Next-generation sequencing technologies have greatly expanded our understanding of cancer genetics. Antisense technology is an attractive platform with the potential to translate these advances into improved cancer therapeutics, because antisense oligonucleotide (ASO) inhibitors can be designed on the basis of gene sequence information alone. Recent human clinical data have demonstrated the potent activity of systemically administered ASOs targeted to genes expressed in the liver. Here, we describe the preclinical activity and initial clinical evaluation of a class of ASOs containing constrained ethyl modifications for targeting the gene encoding the transcription factor STAT3, a notoriously difficult protein to inhibit therapeutically. Systemic delivery of the unformulated ASO, AZD9150, decreased STAT3 expression in a broad range of preclinical cancer models and showed antitumor activity in lymphoma and lung cancer models. AZD9150 preclinical activity translated into single-agent antitumor activity in patients with highly treatment-refractory lymphoma and non-small cell lung cancer in a phase I dose escalation study.

BackgroundThe Janus kinase (JAK) and signal transduction and activation of transcription (STAT) signaling pathway is an attractive target in multiple cancers. Activation of the JAK-STAT pathway is important in both tumorigenesis and activation of immune responses. In diffuse large B-cell lymphoma (DLBCL), the transcription factor STAT3 has been associated with aggressive disease phenotype and worse overall survival. While multiple therapies inhibit upstream signaling, there has been limited success in selectively targeting STAT3 in patients. Antisense oligonucleotides (ASOs) represent a compelling therapeutic approach to target difficult to drug proteins such as STAT3 through of mRNA targeting. We report the evaluation of a next generation STAT3 ASO (AZD9150) in a non-Hodgkin’s lymphoma population, primarily consisting of patients with DLBCL.MethodsPatients with relapsed or treatment refractory lymphoma were enrolled in this expansion cohort. AZD9150 was administered at 2 mg/kg and the 3 mg/kg (MTD determined by escalation cohort) dose levels with initial loading doses in the first week on days 1, 3, and 5 followed by weekly dosing. Patients were eligible to remain on therapy until unacceptable toxicity or progression. Blood was collected pre- and post-treatment for analysis of peripheral immune cells.ResultsThirty patients were enrolled, 10 at 2 mg/kg and 20 at 3 mg/kg dose levels. Twenty-seven patients had DLBCL. AZD9150 was safe and well tolerated at both doses. Common drug-related adverse events included transaminitis, fatigue, and thrombocytopenia. The 3 mg/kg dose level is the recommended phase 2 dose. All responses were seen among DLBCL patients, including 2 complete responses with median duration of response 10.7 months and 2 partial responses. Peripheral blood cell analysis of three patients without a clinical response to therapy revealed a relative increase in proportion of macrophages, CD4+, and CD8+ T cells; this trend did not reach statistical significance.ConclusionsAZD9150 was well tolerated and demonstrated efficacy in a subset of heavily pretreated patients with DLBCL. Studies in combination with checkpoint immunotherapies are ongoing.Trial registrationRegistered at ClinicalTrials.gov: NCT01563302. First submitted 2/13/2012.Electronic supplementary materialThe online version of this article (10.1186/s40425-018-0436-5) contains supplementary material, which is available to authorized users.

Antisense oligonucleotides (ASOs) are a novel therapeutic approach to target difficult-to-drug protein classes by targeting their corresponding mRNAs. Significantly enhanced ASO activity has been achieved by the targeted delivery of ASOs to selected tissues. One example is the targeted delivery of ASOs to hepatocytes, achieved with N-acetylgalactosamine (GalNAc) conjugation to ASO, which results in selective uptake by asialoglycoprotein receptor (ASGR). Here we have evaluated the potential of GalNAc-conjugated ASOs as a therapeutic approach to targeting difficult-to-drug pathways in hepatocellular carcinoma (HCC). The activity of GalNAc-conjugated ASOs was superior to that of the unconjugated parental ASO in ASGR (+) human HCC cells in vitro , but not in ASGR (−) cells. Both human- and mouse-derived HCC displayed reduced levels of ASGR, however, despite this, GalNAc-conjugated ASOs showed a 5- to 10-fold increase in potency in tumors. Systemically administered GalNAc-conjugated ASOs demonstrated both enhanced antisense activity and antitumor activity in the diethylnitrosamine-induced HCC tumor model. Finally, GalNAc conjugation enhanced ASO activity in human circulating tumor cells from HCC patients, demonstrating the potential of this approach in primary human HCC tumor cells. Taken together, these results provide a strong rationale for a potential therapeutic use of GalNAc-conjugated ASOs for the treatment of HCC.

