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The PI3K/Akt survival pathway is often dysregulated in cancer. Our previous studies have demonstrated that coexpression of activated Akt1 with activated ErbB2 or polyoma virus middle T antigen uncoupled from the PI3K pathway (PyVmT Y315/322F) accelerates mammary tumor development but cannot rescue the metastatic phenotype associated with these models. Here we report the generation of transgenic mice expressing activated Akt2 in the mammary epithelium. Like the MMTV-Akt1 strain, mammary-specific expression of Akt2 delayed mammary gland involution. However, in contrast to Akt1, coexpression of Akt2 with activated ErbB2 or PyVmT Y315/322F in the mammary glands of transgenic mice did not impact the latency of tumor development. Strikingly Akt2 coexpresssion markedly increased the incidence of pulmonary metastases in both tumor models demonstrating a unique role in tumor progression. Together these observations argue that these highly conserved kinases have distinct biological and biochemical outputs that play opposing roles in mammary tumor induction and metastasis.

The phosphatidylinositol 3′ kinase/Akt pathway is frequently dysregulated in cancer, which can have unfavorable consequences in terms of cell proliferation, survival, metabolism, and migration. Increasing evidence suggests that Akt1, Akt2, and Akt3 play unique roles in breast cancer initiation and progression. We have recently shown that in contrast to Akt1, which accelerates mammary tumor induction in transgenic mice, Akt2 promotes metastasis of tumor cells without affecting the latency of tumor development. Despite the distinct phenotypic outputs resulting from Akt1 or Akt2 activation, very little is known about the mode by which such unique functions originate from these highly related kinases. Here we discuss potential mechanisms contributing to the differing functional specificity of Akt1 and Akt2 with respect to migration, invasion, and metastasis.

The inducible prostaglandin synthase cyclooxygenase-2 (Cox-2) is overexpressed in f40% of human breast cancers and at higher frequencies in preinvasive ductal carcinoma in situ (DCIS). Cox-2 expression is particularly associated with overexpression of human epidermal growth factor receptor 2 (HER2/neu). To definitively interrogate the role of Cox-2 in mammary neoplasia, we have used a genetic approach, crossing Cox-2-deficient mice with a HER2/neu transgenic strain, MMTV/NDL. At 20 weeks of age, mammary glands from virgin MMTV/NDL females contained multiple focal tumors, or mammary intraepithelial neoplasias, which histologically resembled human DCIS. Mammary tumor multiplicity and prostaglandin E 2 (PGE 2 ) levels were significantly decreased in Cox-2 heterozygous and knockout animals relative to Cox-2 wild-type controls. Notably, the proportion of larger tumors was decreased in Cox-2-deficient mice. HER2/neu-induced mammary hyperplasia was also substantially reduced in Cox-2 null mice. Additionally, mammary glands from Cox-2 knockout mice exhibited a striking reduction in vascularization, and expression of proangiogenic genes was correspondingly reduced. Decreased vascularization was observed both in dysplastic and normal-appearing regions of Cox-2-null mammary glands. Our data provide the first genetic evidence that Cox-2 contributes to HER2/neu-induced mammary tumorigenesis. This finding may help to explain the reduced risk of breast cancer associated with regular use of nonsteroidal antiinflammatory drugs. (Cancer Res 2005; 65(21): 10113-9)

Amplification and elevated expression of the ErbB2 receptor tyrosine kinase occurs in 20% of human breast cancers and is associated with a poor prognosis. We have previously demonstrated that mammary tissuespecific expression of activated ErbB2 under the control of its endogenous promoter results in mammary tumor formation. Tumor development was associated with amplification and overexpression of ErbB2 at both the transcript and protein levels. Here we demonstrate that the EGR2/Krox20 transcription factor and its coactivator CITED1 are coordinately upregulated during ErbB2 tumor induction. We have identified an EGR2 binding site in the erbB2 promoter and demonstrated by chromatin immunoprecipitation assays that EGR2 and CITED1 associate specifically with this region of the promoter. EGR2 and CITED1 were shown to associate, and expression from an erbB2 promoter-reporter construct was stimulated by EGR2 and was further enhanced by CITED1 coexpression. Furthermore, expression of the 14-3-3 tumor suppressor led to downregulation of ErbB2 protein levels and relocalization of EGR2 from the nucleus to the cytoplasm. Taken together, these observations suggest that, in addition to an increased gene copy number and upregulation of EGR2 and CITED1, an elevated erbB2 transcript level involves the loss of 14-3-3, which sequesters a key transcriptional regulator of the erbB2 promoter.

