Faculty Bibliography

Background: Microglandular adenosis (MGA) is a rare borderline lesion of the breast characterized by an infiltrative collection of small glands that characteristically lack a myoepithelial cell layer. MGA is associated with invasive carcinoma in 20-30% of cases, and has been proposed as a non-obligate precursor to basal-like breast cancers. Somatic TP53 mutation of MGA and its associated carcinoma has been previously reported. We identified a case of triple negative metaplastic carcinoma with mesenchymal differentiation with morphologic and immunohistochemical evidence of progression from MGA to atypical MGA (AMGA), carcinoma in situ (CIS) and invasive carcinoma. We performed laser microdissection and whole exome sequencing of each four components (MGA, AMGA, CIS and cancer) with a matched benign sample to characterize the mutational landscape of these foci. Design: We selected a case of a metaplastic carcinoma with mesenchymal differentiation in juxtaposition to foci of MGA, AMGA and CIS. Immunohistochemically, all four foci were negative for estrogen receptor, progesterone receptor and human epidermal growth factor receptor 2 (her2) and positive for S-100. The distinct morphologic components and a matched control lymph node were separately microdissected from formalin-fixed paraffin embedded blocks. DNA extraction was performed and subjected to whole-exome sequencing on Illumina HiSeq platform. The results were demultiplexed and converted to FASTQ format using Illumina bcl2fastq software. Singlenucleotide and small indel somatic variants were called with MuTect2. Copy number profiles were calculated using Control-FREEC. Clonal populations were identified and quantified using PyClone. Results: Sequencing data resulted in mean coverage of 96-134X of targeted exome regions. Our results found a recurrent stop-gain R213* TP53 mutation in MGA, atypical MGA, CIS and metaplastic carcinoma. In addition, through variant allele frequency analysis, we identified two putative clonal clusters shared by all foci indicating a common molecular signature that is preserved in the morphologic spectrum of MGA-AMGA-CIS-metaplastic carcinoma. Conclusions: MGA is a molecularly advanced lesion with somatic mutation of TP53. We postulate that TP53 is an early event in the progression of MGA through AMGA, CIS and its associated metaplastic carcinoma. We report significant genetic overlap between MGA and its associated cancer

Background: Over 250,000 breast cancer cases are diagnosed annually in the US, of which 15% (n=37,500) are triple negative breast cancers (TNBC). TNBC has an aggressive clinical course with limited therapeutic options. Some studies estimate that a small proportion of TNBCs defined by immunohistochemical (IHC) stains alone, i.e. estrogen receptor (ER), progesterone receptor (PR) and human epidermal growth factor receptor 2 (HER2)-negative, can, in fact, show HER2 amplification by fluorescent in situ hybridization (FISH) and, therefore, be reclassified as HER2-enriched. If properly classified, these patients can benefit from anti-HER2 therapy. To capture this subset of patients, we performed a now four-year-long prospective study, in which TNBCs were reflexed to HER2 by FISH. Herein, we present our findings with detailed clinicopathologic analysis of the reclassified cases. Design: TNBC cases and ER/PR-positive, HER2-negative breast cancers from 2014 to 2017 with concurrent IHC and FISH results were analyzed. The pathology slides were reviewed by two pathologists. Stromal tumor infiltrating lymphocytes (sTILs) were defined as the percentage of all mononuclear cells within tumor stroma not in direct contact with tumor cells. Results: Of the 253 TNBCs in the past 4 years, 13 tumors (5%) showed HER2 amplification by FISH (6 surgical excisions and 7 core biopsies). Two patients have previously identified BRCA1 mutation. Twelve of 13 tumors (92%) were grade 3. All tumors were invasive ductal carcinoma of no special type. No apocrine morphology was noted. Notably, five tumors had >=50% sTILs, which can be classified as lymphocyte-predominant breast cancer based on some studies. In the same time period, 41 ER/PR-positive/HER2-negative breast cancers were reflexed to HER2 by FISH, none of which showed amplification. All tumors were fixed in formalin for comparable amount of time (6-72 hours). After reclassification, approximately 40% of patients received anti-HER2 therapy. (Table Presented) Conclusions: Pathologists may consider reflexing HER2 evaluation to FISH in TNBCs with grade 3, high ki67 and high sTILs. If 5% of TNBCs can be reclassified as HER2-enriched tumors, based on the national statistics, annually approximately 1875 patients with TNBCs by IHC may actually benefit from anti-HER2 therapy

