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Objectives Treatment options for malignant pleural mesothelioma (MPM) are limited but some studies on immune checkpoint inhibitors (ICIs) in MPM have reported antitumor activity. Very little is known about immune-related predictive factors. Materials and methods Here we report the case of a 45-year-old woman presenting with dyspnea and evidence of pleural effusion. She was diagnosed with malignant epithelioid pleural mesothelioma with brain metastasis and peritoneal carcinosis, refractory to initial standard chemotherapy treatment. Because of high PDL1 expression (100 %), she was treated with the anti-PD1 agent, pembrolizumab. Results Chemotherapy with cisplatin and pemetrexed was started, imaging studies showing stable disease after 3 treatment cycles. The patient underwent pleural decortication but rapidly progressed and thus started chemotherapy with carboplatin and gemcitabine. After 2 cycles she experienced seizures caused by a brain metastasis. This secondary lesion was surgically removed and confirmed as a metastasis from mesothelioma. Samples from both the primary tumor and the metastasis were molecularly characterized, the pleural sample proving ALK-positive and the brain sample, ALK-negative. PD-L1 was positive in 10 % of tumor cells in the pleural biopsy and 100 % in the brain lesion. Next generation sequencing analysis was negative for both samples. It was decided to start alectinib. Disease progression (peritoneal carcinosis and liver metastases) was documented after one month followed by complete bowel obstruction and recurrence in the site of the brain surgery. Alectinib was stopped and supportive care begun with parenteral nutrition via nasogastric tube. Pembrolizumab was started and after 15 days the patient’s condition had significantly improved, enabling recanalization and restoration of enteral nutrition. Imaging displayed complete response of the brain metastasis, peritoneal carcinosis, bone lesions and mediastinal nodal metastases. A partial response was documented in the pleural and pulmonary nodules, with stable liver metastases. The patient is still undergoing immunotherapy and has no cancer-related symptoms. Conclusions Our findings indicate that the use of immunotherapy in MPM warrants further investigation. Furthermore, the impressive clinical response obtained by our patient suggests that immune checkpoint inhibitors could help in the management of the disease after the failure of other treatments.

Non-small-cell lung cancer (NSCLC) is the primary cause of cancer-related death. Gene rearrangements involving the anaplastic lymphoma kinase (ALK) tyrosine kinase identify a clinical and molecular subset of NSCLC patients, who benefit from the monotherapy with ALK tyrosine kinase inhibitors. Nonetheless, responsiveness to TKIs and prognosis of these patients are influenced by several factors, including resistance mechanisms and mutations affecting genes involved in key molecular pathways of cancer cells. In a cohort of 98 NSCLC patients with ALK gene rearrangements, we investigated the role of Tumor Protein (TP53) gene mutations in predicting patients prognosis. TP53 mutations were evaluated in relation to disease control rate (DCR), objective response rate (ORR), progression-free survival (PFS) and overall survival (OS).Results: In patients with available clinical and TP53 mutation information, we found that 13 patients (20.3%) were affected by TP53 mutations. Considered together, even though showing a trend, TP53 mutations were not associated with PFS and OS. Considering the different TP53 mutations by functionality in terms of disruptive and non-disruptive mutations, we observed that TP53 non-disruptive mutations were able to predict worse OS in the overall case series. Moreover, a worse PFS was seen in the subgroup of patients with TP53 non-disruptive mutation, in first-, second-, and third line of treatment. Our results show that mutations affecting TP53 gene, especially non-disruptive mutations, are able to affect prognosis of ALK-rearranged NSCLC patients.

The microbiota is considered a component of the tumor microenvironment (TME) but its functional role is still not completely clear. In this study, we use both meta-proteomic/transcriptomic and metabarcoding approaches to investigate the composition and the role of microbiota-derived proteins in breast cancer (BC) and microbial diversity, respectively. Three tumoral regions (peripheral tumor, tumor core, in situ carcinoma) and 2 healthy regions (intra-tumoral fibrosis, healthy tissue) were selected on 12 formalin-fixed, paraffin-embedded (FFPE) BC samples. Ten µm tissue slides were used for DNA/RNA extraction and proteomics. The proteomic analysis was performed by nanoLC-MS/MS analysis and data analysis by Perseus and Maxquant, transcriptomic by NEBNext Ultra II Library Prep kit for Illumina and analyzed by Kraken2. Metabarcoding was performed by Ion 16S metagenomics kit for Ion s5 and analyzed by extracting the reads covering the V4 hypervariable region, denoising them with DADA2 and finally taxonomically annotating the obtained ASVs in BioMaS. The proteomic analysis of the 5 areas identified 29 bacterial proteins: 13 chaperonins, 7 ATP synthase subunits, 7 metabolic enzymes, 2 elongation factors. Most of the proteins (16) were located in the intra-tumoral fibrosis. A Glyceraldehyde-3-phosphate dehydrogenase (gapA) and three Chaperone proteins DnaK were overexpressed in the tumor compared to the healthy tissue. Only one of the 4 proteins (DnaK ID:B8IHL3) was associated with a single species: Methylobacterium nodulans. In order to narrow the number of possible species to which the other 3 proteins belong, we have compared the proteomic results with the taxonomic results obtained by metatranscriptomic. The remaining two DnaKs possibly belong to Methylobacterium sp4-46 and Clavibacter michiganensis, while the gapA to Escherichia coli. Overall the investigation based on the V4 hypervariable region of the 16S rRNA gene allowed to highlight a higher alpha diversity in tumor samples than in the healthy ones. In conclusion, the meta-proteomic investigation highlighted the overexpression in tumor tissues of DnaKs, bacterial chaperones able to interfere with important pathways related to DNA damage control/repair and cell-cycle/apoptosis and supposed to promote cancer, and gapA, which plays an important role in multiple cellular functions including metabolism and gene transcription. The most recurrent genera is Methylobacterium which had previously been found as more abundant in BC tissues than healthy tissues. We hypothesize that bacteria supply the tumor cells with proteins that potentially promote cancer progression, and that Methylobacterium could have a key role in breast cancer, but more studies are needed to confirm the hypothesis. Citation Format: Francesca Pirini, Tania Rossi, Soulaimane Aboulouard, Bruno Fosso, Maurizio Puccetti, Sara Ravaioli, Maria Maddalena Tumedei, Michela Cortesi, Michele Zanoni, Luca Magnani, Giovanni Martinelli, Michel Salzet, Sara Bravaccini. A multi-omics approach to study the host-microbiota interaction in breast cancer tissue [abstract]. In: Proceedings of the 2021 San Antonio Breast Cancer Symposium; 2021 Dec 7-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2022;82(4 Suppl):Abstract nr P5-06-04.

