Progression Of Human Neoplasia Research Articles

Abstract 2162: MEK signaling marks resistance to synthetic lethality induced by PI3K/Akt and BCL-XL inhibition in PTEN-mutant cancers

Abstract Background: Loss of PTEN represents one of the most common disruptions of tumor suppressor activity in the progression of human neoplasia. There are no established precision medicine approaches defined in PTEN-deficient neoplasms which include prostate, breast, glioblastoma (GBM) and uterine cancers. In PTEN-mutant prostate cancer cells, the PI3K/Akt pathway collaborates with BCL-XL specifically, to regulate the apoptotic threshold (Ren, MCR 2016). Whereas single agent PI3K/Akt inhibitors are ineffective in inducing cell death, combined inhibition of PI3K/Akt and BCL-XL in PTEN-mutant prostate cancer cells results in synergistic apoptosis suggesting a novel therapeutic strategy with translational potential in this important subset of disease. Hypothesis: We hypothesized that synthetic lethality following combinatorial PI3K/BCL-XL inhibition is conserved across the range of PTEN-mutant cancers. Methods: PTEN-mutant cell lines (ATCC), validated for PTEN-loss and p-Akt expression, were assessed in apoptosis (cleaved Parp) and cell-viability assays following combined PI3K (Buparlisib, SelleckChem) and BCL-XL (A-1331852, AbbVie) inhibition (1µM each, 24h). Since MEK signaling has been associated with resistance to PI3K pathway inhibition, p-Erk was profiled as a candidate marker of resistance. The contribution of MEK signaling to the apoptotic threshold in p-Erk expressing PTEN-mutant cells was tested with pharmacological MEK inhibition (Trametinib, SelleckChem; 1,3,5 µM 48h) in combination. Results: MEK signaling and PI3K/BCL-XL inhibition in PTEN-mutant cancersSourceCell linePTENp-Aktp-ErkSynergistic Apoptosis with PI3K / BCL-XL inhibitionLoss of cell viabilityp-Erk suppression with MEK inhibitionApoptosis with concomitant MEK inhibitionProstatePC3AbsentPresentAbsentYesYesN/AN/AProstateLNCaPAbsentPresentAbsentYesYesN/AN/AProstateC4-2BAbsentPresentAbsentYesYesN/AN/ABreastZR-75-1AbsentPresentWeakYesYesN/AN/AUterineSK-UT-1AbsentPresentPresentNoNoYesYesGBMA-172AbsentPresentPresentNoNoYesNoCervixC33-AAbsentPresentPresentNoNo-- Conclusions: Synthetic lethality in PTEN-mutant tumor cells with PI3K/BCL-XL inhibition was confined to tumor cells without active MEK signaling. MEK inhibition inconsistently overcomes resistance to therapy suggesting heterogeneity of survival pathways in p-erk expressing PTEN-deficient tumors. These data have implications for precision medicine approaches in PTEN-deficient tumors as absence of p-Erk expression may predict for a responsive phenotype. Citation Format: Raghav Joshi, Wenying Ren, Paul Mathew. MEK signaling marks resistance to synthetic lethality induced by PI3K/Akt and BCL-XL inhibition in PTEN-mutant cancers [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 2162. doi:10.1158/1538-7445.AM2017-2162

Recent progress in mouse models for tumor suppressor genes and its implications in human cancer.

Gain-of-function mutations in oncogenes and loss-of-function mutations in tumor suppressor genes (TSG) lead to cancer. In most human cancers, these mutations occur in somatic tissues. However, hereditary forms of cancer exist for which individuals are heterozygous for a germline mutation in a TSG locus at birth. The second allele is frequently inactivated by gene deletion, point mutation, or promoter methylation in classical TSGs that meet Knudson’s two-hit hypothesis. Conversely, the second allele remains as wild-type, even in tumors in which the gene is haplo-insufficient for tumor suppression. This article highlights the importance of PTEN, APC, and other tumor suppressors for counteracting aberrant PI3K, β-catenin, and other oncogenic signaling pathways. We discuss the use of gene-engineered mouse models (GEMM) of human cancer focusing on Pten and Apc knockout mice that recapitulate key genetic events involved in initiation and progression of human neoplasia. Finally, the therapeutic potential of targeting these tumor suppressor and oncogene signaling networks is discussed.

