PIK3CA mutations in the kinase domain (exon 20) of uterine endometrial adenocarcinomas are associated with adverse prognostic parameters

Abstract Background: Rhabdomyosarcoma (RMS) and lung adenocarcinoma (LADC) epitomizes the success of cancer prevention by the development of conventional therapy, but huge challenges remain in the therapy of advanced diseases.Case presentation: We reported two cases of novel BRAF gene fusion. The first case was a 34-year-old female with RMS harboring a BRAF-MAD1L1 fusion. She suffered tumor resection, recurrence and rapid progression. The second case was a 72-year-old female with LADC harboring a BRAF-ZC3H7A fusion, and she gained rapid progression after receiving a first-line course of chemotherapy.Conclusions: These two BRAF fusions retain the intact BRAF kinase domain (exon 11-18) and showed poor prognosis in RMS and LADC.

Download Full-text

Mutation-Associated Phenotypic Heterogeneity in Novel and Canonical PIK3CA Helical and Kinase Domain Mutants

Cells ◽

10.3390/cells9051116 ◽

2020 ◽

Vol 9 (5) ◽

pp. 1116

Author(s):

Arman Ali Ghodsinia ◽

J-Ann Marie T. Lego ◽

Reynaldo L. Garcia

Keyword(s):

Cellular Proliferation ◽

Acquired Resistance ◽

Kinase Domain ◽

Cellular Migration ◽

Regulatory Subunit ◽

Cytoskeletal Reorganization ◽

Apoptosis Resistance ◽

Pik3ca Mutations ◽

In Silico Docking ◽

Mutational Hotspots

Phosphatidylinositol 3-kinase, catalytic subunit alpha (PIK3CA) is an oncogene often mutated in colorectal cancer (CRC). The contribution of PIK3CA mutations in acquired resistance to anti-epidermal growth factor receptor (EGFR) therapy is well documented, but their prognostic and predictive value remain unclear. Domain- and exon-specific mutations are implicated in either favorable or poor prognoses, but there is paucity in the number of mutations characterized outside of the mutational hotspots. Here, two novel non-hotspot mutants—Q661K in exon 13 and C901R in exon 19—were characterized alongside the canonical exon 9 E545K and exon 20 H1047R mutants in NIH3T3 and HCT116 cells. Q661K and E545K both map to the helical domain, whereas C901R and H1047R map to the kinase domain. Results showed variable effects of Q661K and C901R on morphology, cellular proliferation, apoptosis resistance, and cytoskeletal reorganization, with both not having any effect on cellular migration. In comparison, E545K markedly promoted proliferation, survival, cytoskeletal reorganization, migration, and spheroid formation, whereas H1047R only enhanced the first three. In silico docking suggested these mutations negatively affect binding of the p85 alpha regulatory subunit to PIK3CA, thereby relieving PIK3CA inhibition. Altogether, these findings support intra-domain and mutation-specific variability in oncogenic readouts, with implications in degree of aggressiveness.

Download Full-text

Significance of EGFR/HER2 Expression and PIK3CA Mutations in Giant Cell Tumour of Bone Development

BioMed Research International ◽

10.1155/2020/2931784 ◽

2020 ◽

Vol 2020 ◽

pp. 1-11

Author(s):

Raja Amri ◽

Slim Charfi ◽

Mohamed Jemaà ◽

Nabil Miled ◽

Fathia Slimi ◽

...

Keyword(s):

Giant Cell ◽

Giant Cell Tumour ◽

Bone Development ◽

Bone Tumour ◽

Growth Factor Receptor ◽

Kinase Domain ◽

Cell Tumour ◽

Pten Gene ◽

Pik3ca Mutations ◽

The Stability

Giant Cell Tumour of Bone (GCTB) is a rare bone tumour. Locally aggressive and recurrent, it might evolve into pulmonary metastases. Our present work is aimed at investigating the involvement of the epidermal growth factor receptor (ErbB) family and its downstream effectors in the development and recurrence of GCTB. For this purpose, we used a cohort of 32 GCTB patients and we evaluated the clinicohistological features and the expression of RANKL, EGFR, and HER2. The mutation status of KRAS, PI3KCA, and PTEN gene as potential oncogene involved in GCTB was also evaluated. We found a significant correlation between advanced histological stages, overexpression of EGFR/HER2, and tumour recurrence. Moreover, two mutations were found in the PIK3CA gene: a missense mutation, 1634A>C, detected for the first time in GCTB patients, without influencing the stability of the protein, and a frameshift mutation, c.1658_1659delGTinsC, causing the loss of the protein kinase domain. Altogether, these results suggest that overexpression of HER2/EGFR, Campanacci, and histological stages could be used as a novel prognostic marker for GCTB recurrence.