BackgroundCurrent therapy for non-small-cell lung cancer (NSCLC) frequently includes immune checkpoint inhibitors, such as pembrolizumab, and programmed death ligand 1 (PD-L1) positivity is mandatory for its use in this setting. Vimentin plays a role in carcinogenesis through the activation of the epithelial-to-mesenchymal transition (EMT) process. Its prognostic impact in NSCLC has been investigated in numerous studies but little data are available on its relation with PD-L1 expression.Patients and MethodsWe retrospectively retrieved data on patients with advanced NSCLC consecutively enrolled in a clinical trial at our institute. PD-L1 and vimentin expression were determined by immunohistochemistry. Correlations between variables were assessed using the Spearman correlation coefficient. The Kaplan-Meier method was used to estimate overall survival (OS) and the Log-rank test was used to compare survival curves. The association between demographic, clinical and biomarker information and survival was investigated with the Cox model.ResultsFifty-three patients were included in the study. A weak positive correlation was observed between the PD-L1 and vimentin (ρ=0.41, P=0.003). Patients with PD-L1 values P=0.080). No difference in overall survival (OS) was observed on the basis of vimentin expression (HR=1.25; 95% CI: 0.59–2.66; P=0.554). Patients harboring both vimentin and PD-L1 negative expression (P=0.093). No significant associations were observed between gender, age at diagnosis, stage at diagnosis, histology, KRAS or EGFR status, radical surgery or immunotherapy and OS.ConclusionsThe weak positive association between PD-L1 and vimentin suggests a potential interplay between these biomarkers. Further research is warranted to evaluate EMT and immune escape as two components of the same process.

Angiosarcoma of the thyroid is a rare and aggressive primary malignant tumour of the thyroid. We report the case of a 69-year-old woman who presented with a red and sore skin area at the right-anterior region of the neck. Ultrasound examination and computed tomography scan showed a non-homogeneous mass in the right thyroid lobe. Fine needle aspiration cytology was suggestive of atypical vascular proliferation and so the patient underwent right thyroid lobectomy. The specimen measured 6 × 5 × 2.5 cm, and a reddish nodule was found, including a whitish central area of maximum 4 cm in diameter. Immunohistochemistry showed CD31 and ERG positivity, while thyroglobulin, calcitonin and TTF-1 expression were negative, indicating a diagnosis of angiosarcoma.

Liquid biopsy analytes such as cell-free DNA (cfDNA) and circulating tumor cells (CTCs) exhibit great potential for personalized treatment. Since cfDNA and CTCs are considered to give additive information and blood specimens are limited, isolation of cfDNA and CTC in an "all from one tube" format is desired. We investigated whether cfDNA variant sequencing from CTC-depleted blood (CTC-depl. B; obtained after positive immunomagnetic isolation of CTCs (AdnaTest EMT-2/Stem Cell Select, QIAGEN)) impacts the results compared to cfDNA variant sequencing from matched whole blood (WB). Cell-free DNA was isolated using matched WB and CTC-depl. B from 17 hormone receptor positive/human epidermal growth factor receptor 2 negative (HR+/HER2-) metastatic breast cancer patients (QIAamp MinElute ccfDNA Kit, QIAGEN). Cell-free DNA libraries were constructed (customized QIAseq Targeted DNA Panel for Illumina, QIAGEN) with integrated unique molecular indices. Sequencing (on the NextSeq 550 platform, Illumina) and data analysis (Ingenuity Variant Analysis) were performed. RNA expression in CTCs was analyzed by multimarker quantitative PCR. Cell-free DNA concentration and size distribution in the matched plasma samples were not significantly different. Seventy percent of all variants were identical in matched WB and CTC-depl. B, but 115/125 variants were exclusively found in WB/CTC-depl. B. The number of detected variants per patient and the number of exclusively detected variants per patient in only one cfDNA source did not differ between the two matched cfDNA sources. Even the characteristics of the exclusively detected cfDNA variants in either WB or CTC-depl. B were comparable. Thus, cfDNA variants from matched WB and CTC-depl. B exhibited no relevant differences, and parallel isolation of cfDNA and CTCs from only 10 mL of blood in an "all from one tube" format was feasible. Matched cfDNA mutational and CTC transcriptional analyses might empower a comprehensive liquid biopsy analysis to enhance the identification of actionable targets for individual therapy strategies.

Aims The diagnosis of breast cancer (BC) is based on clinical examination in combination with imaging, and confirmed by pathological assessment of core needle biopsy or fine-needle aspiration biopsy (FNAB). The biological profile of the lesion is needed to define the prognosis and establish therapy. Given the importance of an early and minimally invasive diagnosis, we aimed to verify whether the biological features detected in FNAB-derived cytological material reflect the biological characteristics of surgical samples. Methods and results We used immunohistochemistry and fluorescence in-situ hybridisation to study a panel of conventional biomarkers [oestrogen receptor (ER), progesterone receptor (PgR), Ki67, and human epidermal growth factor receptor 2 (HER2)] in FNAB-derived cytological samples included in cell blocks of 93 BC patients, and compared the results with those obtained from histological evaluation of the same parameters in surgical samples. Median immunopositive values of ER, PgR and Ki67 were similar in cell blocks and surgical samples. The concordance rates of ER and PgR between FNAB-derived cell blocks and histological samples were 98% and 84%, respectively. The concordance rates of Ki67 and HER2 between the two sample types were 90% and 96%, respectively. Tumour subtype classification for triple-negative and HER2-positive BCs in FNAB-derived cell blocks was always concordant with the subtype determined in surgical material. Conclusions We demonstrated that biological marker determination in FNAB-derived cell blocks is feasible, and provides useful information and comparable results to those obtained with histological evaluation. Given the low cost of the procedure and its minimal impact on patients, we believe that cytological samples could be used as an alternative to tissue samples for early BC biomarker evaluation.

Background: Molecular diagnostics for non-small cell lung cancer (NSCLC) has become the standard of care for personalized treatment. Epidermal growth factor receptor ( EGFR ) mutation and EML4-ALK translocation represent the two most important alterations in first-line treatment decision-making. However, other potentially targetable alterations are also present. Methods: One thousand consecutive NSCLC patients with EGFR wild type (wt) tumors diagnosed by routine molecular analysis were considered. KRAS, BRAF, ERBB2, PIK3CA, NRAS, ALK, MAP2K1, RET and DDR2 gene mutations were analyzed using the multiparametric Sequenom MassARRAY ® platform. EML4-ALK and ROS1 rearrangements were also assessed by fluorescent in situ hybridization. HER4 status was determined by direct sequencing. Results: Three hundred and forty-eight (34.8%), 31 (3.1%), 39 (4.4%), 14 (1.8%), 6 (0.7%), 16 (1.8%), 5 (0.6%) and 9 (0.9%) patients showed an alteration in KRAS, BRAF, ALK, ROS1, NRAS, PIK3CA, MAPK1/2 and HER2 genes, respectively. Of the 657 patients for whom all markers were determined, 318 (48%) patients had at least one alteration. Eight patients showed overlapping mutations, 4 KRAS mutation/ EML4-ALK translocation, one KRAS mutation/ ROS1 rearrangement, 2 KRAS/PIK3CA mutations, and one BRAF/PIK3CA mutations. Conclusions: About 50% of our patients had a potentially targetable alteration, confirming the usefulness of a multiparametric approach for routine molecular diagnostics aimed at identifying potential therapeutic targets.

Lynch syndrome (LS) is associated with germline mutations in one of the mismatch repair genes or EPCAM. The majority of the causative alterations are point mutations. Large genomic rearrangements represent only 5–20%. Hypothetically, the allelic imbalance, like the loss of heterozygosity, may be another high penetrance risk factor. We describe the case of a patient who developed 5 tumors during her lifetime and with a family history characterized by a high frequency of tumors associated with LS. The proband was tested for mutations and copy number alterations with a panel of hereditary cancer genes and by SNP array. She showed a 187 Kb duplication including EPCAM and the first 7 exons of MSH2, plus two loss of heterozygosity (LOHs) in chromosome 20 and one in chromosome X which include many tumor suppressor genes. We found a novel large EPCAM-MSH2 duplication associated with LS and the presence of LOHs in regions containing numerous tumor suppressors, raising the hypothesis that these alterations could contribute to cancer susceptibility. Our results underline the importance to deepen the knowledge of molecular mechanisms in order to determine the role in cancer predisposition of novel genetic alterations.

