Recurrent loss of heterozygosity correlates with clinical outcome in pancreatic neuroendocrine cancer

Abstract

Pancreatic neuroendocrine tumors (pNETs) are uncommon cancers arising from pancreatic islet cells. Analysis of gene mutation, copy number and RNA expression of 57 sporadic pNETs showed that pNET genomes are dominated by aneuploidy. Remarkably, ~25% of pNETs had genomes characterized by recurrent loss of heterozygosity (LoH) of the same 10 chromosomes, accompanied by bi-allelic MEN1 inactivation, and these cases had generally poor clinical outcome. Another ~25% of all pNETs had chromosome 11 LoH and bi-allelic MEN1 inactivation, lacking the recurrent LoH pattern -these had universally good clinical outcome. Some level of aneuploidy was common, and overall ~80% of pNETs had LoH of ≥1 chromosome. This aneuploidy led to changes in RNA expression at the level of whole chromosomes and allowed pathogenic germline variants (e.g. ATM) to be expressed unopposed, inactivating downstream tumor suppressor pathways. Some pNETs appear to utilize VHL gene methylation or mutation to activate pseudohypoxia. Contrary to expectation neither tumor morphology within well-differentiated pNETs nor single gene mutation had significant associations with clinical outcome, nor did expression of RNAs reflecting the activity of immune, differentiation, proliferative or tumor suppressor pathways. MEN1 was the only statistically significant recurrently mutated driver gene in pNETs. Only one pNET had clearly oncogenic and actionable SNVs (in PTEN and FLCN) confirmed by corroborating RNA expression changes. The two distinct patterns of aneuploidy described here, associated with markedly poor and good clinical outcome respectively, define a novel oncogenic mechanism and the first route to genomic precision oncology for this tumor type. IntroductionPancreatic neuroendocrine tumors (pNETs) are clinically heterogeneous tumors derived from neuroendocrine cells of pancreatic islets, which differ from one other by their primary organ of origin and degree of cellular differentiation. Currently, therapeutic decisions must be made with little knowledge of the biological drivers of individual NETs, underlining the importance of improved genomic understanding of these tumors. Although driver mutations in tumor suppressor genes have been found in pNETs (e.g., MEN1, DAXX, ATRX, VHL, YY1, and mechanistic target of rapamycin (mTOR) pathway genes (1-3)) they are infrequent and are not generally able to indicate specific systemic therapies. In addition to these driver mutations, other genomic changes have been observed in pNETs, including: telomeric dysregulation (4, 5), copy number (CN) changes (6), changes in RNA expression that indicate mTOR pathway activation (7), germline MEN1 and MUTYH inactivation (2, 8). Epigenetic changes in methylation (9) and microRNA expression (10) have been described in pNETs, with insulinomas especially enriched for changes to the sequence, methylation and expression of genes encoding epigenetic modifiers (11).

Epigenetic Dysregulation of 5-hydroxymethylcytosine in Well-Differentiated Pancreatic Neuroendocrine Tumors

“…As such, a direct comparison with recent molecular studies could not be performed. 22,23,31 Also, we were unable to validate the proposed prediction model on a training set in this study or analyze the predictive value of 5-hmC for the rate of subsequent recurrence or distant metastasis in a cohort of patients with localized pancreatic disease. The effect of 5-hmC antibody clones or variation in the immunohistochemistry protocol was not determined in this study.…”

Recurrent Loss of Heterozygosity in Pancreatic Neuroendocrine Tumors

“…1,4 ATRX and DAXX mutations or gene loss are also associated with frequent losses of chromosomes 1, 2, 3, 6, 8, 10, 11, 15, 16, 21, and 22. [4][5][6][7] These losses are termed chromosomal instability, although these losses are recurring, involve entire chromosomes, and do not involve midchromosome or complex rearrangements, fragmentation, or other signs of true instability. 7 ATRX or DAXX genes are mutated in 40% of PanNETs, typically in association with MEN1 mutations, although DAXX and ATRX mutations can occur without MEN1 and MEN1 alterations can also be seen alone.…”

“…[4][5][6][7] These losses are termed chromosomal instability, although these losses are recurring, involve entire chromosomes, and do not involve midchromosome or complex rearrangements, fragmentation, or other signs of true instability. 7 ATRX or DAXX genes are mutated in 40% of PanNETs, typically in association with MEN1 mutations, although DAXX and ATRX mutations can occur without MEN1 and MEN1 alterations can also be seen alone. 2 MEN1 encodes Menin, a tumor suppressor most wellknown for its association with the syndrome termed multiple endocrine neoplasia type 1.…”

“…PanNET with DAXX and ATRX mutations or gene loss are often associated with a worse clinical outcome 1,4 . ATRX and DAXX mutations or gene loss are also associated with frequent losses of chromosomes 1, 2, 3, 6, 8, 10, 11, 15, 16, 21, and 22 4–7. These losses are termed chromosomal instability, although these losses are recurring, involve entire chromosomes, and do not involve mid-chromosome or complex rearrangements, fragmentation, or other signs of true instability 7…”

“…Lawrence et al7 postulated 3 different clinically relevant subtypes of PanNETs. Group 1: a poor prognostic subgroup characterized by recurrent multichromosome losses (specifically the loss of 10 chromosomes: 1, 2, 3, 6, 8, 10, 11, 16, 21, and 22) and mutations in, or loss of, the genes DAXX or ATRX ; group 2: an indolent group characterized by loss of heterozygosity of chromosome 11 only without loss of the other 9 chromosomes and MEN1 mutations alone, without DAXX or ATRX alterations; and group 3: a heterogenous group that did not fit the prior 2 subgroups.…”