The Jak Stat1 Axis, Interferon signaling, Resistance to Cytotoxic Therapy and Metastasis

 

STAT1 is activated by interferon and other ligands such as epidermal growth factor and interleukin-6 (IL-6). Following activation, STAT1 is translocated to the nucleus and activates transcription of interferon stimulated genes (ISGs). Although activation of STAT1 by interferon is associated with anti-viral defense and tumor suppression, data emerging from our laboratory indicates that expression of the STAT1 pathway confers cellular resistance to DNA damaging agents and mediates aggressive tumor growth. Recent advances in the development of Janus-activated kinase/STAT inhibitors and peptide inhibitors specific for the individual STAT proteins provide new insights into the various effects of this pathway on tumor behavior, as well as new and potentially novel targets for cancer therapy.

STAT 1 and Interferon regulated genes confer radio-resistance and resistance to chemotherapy. The term radio-resistance is usually applied to the restoration of wild-type sensitivity to radiation in model organisms that have DNA repair defects. Clinically, the term refers to tumors that are not cured by local radiotherapy. To identify genes that mediate tumor radio resistance, xenografts of a radiosensitive tumor line were subjected to repeated cycles of radiation and serial passage in animals which resulted in a selection of radio-resistant tumors. Expressional profiling of the resistant and the radiosensitive tumors identified a set of genes that were differentially overexpressed in the radio-resistant cells. Of these 49 initially identified genes, 31 were identified as interferon stimulated genes including STAT1, the apical transcription factor of this group of genes. This set of genes was designated as the interferon DNA damage signature (IRDS). Further experiments revealed that members of the IRDS are up regulated by fractionated radiation in vitro and in xenografted tumor models and expressed in human tumors. Enforced expression of STAT1 in radiosensitive lines rendered the lines radio-resistant and knock-down of STAT1 rendered the cells radiosensitive. This work has been confirmed by other investigators.  Over expression of STAT 1 also mediated resistance to doxorubicin. This work was performed in collaboration with Drs. Nikolai Khodarev (Radiation and Cellular Oncology) and Bernard Roizman (Molecular Genetics and Cell Biology)

CLINICAL TRANSLATION STUDIES OF STAT AND INTERFERON STIMULATED GENES

In order to evaluate the clinical significance of these experimental observations, expressional databases from a variety of different cancers were investigated. 37% of head and neck, 48% of lung, 29% of prostate, 48% of breast, and 50% of high-grade gliomas constitutively expressed interferon stimulated genes. Other investigators have similar results showing the expression of interferon stimulated genes in breast, lung, ovarian, and cervical cancer. We developed an IRDS 7-gene classifier applied to a data set of 295 patients with early stage breast cancer. The 243 patients who received adjuvant radiotherapy were analyzed for local-regional failure using Kaplan-Maier survival statistics. Results showed that the IRDS-positive patients suffered a 30% to 40% increase in local failure rate compared with IRDS-negative patients. Further analyses with independent databases are necessary to validate these results.  Chemo-resistance was also associated with overexpression of the STAT1 pathway by us and others. Analysis of the breast cancer databases with the IRDS demonstrated that patients who received adjuvant chemotherapy were significantly more likely to fail with distant metastasis and not respond to adjuvant chemotherapy when they expressed the IRDS. Thus the 7-gene IRDS has potential to be a clinical meaningful classifier in the context of a predictive (as contrasted with prognostic assay). That is, expression of these genes is likely to determine which patients will respond to anti-tumor therapy. This work was performed in collaboration with Drs. Nikolai Khodarev and Andrew Minn (Radiation and Cellular Oncology University of Chicago and Dept of Radiotherapy University of Pennsylvania)

STAT 1 AND INTERFERON STIMULATED GENES AND THE TUMOR MICROENVIRONMENT   

Colleagues have reported that chronic expression of interferons alpha and beta rendered cells radio resistant and similar data were obtained by our laboratory. Interferon can select tumor clones resistant to a toxic tumor microenvironment and concurrently to genotoxic therapy. We demonstrated this by showing that melanoma lines cultivated by serial passage or by prolonged exposure to interferon were resistant to both chemotherapeutic agents as well as radiation. These data suggest that tumor clones with STAT1 dependence resistant to a cytotoxic environment may have selective advantage to promote aggressive tumor growth. We extended our observation of this by demonstrating that melanoma cells selected for high expression of STAT1 were more highly metastatic than melanoma cells with low expression of STAT1 and that these phenotypes could be reversed by suppression of STAT1 (NN Khodarev).

CURRENT RESEARCH

We are currently performing a siRNA screen to determine which of the interferon resistant downstream genes are important and in which tumors specific ISGs mediate resistance to radiotherapy and chemotherapy. Hopefully, these studies will lead to new investigations of mechanisms of tumor resistance and eventually metastasis.

REFERENCES

Weichselbaum RR, Ishwaran H, Yoon T, Nuyten DS, Baker SW, Khodarev N, Su AW, Shaikh AY, Roach P, Kreike B, Roizman B, Bergh J, Pawitan Y, van de Vijver MJ, Minn AJ. An interferon-related gene signature for DNA damage resistance is a predictive marker for chemotherapy and radiation for breast cancer. Proc Natl Acad Sci U S A. 2008 Nov 25;105(47):18490-5. Epub 2008 Nov 10.

Khodarev NN, Roizman B, Weichselbaum RR. Molecular pathways: interferon/stat1 pathway: role in the tumor resistance to genotoxic stress and aggressive growth. Clin Cancer Res. 2012 Jun 1;18(11):3015-21. Epub 2012 May 21.

Pitroda SP, Zhou T, Sweis RF, Filippo M, Labay E, Beckett MA, Mauceri HJ, Liang H, Darga TE, Perakis S, Khan SA, Sutton HG, Zhang W, Khodarev NN, Garcia JG, Weichselbaum RR. Tumor endothelial inflammation predicts clinical outcome in diverse human cancers. PLoS One. 2012;7(10):e46104. doi: 10.1371/journal.pone.0046104. Epub 2012 Oct 4.

Khodarev N, Ahmad R, Rajabi H, Pitroda S, Kufe T, McClary C, Joshi MD, MacDermed D, Weichselbaum R, Kufe D. Cooperativity of the MUC1 oncoprotein and STAT1 pathway in poor prognosis human breast cancer. Oncogene. 2010 Feb 11;29(6):920-9. Epub 2009 Nov 16.

Duarte CW, Willey CD, Zhi D, Cui X, Harris JJ, Vaughan LK, Mehta T, McCubrey RO, Khodarev NN, Weichselbaum RR, Gillespie GY. Expression signature of IFN/STAT1 signaling genes predicts poor survival outcome in glioblastoma multiforme in a subtype-specific manner. PLoS One. 2012;7(1):e29653. Epub 2012 Jan 5.

Khodarev NN, Roach P, Pitroda SP, Golden DW, Bhayani M, Shao MY, Darga TE, Beveridge MG, Sood RF, Sutton HG, Beckett MA, Mauceri HJ, Posner MC, Weichselbaum RR. STAT1 pathway mediates amplification of metastatic potential and resistance to therapy. PLoS One. 2009 Jun 8;4(6):e5821.

Pitroda SP, Wakim BT, Sood RF, Beveridge MG, Beckett MA, MacDermed DM, Weichselbaum RR, Khodarev NN. STAT1-dependent expression of energy metabolic pathways links tumour growth and radioresistance to the Warburg effect. BMC Med. 2009 Nov 5;7:68.