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The development of antibody drug conjugates has provided enhanced potency to tumor-targeting antibodies by the addition of highly potent payloads. In the case of trastuzumab-DM1 (T-DM1), approved for the treatment of metastatic breast cancer, the addition of mertansine (DM1) to trastuzumab substantially increased progression-free survival. Despite these improvements, most patients eventually relapse due to complex mechanisms of resistance often associated with small molecule chemotherapeutics. Therefore, identifying payloads with different mechanisms of action (MOA) is critical for increasing the efficacy of targeted therapeutics and ultimately improving patient outcomes. To evaluate payloads with different MOA, deBouganin, a deimmunized plant toxin that inhibits protein synthesis, was conjugated to trastuzumab and compared with T-DM1 both in vitro and in vivo. The trastuzumab-deBouganin conjugate (T-deB) demonstrated greater potency in vitro against most cells lines with high levels of Her2 expression. In addition, T-deB, unlike T-DM1, was unaffected by inhibitors of multidrug resistance, Bcl-2-mediated resistance, or Her2-Her3 dimerization. Contrary to T-DM1 that showed only minimal cytotoxicity, T-deB was highly potent in vitro against tumor cells with cancer stem cell properties. Overall, the results demonstrate the potency and efficacy of deBouganin and emphasize the importance of using payloads with different MOAs. The data suggest that deBouganin could be a highly effective against tumor cell phenotypes not being addressed by current antibody drug conjugate formats and thereby provide prolonged clinical benefit.

INTRODUCTION AND OBJECTIVE:There is an increasing interest in finding a valid alternative for patients with non-muscle invasive bladder cancer (NMIBC). HIVEC-HR is a pilot trial that aims to compare efficacy and safety between BCG and chemohyperthermia (CHT) with mitomycin C (MMC). Here we present our preliminary results once randomization has been completed.METHODS: Open pilot randomized clinical trial 1:1 including patients with high-risk NMIBC according to EAU Guidelines (EudraCT 2016-001186-85). Patients with CIS, intolerance or contraindication for receiving BCG or MMC were excluded. Patients were randomly assigned to one of the following groups:-BCG (TICE strain): 50 mg diluted in 50 mL of sterile saline held for 2 hours in the bladder, one weekly instillation for 6 weeks and maintenance according to SWOG protocol.-CHT: 40 mg MMC diluted in 40 mL of distilled water at 43 C using COMBATÒ recirculation system for 60 minutes, one weekly instillation for 6 weeks and one monthly instillation for 6 months.Follow-up was performed with cytologyþcystoscopy every 3 months as well as upper urinary tract imaging yearly, as stated by EAU Guidelines. Primary endpoint was recurrence-free survival at 24 months. Secondary objectives: safety, progression rate, overall survival and quality of life.RESULTS: Fifty patients randomized (100% recruitment completed), 48 finally starting treatment. Median age is 73 years, 87.6% males and 83% primary tumours. Baseline characteristics were comparable in both groups. Median follow-up is 24.8 months from TURBT. For the BCG group, 6 recurrences (from which 5 progressions to T2) were reported, and only 3 recurrences (from which 2 progressions) happened in the CHT group. Regarding safety profile, adverse events (AE) appeared in 12 patients from CHT and 10 from BCG group, with no differences in the severity (4 patients in each group for CTCAE grade 3). AE in CHT group were mostly grade 1.CONCLUSIONS: According to our trial preliminary results, CHT in high risk NMIBC patients seems at least not to be inferior to BCG in terms of efficacy. Moreover, patients under CHT have milder side effects than those under BCG treatment.