Genomic technologies have greatly expanded our understanding of cancer biology, although they have not yet been effectively translated into improved cancer therapeutics, partly due to the inability of available therapeutic modalities to target the most promising cancer driver pathways. In contrast to other therapeutic approaches, antisense technology allows the rational design of potent, sequence-specific inhibitors based on genome sequencing information alone. Recent human clinical data have demonstrated the potent activity of antisense oligonucleotides (ASOs) targeted to genes expressed by the liver. Here we describe preclinical and clinical activity of a high affinity, new generation chemistry (constrained ethyl modified) ASO, AZD9150. AZD9150 targets the mRNA coding for the transcription factor STAT3, which is considered a difficult protein to target therapeutically. Systemic delivery of unformulated AZD9150 resulted in strong inhibition of STA3 RNA and protein levels in demonstrates in a broad range of preclinical cancer models in vivo including several patient-derived xenograft models. AZD9150 administration resulted in 2 partial responses highly treatment-refractory diffuse large B-cell lymphoma and in 2 mixed responses in Hodgkin's Lymphoma patients in the phase I dose escalation study. These findings suggest that this technology has the potential to help bridge the pharmacogenomic divide in cancer drug discovery. [D.H. and Y.K. contributed equally to this work.] Citation Format: David Hong, Youngsoo Kim, Anas Younes, John Nemunaitis, Nathan Fowler, Jeff Hsu, Tianyuan Zhou, Luis Fayad, Nancy Zhang, Sarina Piha-Paul, Richard Woessner, Murali VP Nadella, Deborah Lawson, Corinne Reimer, Minji Jo, Joanna Schmidt, Xiaokun Xiao, Sarah Greenlee, Gene Hung, Mason Yamashita, David C. Blakey, Brett P. Monia, A. Robert Macleod, Razelle Kurzrock. Preclinical pharmacology and clinical efficacy of AZD9150 (ISIS-STAT3Rx), a potent next-generation antisense oligonucleotide inhibitor of STAT3. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr LB-227. doi:10.1158/1538-7445.AM2014-LB-227

The worldwide rate of hepatocellular carcinoma (HCC) occurrence is rising rapidly; however, outside of transplantation, few effective treatment options are available. Hepatocellular carcinogenesis is a complex, multistep process involving many genetic alterations, often occurring in targets that are considered “undruggable” by conventional therapeutic modalities. Antisense oligonucleotide (ASO) technology offers the potential to target these undruggable pathways in HCC tumors in vivo and to determine the sensitivity of HCC to the ablation of these pathways. Using either the diethylnitrosamine (DEN) induced mouse HCC model or DEN in combination with the profibrotic agent carbon tetrachloride (CCl4), we first examined gene expression changes in induced HCC tumors. DEN-induced HCC has a very long latency period (∼8 months). We demonstrate that mice injected with DEN and CCl4 developed HCC with a shorter latency and a higher tumor incidence. We also identified several genes of interest that could potentially be HCC drivers and targets for ASO therapy. Conjugation of ASOs with N-acetyl D-galactosamine (GalNAc) has been shown to increase ASO activity >10x in normal liver. Here we demonstrate that GalNAc conjugated ASOs show similar increased potency in HCC tumors and the complete depletion of Myd88 protein levels. Importantly, in addition to the ability to potently inhibit Myd88 levels in tumors, GalNAc-conjugated Myd88 ASOs also resulted in significantly reduced tumor burden. These data demonstrate that GalNAc-conjugated Gen 2.5 ASOs have potent activity in HCC tumors and that Myd88 maybe be an attractive therapeutic target in this disease. Citation Format: Joanna Schmidt, Minji Jo, Tianyuan Zhou, Youngsoo Kim, A. R. MacLeod. Characterization of GalNAc-conjugated generation 2.5 ASOs in DEN and DEN/CCL4-induced HCC tumors. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 4359. doi:10.1158/1538-7445.AM2015-4359