Canine Histiocytic and Hemophagocytic Histiocytic Sarcomas Display KRAS and Extensive PTPN11/SHP2 Mutations and Respond In Vitro to MEK Inhibition by Cobimetinib
Abstract
:1. Introduction
2. Materials and Methods
2.1. Animal Use Statement
2.2. Histiocytic Sarcoma (HS) Samples
2.3. Hemophagocytic Histiocytic Sarcoma (HHS) Samples
2.4. Cell Lines
2.5. Genomic DNA Extraction
2.6. Genotyping using the TaqMan Custom SNP Genotyping Assay
2.7. Genotyping by Sanger Sequencing
2.8. Determination of IC50 Values
2.9. Analysis of Amino Acid Substitution Effects
2.10. Statistical Methods
3. Results
3.1. PTPN11/SHP2 and KRAS Mutational Status in HS Cases
3.2. PTPN11/SHP2 and KRAS Mutational Status in HHS Cases
3.3. Age and Sex Status in HS and HHS Cases
3.4. Cobimetinib Inhibits HS and HHS Cell Growth
4. Discussion
5. Conclusions
Supplementary Materials
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Acknowledgments
Conflicts of Interest
References
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Gene (Protein) Name | Mutation | Assay ID | Genomic Coordinates of Mutations * | Location in cDNA |
---|---|---|---|---|
PTPN11 (SHP2) | E76K | ANT2AD7 (Lot P180405-000 B02) | Chr26: 10,340,007 | c.440G>A (XM_038575149.1) |
G503V | ANDJ2M3 (Lot P180403-001 E05) | Chr26: 10,377,939 | c.1,508G>T (XM_038575149.1) | |
KRAS | Q61H | AN47YG4 (Lot P180307-008 A08) | Chr27: 24,263,793 | c.183A>C (XM_038577089.1) |
Gene Name | Forward Primer Sequence (5′ -> 3′) | Reverse Primer Sequence (5′ -> 3′) | Amplicon Size | Amplicon Coordinates * |
---|---|---|---|---|
PTPN11 (exon 3) | GGAAAGGAGCCAGGCAACAA | TGGCATGGAAGAGGTGCATT | 395 bp | chr26: 10339752+10340146 |
KRAS (exon 2) | AAAGGTGTTGATAGAGTGGGT | AGCCAATGGAACCCAAGTACA | 380 bp | chr27: 24279858-24280237 |
Frequency of All Mutations in PTPN11/SHP2 | 55.8% (72/129) | ||||||||
---|---|---|---|---|---|---|---|---|---|
Frequency of PTPN11/SHP2 Mutations in 2 Exons | Exon 3 47.3% (61/129) | Exon 13 8.5% (11/129) | |||||||
PTPN11/SHP2 Mutations | G60V | D61V | E69K | E76A | E76G | E76K | E76Q | A72T | G503V |
Number of dogs | 2 | 1 | 4 | 4 | 3 | 44 | 1 | 2 | 11 |
Frequency in 129 HS cases (current study) | 1.6% | 0.8% | 3.1% | 3.1% | 2.3% | 34.1% | 0.8% | 1.6% | 8.5% |
Status: New or previously reported [reference] (species) | [15] (dog) | New | [15] (human) | [16] (dog) | [15] (dog) | [9] (dog) | [16] (dog) | New | [10] (dog) |
Reported in human cancers: (per cBioportal) | yes | no | yes | yes | no | yes | no | yes | yes |
Amino Acid Substitution Effects: | |||||||||
AlphaMissense (human) | 1.000 | 0.999 | 0.998 | 0.998 | 0.997 | 1.000 | 0.997 | 0.998 | 1.000 |
PolyPhen2 (dog protein) | 1.000 | 0.997 | 0.012 | 0.999 | 1.000 | 0.997 | 0.997 | 0.967 | 1.000 |
Frequency of All Mutations in PTPN11/SHP2 | Total Percentage: 46.2% (12/26 dogs) | ||||
---|---|---|---|---|---|
Frequency of PTPN11/SHP2 Mutations in 2 Exons | Exon 3 Frequency: 26.9% (7/26) | Exon 13 Frequency: 19.2% (5/26) | |||
PTPN11/SHP2 Mutations | E69K | A72V | E76K | E76V | G503V |
Number of dogs | 1 | 1 | 4 | 1 | 5 |
Frequency in 26 HHS cases | 3.8% | 3.8% | 15.4% | 3.8% | 19.2% |
[reference] (species) | [15] (human) | New variant | [9] (dog) | New variant | [10] (dog) |
Reported in human cancers: (per cBioportal) | yes | yes | yes | no | yes |
Amino Acid Substitution Effects: | |||||
AlphaMissense (human) | 0.998 | 0.999 | 1.000 | 1.000 | 1.000 |
PolyPhen2 (dog protein) | 0.012 | 0.962 | 0.997 | 0.998 | 1.000 |
Groups | N | Avg Age | PTPN11/SHP2 Mutant | E76K | G503V | Other Variants |
---|---|---|---|---|---|---|
HS BMD | 129 | 8.4 | 72 (56%) | 44 (34%) | 11 (9%) | 17 (13%) |
HHS all cases | 26 | 7.7 | 12 (46%) | 4 (15%) | 5 (19%) | 3 (12%) |
HHS BMD | 14 | 6.6 | 9 (64%) | 4 (29%) | 4 (29%) | 1 (7%) |
HHS other breeds | 12 | 8.9 | 3 (25%) | 0 | 1 (8%) | 2 (17%) |
Groups | N | KRAS Mutant | Q61H | G12A | G12D |
---|---|---|---|---|---|
HS BMD | 129 | 4 (3%) | 3 (2%) | 1 (0.8%) | 0 |
HHS all cases | 26 | 1 (4%) | 0 | 0 | 1 (4%) |
HHS BMD | 14 | 1 (7%) | 0 | 0 | 1 (7%) |
HHS other breeds | 12 | 0 | 0 | 0 | 0 |
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Yang, Y.-T.; Engleberg, A.I.; Kapoor, I.; Kitagawa, K.; Hilburger, S.A.; Thaiwong-Nebelung, T.; Yuzbasiyan-Gurkan, V. Canine Histiocytic and Hemophagocytic Histiocytic Sarcomas Display KRAS and Extensive PTPN11/SHP2 Mutations and Respond In Vitro to MEK Inhibition by Cobimetinib. Genes 2024, 15, 1050. https://doi.org/10.3390/genes15081050
Yang Y-T, Engleberg AI, Kapoor I, Kitagawa K, Hilburger SA, Thaiwong-Nebelung T, Yuzbasiyan-Gurkan V. Canine Histiocytic and Hemophagocytic Histiocytic Sarcomas Display KRAS and Extensive PTPN11/SHP2 Mutations and Respond In Vitro to MEK Inhibition by Cobimetinib. Genes. 2024; 15(8):1050. https://doi.org/10.3390/genes15081050
Chicago/Turabian StyleYang, Ya-Ting, Alexander I. Engleberg, Ishana Kapoor, Keita Kitagawa, Sara A. Hilburger, Tuddow Thaiwong-Nebelung, and Vilma Yuzbasiyan-Gurkan. 2024. "Canine Histiocytic and Hemophagocytic Histiocytic Sarcomas Display KRAS and Extensive PTPN11/SHP2 Mutations and Respond In Vitro to MEK Inhibition by Cobimetinib" Genes 15, no. 8: 1050. https://doi.org/10.3390/genes15081050