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The Notch signaling pathway, particularly endothelial Notch signaling, plays a key role in tumor formation and angiogenesis. Inhibiting Notch1 or its ligand, Delta-like ligand 4 (Dll4), can induce hypersprouting of the tumor vasculature and thus can reduce tumor perfusion and progression. However, due to the expression of Notch1 in multiple healthy cell types, pharmacologic blockade poses the challenge of inducing tissue hypoxia, vascular neoplasms, and toxicity in non-target tissues. Unlike Notch1, Notch4 expression is largely restricted to endothelium and is upregulated in models of triple negative breast cancer (TNBC). Loss of Notch4 in mice results in delayed developmental angiogenesis and reduced mammary tumor angiogenesis and perfusion. Therefore, Notch4 is an attractive potential pharmacologic target to interfere with tumorigenesis and minimize nonspecific toxicity. We determined that Notch4 is expressed primarily in tumor endothelium in both human and mouse mammary tumors, including murine syngeneic Py8119 tumors, as well as a selection of other human and syngeneic mouse tumor types such as B16F10 melanoma tumors. We explored the effects of Notch4 inhibition on the progression of murine mammary epithelial carcinoma using newly developed anti-Notch4 antibodies, 6-3-A6 and its humanized derivative E7011. Administration of E7011 or 6-3-A6 to orthotopically implanted Py8119 murine breast carcinoma and B16F10 melanoma significantly reduced tumor size in comparison to IgG-treated controls. We performed single cell RNA sequencing of Py8119 tumors treated with E7011 and found that although Notch4 transcripts were found specifically in the endothelium, the most notable effect of E7011 was a dramatic increase in the number of macrophages and cancer-associated fibroblasts, suggesting a non-cell autonomous effect. To examine the potential role of these immune cell population changes in facilitating E7011’s anti-tumor activity, we investigated treatment of Py8119 murine mammary carcinomas in the absence of macrophages. Administration of clodronate liposomes to eliminate macrophages blocked the anti-tumor effects of E7011 treatment. Taken together, our data suggests that tumor endothelial Notch4 promotes TNBC growth by altering the immune landscape and increasing anti-tumor macrophage recruitment, and that blockade via the novel anti-Notch4 E7011 antibody has potential therapeutic efficacy. Citation Format: Jason W. Eng, L.A. Naiche, Bhairavi Swaminathan, Yu Kato, Krishna Thakkar, Katherine Ann Alexander, Rahul Vadakath, James H. Herts, Rajyasree Emmadi, Junji Matsui, Jan K. Kitajewski. Notch4 inhibition by a novel neutralizing antibody reduces tumor progression and increases macrophage recruitment within the tumor microenvironment [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 2 (Clinical Trials and Late-Breaking Research); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(8_Suppl):Abstract nr LB009.

Objectives Insulin-like growth factor-methotrexate (IGF-MTX) is a conjugate of methotrexate and 765IGF, a variant of IGF-1 with high affinity for insulin-like growth factor type 1 receptor. The study aim was to determine the maximum tolerated dose of IGF-MTX in refractory solid organ and hematologic malignancies expressing insulin-like growth factor type 1 receptor. Materials and methods This phase I trial used a modified toxicity probability interval design with 5 cohort dose levels, and expansion cohort at maximum tolerated dose. IGF-MTX was given intravenously over 90 minutes on days 1, 8, and 15 of a 28-day cycle. Results A total of 17 patients were enrolled. The highest tolerated dose tested was 0.80 µEq/kg with dose-limiting toxicity of grade 3 hypoglycemia. Drug-related grade 3 and 4 toxicities included abdominal pain (26%), hypoglycemia (10%), and hypotension (10%). Of the 15 evaluable for response, 3 patients (20%) had stable disease, including the patient with Hodgkin lymphoma with stable disease for 12 cycles of therapy. IGF-MTX concentrations declined rapidly, with half-lives of 5.2 to 14 minutes for the initial distribution phase and 6.5 to 7.5 hours for the terminal elimination phase. Higher IGF-R1 expression did not correlate with better outcome. Conclusions IGF-MTX is well tolerated. IGF-MTX pharmacokinetics suggest rapid cellular uptake. The activity of IGF-MTX in Hodgkin lymphoma should be explored.