Background: In the Italian Breast Cancer Intergroup Studies (IBIS) 3 phase III trial, we compared cyclophosphamide, methotrexate, 5-fluorouracil (CMF) alone to sequential epirubicin/CMF regimens in patients with rapidly proliferating early breast cancer (RPEBC). We performed a post hoc analysis in the subgroup of patients with hormone-receptor-positive RPEBC on the prognostic role of progesterone receptor (PgR) status. Methods: RPEBC was defined by thymidine labeling index (TLI) >3% or grade 3 or S-phase >10% or Ki67 >20%. We analyzed 466 patients with hormone-receptor-positive RPEBC receiving sequential epirubicin/CMF regimens followed by tamoxifen, and for whom the status of ER and PgR was available. Results: Considering both cut-off values of 10% and 20%, PgR expression was significantly associated with age, menopausal status, and ER expression; HER2 status was associated with PgR status only at a cutoff value of 20% PgR. Upon univariate analysis, tumor size, nodal status, and PgR were significantly associated with disease-free survival (DFS) and overall survival (OS), while age class and local treatment type were associated only with DFS. Patients with PgR

Tumor-infiltrating macrophages (TIMs) often outnumber other immune cells and across different tumor histologies have been linked to resistance to therapies, including immunotherapy. Despite the huge steps headed in the characterization of these cells, their phenotyping and quantification within the tumor micro-environment (TME) still remain challenging. Multiplex Immunohistochemistry/Immunofluorescence (mIHC/IF) technologies, which allow the simultaneous detection of multiple markers on a single tissue section have empowered the approach to study the TME, allowing the analysis of co-expressing molecules and the spatial relationship between distinct immune and non-immune cells. However, this technique well performed for lymphoid cells and nuclear markers, needs to be optimized for TIMs, often characterized by an elongated shape and an enlarged cytoplasm. mIHC by iterative immunostaining on a single paraffin-embedded (FFPE) tissue slide followed by whole slide digital pathology analysis was performed. Different TIM-related antibodies were used, including: CD68, CD206, CD163, CD14, MS4A4A, HLA-DR. Moreover, an array of checkpoint molecules (B7-H3, VISTA, TIM-3, PD-L1), an inflammation-related marker (STING), a pan-leukocyte marker (CD45) and lymphoid related markers (CD56, CD8, CD3, CD4) have been also validated. Together with the staining procedure, we developed computational pipelines for the processing of the whole-slide images. To perform a precise ascription of the staining intensity, we used a customized CellProfiler pipeline for segmenting single cells after using the Color Deconvolution ImageJ plugin for the generation of combined pseudo-fluorescent images. This segmentation pipeline permits the conversion of microscopy data into CSV files reporting the single-cell feature values (e.g. staining intensity and reciprocal position). With this approach IHC images can be visualized in user-friendly multi-parameter cytometric-like outputs allowing a comprehensive in situ TIM quantification and phenotyping. Nonetheless, our approach does not require specialized equipment and makes usage of commonly reagents, allowing its easy translation also in real-life clinical practice for the tailoring of therapeutic choices. In conclusion, in this study we discuss critical aspects and computational tools for data visualization and analysis of mIHC TIM data together with results obtained in different clinical setting (melanoma, breast cancer and lymphoma). We believe that this approach can be useful for a deeper characterization of the TME. Citation Format: Maria Maddalena Tumedei, Filippo Piccinini, Jenny Bulgarelli, Massimo Guidoboni, Francesco Limarzi, Barbara Vergani, Biagio Eugenio Leone, Maurizio Puccetti, Sara Bravaccini, Giovanni Martinelli, Toni Ibrahim, Marcella Tazzari. Sequential immunohistochemistry and computational image analysis for a deeper characterization of tumor-infiltrating myeloid cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 2776.

The aim of our study was to assess the quality of Tanzanian cervical cancer specimens, evaluating telomerase alterations and human papilloma virus (HPV) infection in relation to histopathological characteristics since these biomarkers are not routinely analyzed. Thirty-two Tanzanian women with invasive cervical cancer were included in the study. Histopathological classification and all the analyses on tissue, including TERT immunohistochemistry, were performed at IRST IRCCS (Meldola, Italy). HPV typization was performed by pyrosequencing. FHACT™ was used to identify chromosomal aberrations. Nonparametric ranking statistics were used. The majority (75 %) of the cases analyzed were squamous carcinoma, while 12.5 % were adenocarcinoma. The presence of HPV infection was confirmed in 26/27 (96.3 %) cases. A high percentage of patients (88 %) were infected with HPV16 of whom 12 (44.4 %) with African type 1, and 4 (14.8 %) with African type 2. TERT expression evaluated in the entire case series showed a median H-score of 130 (range 3−270), with only one negative case. 88 % of the FISH-evaluable samples showed an amplification of the chromosomal region 3q26 (TERC) and/or 5p15, and 20q13, associated with a higher median expression of TERT (P = 0.0226). Despite pre-analytical problems in terms of sample fixation, we showed that the search for biomarkers such as HPV and telomerase is feasible in Tanzanian tissue. These markers could be important risk-stratification tools in this population.

Objectives Amplification of human telomerase is known to be associated with cervical tumorigenesis, although its role in tumor progression of cervical lesions is still unclear. We aimed to evaluate the role of telomerase in predicting the evolution of cervical lesions. Methods A total of 50 tissue samples taken by biopsy or conization once or repeatedly from 17 patients with cervical lesions over a 14-year follow-up was analyzed using fluorescence in situ hybridization (FISH) for hTERC gene alterations and immunohistochemistry (IHC) for hTERT expression. The accuracy of the biomarkers was measured using the area under the curve. Results Telomerase gene amplification is highly indicative of cervical lesion evolution and seems to be a more reliable biomarker than the protein expression detected by IHC. In fact, patients with benign lesions or cervical intraepithelial lesions (CINs) showing hTERC amplification relapsed or progressed into CIN 2 and CIN 3 more frequently than those without any gene amplification. FISH and IHC assays had both 86% sensitivity on conized material and 78% and 40% specificity, respectively. Conclusions We demonstrated that the most accurate method to evaluate telomerase alterations as prognostic markers in cervical lesions was FISH assay on hTERC gene. The best accuracy was obtained using conized materials.

