High levels of activated Stat3 are often found in human breast cancers and can correlate with poor patient outcome. We employed an activated ErbB2 mouse model of breast cancer to investigate the in vivo role of Stat3 in mammary tumor progression and found that Stat3 does not alter mammary tumor initiation but dramatically affects metastatic progression. Four-fold fewer animals exhibited lung metastases in the absence of Stat3 and a 12-fold reduction in the number of lung lesions was observed in animals bearing Stat3-null tumors when compared with the wild-type cohort. The decreased malignancy in Stat3-deficient tumors is attributed to a reduction in both angiogenic and inflammatory responses associated with a Stat3-dependent transcriptional cascade involving CCAAT/enhancer binding protein D. [Cancer Res 2009;69(17):6823-30]

Immunosurveillance constitutes the first step of cancer immunoediting in which developing malignant lesions are eliminated by anti-tumorigenic immune cells. However, the mechanisms by which neoplastic cells induce an immunosuppressive state to evade the immune response are still unclear. The transcription factor Stat3 has been implicated in breast carcinogenesis and tumor immunosuppression in advanced disease, but its involvement in early disease development has not been established. Here, we genetically ablated Stat3 in the tumor epithelia of the inducible PyVmT mammary tumor model and found that Stat3-deficient mice recapitulated the three phases of immunoediting: elimination, equilibrium, and escape. Pathological analyses revealed that Stat3-deficient mice initially formed hyperplastic and early adenoma-like lesions that later completely regressed, thereby preventing the emergence of mammary tumors in the majority of animals. Furthermore, tumor regression was correlated with massive immune infiltration into the Stat3-deficient lesions, leading to their elimination. In a minority of animals, focal, non-metastatic Stat3-deficient mammary tumors escaped immune surveillance after a long latency or equilibrium period. Taken together, our findings suggest that tumor epithelial expression of Stat3 plays a critical role in promoting an immunosuppressive tumor microenvironment during breast tumor initiation and progression, and prompt further investigation of Stat3 inhibitory strategies that may reactivate the immunosurveillance program.

Bcl3 is a putative proto-oncogene deregulated in hematopoietic and solid tumors. Studies in cell lines suggest that its oncogenic effects are mediated through the induction of proliferation and inhibition of cell death, yet its role in endogenous solid tumors has not been established. Here, we address the oncogenic effect of Bcl3 in vivo and describe how this Stat3-responsive oncogene promotes metastasis of ErbB2-positive mammary tumors without affecting primary tumor growth or normal mammary function. Deletion of the Bcl3 gene in ErbB2-positive (MMTV-Neu) mice resulted in a 75% reduction in metastatic tumor burden in the lungs with a 3.6-fold decrease in cell turnover index in these secondary lesions with no significant effect on primary mammary tumor growth, cyclin D1 levels, or caspase-3 activity. Direct inhibition of Bcl3 by siRNA in a transplantation model of an Erbb2-positive mammary tumor cell line confirmed the effect of Bcl3 in malignancy, suggesting that the effect of Bcl3 was intrinsic to the tumor cells. Bcl3 knockdown resulted in a 61% decrease in tumor cell motility and a concomitant increase in the cell migration inhibitors Nme1, Nme2, and Nme3, the GDP dissociation inhibitor Arhgdib, and the metalloprotease inhibitors Timp1 and Timp2. Independent knockdown of Nme1, Nme2, and Arhgdib partially rescued the Bcl3 motility phenotype. These results indicate for the first time a cell-autonomous disease-modifying role for Bcl3 in vivo, affecting metastatic disease progression rather than primary tumor growth. Cancer Res; 73(2); 745-55. Ó2012 AACR.