Download Full-text

Highly sensitive determination of PIK3CA exon 9 and 20 hotspot mutations in breast tumors.

Journal of Clinical Oncology ◽

10.1200/jco.2012.30.15_suppl.tps10634 ◽

2012 ◽

Vol 30 (15_suppl) ◽

pp. TPS10634-TPS10634

Author(s):

Alexandre Harle ◽

Nicolas Lozano ◽

Cyrielle Nicolaizeau ◽

Maeva Lion ◽

Marie Husson ◽

...

Keyword(s):

Breast Tumors ◽

Amplification Refractory Mutation System ◽

Breast Carcinomas ◽

Pik3ca Mutations ◽

Pi3k Inhibitors ◽

Highly Sensitive ◽

Exon 9 ◽

Hotspot Mutations ◽

Ductal Carcinomas ◽

Exon 20

TPS10634 Background: Hotspot mutations in exons 9 (E542K and E545K) and 20 (H1047L and H1047R) of PIK3CA gene encoding for the p110α catalytic subunit of Phosphatidylinositol 3-kinases (PI3K) are found in approximately 25% of breast carcinomas. PIK3CA mutations cause the overactivation of PI3K/AKT/mTOR pathway and lead to resistance to anti-HER2 targeted therapies in breast cancers, are involved in response to anti-mTOR agents and are proposed as molecular diagnosis marker for PI3K inhibitors. A highly sensitive method is required to detect PIK3CA mutations in paraffin-embedded tumor tissues. Methods: We developed a real-time PCR assay using allele-specific scorpion primers and amplification refractory mutation system (PCR-ARMS) to detect hotspot mutations in exons 9 (E542K and E545K) and 20 (H1047L and H1047R) of PIK3CA of PI3K. Results: PIK3CA mutations were analyzed in 102 paraffin embedded breast tumors (88 invasive ductal carcinomas and 14 lobular ductal carcinomas). All specimens were validated by pathologist examination and processed for DNA extraction and PCR. PIK3CA exon 9 and 20 mutations were found in 22.5% of the specimens, 39.1% in exon 9 (26.1% for E542K, 13.0% for E545K) and 60.9% in exon 20 (17.4% for H1047L and 43.5% for H1047R). These results were comparable to the literature data (χ²=0.327 ; p<0.05). PCR ARMS was found to be highly sensitive, able to detect 0.5 % mutated DNA. Conclusion: PCR ARMS assay is highly sensitive for PIK3CA exon 9 and exon 20 hotspot mutations analysis in breast carcinomas as molecular marker for anti-HER2 and PI3K inhibitors response prediction.

Download Full-text

PIK3CA mutations in primary HER2-positive and triple negative breast cancer.

Journal of Clinical Oncology ◽

10.1200/jco.2013.31.15_suppl.11061 ◽

2013 ◽

Vol 31 (15_suppl) ◽

pp. 11061-11061

Author(s):

Sibylle Loibl ◽

Carsten Denkert ◽

Sherene Loi ◽

Fabrice Andre ◽

Berit Mueller ◽

...

Keyword(s):