In lung cancer patients, the only available diagnostic material often comes from biopsy or from cytological samples obtained by fine needle aspiration (FNA). There is a lack of easily detectable cytomorphological features for rapid on-site evaluation (ROSE) to orient lung cancer diagnosis towards a specific tumor histotype. We studied the cytological features evaluated on site to define tumor histotype and to establish the number of specimens to be taken. Cytological specimens from 273 consecutive patients were analyzed with ROSE: bronchoscopy with transbronchial needle aspiration (TBNA) had been performed in 72 patients and with endobronchial ultrasound (EBUS)-TBNA in 201. Cytomorphological features were correlated with the final diagnosis and diagnostic accuracy was measured. Analysis of the different cytomorphological parameters showed that the best sensitivity and specificity were obtained for adenocarcinoma by combining the presence of nucleoli and small/medium cell clusters, and for squamous cell carcinoma by considering the presence of necrosis ≥50% and large cell clusters. For small cell carcinoma, the best diagnostic accuracy was obtained by combining moderate necrosis (

Follicular lymphoma (FL) still remains an incurable disease, with most of the patients undergoing subsequent phases of remission and relapse. In the era of immunotherapy, understanding the immunobiology of FL patients who experience progression of disease within 24 months (POD24) and show chemotherapy resistance remains a priority and an unmet clinical need. Tumor-associated macrophages (TAM) are multifaceted cellular components of the tumor microenvironment (TME). In concert with lymphoid-lineage B and T cells at various developmental stages, TAM can mediate enhanced tumor progression, often leading to poor clinical prognosis and impact the clinical response to chemotherapy. Recently, Joshua and colleagues demonstrated that a subset of FL patients with low immune infiltration was enriched in POD24 events. In our work, we characterized the immune repertoire of lymph node biopsies collected retrospectively from 30 patients with histological diagnosis of FL according WHO criteria. Median follow up was 8 years. The expression levels of TAM (CD68, CD163, MS4a4a) and tumor-infiltrating T-lymphocytes (CD8, PD-1 positive subsets) were assessed by immunohistochemistry and summarized using descriptive statistics. Our data highlighted that the subset of patients with a high extrafollicular CD163/CD8 ratio was enriched in POD24 events (p = 0.01). In addition, patients who showed an higher number of intrafollicular CD68+ macrophages, showed a longer disease free survival (p= 0.04). Another difference was related to the number of CD68, CD163, MS4A4A-positive polarized macrophages that resulted higher in bcl-2 negative than bcl-2 positive cases (p< 0.05 for all markers). In the evaluation of T lymphocytes, most relevant associations were found between the content and distribution of PD1+ cells and the treatment response: the higher the number of intrafollicular PD1+ lymphocytes the lower chemotherapy response rates (p= 0.04). No association was found between the number of positive elements in extrafollicular areas, where instead the PD1/CD8 ratio seems to be related with therapy response: the higher the ratio the lower response rate (p= 0.01). This finding could be explained assuming that there may be different types of cellular interactions inside and outside the neoplastic follicles. No other statistically significant difference in terms of expression of these markers was observed in relation to clinical pathological features such as staging, grade and FLIPI score. In conclusion, extrafollicular high CD163/CD8 ratio is associated with POD24 in FL patients, this finding underlines the pathological significance of CD163-expresing macrophages in TME, suggesting this biomarker as a potential therapeutic target in this disease. Citation Format: Clara Bertuzzi, Maria Maddalena Tumedei, Sara Ravaioli, Claudio Agostinelli, Maurizio Puccetti, Sara Bravaccini, Simona Righi, Beatrice Casadei, Andrea Pession, Giovanni Martinelli, Giorgia Simonetti, Elena Sabattini, Serena De Matteis. Extrafollicular high CD163/CD8 ratio is associated with progression of disease within 24 months in follicular lymphoma patients [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 2682.

Molecular diagnostics for non-small cell lung cancer (NSCLC) has become the standard of care for personalized treatment. Epidermal growth factor receptor (One thousand consecutive NSCLC patients withThree hundred and forty-eight (34.8%), 31 (3.1%), 39 (4.4%), 14 (1.8%), 6 (0.7%), 16 (1.8%), 5 (0.6%) and 9 (0.9%) patients showed an alteration inAbout 50% of our patients had a potentially targetable alteration, confirming the usefulness of a multiparametric approach for routine molecular diagnostics aimed at identifying potential therapeutic targets.

The incidence of breast cancer (BC) in sub-Saharan Africa is projected to double between 2016 and 2035. The control of the disease in the region will become an increasingly critical challenge.1 Comprehensive prevention, detection and treatment programmes are lacking; the health infrastructure is insufficient; the technical workforce is limited, and the diagnostic and treatment capacity is poor.2 Over the last 20 years, a pool of Italian cancer research centres, cancer volunteer associations and scientific societies in collaboration with the Tanzanian political and health authorities have created a pathology laboratory (that they considered to be a core element of a cancer care and control programme) and a medical oncology unit at the Bugando Medical Centre (BMC) of Mwanza, the largest referral hospital in the Lake zone of north-western Tanzania. This endeavour is known as the Mwanza Cancer Project.3 Based on the idea that offering scientific tutoring and access to research programmes can increase the capacity building of local healthcare professionals, these have been involved in several research projects under way in Italy. One of these has been designed to perform a comparative biological characterisation of BC from a series of Tanzanian patients and a series of Italian patients.4 For the former, the pathological processing and reporting of surgical specimens have been performed at the pathology laboratory of the BMC, while oestrogen receptor (ER), progesterone receptor (PR) and …

Ductal carcinoma in situ (DCIS) is a highly heterogenous tumor that is now more frequently diagnosed because of the increased number of screening programs. Women with DCIS are mainly treated with conservative surgery almost always followed by radiotherapy. Although conventional biomarkers, i.e. ER, PgR, Ki67, and HER2, have been extensively investigated in invasive tumors, little is known about their role in DCIS, especially that of the androgen receptor (AR). In the present study, the expression of conventional biomarkers and AR was determined by immunohistochemistry in 85 DCIS samples from patients monitored for a maximum of 13 years: 43 patients were treated with quadrantectomy and 42 patients underwent quadrantectomy and radiotherapy. Of these, 5 and 11 patients relapsed, respectively. Our findings showed that ER and PgR were higher in nonrelapsed than in relapsed patients (P=0.025 and 0.0038). In contrast, AR expression and the AR/ER ratio were higher in relapsed patients than in the nonrelapsed group (P=0.0069 and 0.0012). At the best cut-off value of 1.1, the AR/ER ratio showed an overall accuracy of 92% and 80% in predicting in situ relapse or progression to invasive carcinoma in DCIS patients treated with surgery and those treated with surgery plus radiotherapy, respectively. AR would therefore appear to be an independent prognostic biomarker in the latter group. Our preliminary results highlight the potentially important role of the AR/ER ratio as a predictive indicator of DCIS relapse, independently of treatment.