Many colorectal cancers (CRCs) that exhibit microsatellite instability (MSI) are not explained by MLH1 promoter methylation or germline mutations in mismatch repair (MMR) genes, which cause Lynch syndrome (LS). Instead, these Lynch-like syndrome (LLS) patients have somatic mutations in MMR genes. However, many of these patients are young and have relatives with cancer, suggesting a hereditary entity. We performed germline sequence analysis in LLS patients and determined their tumor's mutational profiles using FFPE DNA. Six hundred fifty-four consecutive CRC patients were screened for suspected Lynch syndrome using MSI and absence of MLH1 methylation. Suspected LS cases were exome sequenced to identify germline and somatic mutations. Single nucleotide variants were used to characterize mutational signatures. We identified 23 suspected LS cases. Germline sequence analysis of 16 available samples identified 5 cases with LS mutations and 11 cases without LS mutations, LLS. Most LLS tumors had a combination of somatic MMR gene mutation and loss of heterozygosity. LLS patients were relatively young and had excess first-degree relatives with cancer. Four of the 11 LLS patients had rare likely pathogenic variants in genes that maintain genome integrity. Moreover, tumors from this group had a distinct mutational signature compared to tumors from LLS patients lacking germline mutations in these genes. In summary, more than a third of the LLS patients studied had germline mutations in genes that maintain genome integrity and their tumors had a distinct mutational signature. The possibility of hereditary factors in LLS warrants further studies so counseling can be properly informed.

Endothelial Notch signaling is critical for tumor angiogenesis and growth. Inhibition of Notch1 or its ligand, Dll4, can induce tumor vessel hypersprouting, which interferes with perfusion and reduces tumor development. Unfortunately, blocking Notch1 in endothelial cells of normal tissue can lead to tissue hypoxia and vascular neoplasms. Notch4, another member of the Notch family, is upregulated and increased in the endothelium of certain tumor types compared to normal vasculature. Reducing Notch4, unlike Notch1, causes reduction in developmental and tumor angiogenesis. Using a newly developed anti-Notch4 antibody, E7011, we assessed the effects on Notch4 inhibition on developmental retinal angiogenesis. E7011 treatment did not alter capillary density within the vascular plexus or endothelial tip cell number, but did decrease radial outgrowth and vein diameter, supporting the hypothesis that Notch4 functions as a pro-angiogenic signal. These findings suggest that the targeting of Notch4 could interfere with tumor angiogenesis and offer a novel therapeutic avenue. To establish the role of Notch4 within the tumor microenvironment, we analyzed its expression in both human and murine tumor samples. Immunofluorescent staining revealed a range of Notch4 expression in both tumor cells and endothelium in human triple negative breast cancers. Several murine syngeneic tumors examined expressed Notch4 primarily in the endothelium with only rare tumor expression. We treated orthotopically implanted Py8119 murine breast carcinomas and B16F10 melanomas with E7011, which significantly reduced tumor size compared to IgG treated controls, indicating that suppression of Notch4 is anti-tumorigenic. To determine mechanistically how E7011 delays tumor growth, we performed single cell RNA sequencing on E7011 and control treated Py8119 tumors. Surprisingly, we found that while Notch4 transcripts were present only in endothelial cells, E7011 administration dramatically altered macrophage and monocyte populations within the tumor. Taken together, these findings indicate that inhibition of tumor endothelial Notch4 by E7011 can regulate the presence of immune cells within the tumor microenvironment and improve tumor control. Citation Format: Jason Eng, L.A. Naiche, Bhairavi Swaminathan, Yu Kato, Krishna Thakkar, Rahul Vadakath, James Herts, Rajyasree Emmadi, Junji Matsui, Jan Kitajewski. Targeting Notch4 with a novel neutralizing antibody inhibits tumor growth and alters immune cell populations within the tumor microenvironment [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 247.