Breast Cancer ◽

Hormone Receptors ◽

Triple Negative ◽

Response To Therapy ◽

Her2 Positive ◽

Pik3ca Mutations ◽

Formalin Fixed Paraffin Embedded ◽

Exon 9 ◽

Exon 20 ◽

Reported Data

11061 Background: Phosphatidylinositol 3-kinase (PI3K)/AKT pathway aberrations are common in breast cancer (BC), PIK3CA mutations being the most common. Mutations are frequently found in hot-spots located in the helical and kinase domains (exons 9 and 20). Reported data is discrepant with regard to prognostic or predictive value of PIK3CA mutations especially in HER2+ve BC. We therefore investigated the frequency and prognostic associations of PIK3CA mutations in HER2+ve and triple negative (TN) primary BC by treated with neoadjuvant therapy. Methods: We prospectively evaluated PIK3CA mutations in the 595 participants of the neoadjuvant Geparsixto study (NCT01426880). The study investigates the effect of adding carboplatin to a liposomal doxorubicin/taxane combination for the treatment of patients with HER2+ve and TN primary BC. All HER2+ve patients received trastuzumab and lapatinib, the TN patients received bevacizumab. HER2, hormone receptors (HR), and Ki67 were centrally assessed. PIK3CAwas genotyped in tumor material from formalin-fixed, paraffin embedded core biopsies taken before therapy using classical Sanger sequencing of exon 9 and 20. Results: From 09/2011 to 11/2012, 595 patients with HER2+ve or TN primary BC have been randomized in the Geparsixto study. Median age was 47 years (range 21-78); most tumors were cT2 (65%); cN0 (57%); ductal invasive (93%), grade 3 (65%); within the HER2+ve group 28% were HR positive. So far, PIK3CA genotype was evaluated in 395 randomized patients - 176 with HER2+ve and 219 with TN disease. Overall, 11.1% were found to have at least one mutation, in HER2+ve: 18.2% and TN BC: 5.5%. An exon 9 mutation was detected in 6.3% of the HER2+ve and 2.7% of the TNBC cases and an exon 20 mutation in 11.9% of the HER2+ve and 3.6% of the TN cases. A mutation in both exons was detected in 2 TN cases. PIK3CAmutations were more frequent in the HER2+ve/HR+ve compared to the HER2+ve/HR-ve group: 22.8% vs 10.6% respectively (p=0.047). Conclusions: PIK3CAmutations are more frequent in HER2+ve then in TN BC which is in line with previous reports. Results on all 595 patients and the correlation with response to therapy (pCR) will be presented at the meeting. The project has been funded within the EU-FP7 project RESPONSIFY No 278659. Clinical trial information: NCT01426880.

Download Full-text

Frequency of PIK3CA mutations in HER2+ surgically resected breast cancers in a Hispanic cohort.

Journal of Clinical Oncology ◽

10.1200/jco.2012.30.15_suppl.e11066 ◽

2012 ◽

Vol 30 (15_suppl) ◽

pp. e11066-e11066

Author(s):

Carlos Munive ◽

Aly Gallo ◽

Silvia P. Neciosup ◽

Franco Doimi ◽

Richard Dyer ◽

...

Keyword(s):

Targeted Therapy ◽

Pik3ca Mutation ◽

Breast Tumors ◽

Breast Cancers ◽

Pik3ca Gene ◽

Pik3ca Mutations ◽

Mutational Status ◽

Pi3k Signaling Pathway ◽

Exon 9 ◽

Exon 20

e11066 Background: PI3K signaling pathway is responsible for balancing cell survival and apoptosis, and plays an important role in the pathogenesis and progression of human breast cancers. Alterations in this pathway could influence the response to targeted therapy. Our aim is to know the frequency of mutations in the PIK3CA gene in HER2+, operable breast tumors in an Andean population. Methods: We include a cohort of 59 patients with HER2+ surgically resected breast cancer that received adjuvant anti-HER2 targeted therapy at the Instituto Nacional de Enfermedades Neoplásicas. Genomic DNA was extracted from Paraffin embedded-formalin fixed tumor samples and mutational status of PIK3CA gene was using PCR-based DNA sequencing Results: At the current date, the median of follow-up in our cohort is 2.4 years and two recurrences have been registered. Four samples were not evaluable for molecular analysis. Mutation in PIK3CA gene was detected in 12 cases (20.3%). In cases where PIK3CA mutation was detected, eight cases (66.7%) had mutations in exon 9 (A1634C/E545A in 6 cases; G1624A/E542K in 1 case and G1633A/E545K in 1 case). Four cases (33.3%) had exon 20 mutated (A3140G/H1047R in all cases). There was not coexistence of mutations in both exons. One case of tumor recurrence had mutation in exon 9. Conclusions: We found similar rates of PIK3CA mutations as described previously to HER2+, breast tumors. A longest follow upwill show the prognostic value of PIK3CA status in our cohort treated with targeted anti-HER2 therapy. [Table: see text]

Download Full-text

Rapid identification of PIK3CA-mutations in lung cancer by using pyrosequencing.

Journal of Clinical Oncology ◽

10.1200/jco.2012.30.15_suppl.e18000 ◽

2012 ◽

Vol 30 (15_suppl) ◽

pp. e18000-e18000

Author(s):

Verena Schildgen ◽

Jessica Lusebrink ◽

Walburga Engel-Riedel ◽

Erich Stoelben ◽

Oliver Schildgen ◽

...