Background: Traditionally, the American Joint Committee on Cancer (AJCC) staging uses Tumor size, Node status and distant Metastasis (TNM) to determine breast cancer prognosis, known as anatomic stage (AS). In the eighth edition, new parallel prognostic stage (PS) groups incorporate tumor biomarkers such as estrogen receptor (ER), progesterone receptor (PR), human epidermal growth factor receptor 2 (HER2), tumor grade and Recurrence Score (RS). The new PS system attempts to refine the prognostic groups tailored to the tumor biology. Starting 2018, both AS and PS will be available to patients. In anticipation of the change, we performed a one-year retrospective study to look at how the breast cancer cases would be staged according to the new PS compared to the traditional AS. Design: Patients diagnosed with primary breast cancer in 2016 were identified using pathology laboratory software system. Only the cases with positive ER, PR and negative HER2 and concurrent Oncotype Dx RS were included. The cases were reviewed and stratified using the traditional TNM AS groups and also into the new PS groups. The change of each case's AS and PS was analyzed. Results: 106 cases were reviewed; 27 cases had Oncotype Dx RS less than 11 and 79 had equal to and greater than 11. Most common AS and PS were IA (67 and 65 cases, respectively). PS remained unchanged in 58.5% of cases, of which 91% were stage IA. Seventeen (16%) cases were upstaged and 27 (25%) cases were downstaged (see table 1). Conclusions: The new PS staging will bring about significant change to the traditional AS system-nearly half (42%) of cases reviewed at our institution would see a change in new prognostic stage grouping. We, as pathologists, need to reach a consensus as to how to integrate the results of the ancillary tests into our pathology reports in preparation for the upcoming PS to better serve our breast cancer patients

Bromodomain and extraterminal domain inhibitors (BETi) represent promising therapeutic agents for metastatic melanoma, yet their mechanism of action remains unclear. Here we interrogated the transcriptional effects of BETi and identified AMIGO2, a transmembrane molecule, as a BET target gene essential for melanoma cell survival. AMIGO2 is upregulated in melanoma cells and tissues compared to human melanocytes and nevi, and AMIGO2 silencing in melanoma cells induces G1/S arrest followed by apoptosis. We identified the pseudokinase PTK7 as an AMIGO2 interactor whose function is regulated by AMIGO2. Epigenomic profiling and genome editing revealed that AMIGO2 is regulated by a melanoma-specific BRD2/4-bound promoter and super-enhancer configuration. Upon BETi treatment, BETs are evicted from these regulatory elements, resulting in AMIGO2 silencing and changes in PTK7 proteolytic processing. Collectively, this study uncovers mechanisms underlying the therapeutic effects of BETi in melanoma and reveals the AMIGO2-PTK7 axis as a targetable pathway for metastatic melanoma.

OBJECTIVE: To report a case of refractory ulcerated necrobiosis lipoidica (NL) with significant response to treatment with topical tacrolimus. SUBJECT: A 55-year-old woman without diabetes and with a previous history of NL presented to the Helen L. and Martin S. Kimmel Hyperbaric and Advanced Wound Healing Center of NYU Langone Medical Center, New York, with bilateral lower-leg ulcerations resistant to wound healing techniques at other institutions. MATERIALS AND METHODS: Repeat biopsy performed at the author's institution confirmed the diagnosis of NL. Initial therapy was based on reports of other successful treatment methods, which included collagen wound grafts and collagen-based dressings coupled with compression. These methods initially showed promising results; however, the wounds reulcerated, and any gains in wound healing were lost. Alternative options were initiated, including topical clobetasol and narrowband ultraviolet B; however, no significant improvement was observed. The patient's lower-extremity wounds began to deteriorate. The patient also refused systemic therapy. Treatment was changed to topical 0.1% tacrolimus ointment and was applied daily for 10 months with multilayer compression wraps. RESULTS: Both lower-extremity ulcerations began to show significant improvement, with the ulcers progressing toward closure except for 1 very small area on the left lower extremity. CONCLUSIONS: Topical tacrolimus seems to be an effective treatment option for patients with refractory chronic ulcerated NL who do not want systemic oral therapy. The authors found that successful wound closure may require a multimodal approach, which promotes wound healing, but also concurrently addresses the underlying disease process.