IntroductionEffective in vivo models of breast cancer are crucial for studying the development and progression of the disease in humans. We sought to engineer a novel mouse model of polyomavirus middle T antigen (PyV mT)-mediated mammary tumourigenesis in which inducible expression of this well-characterized viral oncoprotein is coupled to Cre recombinase (TetO-PyV mT-IRES-Cre recombinase or MIC).MethodsMIC mice were crossed to the mouse mammary tumour virus (MMTV)-reverse tetracycline transactivator (rtTA) strain to generate cohorts of virgin females carrying one or both transgenes. Experimental (rtTA/MIC) and control (rtTA or MIC) animals were administered 2 mg/mL doxycycline beginning as early as eight weeks of age and monitored for mammary tumour formation, in parallel with un-induced controls of the same genotypes.ResultsOf the rtTA/MIC virgin females studied, 90% developed mammary tumour with complete penetrance to all glands in response to doxycycline and a T50 of seven days post-induction, while induced or un-induced controls remained tumour-free after one year of induction. Histological analyses of rtTA/MIC mammary glands and tumour revealed that lesions followed the canonical stepwise progression of PyV mT tumourigenesis, from hyperplasia to mammary intraepithelial neoplasia/adenoma, carcinoma, and invasive carcinoma that metastasizes to the lung; at each of these stages expression of PyV mT and Cre recombinase transgenes was confirmed. Withdrawal of doxycycline from rtTA/MIC mice with end-stage mammary tumours led to rapid regression, yet animals eventually developed PyV mT-expressing and -non-expressing recurrent masses with varied tumour histopathologies.ConclusionsWe have successfully created a temporally regulated mouse model of PyV mT-mediated mammary tumourigenesis that can be used to study Cre recombinase-mediated genetic changes simultaneously. While maintaining all of the hallmark features of the well-established constitutive MMTV-PyV mT model, the utility of this strain derives from the linking of PyV mT and Cre recombinase transgenes; mammary epithelial cells are thereby forced to couple PyV mT expression with conditional ablation of a given gene. This transgenic mouse model will be an important research tool for identifying synthetic viable genetic events that enable PyV mT tumours to evolve in the absence of a key signaling pathway.

While silicone elastomers generally have excellent biomaterials properties, their hydrophobicity can elicit undesired local biological responses through adsorption and denaturation of proteins. Surface-bound poly(ethylene glycol) (PEG) can ameliorate the situation by preventing contact between the external biology and the silicone elastomer. It is further possible to manipulate the biocompatibility of the surface by linking peptides, proteins or other biological entities to the PEG. Previous synthetic approaches to PEG-protected surfaces are compromised by issues of reproducibility. We describe two rapid and efficient approaches to silicone surface modification by PEG-linked adhesion peptides that overcome this problem: SiH groups are introduced throughout a silicone elastomer during elastomer synthesis or only at the surface after cure; then, in either case, protein-repellent PEG brushes at the surface are introduced by hydrosilylation to give surfaces that can be stored for extensive periods of time without degradation. Activation of the free alcohol with an NSC group followed by immediate conjugation to relevant biological molecules occurs in high yields, as shown for RGDS and GYRGDS. High surface grafting density of the peptides was demonstrated using radiolabeling techniques. Biological activity was demonstrated by a 5-fold increase in cell adhesion on the peptide-modified surfaces when compared to unmodified PDMS control surfaces.