Neuroendocrine Lung Tumors: Grade Correlates with Proliferation but not Angiogenesis

Angiogenesis has been implicated in the progression of human neoplasia from benign precursor to invasive and metastatic phenotypes. The acquisition of dominant oncogenes in preneoplastic cells in vitro and in vivo has been associated with the increased ability of tumor cells to secrete angiogenic mediators and recruit blood vessels. However, in a subset of benign lesions, high levels of angiogenesis have been found before the conversion to invasive and metastatic phenotypes. In many of these benign lesions, dominant oncogenic pathways are activated first; then as malignant potential is acquired, there is a loss of nuclear tumor suppressor genes, such as p53 and p16. We studied neuroendocrine lung tumors (NLT) ranging from typical and atypical carcinoid tumors to large cell neuroendocrine and small cell carcinomas in order to determine whether angiogenesis (as assessed by mean vessel density) and proliferation rates (as assessed by MIB-1 nuclear immunohistochemical staining) correlate with tumor type. We found that increased rates of proliferation, but not angiogenesis, correlate with tumor type. The association of increased proliferation and tumor type may prove to be clinically useful and shed light on the role of sequential oncogenic alterations in NLT.

Distinct characteristics of heregulin signals mediated by HER3 or HER4.

Members of the epidermal growth-factor-receptor tyrosine-kinase (EGFR) family play important roles both in normal growth regulation/cell differentiation and in the genesis and progression of human neoplasia. In the present study, we analysed distinct heregulin (HRG) signals mediated by the HRG receptors HER3 and HER4. In overexpression cell systems, we demonstrate that HRG-induced transformation by "kinase-impaired" HER3 is dependent on coexpression of kinase active HER2. In cells coexpressing HER2 and HER4, however, both kinases significantly contribute to the HRG-induced mitogenic stimulus. In addition, we show that HER3 is no substrate of HRG-activated HER4. Analysis of EGFR crosstalk in a panel of human carcinoma cell lines revealed mainly HRG-induced activation of HER2/HER3, whereas HER4 activation is also detectable to various extents. Evidence for HRG-induced activation of HER3 and/or HER4 indicates relevance of cell-specific expression patterns of these high- and low-affinity HRG receptors in the modulation of a ligand-induced stimulus. Specific signal modulation and definition can be demonstrated further by distinct time courses of mitogen-activated protein (MAP) kinase (MAPK) activation, which are induced by distinct HRG isotypes via differential binding to HER2/HER3 versus HER2/HER4. In concert, these mechanisms of signal modulation may be decisive for the diverse biological activities of HRG in different cell types.

Proto-oncogene expression in dermal naevi and melanomas

Overexpression of proto-oncogenes (c-onc) may be involved in the initiation and progression of human neoplasia. We investigated the mRNA expression of the proto-oncogenes c-myc, c-fos, c-neu (erb B-2) and Ha-ras in 12 cutaneous melanocytic lesions (four dermal naevi, seven melanomas; one cutaneous metastatis of melanoma) and in normal skin (five cases) by Northern blot analysis; mRNA expression levels were quantified by densitometry of the specific bands. The expression of c-myc mRNA was higher in naevi than in melanomas, whereas c-fos mRNA expression was significantly higher in melanomas than in naevi. No significant differences were detected in the c-neu and Ha-ras mRNA levels in naevi and melanomas. The expression of c-neu mRNA was higher in normal skin than in the melanocytic lesions examined. Our results suggest that enhanced c-myc and c-fos expression may play an important role in the growth of melanocytic naevi and melanomas. The overexpression of c-fos may be involved in the progression of a particular subset of melanoma. Activation of Ha-ras and the c-neu gene, as determined by mRNA overexpression, does not seem to play a significant role in the progression of cutaneous pigmented lesions. Expression of c-neu may have an important function in the proliferation and/or differentiation of normal cells of the epidermis.

Strategies for the Analysis of Oncogene Overexpression: Studies of the neu Oncogene in Breast Carcinoma

The development of a consistent strategy for the analysis of oncogene expression at the cellular level is essential for understanding the roles of these genes in the development and progression of human neoplasia. Detection of the neu oncogene products in breast carcinoma was selected as a model for analysis of oncogene expression. Fifty-two primary human breast carcinomas were evaluated by quantitation of neu DNA amplification and mRNA expression and by localization of neu mRNA and protein (p 185) at the cellular level by in situ hybridization (ISH) and immunohistochemistry (IHC). The specificity and sensitivity of the molecular and immunologic probes for neu were established with the use of genetically engineered cell lines that overexpressed either neu or epidermal growth factor receptor (EGFR). Twenty-nine percent of breast carcinomas demonstrated neu DNA amplification and mRNA overexpression, and there was close correlation between the level of neu mRNA expression and detection of neu gene products by ISH and IHC. Thirty-two percent of carcinomas demonstrated neu mRNA overexpression by ISH. The immunohistochemical method using TA1 monoclonal antibody for p185 was exquisitely sensitive in acetone-fixed frozen sections and provided an excellent approach for judging overexpression as confirmed by the various molecular analyses. All areas of nonmalignant breast epithelium stained weakly, and a wide range of staining intensity was observed in malignant breast epithelium, with 31% of carcinomas judged to be p185 overexpressors. Heterogeneous expression of p185 was seen in some carcinomas. This study provides a strategic approach for the evaluation of oncogene expression in human tumors.