Keyword(s):

Lung Cancer ◽

Lung Tumor ◽

Rapid Identification ◽

Pik3ca Gene ◽

Pik3ca Mutations ◽

Large Patient ◽

Oncogenic Mutations ◽

Cell Processes ◽

Exon 9 ◽

Exon 20

e18000 Background: Phospatidylinositol-3-kinases (PI3K) play an important role in various cell processes. Oncogenic mutations in the PIK3CA gene which codes for the catalytic subunit have been identified in various malignancies and activate the PI3K/AKT/mTOR pathway which is a critical driver of tumorigenesis. Methods: We tested 41 tumor samples with known KRAS, BRAF, and EGFR mutation status for the occurence of mutations in the PIK3CA gene using a new commercial pyrosequencing kit. Results: Pyrosequencing revealed 2 mutations (4.9 %) in the PIK3CA gene, one in exon 9 and one in exon 20. Both mutations have not yet been identified in lung tumor tissue. Conclusions: The screening of our small patient cohort by pyrosequencing identified two mutations (4.8 %) in PIK3CA, on in exon 9 (Q546H) and on in exon 20 (M1043T). Both mutations have not yet been described in lung tumours and seem to be rather uncommon mutations. Future Screening of large patient cohorts with pyrosequencing may contribute to the detection of more mutations in lung cancer due to the low limit of detections of this method and may contribute to a better understanding of the interaction of mutations and tumorigenesis.

Download Full-text

Sensitivity for detecting PIK3CA mutations in early-stage breast cancer with droplet digital PCR.

Journal of Clinical Oncology ◽

10.1200/jco.2013.31.15_suppl.11019 ◽

2013 ◽

Vol 31 (15_suppl) ◽

pp. 11019-11019 ◽

Cited By ~ 1

Author(s):

Julia A. Beaver ◽

Sasidharan Balukrishna ◽

Danijela Jelovac ◽

Michaela Jane Higgins ◽

Stacie Jeter ◽

...

Keyword(s):

Breast Cancer ◽

Digital Pcr ◽

Droplet Digital Pcr ◽

Wild Type ◽

Pik3ca Mutations ◽

Tumor Specimen ◽

Ffpe Samples ◽

Exon 9 ◽

Hotspot Mutations ◽

Exon 20

11019 Background: PIK3CA is mutated in up to 30% of breast cancers. Classically somatic mutations are identified by Sanger sequencing of the primary tumor specimen. However third generation droplet digital PCR technologies offer a novel platform for quantitative mutation detection with improved sensitivity. Methods: Thirty stage I-III breast cancer patients were consented on an IRB-approved prospective repository study at Johns Hopkins for collection of their primary breast tumor specimen. Formalin-fixed paraffin embedded (FFPE) samples were analyzed by standard sequencing for three PIK3CA hotspot mutations. The DNA from these samples was then analyzed using the RainDrop digital PCR platform with TaqMan probes in a triplex format to simultaneously detect and quantitate hotspot mutations and genome equivalents. Results are expressed as a percentage of mutant to wild-type PIK3CA molecules for each sample. Results: Standard sequencing of all tumors (n=30) identified seven PIK3CA Exon 20 mutations (H1047R) and three Exon 9 mutations (E545K). Samples were scored as PIK3CA mutation positive by digital PCR if the tumor DNA contained at least 5% mutant molecules. All ten mutations identified by sequencing were verified by digital PCR with quantities of mutant molecules ranging from 20.3-55.6% in a given sample. Digital PCR identified additional PIK3CA mutations that were wild type by standard sequencing including three mutant Exon 20 samples, two mutant Exon 9 samples and one sample with an Exon 20 and Exon 9 mutation. Quantities of mutant molecules in these additional samples ranged from 5-28.9%. Conclusions: RainDrop digital PCR offers improved sensitivity and quantification for detecting PIK3CA mutations in FFPE samples using nanograms of DNA. Additional mutations identified by digital PCR may reflect genetic heterogeneity or possibly tissue contamination. The clinical utility of identifying a small proportion of mutations is unknown but may impact eligibility for targeted therapies and clinical trials. Ongoing studies will also address whether the identification of solid tumor mutations in circulating cell-free plasma DNA by digital PCR can improve diagnostics and aid in therapeutic decisions.

Download Full-text

Structural basis of the effect of activating mutations on the EGF receptor

eLife ◽

10.7554/elife.65824 ◽

2021 ◽

Vol 10 ◽

Author(s):

Ioannis Galdadas ◽

Luca Carlino ◽

Richard A Ward ◽

Samantha J Hughes ◽

Shozeb Haider ◽

...

