Cancer development and growth involves the coordinated dysregulation of multiple cellular processes, including proliferation, apoptosis, migration, immune evasion, genome stability, metabolism, and angiogenesis. As global regulators capable of simultaneously regulating hundreds of genes, microRNAs (miRs) are lead candidates for coordination of the oncogenic process, with multiple miR being identified as tumor-suppressor-miR, capable of preventing oncogenesis, and oft downregulated in tumors, or onco-miR, with tumor-promoting properties and overexpression in certain tumors (1). As the technology to either supplement or suppress miR with a high degree of specificity is available and readily translated from one miR to the next, miR represent highly tempting therapeutic targets in cancer (2). Pancreatic cancer represents an important therapeutic target, with radically different expression of several miRs (3) and a distinct paucity of effective treatments leading to poor prognosis (4). It is therefore a promising line of research to functionally investigate the known miR expression changes in pancreatic cancer.

miR-342 is an intronic miR located within the host gene EVL. Both miR-342 and EVL are poorly characterized; however, miR-342 is proposed to have important oncogenic functions, with differential expression in pancreatic cancer (5) as well as acute myeloid leukemia (6, 7), breast cancer (triple-negative (8), inflammatory (9), or recurrent (10) subtypes), cervical cancer (11), colorectal cancer (12, 13), lung adenocarcinoma (14), lymphoma (15), and metastatic melanoma (16). The breadth of cancers observed to show expression changes in miR-342 suggests a common oncogenic function for these changes, mediated by altered regulation of the target mRNA. It should, however, be noted that miR-342 expression is coordinated with the host gene, EVL (17), and thus even if the observed correlation is functional it may not indicate an oncogenic role specific to miR-342.

Based on expression changes in cancer, miR-342 has been proposed as both an onco-miR and, more commonly, a tumor-suppressor-miR. During functional testing, the roles identified for miR-342 have been largely tumor-suppressing in nature, with repression of key oncogenic proteins. For example, miR-342 inhibits proliferation and invasiveness of non-small cell lung cancer cell lines after transfer into nude mice via regulation of RAP2B (18). Likewise, miR-342 inhibits Myc activity via regulation of E2F1 (19) and inhibits apoptosis in breast cancer cell lines via regulation of Apollon/BRUCE (20) or the human epidermal growth factor receptor 2 (HER2) pathway and inhibited the proliferation of HER2-positive cell lines (21). miR-342 has been demonstrated to inhibit the tumorigenic capacity in transplanted colon cancer cell lines, a role attributed to regulation of NAA10 (22), DNA methyltransferase 1 (23), or FOXM1 and FOXQ1 (24). The regulation of FOXM1 by miR-342 has also been proposed to be important for cervical cancer, with a function in inhibiting growth and metastasis in HeLa transplant models (25). In hepatocellular carcinoma cell lines, miR-342 inhibits proliferation by regulating the NF-κB pathway (26). Finally, miR-342 expression in the surrounding stroma may provide a tumor-suppressing function via enhancing the inhibitory effect of TGFβ on angiogenesis (27).

Together, the expression and functional data indicate miR-342 as a tumor-suppressor-miR. However, notably, to our knowledge, all functional testing has been performed in cell lines, either in vitro (19–21, 26) or in xenogenic transplant models (18, 22–25). Here, we sought to determine the in vivo functional role of miR-342 in a spontaneous murine model of pancreatic acinar carcinoma. Through longitudinal magnetic resonance imaging (MRI) assessment, we found no evidence for a tumor-suppressor role for miR-342 in the Ela1-TAg transgenic model.

The function of miR-342 in the development, growth, and pathogenicity of pancreatic cancer was tested by newly generating miR-342^0/0 mice and intercrossing with the Ela1-TAg transgenic mouse (28). The resulting miR-342^0/0 TAg⁺ mice, and the parental TAg⁺ strain, develop spontaneous pancreatic acinar carcinoma (Figure 1A), in which miR-342 expression was specifically lost in the KO (Figure 1B). The presence and size of tumors were assessed through longitudinal MRI assessment of tumor size from 7 weeks of age (Figure 2). Onset of cancer development was assessed in each group through analysis of first tumor detection by MRI. No significant difference in age of first tumor detection was induced by heterozygous or homozygous loss of miR-342 (Figure 3A). This resulted in normal cumulative incidence of pancreatic cancer in both female (Figure 3B) and male (Figure 3C) mice, indicating no significant effect of miR-342 in pancreatic cancer development in this model.

In order to measure the function of miR-342 on the growth rates of pancreatic cancer, we normalized the longitudinal MRI assessments of tumor volume (Figure 2) to the point of first detection. Tumor volumes were square root transformed to take into account the exponential growth, providing a measurement of the growth rate observed in each individual mouse (Figures 4A–F). Growth rate curves were then calculated as the percentage of tumor volume increased observed per 14 days, averaged across the entire observation period. In miR-342-sufficient mice, tumors increased in size ±500% every 14 days, with no significant change in the growth rate observed with miR-342-deficiency, in either males or females (Figure 4G).

Finally, the pathogenic impact of pancreatic cancer was analyzed in each strain. Total survival was measured out to 21 weeks of age, with only limited mortality observed in TAg⁺ mice (Figure 5). No excess mortality was observed within this time period in miR-342^+/0 TAg⁺ or miR-342^0/0 TAg⁺ mice (Figure 5), although the limited time frame of observation does not exclude effects on survival after 21 weeks of age. Together, these results indicate that there is no major function for miR-342 in the development, growth, or pathogenicity of pancreatic cancer in the Ela1-TAg model.

While these results do not encourage the exploration of miR-342 as a clinical target in pancreatic cancer, neither do they disqualify it. The expression changes observed in miR-342 in multiple cancers may still serve as diagnostic targets with clinical benefits; indeed, expression of miR-342 has been observed to correlate with postoperative survival in pancreatic cancer (29) and response to tamoxifen treatment in breast cancer (30). Furthermore, our results, while negative, do not exclude a functional role for miR-342 in pancreatic cancer—the TAg model is just one of many increasingly sophisticated pancreatic cancer models, which more closely model the most common genetic insults observed in patient specimens (31). In addition, even the extent to which murine miR physiology mimics that of the human is unclear, with mRNA exhibiting lower levels of miR recognition conservation than protein sequence conservation (32) [although many of the predicted oncogenic targets of miR-342 are conserved from mouse to human (33)]. It therefore remains distinctly possible that further preclinical exploration will support miR-342 as a valid target for clinical intervention in a subset of pancreatic cancer patients with the appropriate molecular distortions (2).

The study was approved by the University of Leuven Animal Ethics Committee, and all mice were used in accordance with the approved protocol.

JD, ML, and AL designed the study. HP generated KO mice. JD and EP performed experiments. JD, UH, and VL analyzed the data. AL wrote the study.

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.