Transcriptomic analysis by RNA sequencing characterises malignant progression of canine insulinoma from normal tissue to metastatic disease

Abstract

insulinomas (inS) are the most common human and canine functioning pancreatic neuroendocrine tumours. The long-term prognosis for malignant INS is poor, because micrometastases are frequently missed during surgery. As human and canine malignant inS share clinical and histopathological features, dogs have been proposed as models for INS research. Using RNA-sequencing, we conducted a pilot study to better understand the underlying molecular mechanisms of canine inS. normal canine pancreas and lymph node control tissues were compared with primary INS and INS-metastatic lymph nodes, revealing more than 3,000 genes differentially expressed in normal pancreas compared to primary INS. Only 164 genes were differentially expressed between primary INS and INS-metastatic lymph nodes. Hierarchical clustering analysis demonstrated similar genetic profiles in normal pancreas and early clinical stage primary INS, whereas late clinical stage primary INS resembled the genetic profile of INS-metastatic lymph nodes. These findings suggest that markers of malignant behaviour could be identified at the primary site of the disease. Finally, using the REACTOME pathways database, we revealed that an active collagen metabolism, extracellular matrix remodelling, betacell differentiation and non-beta-cell trans-differentiation might cause disease progression and hyperinsulinism in INS, identifying major pathways worthy of future research in this currently poorly controlled disease. Insulinomas (INS) are the most commonly diagnosed functioning pancreatic neuroendocrine tumours in humans and dogs 1,2. They are insulin-producing tumours that can arise from pancreatic beta-cells 1,2. Diagnosis and treatment of human and canine INS can be challenging and prognosis highly variable depending on therapeutic decisions 3-5. Currently, surgical resection of primary localised benign INS is considered the treatment of choice. However, the prognosis after surgery for advanced-stage disease is still poor and adjuvant therapy options are limited 5,6. Thus, better therapeutic tools are urgently required. Traditional benign versus malignant classification of these tumours is often difficult at initial diagnosis. Grade and stage are the major determinants of prognosis in humans 4,7. Similarly, in dogs, TNM stage, increased nuclear size, pleomorphism, Ki67 and DNA content are considered valuable histological features to predict the biological behaviour of INS 6, 8. Still, grading and staging have been so far insufficient to fully assess the degree of malignancy of human and canine INS. In both humans and dogs, these tumours can have heterogeneous microscopic findings and the presence of metastases, mainly located in the liver, represents so far the only definitive feature that characterises individual tumours as malignant 3,8-11. Canine INS are considered typically malignant as they

Fluorine‐18 fluorodeoxyglucose positron emission tomography‐computed tomography for staging of canine insulinoma: 3 cases (2019–2020)

“…2017, Capodanno et al . 2020). Gene expression and extent of metastatic potential have been shown to vary between subsets of insulinomas in dogs, feasibly accounting for this discrepancy (Buishand et al .…”

“…The genetic profile of canine insulinomas is also dictated by clinical stage, in that late stage primary insulinomas have genetic profiles more similar to their metastatic lesions than do early stage primary insulinomas to their metastases (Capodanno et al . 2020). We speculate that the confounding clinicopathological features and multifaceted transformation of canine insulinomas may influence the glucose metabolism and subsequent degree of avidity of these tumours.…”

“…2019, Capodanno et al . 2020). Misregistration artefact from respiratory motion or peristalsis of adjacent gastrointestinal segments also has the potential to obscure assessment of pancreatic lesions, though this did not significantly interfere with interpretation in our study.…”

“…2019, Capodanno et al . 2020). Canine insulinomas predominantly demonstrate malignant behaviour with a high rate of metastasis, which is presumed in almost 100% of cases at the time of diagnosis and most frequently involves the liver and regional lymph nodes (Robben et al .…”

“…2019, Capodanno et al . 2020). In comparison, the majority of human insulinomas are classified as benign and only 5 to 16% of cases develop metastatic disease (Goutal et al .…”