Background: Immunohistochemistry (IHC) is the method conventionally used in clinical practice to define breast cancer (BC) subtypes. This technique is subjective and semi-quantitative. Molecular approaches like multigene panels are increasingly used in clinical practice for their reliability and accuracy. This preliminary study was performed to assess the applicability of the NanoString Breast Cancer 360™ panel, which analyze 770 transcripts of genes involved in a number of signatures useful to define the molecular subtype. We evaluated the correlation between the gene expression levels with the IHC values of conventional biomarkers on 12 BC formalin-fixed paraffin-embedded (FFPE) samples. Methods: Immunostaining was performed by using Ventana Benchmark Ultra system. Anti-ER, -PR, -Ki67 and -HER2 Ventana antibodies were used. Tumors were classified according to the most recent St. Gallen classification. RNA was isolated from FFPE tumor with AllPrep DNA/RNA FFPE Kit and quantified by Nanodrop. The Breast Cancer 360™ panel assay was performed according to manufacturer’s instructions. The normalized count for each gene analyzed using nSolver analysis software was compared with the respective IHC biomarker status by Mann-Whitney test to assess the correlation between the two methods. The mean mRNA count was compared between luminal and triple negative BC by unpaired T test. All statistical analyses were performed by Graphpad Prism software 5. Results: The 12 BC samples were classified in 3 luminal A, 3 luminal B (HER2+) and 6 triple negative tumors by IHC. The expression level of ESR1, PGR, MKI67 and ERBB2 genes showed a statistically significant correlation with the corresponding protein expression (p=0.0022, p=0.0054, p=0.0025 and p=0.0091, respectively). A significant difference in the expression levels of ERBB2 gene between HER2+ and HER2- BC was observed (mean mRNA counts 16848 vs 1307, p=0,035). Furthermore, ESR1 expression was significantly higher in ER+ than in ER- BC (mean mRNA counts 9105 vs 195, p=0,021). We also evaluated the differential expression between luminal and triple negative BC for AR, EGFR, HIF1A, MKI67, MYC, NOTCH1, RAD51 genes. The expression levels for all these genes were significantly higher in triple negative cases than luminal BC, except for AR that was higher in luminal tumors (p Conclusions: In this study Breast Cancer 360™ panel showed high concordance with IHC data. We are aware that our preliminary results require validation in a larger case series to establish the real value of this multi-gene expression profile in BC. However, our results highlight the potential of this molecular approach to properly identify tumor subtypes, allowing a better management of BC patients. Citation Format: Sara Ravaioli, Francesca Pirini, Andrea Rocca, Maurizio Puccetti, Massimiliano Bonafè, Giovanni Martinelli, Sara Bravaccini. Breast cancer subtype classification using a multi-gene expression profile [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 5255.

e20688 Background: The immune modulation is emerging as a key strategy in non-small cell lung cancer (NSCLC) patients management. PD-L1 has been the only biomarker validated for immunotherapy but it holds many limitations. Vimentin is one of the marker for epithelial-to-mesenchymal transition (EMT), a biological process which favors immune escape. We explored two issues: the correlation between PD-L1 and Vimentin expression in NSCLC samples and the prognostic value of PD-L1 and vimentin concordance in NSCLC patients. Methods: We collected all the NSCLC samples evaluable for both PD-L1 and Vimentin and recorded the relative available clinical and survival data, if written informed consent was present. PD-L1 and Vimentin were retrospectively assessed through immunohistochemistry (IHC) by using prediluted antibodies clones SP263 (Ventana Medical Systems, Tucson, AZ, USA) and anti Vimentin V9 (Ventana Medical Systems) on Benchmark XT Platform (Ventana Medical Systems). Biomarker expression was detected and semiquantitatively quantified as the percentage of immunopositive tumor cells on the total of tumor cells. We correlated PD-L1 and Vimentin expression through linear regression. We compared survival of patients with both PD-L1 and Vimentin 0% and all the others via Kaplan-Meier method by using MedCalc software version 18.11.3. Results: Ninety-nine samples underwent PD-L1 and Vimentin IHC analysis. A weak positive statistically significant correlation was found between the 2 biomarkers (r = 0.41; p < 0.001). For 40 patients survival data (from first NSCLC diagnosis) were available. Survival analysis showed that PD-L1 and Vimentin negative patients (n = 12) experienced not statistically significant longer survival (HR = 0.35; 95%CI: 0.08-1.51; p = 0.16). Conclusions: These preliminary findings suggest that an interplay can exist between PD-L1 and vimentin in NSCLC, but analysis in a wider population is necessary. Studies on the significance of this interplay for immunotherapy efficacy should be designed.

545 Background: The prognostic role of progesterone receptor (PgR) in highly proliferating early breast cancer (BC) is not well established. We retrospectively explored this biomarker in a cohort of patients with highly proliferating tumors enrolled in a phase III trial of adjuvant therapy. Methods: 1066 patients with N- or 1-3 N+ BC were randomized to receive: epirubicin followed by CMF, CMF followed by epirubicin, or CMF alone. Rapidly proliferating tumors were defined by thymidine labeling index (TLI) > 3% or histological grade 3 or S-phase > 10% or Ki67 > 20%. We analyze the subgroup of 466 patients with hormone receptor (HR)-positive tumors treated with sequential epirubicin/CMF regimens followed by tamoxifen and for whom immunohistochemical assessments of estrogen receptor (ER), PgR, HER2 and Ki67 were available. Disease-free (DFS) and overall survival (OS) curves were built with the Kaplan–Meier estimator and compared by logrank test and Cox regression models. Results: PgR expression was significantly associated with ER expression, HER2 status, age and menopausal status, but not with Ki67, tumor size and nodal status. PgR cutoff values of 10% and 20% were chosen based on a Receiver Operating Characteristics analysis and the literature data. DFS and OS figures at 5 and 10 years, as well as the relative hazard ratios, according to the different PgR cutoff values, are reported in the table. Conclusions: Our results confirm the prognostic relevance of PgR expression in a cohort of patients with highly proliferating, HR-positive early BC treated with adjuvant chemotherapy and endocrine therapy. [Table: see text]

Background The diagnosis of breast cancer (BC) is based on clinical examination in combination with imaging and confirmed by pathological assessment of core needle biopsy (CNB) or fine needle aspiration (FNA). The biological profile of the lesion is needed to define prognosis and guide therapy. Given the importance of an early and minimally invasive diagnosis, we aimed to verify whether the biological features detected in FNA-derived cytological material reflect the biological characteristics of surgical specimens. Methods We used immunohistochemistry and fluorescence in situ hybridization (FISH) to study a panel of biomarkers (ER, PgR, Ki67 and HER2 in 93 patients, programmed death-ligand 1 (PD-L1) in 20 patients) in FNA-derived cell blocks of BC, comparing the results with those obtained on the histological specimens.Immunostaining was performed with the Ventana Benchmark XT system and the Ultraview DAB Detection Kit (Ventana Medical Systems). Confirm anti-ER (clone SP1, Ventana), Confirm Anti-PgR (clone 1E2, Ventana) and Ki67 (clone Mib-1, Dako, Carpinteria, CA, US) antibodies were used. Ventana PD-L1 (SP263) assay (Ventana Medical Systems) was used for PD-L1 immunostaining. HER2 status was analyzed by FISH using PathVysion kit (Abbott Molecular, Abbott Park, Illinois, IL, USA). Results Median immunopositive values of ER, PgR Ki67, and PD-L1 were similar in cell blocks and surgical samples. Concordance for ER and PgR between FNA and histological samples was 98% and 84%, respectively. With regard to Ki67 and HER2 status, concordance between the two specimen types was 90% and 96%, respectively. PD-L1 expression analyzed in FNA-derived samples was 100% concordant with that of surgical specimens. Tumor subtype classification for triple-negative and HER2-positive tumors in FNA samples was always concordant with the subtype determined in surgical material. Table 1.Concordance of tumor subtype classification between FNAB-derived and histological samples.Overall series (n=93)87%Luminal A (n=26)81%Luminal B (n=47)86%HER2-positive (n=6)100%Triple-negative (n=14)100% Conclusions We showed that biological marker determination in FNA-derived cell blocks is feasible and provides useful information and comparable results with those obtained by histological evaluation. Given the low cost of the procedure and its minimal impact on patients, that cytological samples could be used as an alternative to tissue samples for early BC biomarker evaluation to facilitate the planning of tailored neoadjuvant therapy. Citation Format: Bravaccini S, Ravaioli S, Rocca A, Tumedei MM, Bucchi E, Malmesi M, Medri L, Bonafè M, Puccetti M. Are fine-needle aspiration (FNA)-derived cell blocks a useful tissue sample surrogate for testing conventional biomarkers and PD-L1 in breast cancer? [abstract]. In: Proceedings of the 2018 San Antonio Breast Cancer Symposium; 2018 Dec 4-8; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2019;79(4 Suppl):Abstract nr P4-02-10.

