Genome Aberrations in Canine Mammary Carcinomas and Their Detection in Cell-Free Plasma DNA

Abstract

Mammary tumors are the most frequent cancers in female dogs exhibiting a variety of histopathological differences. There is lack of knowledge about the genomes of these common dog tumors. Five tumors of three different histological subtypes were evaluated. Massive parallel sequencing (MPS) was performed in comparison to the respective somatic genome of each animal. Copy number and structural aberrations were validated using droplet digital PCR (ddPCR). Using mate-pair sequencing chromosomal aneuploidies were found in two tumors, frequent smaller deletions were found in one, inter-chromosomal fusions in one other, whereas one tumor was almost normal. These aberrations affect several known cancer associated genes such as cMYC, and KIT. One common deletion of the proximal end of CFA27, harboring the tumor suppressor gene PFDN5 was detected in four tumors. Using ddPCR, this deletion was validated and detected in 50% of tumors (N = 20). Breakpoint specific dPCRs were established for four tumors and tumor specific cell-free DNA (cfDNA) was detected in the plasma. In one animal tumor-specific cfDNA was found >1 year after surgery, attributable to a lung metastasis. Paired-end sequencing proved that copy-number imbalances of the tumor are reflected by the cfDNA. This report on chromosomal instability of canine mammary cancers reveals similarities to human breast cancers as well as special canine alterations. This animal model provides a framework for using MPS for screening for individual cancer biomarkers with cost effective confirmation and monitoring using ddPCR. The possibility exists that ddPCR can be expanded to screening for common cancer related variants.

Chromosomal Instability in Cell-Free DNA Is a Serum Biomarker for Prostate Cancer

“…No separation of cfDNA chromosomal markers was observed on the basis of the Gleason score that would allow identification of aggressive from indolent prostate cancer. This observation is consistent with the lack of the hemizygous 3-Mb deletion generated by the fusion of TMPRSS2 (transmembrane protease, serine 2) 7 and ERG (v-ets erythroblastosis virus E26 oncogene homolog) on chromosome 21 within the confines of our 20-loci hotspot model, al- 20; HIST1H2BPS3, HIST1H2B pseudogene 3; MICU2, mitochondrial calcium uptake 2; FNTAP2, farnesyltransferase, CAAX box, alpha pseudogene 2; RNU6 -59P, RNA, U6 small nuclear 59, pseudogene; RPS7P10, ribosomal protein S7 pseudogene 10; FGF9, fibroblast growth factor 9; LINC00424, long intergenic non-protein coding RNA 424; NME1P1, NME/NM23 nucleoside diphosphate kinase 1 pseudogene 1; IFT88, intraflagellar transport 88; IL17D, interleukin 17D; N6AMT2, N-6 adenine-specific DNA methyltransferase 2 (putative); XPO4, exportin 4; PPIAP27, peptidylprolyl isomerase A (cyclophilin A) pseudogene 27; LATS2, large tumor suppressor kinase 2; SAP18, Sin3A-associated protein, 18 kDa; SKA3, spindle and kinetochore associated complex subunit 3; MRPL57, mitochondrial ribosomal protein L57; EEF1A1, eukaryotic translation elongation factor 1 alpha 1; FAM214A, family with sequence similarity 214, member A; RPSAP55, ribosomal protein SA pseudogene 55; VPS13C, vacuolar protein sorting 13 homolog C (S. cerevisiae); MON1B, MON1 secretory trafficking family member B; SYCE1L, synaptonemal complex central element protein 1-like; VN2R10P, vomeronasal 2 receptor 10 pseudogene; PTPRT, protein tyrosine phosphatase, receptor type, T; PPIAP21, peptidylprolyl isomerase A (cyclophilin A) pseudogene 21; IFT52, intraflagellar transport 52; MYBL2, v-myb avian myeloblastosis viral oncogene homolog-like 2; GTSF1L, gametocyte specific factor 1-like; JPH2, junctophilin 2; FITM2, fat storage-inducing transmembrane protein 2; R3HDML, R3H domain containing-like; HNF4A, hepatocyte nuclear factor 4, alpha; MIR3646, microRNA 3646; TTPAL, tocopherol (alpha) transfer protein-like; SERINC3, serine incorporator 3; PKIG, protein kinase (cAMP-dependent, catalytic) inhibitor gamma; ADA, adenosine deaminase.…”

“…Despite the potential for bias introduced by wholegenome sequencing, cancer-derived cfDNA has been demonstrated to recapitulate genomic tumor DNA (11,20,21,37 ). Individual tumor genomic analysis brings significant advantages to the power of cfDNA diagnostics.…”

Tumor Cell–Free DNA Copy Number Instability Predicts Therapeutic Response to Immunotherapy

“…One hallmark of cancer biology is that most cancer cells have chromosomal instability often expressed as large physical or functional somatic gains and losses in tumor cfDNA (11)(12)(13). This report describes a novel application of the liquid biopsy by quantifying cancer-related chromosomal instability in cfDNA, an approach minimally influenced by clonal heterogeneity, which remains a problem for targeted "actionable" genes (10).…”

Molecular Homology and Difference between Spontaneous Canine Mammary Cancer and Human Breast Cancer

“…However, unlike human breast cancer, canine MC is poorly characterized at the genome-wide level. For example, only five canine MC cases have recently undergone ~2X whole genome sequencing (WGS) (14), and a limited number have been analyzed with gene expression microarray (15-17). This drastically differs from their human counterparts, where thousands of breast cancer genomes and transcriptomes are characterized, with several studies cited here (18-23).…”