Pyrosequencing-Based Assays for Rapid Detection of HER2 and HER3 Mutations in Clinical Samples Uncover an E332E Mutation Affecting HER3 in Retroperitoneal Leiomyosarcoma.
Mutations in Human Epidermal Growth Factor Receptors (HER) are associated with poor prognosis of several types of solid tumors. Although HER-mutation detection methods are currently available, such as Next-Generation Sequencing (NGS), alternative pyrosequencing allow the rapid characterization of specific mutations. We developed specific PCR-based pyrosequencing assays for identification of most prevalent HER2 and HER3 mutations, including S310F/Y, R678Q, L755M/P/S/W, V777A/L/M, 774-776 insertion, and V842I mutations in HER2, as well as M91I, V104M/L, D297N/V/Y, and E332E/K mutations in HER3. We tested 85 Formalin Fixed and Paraffin Embbeded (FFPE) samples and we detected three HER2-V842I mutations in colorectal carcinoma (CRC), ovarian carcinoma, and pancreatic carcinoma patients, respectively, and a HER2-L755M mutation in a CRC specimen. We also determined the presence of a HER3-E332K mutation in an urothelial carcinoma sample, and two HER3-D297Y mutations, in both gastric adenocarcinoma and CRC specimens. The D297Y mutation was previously detected in breast and gastric tumors, but not in CRC. Moreover, we found a not-previously-described HER3-E332E synonymous mutation in a retroperitoneal leiomyosarcoma patient. The pyrosequencing assays presented here allow the detection and characterization of specific HER2 and HER3 mutations. These pyrosequencing assays might be implemented in routine diagnosis for molecular characterization of HER2/HER3 receptors as an alternative to complex NGS approaches.
Group of Cancer Biomarkers, Pathology Department, Health Research Institute Fundación Jiménez Díaz (IIS-FJD, UAM), Avda. Reyes Católicos. paula.galonso@fjd.es.
Paula
P
González-Alonso
Group of Cancer Biomarkers, Pathology Department, Health Research Institute Fundación Jiménez Díaz (IIS-FJD, UAM), Avda. Reyes Católicos. chamizomunoz@gmail.com.
Cristina
C
Chamizo
Madrid, Spain. victor.moreno@fjd.es.
Víctor
V
Moreno
Group of Cancer Biomarkers, Pathology Department, Health Research Institute Fundación Jiménez Díaz (IIS-FJD, UAM), Avda. Reyes Católicos. jmadoz@fjd.es.
Juan
J
Madoz-Gúrpide
Group of Cancer Biomarkers, Pathology Department, Health Research Institute Fundación Jiménez Díaz (IIS-FJD, UAM), Avda. Reyes Católicos. nerea.carvajal@fjd.es.
Nerea
N
Carvajal
Group of Cancer Biomarkers, Pathology Department, Health Research Institute Fundación Jiménez Díaz (IIS-FJD, UAM), Avda. Reyes Católicos. daoudlina@gmail.com.
Lina
L
Daoud
Group of Cancer Biomarkers, Pathology Department, Health Research Institute Fundación Jiménez Díaz (IIS-FJD, UAM), Avda. Reyes Católicos. szazo@fjd.es.
Sandra
S
Zazo
Group of Cancer Biomarkers, Pathology Department, Health Research Institute Fundación Jiménez Díaz (IIS-FJD, UAM), Avda. Reyes Católicos. ester.martin@fjd.es.
Ester
E
Martín-Aparicio
Madrid, Spain. ion.cristobal@fjd.es.
Ion
I
Cristóbal
Group of Cancer Biomarkers, Pathology Department, Health Research Institute Fundación Jiménez Díaz (IIS-FJD, UAM), Avda. Reyes Católicos. rrincon23@gmail.com.
Raúl
R
Rincón
Madrid, Spain. jmadoz@fjd.es.
Jesús
J
García-Foncillas
Group of Cancer Biomarkers, Pathology Department, Health Research Institute Fundación Jiménez Díaz (IIS-FJD, UAM), Avda. Reyes Católicos. frojo@fjd.es.
Federico
F
Rojo
D004252:DNA Mutational Analysis / Q000191:economics / Q000379:methods; D005260:Female; D059014:High-Throughput Nucleotide Sequencing / Q000191:economics / Q000379:methods; D006801:Humans; D007890:Leiomyosarcoma / Q000175:diagnosis / Q000235:genetics*; D008297:Male; D009154:Mutation; D017354:Point Mutation; D018719:Erb-b2 Receptor Tyrosine Kinases / Q000235:genetics*; D020893:Receptor, ErbB-3 / Q000235:genetics*; D012186:Retroperitoneal Neoplasms / Q000175:diagnosis / Q000235:genetics*; D012187:Retroperitoneal Space / Q000473:pathology
D018719:Erb-b2 Receptor Tyrosine Kinases; D020893:Receptor, ErbB-3; C508053:ERBB2 protein, human; C581292:ERBB3 protein, human
HER2; HER3; cancer; pyrosequencing