Objectives To report the results of a pilot study for a service for cervical cancer screening and diagnosis in north-western Tanzania. Methods The pilot study was launched in 2012 after a community-level information campaign. Women aged 15–64 years were encouraged to attend the district health centres. Attendees were offered a conventional Pap smear and a visual inspection of the cervix with acetic acid (VIA). Results The first 2500 women were evaluated. A total of 164 women (detection rate 70.0/1000) were diagnosed with high-grade cervical intraepithelial neoplasia and invasive cervical cancer. The performance of VIA was comparable to that of Pap smear. The district of residence, a history of untreated sexually transmitted disease, an HIV-negative status (inverse association), and parity were independently associated with the detected prevalence of disease. The probability of invasive versus preinvasive disease was lower in HIV-positive women and in women practicing breast self-examination. Conclusions The diagnostic procedure had an acceptable level of quality. Factors associated with the detected prevalence of disease will allow for a more targeted promotion of the service. Cervical screening should be coordinated with sexually transmitted disease and HIV infection control activities.

Background Epidermal growth factor receptor ( EGFR ) and v-Ki-ras2 Kirsten rat sarcoma viral oncogene homolog ( KRAS) mutations, and echinoderm microtubule-associated protein-like 4 ( EML4 ) anaplastic lymphoma kinase ( ALK ) translocation are generally considered to be mutually exclusive. However, concomitant mutations are found in a small number of patients and the effect of these on response to targeted therapy is still unknown. Patients and Methods We considered 380 non–small-cell lung cancer (NSCLC) patients who underwent nonsequential testing for EGFR and EML4-ALK translocation. KRAS mutation analysis was also performed on 282 patients. Results We found 1.6%, 1.1%, and 2.5% of patients who showed a double mutation comprising EGFR and EML4-ALK , EGFR and KRAS , and EML4-ALK and KRAS , respectively. Twenty-eight patients with EGFR mutation underwent first-line therapy with a tyrosine kinase receptor; a clinical benefit was observed in 81.8% of patients with EGFR mutations only and in 67% of those who also showed an EML4-ALK translocation. Twelve patients with an EML4-ALK translocation received crizotinib and 7 of these had disease progression within 3 months (2 had a concomitant KRAS mutation and 1 had a concomitant EGFR mutation). Two patients showed stable disease, 1 of whom also had a KRAS mutation. Two patients obtained a partial response and 1 had a complete response; all harbored an EML4-ALK translocation only. The median overall survival of patients who carried an EML4-ALK translocation alone or concomitant with a KRAS mutation was 57.1 (range, 10.7-not reached) and 10.7 (range, 4.6-not reached) months, respectively. Conclusion Concomitant EGFR, EML4-ALK , or KRAS mutations can occur in NSCLC. Concomitant KRAS mutation and EML4-ALK translocation represents the most common double alteration and confers a poor prognosis.

Background Ductal carcinoma in situ (DCIS) is a heterogeneous disease that has not been investigated as widely as invasive breast cancer. Thus, the search for biomarkers capable of identifying DCIS lesions that may recur or progress to invasive cancer is ongoing. Although conventional steroid hormone receptors, cell proliferation and other important tumor markers have been extensively studied in invasive tumors, little is known about the role played by androgen receptors (ARs), widely expressed in breast cancer, in DCIS. Methods We performed a retrospective study in a series of 43 DCIS patients treated with quadrantectomy only and followed up for a period ranging from 5 to 13 years, to evaluate the prognostic relevance of conventional biomarkers (estrogen receptor [ER], progesterone receptor [PgR], Ki67, human epidermal growth factor receptor 2 [HER2]) and AR. Results Our findings showed that AR and ER were not independent prognostic variables and that an AR/ER ratio cutoff of 1.13 showed a sensitivity of 75% and a specificity of 94% in predicting in situ relapse or progression to the invasive phenotype. Moreover, while the variables considered singly showed area under the curve (AUC) values ranging from 0.52% to 0.77%, the AR/ER ratio reached a very high AUC (0.92%). Conclusions These preliminary results highlight the potentially important role of AR and ER and, in particular, of their ratio, as prognostic indicators of DCIS evolution.

To the Editors: Mutations in the epidermal growth factor receptor (EGFR) gene in patients with nonsmall cell lung cancer (NSCLC) have been correlated with tumour response to treatment with targeted tyrosine kinase inhibitors (TKIs), especially gefitinib and erlotinib [1, 2]. A large number of studies have reported a significantly higher overall response rate, overall survival and time-to-progression in patients with EGFR-activating mutations compared with those with wild-type tumours [3–5]. It has thus become mandatory to perform EGFR mutation analysis in all adenocarcinoma patients to improve treatment opportunities. In addition to EGFR alterations, K-ras mutations are found in ∼30% of adenocarcinomas [6] and each is usually mutually exclusive. About 70% of NSCLC patients present at first diagnosis with advanced disease, and in such cases, morphologic and molecular diagnoses are necessarily based on cytologic samples obtained by different methodological procedures [7]. It has been demonstrated that EGFR mutations can be detected in cytologic specimens containing >50% of neoplastic cells, and that analysis can be performed in samples with as little as 25% tumour cellularity [8]. Moreover, although the most reliable results have been obtained in samples from which ≥100 cells were analysed, it has also been noted that mutation analysis can be performed in as few as 30 isolated cells [9]. In the present study, 66 consecutive NSCLC patients diagnosed between …

1042 Background: The androgen receptor (AR) is widely expressed in breast cancers but its role in estrogen receptor (ER)-positive tumors is still controversial. However, the AR/ER ratio may impact prognosis and the response to antiestrogen endocrine therapy (ET). Methods: We assessed whether AR in primary tumors and/or matched metastases is a predictor of efficacy of first-line ET in advanced breast cancer (ABC). We evaluated patients treated with first-line ET (2002–2011), excluding those receiving concomitant chemotherapy or trastuzumab or pretreated with > 2 lines of chemotherapy. ER, progesterone receptor (PgR), Her2, Ki67 and AR expression was determined by immunohistochemistry. A cut-off of < 1% immunostained cells was used to categorize AR expression. AR expression was analyzed in relation to the other conventional biomarkers (ER, PgR, Her2 and Ki67), best response (CR, PR, SD, PD), and time to progression (TTP) (months). TTP was estimated using the Kaplan-Meier method and compared with the log-rank test. Hazard ratios and their 95% confidence intervals (95% CI) were estimated using the Cox regression model. The Chi-square test was used to evaluate correlations between categorical variables and best response. p values < 0.05 were considered statistically significant. Results: Of the 102 evaluable patients (93% were treated with an aromatase inhibitor), biomarkers were assessed in primary tumors in 70 cases, in metastases in 49 and in 17 in both). Median TTP was 17 months (95% CI 14-21.5, median follow-up 75 months). The overall concordance rate between primary tumors and metastases was 64.7% (95% CI 42%-87.4%) for AR expression. Differences in TTP according to AR status were not statistically significant. AR/PgR ≥ 0.96 was associated with a significantly shorter TTP (HR = 1.65, 95% CI 1.05-2.61, p = 0.030). AR status in primary tumors or metastases was not associated with PD as best response. In contrast, Ki67 > 20% and PgR < 10% showed a significant association with PD as best response. Using a cut off of ≤10% for AR expression, results did not change. Conclusions: AR expression does not appear to be useful to predict the efficacy of ET in ABC. Ki67 and PgR exert a greater impact on the efficacy of hormone therapy than AR.