Renal cell neoplasia are heterogeneous with diverse histology, genetic alterations, and clinical behavior that are diagnosed mostly on morphologic features. The Renal Cell Neoplasia Workgroup of the Cancer Genomics Consortium systematically evaluated peer-reviewed literature on genomic studies of renal cell carcinoma (RCC), including clear cell RCC, papillary RCC, chromophobe RCC, and the translocation RCC involving TFE3, TFEB and MITF rearrangements, as well as benign oncocytoma, which together comprise about 95% of all renal cell neoplasia. The Workgroup curated recurrent copy number alterations (CNAs), copy-neutral loss-of-heterozygosity (cnLOH), rearrangements, and mutations, found in each subtype and assigned clinical relevance according to established criteria. In clear cell RCC, loss of 3p has a disease-initiating role and most likely also in progression with mutations detected in VHL and other genes mapped to this arm, and loss of 9p and/or 14q has well-substantiated prognostic utility. Gain of chromosomes 7 and 17 are hallmark CNAs of papillary RCC, but patterns of other CNAs as detected by chromosomal microarray analysis (CMA) afford sub-classification into Type 1 and 2 with prognostic value, and for further sub-stratification of Type 2. Inherent chromosome loss in chromophobe RCC as detected by CMA is useful for distinguishing the eosinophilic variant from benign oncocytoma which in contrast exhibits few CNAs or rearranged CCND1, but share mitochondrial DNA mutations. In morphologically atypical RCCs, rearrangement of TFE3 and TFEB should be considered in the differential diagnosis, portending an aggressive RCC subtype. Overall, this evidence-based review provides a validated role for assessment of CNAs in renal cell neoplasia in the clinical setting to assist in renal cell neoplasm diagnosis and sub-classification within subtypes that is integral to the management of patients, from small incidentally found renal masses to larger surgically resected specimens, and simultaneously identify the presence of key alterations portending outcome in malignant RCC subtypes.

Background Genomic abnormalities in breast cancer have been described according to diverse conceptual frameworks, including histologic subtypes, clinical molecular subtypes, intrinsic DNA, RNA, and epigenetic profiles, and activated molecular pathways. Methods The Cancer Genomics Consortium (CGC) Breast Cancer Workgroup performed an evidence based literature review to summarize current knowledge of clinically significant genomic alterations in breast cancer using CGC levels of evidence. Targetable or disease-defining alterations were prioritized. Results We summarized genomic alterations in breast cancer within a framework of existing clinical tools for diagnosis, risk stratification, and therapeutic management. Using CGC levels of evidence, we catalog copy number profiles, gene expression profiles, and mutations in clinically significant genes. We also describe emerging molecular markers such as methylation profiling and immunotherapy biomarkers. Conclusion A summary of currently available information on breast cancer genomics will enhance precision medicine by serving as an interpretive resource for clinical laboratory geneticists, providing a foundation for future practice guidelines, and identifying knowledge gaps to address in future research.

Endothelial Notch signaling is critical for tumor angiogenesis and growth. Inhibitors of Notch1 or its ligand Dll4 cause tumor vessel hypersprouting but interfere with vessel perfusion causing reduced tumor growth. Unfortunately, normal tissue vessels respond to these inhibitors causing tissue hypoxia or vascular neoplasms. Mice null for Notch4, unlike Notch1 mutants, exhibit reduced developmental and tumor angiogenesis. Notch4 expression is increased in the endothelium of some tumor types. These results suggest that specifically targeting Notch4 could reduce tumor angiogenesis with fewer effects on normal tissue and represents an unexplored therapeutic opportunity. We use a newly developed anti-Notch4 antibody, E7011, and show that in endothelial cells, inhibition of Notch4 signaling suppresses expression of a subset of canonical Notch target genes. Treatment with E7011 also suppresses expression of a unique set of genes not regulated by canonical Dll4-Notch1 signaling, which appear to be distinct Notch4 targets. We administered E7011 to neonatal mice to examine the effects on developmental retinal angiogenesis. A single dose of E7011 significantly reduced retinal vascular outgrowth, demonstrating that E7011 has anti-angiogenic activity in vivo. E7011 treatment did not alter capillary density within the vascular plexus or endothelial tip cell number, both of which are typically increased by loss of Notch1 signaling. Because the E7011 phenotype was markedly distinct from the Notch1 loss of function, we combined E7011 treatment with endothelial-specific loss of Notch1 (Notch1ECKO) to compare Notch1 to Notch4 function. E7011-treated Notch1ECKO retina showed a compound phenotype, exhibiting both the loss of vascular outgrowth characteristic of E7011/Notch4 and the hypersprouting characteristic of Notch1 loss of function. These results suggest that Notch1 and Notch4 have distinct roles in developmental angiogenesis, with Notch4 being a pro-angiogenic signaling pathway. We evaluated Notch4 expression in the breast cancer tumor microenvironment to determine if Notch4 functions in tumor vessels. Human triple negative breast cancer (TNBC) shows a range of expression of Notch4 in both tumor cells and tumor endothelium. The anti-angiogenic effects that we observe in the retina lead us to hypothesize that E7011 will show anti-tumor activity against tumors that induce Notch4 expression in neighboring endothelium, and that targeting endothelial Notch4 is a novel anti-angiogenic approach. Citation Format: L. A. Naiche, Bhairavi Swaminathan, Yu Kato, Sarita Das, Jason W. Eng, Qanber Raza, Krishna Thakkar, James H. Herts, Rajyasree Emmadi, Junji Matsui, Jan K. Kitajewski. A novel Notch4 neutralizing antibody inhibits angiogenesis and tumor growth via a distinct mechanism from endothelial Notch1 inhibition [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 1488.

