VCAM-1 expression, and sVCAM-1 did not vary between groups. CONCLUSIONS: Mesothelium VCAM-1 expression is negatively associated with progression-free and overall survival in EOC. This is especially compelling in light of previous data suggesting that persistent VCAM-1 expression during treatment is an indicator of platinum resistance. Our pilot study had insufficient cases to determine whether sVCAM-1 would substitute for mesothelium expression. Cancer 2017;123:977-84. © 2016 American Cancer Society.

Fine Needle Aspiration Cytology of Diffuse-Type Tenosynovial Giant Cell Tumors.

BACKGROUND: Tenosynovial giant cell tumor (TSGCT), also known as giant cell tumor of tendon sheath or pigmented villonodular synovitis, is the most common benign tumor of the tendon and synovium. The intra-articular diffuse type can present as a large infiltrative mass involving adjacent soft tissue and sometimes causes secondary destruction of bone, which leads to radiographic and clinical concern for malignancy. The tumor may also be purely extra-articular. CASE: Here, we report the fine needle aspiration cytology findings of 2 cases of diffuse-type TSGCT with large mononuclear cells with eccentric nuclei, finely granular cytoplasm, and a peripheral well-defined cytoplasmic rim of hemosiderin ("ladybird cells"). CONCLUSION: Although the presence of ladybird cells has been described in tissue sections of TSGCT, their identification in cytological specimens has not been reported to our knowledge. When observed, their presence may aid in differentiating TSGCT from other lesions with multinucleated osteoclast-type giant cells occurring at or near joints.