Background: Efforts to identify targeted therapies that can improve treatment outcome in metastatic ALM have been unsuccessful. In a previous genomic screening, we identified copy number amplification of the histone methyltransferase EZH2 in 47% of ALM cases, a higher frequency than previously reported in cutaneous melanomas (CM) (5%). Here, we tested the hypothesis that increased EZH2 expression contributes to ALM progression and may confer selective sensitivity to EZH2 inhibition. Methods: EZH2 expression was examined by immunohistochemistry (IHC) in 51 primary (21 stage I, 13 Stage II and 17 Stage III) and 23 metastatic (11 in transit, 8 nodal and 4 visceral) ALM cases with extensive clinicopathological data and protocol-driven follow up. Colony formation and cell proliferation was assessed following treatment of ALM and CM cell lines with three EZH2 inhibitors, including GSK126, currently in clinical trials. The effect of GSK126 on H3K27me3 and downstream EZH2 targets was analyzed by western blotting. Results: EZH2 is commonly overexpressed in both primary (30/51; 65%) and metastatic (20/23; 87%) ALM cases, with a significant increase in mean IHC score between primary and metastatic tumors (1.9 vs. 2.7, respectively, p = 0.047). EZH2 expression increased in 6/10 metastatic ALM tumors compared to their matched primary tumors. ALM tumors with EZH2 gene amplification showed increased EZH2 protein expression; however more cases showed overexpression with no amplification suggesting a potential epigenetic component of EZH2 regulation. GSK126 significantly suppressed ALM colony formation at lower doses compared to CM (1 muM vs. 5 muM, respectively). EZH2 inhibition also increased expression of the downstream tumor suppressor E-cadherin in ALM but not in CM cell lines. Finally, ALM cell lines had significantly lower basal H3K27me3 levels than CM cell lines, suggesting an additional, histone methyltransferase-independent function of EZH2 in ALM. Conclusions: Our data demonstrate thatEZH2 upregulation is common in ALM, and suggest that it may play a role in ALM's metastatic progression that requires further investigation. Selective sensitivity of ALM cell lines to EZH2 inhibitors supports the therapeutic potential of EZH2-targeted therapy in ALM

Association of aberrant glycosylation with melanoma progression is based mainly on analyses of cell lines. Here we present a systems-based study of glycomic changes and corresponding enzymes associated with melanoma metastasis in patient samples. Upregulation of core fucosylation (FUT8) and downregulation of alpha-1,2 fucosylation (FUT1, FUT2) were identified as features of metastatic melanoma. Using both in vitro and in vivo studies, we demonstrate FUT8 is a driver of melanoma metastasis which, when silenced, suppresses invasion and tumor dissemination. Glycoprotein targets of FUT8 were enriched in cell migration proteins including the adhesion molecule L1CAM. Core fucosylation impacted L1CAM cleavage and the ability of L1CAM to support melanoma invasion. FUT8 and its targets represent therapeutic targets in melanoma metastasis.

Patients with multifocal breast cancers (MBCs) have a poorer prognosis than patients with unifocal breast cancers. Studies have attributed this to tumor size underestimation in MBC. An alternative hypothesis is that some MBCs behave in a fashion analogous to the "satellite" and "in-transit metastasis" observed in melanoma and, thereby, are more clinically aggressive. We identified 79 cases of MBC, which we classified into 2 groups: study cases defined as >/=2 morphologically similar tumor foci with >/=1 focus without in situ carcinoma (n = 21); and a control group defined as >/=2 morphologically similar or dissimilar foci with associated in situ carcinoma in all foci (n = 58). The odds of being a study case is 1.86 (95% confidence interval [CI] 1.26-2.74) times greater per unit increase in number of tumor foci (median of 4 tumor foci; P = .002). Study cases were 73.33 (95% CI = 8.91-603.16) times more likely to have lymphovascular invasion (LVI) and 14.72 (95% CI = 4.37-49.61) times more likely to have nodal metastases. Grade I/II tumors were 0.20 (95% CI = 0.07-0.59) times less likely to be study cases. There was a significant positive interaction (P < 0.001) indicated by the relationship of LVI status and nodal status with the study case and control group. We conclude that there is a subset of MBC that presents with more numerous tumor foci and a higher rate of nodal metastasis. The aggressive behavior of these cases may be attributed to their proclivity for LVI.