African Americans (AAs) have higher incidence and mortality rates of colorectal cancer (CRC) compared with other US populations. They present with more right-sided, microsatellite stable disease and are diagnosed at earlier ages compared with non-Hispanic Whites (NHWs). To gain insight into these trends, we conducted exome sequencing (n = 45), copy number (n = 33) and methylation analysis (n = 11) of microsatellite stable AA CRCs. Results were compared with data from The Cancer Genome Atlas (TCGA). Two of the 45 tumors contained POLE mutations. In the remaining 43 tumors, only 27 (63%) contained loss-of-function mutations in APC compared with 80% of TCGA NHW CRCs. APC-mutation-negative CRCs were associated with an earlier onset of CRC (P = 0.01). They were also associated with lower overall mutation burden, fewer copy number variants and a DNA methylation signature that was distinct from the CpG island methylator phenotype characterized in microsatellite unstable disease. Three of the APC-mutation-negative CRCs had loss-of-function mutations in BCL9L. Mutations in driver genes identified by TCGA exome analysis were less frequent in AA CRC cases than TCGA NHWs. Genes that regulate the WNT signaling pathway, including SOX9, GATA6, TET1, GLIS1 and FAT1, were differentially hypermethylated in APC-mutation-negative CRCs, suggesting a novel mechanism for cancer development in these tumors. In summary, we have identified a subtype of CRC that is associated with younger age of diagnosis, lack of APC mutation, microsatellite and chromosome stability, lower mutation burden and distinctive methylation changes.

Background Activating mutations in the PIK3CA gene represent the most common molecular aberration in luminal breast cancers. Three hot-spot mutations (E542K, E545K, H1047R) account for approximately 70% of the PIK3CA mutations identified. The presence of a mutation is likely to become an important predictive biomarker to select patients for treatment with PIK3CA inhibitors. Tumor-derived DNA can be detected in the systemic circulation in the form of cell-free plasma DNA (cfpDNA), and the ability to interrogate tumor-derived DNA from an easily acquired biospecimen like cfpDNA has practical advantages over analysis of tissue specimens. Furthermore, cfpDNA can be monitored serially and could provide a dynamic measure of tumor progression and response to therapy. Droplet digital PCR (ddPCR) is a high-throughput technique that provides highly precise and sensitive detection of rare alleles in samples with ultra-low abundance of target DNA. Methods A nested case-control study was conducted with plasma samples collected as part of a study to identify blood-based biomarkers. Blood was collected from women scheduled for a breast biopsy at one of seven participating mammography units. All blood samples were collected prior to the biopsy procedure and were processed using a standard protocol. Cases are women diagnosed with invasive ductal carcinoma, stage I-III, that is positive for the estrogen receptor (ER+) and negative for HER2/neu (0 or 1+ by IHC). Controls are women with a benign, non-proliferative lesion, matched by collection site, year of collection and age. cfpDNA was isolated using a commercial kit (Macherey-Nagel, Bethlehem, PA). DNA was extracted from FFPE biopsy specimens by standard techniques. Central pathologic review confirmed at least 10% tumor cellularity in biopsy specimens from cases. A ddPCR assay was developed on a BioRad QX100 Droplet Digital PCR instrument using standard TaqManR assays with probe pairs designed to detect the 3 PIK3CA hotspot mutations and corresponding wild type (wt) alleles. The assay was validated using DNA from four cell lines, each positive for one of the hotspot mutations or wt at all 3 loci (Horizon Diagnostics, Cambridge, UK). ddPCR was performed on cfpDNA and DNA extracted from FFPE biopsy specimens in cases and controls. Next generation DNA sequencing (NGS) of exons 9 and 20 of PIK3CA was performed on biopsy specimens from cases using an Ion Torrent Personal Genome Sequencer. Results Validation of the ddPCR assay with serial dilutions of cell line DNA demonstrated the assay is highly sensitive with mutant alleles detected at an allele frequency down to 25% when input DNA template is as low as 2 ng. With DNA template input of 20 ng (typical recovery of cfpDNA), mutations can be detected down to an allele frequency of 0.78%. We will present data on the rate of hotspot mutation detection with the ddPCR assay performed on cfpDNA and FFPE samples in cases (n=80) and controls (n=80), and we will report performance characteristics of the assay. Mutation status of tumors determined by NGS will serve as the gold standard. Conclusion We have developed a ddPCR assay to detect PIK3CA hotspot mutations in cfpDNA, and will report the performance characteristics of the assay in 80 cases of untreated, ER+ early breast cancer. Citation Format: Kent F Hoskins, Amer Sidani, Tiffany Jones, Stefan Green, Rajyasree Emmadi, Anjen Chenn. Detection of PIK3CA mutations in cell-free plasma DNA from women with early breast cancer [abstract]. In: Proceedings of the Thirty-Seventh Annual CTRC-AACR San Antonio Breast Cancer Symposium: 2014 Dec 9-13; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2015;75(9 Suppl):Abstract nr P3-06-42.