S. Bravaccini, M. M. Tumedei, P. Ulivi, W. Zoli, D. Calistri, P. Candoli, D. Amadori and M. PuccettiALK translocation detection in non-small cell lung cancer cytological samples obtained by EBUS-TBNAObjective: As the diagnosis of non-small cell lung cancer (NSCLC) is based on cytology in around 70% ofcases, it is important to use the same material for molecular analyses. Fluorescence in situ hybridization (FISH)is the only approved test for the detection of the translocation and inversion of anaplastic lymphoma kinase(ALK), but the optimal procedures for the fixation or staining of the sample before FISH evaluation have notbeen established. We investigated whether ALK gene status determined by FISH in a prospectively enrolledcase series of patients was affected by fixation and staining.Methods: One hundred and fifteen cytological samples were obtained by transbronchial needle aspiration(TBNA) or endobronchial ultrasound (EBUS)-TBNA from 109 patients with NSCLC. All samples were evalu-ated for epidermal growth factor receptor (EGFR) mutation by pyrosequencing and for ALK rearrangement byFISH. Specimens for ALK determination had been fixed with Cytofix and/or Carnoy’s solution or 10% for-malin (cell blocks) and variously stained.Results: Sixteen (14%) of the 115 samples were mutated for EGFR and 99 (86%) showed wild-type EGFRstatus. Of these 115 samples, 79 (69%) were negative for echinoderm microtubule-associated protein like 4(EML4)-ALK translocation, nine (8%) were positive and 27 (23%) were unevaluable. In particular, 19 (26%)of the 72 Papanicolaou-stained smears fixed with Cytofix were unevaluable because of inadequate samples orcell overlapping; neither of the two May–Gr€unwald –Giemsa-stained samples were evaluable. Ten of 17 smearsused for rapid on-site evaluation (ROSE) and immediately post-fixed in Carnoy’s solution or 80% alcoholwere evaluable.Conclusions: In this series, smears were unevaluable as a result of inadequate samples, cell overlapping orlack of fixation performed immediately after FNA.Keywords: cytological samples, ALK translocation, FISH, EBUS-TBNA, cell fixation, non-small cell lungcancerIntroductionSeveral new targeted drugs have shown promisingresults for the treatment of non-small cell lung can-cer (NSCLC), and some have already been intro-duced into clinical practice. Crizotinib, an orallyactive, small-molecule inhibitor of anaplastic lym-phoma kinase (ALK) and MNNG HOS transforminggene (c-MET), has produced high overall survivalrates in patients with NSCLC whose tumours har-bour the echinoderm microtubule-associated proteinlike 4 (EML4)-ALK rearrangement, indicating theimportance of checking for the presence of thesealterations before making treatment decisions.

Background Although fine needle aspiration (FNA) is the standard diagnostic test for the characterization of a suspicious thyroid nodule, in some cases cytological evaluation is inconclusive. The aim of this study was to determine the role of BRAF mutation in aiding diagnosis and to verify whether archival cytological samples could be suitable for molecular analysis. Methods Eighty-five patients with suspicious (Thy4) or follicular (Thy3) lesions on cytology were resubmitted to a second FNA for BRAF mutation analysis. Of these, 56 subsequently underwent surgery. The usefulness of archival samples for molecular analysis was also studied in a second cohort of 42 patients with a confirmed diagnosis of papillary thyroid carcinoma for whom both archived paraffin-embedded histological samples and cytological smears were available. A further 15 patients with paired fresh FNA and archived cytological and histological samples were recruited. Results BRAF mutation was found in the fresh FNA samples from 10 of 56 patients who had surgery with previous inconclusive cytology (4/45, 9%, Thy3 and 6/11, 55%, Thy4). The BRAF test showed a specificity and positive predictive value of 100% (26/26 and 10/10, respectively), sensitivity of 33% (10/30) and negative predictive value of 57% (26/46). There was absolute concordance between the BRAF results obtained with 42 histological and cytological archived samples. BRAF analysis on 15 archived cytological samples showed absolute concordance with histology, whereas there was one false negative on the matched fresh FNA. Conclusion BRAF analysis is a highly specific test that can facilitate cytological diagnosis in some cases and can also be performed on archived cytological samples.

Information on hormone receptor and human epidermal growth factor receptor-2 (HER2) expression in breast cancer is acknowledged as mandatory for prognostic stratification and treatment planning. Data on the biological features of African breast cancers are poor. We decided to compare histopathological and biomolecular characteristics (estrogen and progesterone receptor—ER, PgR, and HER2) of Tanzanian and Italian breast cancers. Differences in proliferating index and androgen receptor (AR) expression in triple-negative patients from the two case series were also assessed. Of the 103 consecutive patients seen at the Bugando Medical Center (Mwanza, Tanzania) from 2003 to 2010, who underwent biopsy or surgical resection of primary breast cancer, 69 patients had tissue samples that were evaluable for estrogen receptor (ER), progesterone receptor (PgR), and HER2. Histopathological assessment and biomolecular determinations were performed at the Cancer Institute of Romagna (IRST IRCCS, Meldola, Italy). Caucasian breast cancers were randomly extracted from an electronic database and matched (1:2 ratio) for year of diagnosis and age at diagnosis. Median age of both populations was 51 years (range 27–84). With respect to Caucasian tumors, Tanzanian breast cancers at diagnosis more frequently showed high histological grade (mainly grade 3) (P = 0.03), advanced clinical stage (III or IV) (P\0.001), ER negativity (52.2 %, P\0.001) and high proliferation (P = 0.0002). Triple-negative tumors were over-represented in Tanzanian women. AR was positive in 38.5 and 38 %of triple-negative Tanzanian and Italian breast cancers, respectively. Our results show that histopathological and biomolecular characteristics in Tanzanian and Italian breast cancers differ substantially. The high frequency of poorly differentiated, ER-negative, highly proliferating tumors, together with advanced stage at presentation, could be considered as the main prognostic factors linked to the high mortality rates for breast cancer in the African population.

