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Aberrant constitutive activation of STAT3 (signal transducer and activator of transcription) leads to cellular transformation and aids tumorigenesis in pancreatic cancer and its inhibition can lead to growth arrest.1,2,3,4 The exact mechanism of this constitutive activation has not been elucidated, although downstream mediators may include ERK and p21 signalling.1,2,3,4 Blocking JAK2 (Janus Kinase), a known upstream activator of STAT3, by AG490, a tryphostin inhibitor, reduced phosphorylation of STAT3 and produced comparable in vitro effects, suggesting a role for JAK2 in pancreatic carcinogenesis.3
JAK2 V617F missense mutation has recently been shown by several independent groups to play an important role in myeloproliferative disorders5 as well as prothrombotic states such as Budd‐Chiari syndrome (which may be due to latent myeloproliferative disorders).5,6 We postulated that JAK2 V617F mutation may play a role in pancreatic cancer, in which the prothrombotic state is well recognised.
We analysed genomic DNA (all from unstained slides of representative surgical specimens) from 26 patients undergoing surgery for various pancreatic diseases (table 11)) along with genomic DNA from 10 cell lines (Panc1, Paca3, MiaPaCa2, Capan1 and 2, Suit2, AsPc1, 818.4, Hpaf2, H766T) for JAK2 V617F mutation using phenol‐choloroform extraction with ethanol precipitation, using appropriate positive and negative controls as previously described.6 Briefly, all cells (>70% tumour tissue in cancer specimens, no microdissection) were scraped from unstained slides into high salt buffer and samples were made up to 200 μl with proteinase K, RNase A and sodium dodecyl sulfate, incubated at 37°C overnight before phenol extraction and ethanol precipitation. Genomic DNA from cell lines was extracted from 70% confluent million cells using Trizol (Invitrogen Ltd, Paisley, UK) reagent digestion followed by ethanol precipitation. A highly sensitive allele‐specific PCR was used to detect the G to T mutation in exon 14 of JAK2 (corresponding to amino acid position 617) as previously described.6 Briefly, a three primer PCR was performed which, in the absence of a mutation, produces an internal amplification control band. Where a mutation is present, an additional band is also amplified and is specific to the mutant sequence. The sensitivity of the assay is 1 cell in 100, determined by mixing positive control genomic DNA (extracted from HEL cells) with non‐mutated genomic DNA. We have previously validated this methodology by conventional Sanger sequencing and pyrosequencing.6 No such mutations were detected at the JAK2 617 codon for pancreatic cancer or cell samples.
We propose that STAT3 activation, as previously described in pancreatic cancer,1,2,3,4 is by mechanisms other than constitutive activation of JAK2 by missense mutation at the V617F site. Similarly, while JAK2 mutation is absent in acute myeloid leukaemia, STAT3 activation is common and is being explored.7 In glioblastoma, for example, it has been suggested that STAT3 is activated by exogenous interleukin (IL)‐4.8 IL‐4 interacts with IL‐13Rα2 receptor and, although IL‐13Rα2 does not bind with STAT3 directly, it blocks STAT6 activation and excludes this downstream event. Thus, IL‐4 can have an anti‐apoptotic effect in glioblastoma cells via STAT3 signalling. Conversely, deletion of the SOC3 (suppressor of cytokine signalling 3) gene in liver cancer9 and silencing of the SHP1 gene (by methylation) in lymphoma10 can activate STAT3 signalling. It has recently been suggested that there may be novel mutations in exon 12 which may account for a minority of patients with polycythemia vera and idiopathic erythrocytosis, who are V617F‐negative.11 The exact nature of STAT3 activation in pancreatic cancer remains to be elucidated.
Competing interests: None.