Protein Tyrosine kinase 6 (PTK6/BRK) is overexpressed in the majority of human breast tumors and breast tumor cell lines. It is also expressed in normal epithelial linings of the gastrointestinal tract, skin, and prostate. To date, expression of PTK6 has not been extensively examined in the normal human mammary gland. We detected PTK6 mRNA and protein expression in the immortalized normal MCF-10A human mammary gland epithelial cell line, and examined PTK6 expression and activation in a normal human breast tissue microarray, as well as in human breast tumors. Phosphorylation of tyrosine residue 342 in the PTK6 activation loop corresponds with its activation. Similar to findings in the prostate, we detect nuclear and cytoplasmic PTK6 in normal mammary gland epithelial cells, but no phosphorylation of tyrosine residue 342. However, in human breast tumors, striking PTK6 expression and phosphorylation of tyrosine 342 is observed at the plasma membrane. PTK6 is expressed in the normal human mammary gland, but does not appear to be active and may have kinase-independent functions that are distinct from its cancer promoting activities at the membrane. Understanding consequences of PTK6 activation at the plasma membrane may have implications for developing novel targeted therapies against this kinase.

Purpose: African Americans (AA) have the highest incidence of colorectal cancer compared with other U.S. populations and more proximal colorectal cancers. The objective is to elucidate the basis of these cancer disparities. Experimental design: Of note, 566 AA and 328 non-Hispanic White (NHW) colorectal cancers were ascertained in five Chicago hospitals. Clinical and exposure data were collected. Microsatellite instability (MSI) and BRAF (V600E) and KRAS mutations were tested. Statistical significance of categorical variables was tested by the Fisher exact test or logistic regression and age by the Mann–Whitney U test. Results: Over a 10-year period, the median age at diagnosis significantly decreased for both AAs (68–61; P < 0.01) and NHWs (64.5– 62; P = 0.04); more AA patients were diagnosed before age 50 than NHWs (22% vs. 15%; P = 0.01). AAs had more proximal colorectal cancer than NHWs (49.5% vs. 33.7%; P < 0.01), but overall frequencies of MSI, BRAF and KRAS mutations were not different nor were they different by location in the colon. Proximal colorectal cancers often presented with lymphocytic infiltrate (P < 0.01) and were diagnosed at older ages (P = 0.02). Smoking, drinking, and obesity were less common in this group, but results were not statistically significant. Conclusions: Patients with colorectal cancer have gotten progressively younger. The excess of colorectal cancer in AAs predominantly consists of more proximal, microsatellite stable tumors, commonly presenting lymphocytic infiltrate and less often associated with toxic exposures or a higher BMI. Younger AAs had more distal colorectal cancers than older ones. These data suggest two different mechanisms driving younger age and proximal location of colorectal cancers in AAs. Clin Cancer Res; 20(18); 4962–70. ©2014 AACR.