There are no validated predictors of benefit from anthracyclines. We compared cyclophosphamide, methotrexate, 5-fluorouracil (CMF), and epirubicin in different sequences with CMF alone in a phase III trial on operable breast cancers. Outcomes were analyzed in relation to tumor biological profiles to identify potential predictors of the efficacy of different treatments/drug combinations. Patients with N− or 1–3N+ tumors, were randomized to receive (a) epirubicin (4 cycles) followed by CMF (4 cycles); (b) CMF (4 cycles) followed by epirubicin (4 cycles), or (c) CMF (6 cycles) alone. Immunohistochemical assessments of estrogen (ER) and progesterone (PgR) receptors, HER2 and Ki67 were available for 705 patients (arm A/B/C: 276/269/160). Prognostic and predictive relevance was analyzed by log-rank tests and Cox models. Ki67 > 20 % and absent/low expression of ER and PgR were associated with worsen disease-free (DFS) and overall survival (OS). In patients with triple negative tumors (ER−, PgR−, HER2−), epirubicin-containing regimens yielded better DFS (HR 0.33, 95 % CI 0.17–0.62, P = 0.0007) and OS (HR 0.24, 95 % CI 0.10–0.57, P = 0.001) compared with CMF alone, whereas no differences were found in patients with HER2-positive (HER2+, ER−, PgR−) subtype. Treatment by subtype interaction (HER2-positive vs. others) was significant for DFS (χ2 = 6.72, P = 0.009). In triple unfavorable (ER−, PgR−, Ki67 > 20 %) tumors, the use of epirubicin yielded better DFS (HR 0.45,95 % CI 0.26–0.78, P = 0.005) and OS (HR 0.30, 95 % CI 0.15–0.63, P = 0.001). Epirubicin-containing regimens seem to be superior to CMF alone in patients with highly proliferating, triple negative or triple unfavorable tumors .

Introduction Tyrosine kinase inhibitors (TKIs) and anti-anaplastic lymphoma kinase (ALK) agents are highly effective for the treatment of non-small cell lung cancer (NSCLC) patients harbouring specific alterations, and molecular characterisation of the tumour is needed even when limited tumour material is available. Methods 20 patients with a known epidermal growth factor receptor ( EGFR ) gene status were enrolled: 10 had mutated and 10 had wild type tumours. FISH analysis was performed on one cytological or histological sample to determine EML4-ALK status, after which the same cells scraped off each slide were used to evaluate the EGFR status. Results In the 10 EGFR mutated patients, molecular analysis showed the same results as those obtained before the FISH test. One patient with an EGFR mutation also showed an EML4-ALK translocation, and both FISH-positive and FISH-negative cells maintained the EGFR mutation. Conclusions EGFR mutation analysis can be performed on the same sample previously submitted to the EML4-ALK FISH procedure.

Introduction: After an enthusiastic and promising start to the large scale use of screening programs for the detection of small in situ breast cancers, an analysis of results from different screening studies has led to disappointing and somewhat alarming conclusions. A problem of over diagnosis has emerged, involving, in many cases, unnecessary treatments, and causing a negative psychological and social impact on patients and a considerable economic burden on Health Services. Ductal carcinoma in situ (DCIS) is a heterogeneous disease that has not been investigated as widely as invasive breast cancer. Today women with in situ breast cancer are mainly treated with conservative surgery and often with radiotherapy, even though the real impact of this treatment is still not known. New diagnostic and therapeutic information is thus needed. Although conventional steroid hormone receptors, cell proliferation and other important tumor markers have been extensively studied in invasive tumors, little is known about the role played by androgen receptors (ARs), widely expressed in DCIS. We previously demonstrated in a series of DCIS patients treated with surgery alone that the AR/estrogen receptor (ER) ratio is an important prognostic marker. In the present study, we analyzed these markers in a series of DCIS patients treated with surgery and radiotherapy. Methods: We performed a retrospective study on a series of 42 DCIS patients treated with quadrantectomy and radiotherapy and followed up for a period ranging from 5 to 13 years to evaluate the prognostic relevance of ER and ARs determined by immunohistochemistry. Results: Our findings showed that AR expression was significantly higher in relapsed patients than in non relapsed patients, and that ER levels were higher in patients who did not relapse with respect to those who did. The AR/ER ratio was significantly different in the two subgroups. Moreover, when the variables were considered singly, area under the curve (AUC) values were 0.85 for ARs, 0.62 for ER and 0.79 for the AR/ER ratio. Conclusions: In agreement with our previous work on a series of DCIS patients treated with surgery alone, the preliminary results from the present study highlight the important role of AR and of the AR/ER ratio as prognostic indicators of relapse in patients treated with surgery and radiotherapy. Our findings also suggest that the treatment used was not capable of controlling disease or of eliminating the risk of recurrence. New avenues for tailored therapy must be sought. Citation Format: Sara Bravaccini, Sara Ravaioli, Maria Maddalena Tumedei, Rosella Silvestrini, Flavia Foca, Ilaria Massa, Roberta Maltoni, Andrea Rocca, Secondo Folli, Federico Buggi, Annalisa Curcio, Maurizio Puccetti, Luigi Serra, Elisabetta Pietri, Daniele Andreis, Alberto Farolfi, Dino Amadori. Clinical relevance of androgen and estrogen receptors in patients with ductal carcinoma in situ of the breast treated with surgery and radiotherapy. [abstract]. In: Proceedings of the AACR Special Conference on Advances in Breast Cancer Research; Oct 17-20, 2015; Bellevue, WA. Philadelphia (PA): AACR; Mol Cancer Res 2016;14(2_Suppl):Abstract nr B44.

This is a report of an angiolipoma of the thyroid gland, an extremely rare entity. A thorough search of the literature revealed only one previously reported example. The patient was a 77-year-old woman with a history of a nodular lesion of the thyroid in the context of a multinodular goiter. A fine needle aspiration highlighted the presence of abundant adipocytes associated with numerous dilated vessels. Histopathologic examination of the lobectomy specimen documented the presence of a tumor composed of two main elements, namely, mature adipocytes and proliferating vessels, some of the latter containing fibrin thrombi.

e22227 Background: Ductal carcinoma in situ (DCIS) is a heterogeneous disease that has not been investigated as widely as invasive breast cancer. Thus, the search for biomarkers capable of identify...

e19086 Background: Tyrosine kinase inhibitors (TKIs) are the first choice of treatment in a subset of patients with non- small cell lung cancer (NSCLC) harboring specific epidermal growth factor re...

5594 Background: In Sub-Saharan Africa cervical cancer represents 24% of all cancers and accounts for 23% of all cancer deaths in women. An early diagnosis program for breast and cervical cancer (Vanda Project) is ongoing in Mwanza and the surrounding lake area (12 districts with a population of 14,000,000). The aim of this project was to screen women aged 15-64 years living in the 12 districts. Methods: Women were invited to participate through local media and a mobile unit operating within the districts. A multidisciplinary team including medical oncologists was involved. Interventions consisted in Pap smear, clinical breast examination, breast self-examination training and training of district physicians to perform Pap smear and breast examination. Results: From May to December 2012, 2155 women from the districts of Shinyanga, Bukumbi, Kibara, Serengema and Musoma took part in the program: of these 91 (4%) had clinically evident cervical cancer. Age distribution classes were: < 18 years, 12% ; 18-35, 38%; 36-50, 41%; > 50, 9%. As expected a high stage distribution at diagnosis was observed: 30% stage III and 20% stage IV. Among the women with no clinical evidence of cancer, 408 samples were analyzed by cytology and 4% consisted of inadequate material. Of the remaining 392 samples, 85 (22%) were normal, 216 (55%) were infections (chiefly mycotic), 72 (18%) were precancerous lesions (50% H-SIL according to Bethesda classification) and 19 (5%) were positive for cancer (mainly stages III-IV). Precancerous lesions turned out to be cancer at histology in 44% of cases. 22% of precancerous lesions and 8% of clinically evident cancer were HIV-positive. Conclusions: This experience shows the high feasibility, good compliance and usefulness of a screening program for the early detection of cervical cancer in this high-risk population. [Table: see text]