Canine Histiocytic and Hemophagocytic Histiocytic Sarcomas Display KRAS and Extensive PTPN11/SHP2 Mutations and Respond In Vitro to MEK Inhibition by Cobimetinib

Abstract

Histiocytic sarcoma (HS) is a rare and highly aggressive cancer in humans and dogs. In dogs, it has a high prevalence in certain breeds, such as Bernese mountain dogs (BMDs) and flat-coated retrievers. Hemophagocytic histiocytic sarcoma (HHS) is a unique form of HS that presents with erythrophagocytosis. Due to its rareness, the study of HHS is very limited, and mutations in canine HHS patients have not been studied to date. In previous work, our research group identified two major PTPN11/SHP2 driver mutations, E76K and G503V, in HS in dogs. Here, we report additional mutations located in exon 3 of PTPN11/SHP2 in both HS and HHS cases, further supporting that this area is a mutational hotspot in dogs and that mutations in tumors and liquid biopsies should be evaluated utilizing comprehensive methods such as Sanger and NextGen sequencing. The overall prevalence of PTPN11/SHP2 mutations was 55.8% in HS and 46.2% in HHS. In addition, we identified mutations in KRAS, in about 3% of HS and 4% of HHS cases. These findings point to the shared molecular pathology of activation of the MAPK pathway in HS and HHS cases. We evaluated the efficacy of the highly specific MEK inhibitor, cobimetinib, in canine HS and HHS cell lines. We found that the IC₅₀ values ranged from 74 to 372 nM, which are within the achievable and tolerable ranges for cobimetinib. This finding positions cobimetinib as a promising potential candidate for future canine clinical trials and enhances our understanding of the molecular defects in these challenging cancers.

Keywords: KRAS; MAPK pathway; MEK inhibitors; PTPN11; SHP2; cancer genetics; cobimetinib; comparative oncology; hemophagocytic histiocytic sarcoma; histiocytic sarcoma; myeloid tumors.

Figure 1

A lollipop plot of somatic mutations found in the HS samples diagramed on a PTPN11/SHP2 protein domain map, modeled after similar plots on cbioportal.org (accessed on 6 January 2024). The two Src homology domains are indicated with the label SH2, the N-terminal SH2 domain is indicated in blue, the C-terminal SH2 domain is indicated in green, and the phosphatase domain is indicated in red. The numbers along the x axis indicate the amino acid (aa) residues from the N to the C terminal of the SHP2 protein.

Figure 2

A lollipop plot of mutations found in the HHS samples diagramed on a PTPN11/SHP2 protein domain map, modeled after similar plots on cbioportal.org (accessed on 6 January 2024). The two Src homology domains are indicated with the label SH2, the N-terminal SH2 domain is indicated in blue, the C-terminal SH2 domain is indicated in green, and the phosphatase domain is indicated in red. The numbers along the x axis indicate the amino acid (aa) residues from the N to the C terminal of the SHP2 protein.

Figure 3

Cartoon and 3D model representation of the canine SHP2 protein. The protein transcript used is the 593 amino acid peptide (A0A4D6PF96). (A) A schematic domain map depicting the locations of the N- and C-terminal SRC homology 2, or SH2, domains (blue and green) and phosphatase, or PTP, domain (red), with depictions of the auto-inhibited inactive and “open” active conformations of the SHP2 protein. In its inactive state, the N-SH2 domain binds to the PTP domain around the PTP catalytic site. Mutations in the N-SH2 and PTP domains help to promote the “open” conformation. Pink stars represent SNPs that result in the promotion of the active state and increased catalytic activity. Additionally, an exon map of the SHP2 protein highlights the exon 3 mutational hotspot. (B) 3D model of the SHP2 protein depicting the auto-inhibited conformation. The catalytic cleft is bounded by a red box. (C) Zoomed in view of a portion (highlighted in panel (B)) of the cleft encompassing numerous known mutation sites, including the mutations presented here. The residues G60, D61, A72, and E76 are located in the N-SH2 domain (E69 not captured in panel (C)). The G503 residue is located in the PTP domain. Cartoon inspired by Chen et al. [25]. 3D protein models were generated from AlphaFold [26,27] and UCSF ChimeraX [28].