Keyword(s):

Growth Factor ◽

Egf Receptor ◽

Growth Factor Receptor ◽

Kinase Domain ◽

Structural Basis ◽

Kinase Activation ◽

Activating Mutations ◽

Dimeric Interface ◽

Exon 20 ◽

Dynamics Simulations

Mutations within the kinase domain of the epidermal growth factor receptor (EGFR) are common oncogenic driver events in non-small cell lung cancer. Although the activation of EGFR in normal cells is primarily driven by growth-factor-binding-induced dimerization, mutations on different exons of the kinase domain of the receptor have been found to affect the equilibrium between its active and inactive conformations giving rise to growth-factor-independent kinase activation. Using molecular dynamics simulations combined with enhanced sampling techniques, we compare here the conformational landscape of the monomers and homodimers of the wild-type and mutated forms of EGFR _ELREA and L858R, as well as of two exon 20 insertions, D770-N771insNPG, and A763-Y764insFQEA. The differences in the conformational energy landscapes are consistent with multiple mechanisms of action including the regulation of the hinge motion, the stabilization of the dimeric interface, and local unfolding transitions. Overall, a combination of different effects is pronounced in the different mutants and leads to the observed aberrant signaling.

Download Full-text

The role of KRAS, BRAF, NRAS, and PIK3CA mutations as markers of resistance to cetuximab in chemorefractory metastatic colorectal cancer

Journal of Clinical Oncology ◽

10.1200/jco.2009.27.15_suppl.4020 ◽

2009 ◽

Vol 27 (15_suppl) ◽

pp. 4020-4020 ◽

Cited By ~ 1

Author(s):

D. Lambrechts ◽

W. De Roock ◽

H. Prenen ◽

J. De Schutter ◽

B. Jacobs ◽

...

Keyword(s):

Colorectal Cancer ◽

Metastatic Colorectal Cancer ◽

Objective Response ◽

Pik3ca Mutation ◽

Braf V600e ◽

Kras Mutations ◽

Exact Test ◽

Pik3ca Mutations ◽

Exon 20

4020 Background: KRAS mutations (MUT) negatively affect outcome after cetuximab (CTX) in metastatic colorectal cancer (mCRC). As only 40% of KRAS wild-type (WT) respond it is possible that other MUT, constitutively activating the Ras/Erk or PI3K/Akt pathways, are present in the non-responding KRAS WT. We analyzed the KRAS, BRAF, NRAS & PIK3CA MUT status in 276 chemorefractory CRC treated with CTX +- irinotecan and correlated the MUT status with outcome. Methods: KRAS codon 12,13, 61&146, BRAF V600E, NRAS codon 12&13, PIK3CA E542K, E545K, A, G, V (exon 9), H1047Y, R, L (exon 20), N345K, R88Q and Q546K MUT were evaluated on FFPE primary CRC using the Sequenom MALDI TOF MassArray system. A two- sided Fisher's exact test was used to evaluate the association between PIK3CA, KRAS, BRAF & NRAS MUT and objective response (OR). Progression-free (PFS) and overall survival (OS) were estimated by the Kaplan-Meier method. Results: 116/276 (42%) CRC had a KRAS MUT, 96% of which occurred in codon 12 or 13. KRAS WT was associated with OR (p<.0001), longer median PFS (p<.0001) and OS (p<.0001). 15/153 (9.8%) KRAS WT had a BRAF MUT. BRAF WT was associated with OR (p=.01), longer PFS (p<.0001) and OS (p=.007). 5/98 (5%) KRAS WT had an NRAS MUT and none of these showed OR. KRAS, BRAF and NRAS MUT were mutually exclusive. The combined KRAS/BRAF/NRAS WT state was associated with OR (p<.0001), longer PFS (p<.0001) and OS (p<.0001). 23/200 (12%) CRC carried a PIK3CA mutation: 5/39 (13%) of responders and 18/160 (11%) of non-responders (p=.781). Median PFS and OS were not associated with PIK3CA MUT state (p=.760 & p=.698) overall, nor in the KRAS/BRAF/NRAS WT subgroup (p=.946 & p=.509). 5/13 (38.5%) PIK3CA MUT KRAS/BRAF/NRAS WT CRC showed an OR. 13/107 (12%) of KRAS/BRAF/NRAS WT and 10/93 (11%) of KRAS/BRAF/NRAS MUT tumors harbored a PIK3CA MUT (p=.826). Conclusions: KRAS, BRAF & NRAS MUT are mutually exclusive and occur in at least 47% of CRC. Like KRAS WT, BRAF WT state of the primary is significantly associated with outcome in mCRC treated with CTX. The combined KRAS/BRAF/NRAS WT state is significantly associated with outcome. PIK3CA MUT occur independently of the KRAS/BRAF/NRAS MUT status. We cannot provide any evidence for a strong role of PIK3CA MUT as a marker in determining outcome to CTX. [Table: see text]

Download Full-text