Introduction: The purpose of this study was to create the first ever breast cancer patient-derived xenograft (PDX) model at the University of Illinois at Chicago (UIC) Cancer Center available to all breast cancer investigators. The intention is to provide opportunities to perform preclinical trials of lead cancer drugs as well as to conduct racial disparity studies. Methods: All patient tissues were collected from informed, consented patients with an IRB approved protocol (#14-1078) and with compliance of HIPAA law. None of the patients received neoadjuvant chemotherapy. Within 30 minutes of surgical excision, each tumor specimen was bilaterally implanted into the 2nd thoracic mammary fat pad of two NOD/SCID/IL2γ-receptor null (NSG) mice. Additional tissue was placed in RNAlater for gene expression analysis in successful engraftments. Transplantation sites were monitored daily for one week and measured weekly with calipers using the formula l x w2/2. Formalin-fixed and paraffin embedded sections (5 μm) were stained with hematoxylin and eosin (H&E) for histological examination. Immunohistochemical (IHC) staining was performed with ERα (SP1), CD20 (L26), and keratin cocktail (AE1/3) antibodies (Ventana). Results: Between April and August 2015, a total of eleven primary breast cancer PDXs were engrafted from eight African-American, two Caucasian and one patient of unknown race. The subtypes represented in these primary engraftments included one triple-negative breast cancer (TNBC), eight estrogen receptor alpha positive (ERα+) and two HER2 positive (HER2+) PDXs. Five primary PDXs engrafted successfully and were passaged to second generation mice including one TNBC and four ERα+ PDXs. After 12 weeks of primary engraftment, UIC-PDX#6 (ERα+) PDX grew at a much faster rate than any of the other PDXs and was passaged twice. Following the second passage, tumor tissue was processed for histological examination when it was discovered the PDX was not a tumor of epithelial origin, but was in fact a tumor of B-cell origin. This finding is in agreement with the recent report by Bondarenko et. al. (Neoplasia, 2015) that B-cell lymphomas frequently occur in PDXs engrafted into NSG mice. At the time of this writing, the remaining TNBC and ERα+ PDXs are growing at a slower pace in the second passage NSG mice and two additional primary PDXs are growing and nearing secondary passage. Conclusions: A breast cancer PDX effort was initiated at UIC from a racially diverse patient population. Despite engraftment of a B-cell tumor in one PDX, four promising stable breast cancer PDXs are currently being maintained. We have plans to test and validate novel endocrine therapies as well as other promising strategies to treat breast cancer. Citation Format: Huiping Zhao, Thao Pham, Rajyasree Emmadi, Elizabeth Wiley, Michael Warso, Alejandra Perez-Tamayo, George Salti, Kent Hoskins, Debra A. Tonetti. Breast cancer patient-derived xenografts: The University of Illinois at Chicago Cancer Center experience. [abstract]. In: Proceedings of the AACR Special Conference: Patient-Derived Cancer Models: Present and Future Applications from Basic Science to the Clinic; Feb 11-14, 2016; New Orleans, LA. Philadelphia (PA): AACR; Clin Cancer Res 2016;22(16_Suppl):Abstract nr B19.

TPS2635 Background: The insulin-like growth factor 1 receptor (IGF-1R) is overexpressed in multiple malignancies, including breast, prostate, lung, colorectal, CLL, T-ALL, and mantle cell lymphoma....

Breast cancer risk increases transiently in the period following pregnancy; pregnancy-associated breast cancers (PABC) are more aggressive than cases diagnosed in nulliparous women. We have previously reported that in the normal human breast pregnancy results in the upregulation of a number of inflammation related genes, suggesting a pro-tumorigenic environment as well as downregulation of ESR1 (ERα) and ERBB2 (HER2) and upregulation of ESR2 (ERβ), suggesting a protective effect. In this study, we aimed to investigate the possibility of differential regulation of the same gene set modulated in the normal breast, in human breast tumors following pregnancy. Gene expression was measured by real-time PCR on tumor regions isolated by laser capture microdissection from paraffin sections. Immunohistochemistry was performed on tissue microarrays (TMA) for protein expression. Hierarchical clustering was performed using the average linkage method to determine coordinate expression of sets of genes. We find that breast cancers detected within 10 years following pregnancy display a different gene expression pattern than those detected in nulliparous breast cancer patients. The gene expression difference is mainly attributable to a triple negative (TNBC) subgroup found to be more frequent in PABCs up to 10 years following a pregnancy. We also show that protein and mRNA expression levels correlate in half of the proteins tested by TMA. Despite the fact that this is a small study of 53 patients, we identified a gene expression signature that is differentially expressed in pregnancy-associated TNBC. Electronic supplementary material The online version of this article (doi:10.1186/s40064-015-1512-7) contains supplementary material, which is available to authorized users.