400 Background: Since 2011, options for treatment of metastatic pancreatic cancer (mPC) have improved with the use of nab-paclitaxel plus gemcitabine (n-PGEM) or FOLFIRINOX (FFX) as first line treatments (1LTx). In 2016, Nanoliposomal irinotecan plus 5FU (nal-IRI-5FU) demonstrated efficacy in 2LTx. Yet, optimal sequential Tx has not been established. Methods: We evaluated oncologist-selected Tx algorithms and resultant progression free & overall survival (PFS, OS) for pts with mPC from 2010 and 2018 at the Jewish General Hospital, Montreal, QC. This retrospective study included 203 pts with mPC (33 to 89 years, 54% male). Results: 1LTx included 66 pts on FFX, 60 pts on n-PGEM, and 66 pts on single-agent GEM. The remaining pts received Capecitabine (CAP) or another Tx (N = 11). Mean PFS in FFX, n-PGEM, and GEM groups was 5.07, 5.52, and 4.10 months, respectively (progression was 1˚ or 2˚ disease progression or a change of Tx due to adverse events or intolerance). Only the FFX and GEM groups were significant when compared (p = 0.049). Only 43.8% of pts (N = 89/203) advanced to 2LTx most receiving GEM (N = 27), n-PGEM (N = 21), or FFX (N = 11), PFS 3.87, 7.04, and 2.30 months, respectively. FFX and n-PGEM groups were significant when compared (p=0.011). CAP and nal-IRI-5FU were 2LTx options for 25.8% (N=23/89) and 7.9% (N = 7/89) of pts, respectively. For 30 pts in 3LTx, Txs included: nal-IRI-5FU (N = 8), clinical trials (CT) (N = 7), GEM (N = 5), FFX (N = 4), n-PGEM (N = 2), CAP (N = 2) and Irinotecan (IRI) (N = 2). Only 7 pts received 4LTx: GEM (N = 3), CAP (N = 2), CT (N = 1), and IRI (N = 1). Median OS from start of 1LTx for pts in FFX (N = 60), n-PGEM (N = 41), and GEM (N = 60) groups was 11.42, 9.50, and 6.23 months, respectively. (Excluding pts on ongoing tx and other censored data points). GEM tx was a significant prognostic factor for shorter OS, GEM verus FFX, HR 1.673 (1.165 to 2.402, p = 0.0053), GEM versus n-PGEM, HR 1.511 (1.012 to 2.258, p = 0.0437). No difference in survival was seen between FFX and n-PGEM groups, HR 0.903 (95% CI 0.605-1.349, p = 0.6196). Conclusions: Though FFX and n-PGEM are considered mainstays of 1LTx, GEM was chosen by physicians in ~1/3 of cases despite reduced PFS. Pts on FFX or n-PGEM had better OS compared to GEM alone, as expected. Further investigation into Tx sequencing in this and larger cohorts, is needed.

414 Background: The efficacy of FFX and GNP as 1LTx of mPC were established in phase 3 trials against Gemcitabine alone. However, no head‐to‐head trial has been performed. This analysis was conducted to compare the use of the two regimens in the 1LTx of mPC patients (pts). Methods: Retrospective study collected data of pts diagnosed with mPC (ECOG 0‐1) that received FFX or GNP as 1LTx at the Jewish General Hospital between 2010‐2016. Pt selection for 1LTx was based in ASCO Guidelines 2016 Criteria (AGC2016). Progression free survival (PFS) and overall survival (OS) were estimated using a Kaplan Meier method. Rate of 1LTx discontinuation and start of 2LTx was compared using two‐sided Fisher's exact test. Results: Among 75 pts with mPC (median age 69), 44 (59%) received FFX and 31 (41%) received GNP. In the FFX group 57% were male and 24 pts (55%) had primary tumors localized in the pancreatic head (PTPH). The majority of patients [n = 36 (82%)] had ECOG 1 at the start of FFX. The most common grade 3‐4 adverse events (AEs) were gastrointestinal symptoms (GI) [n = 12 (27%)], neutropenia (N) [n = 9 (20%)], fatigue (F) [n = 5 (11%)], and peripheral sensory neuropathy (PSN) [n = 2 (4%)]. In the GNP group 61% were male and 20 pts (65%) had PTPH. The majority of pts [n = 23 (74%)] had ECOG 1 at the start of GNP. The most common grade 3‐4 AEs were F [n = 8 (26%)], N [n = 4 (13%)], GI [n = 3 (10%)], and PSN [n = 2 (7%)]. Similar rates of 1LTx discontinuation due to AEs were seen in both groups: 5 pts (11%) in the FFX group due to GI, 2 pts (6.5%) in the GNP group due to F (p = 0.69). In the FFX cohort, 68.2% (30/44) went on to 2LTx whereas in the GNP cohort, 32% (10/31) received 2LTx (p = 0.0001). Of the FFX cohort receiving 2LTx, 40% (12/30) received GNP. The median PFS for the FFX and GNP groups were 5.75 and 4.63 months, respectively, and were not statistically significant (p = 0.523). The OS with FFX and GNP was 9.23 vs. 6.6 months (p = 0.09). Conclusions: For pts selected as per ASC2016, FFX and GNP cohorts showed similar PFS, OS, AEs, and 1LTx discontinuation rate. Our data highlight the importance of optimal therapeutic sequencing to prolong OS. A randomized trial will be needed to confirm 1LTx in mPC.

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