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  Table of Contents  
CASE REPORT
Year : 2019  |  Volume : 56  |  Issue : 2  |  Page : 176-179
 

Concomitant RAS and BRAF mutation in colorectal cancer - A report of 7 cases


1 Department of Medical Oncology, Health Sciences University, Dr. A.Y. Ankara Oncology Hospital, Ankara, Turkey
2 Department of Medical Oncology, Hacettepe University Cancer Institute, Ankara, Turkey

Date of Web Publication2-May-2019

Correspondence Address:
Ozturk Ates
Department of Medical Oncology, Health Sciences University, Dr. A.Y. Ankara Oncology Hospital, Ankara
Turkey
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Source of Support: None, Conflict of Interest: None


DOI: 10.4103/ijc.IJC_430_18

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 » Abstract 


Rat sarcoma viral oncogene homolog (RAS) and B-Raf murine sarcoma viral oncogene homolog B1 (BRAF) are members of the same signaling pathway (RAS-RAF-mitogen-activated protein kinase (MAPK) in colorectal cancer (CRC). It is generally assumed that BRAF mutations are seen only with wild-type RAS in CRC. But RAS and BRAF are not mutually exclusive. We have identified concomitant BRAF and RAS mutations in seven patients. DNA was extracted from formalin-fixed paraffin-embedded tumor tissue and the mutation status of the RAS gene (exons 2, 3, 4) and BRAF (exon 15 V600, V597) was assessed using a polymerase chain reaction enzyme-linked mini sequence assay-based DNA sequencing method. Three patients harbored Kirsten rat sarcoma viral oncogene homolog (KRAS) with a codon 13 mutation (gly13asp) along with a BRAF variation of L597V in exon 15 (p. leu597val, c.1789C>G (CTA>GTA). Two patients harbored KRAS with codon 12 mutations; one harbored the gly12val mutation with a variation of leu597val in the BRAF exon 15 codon, the other harbored a gly12asp mutation with p. leu597val, c.1789C>G (CTA>GTA) in the BRAF exon 15 codon. One patient harbored a codon 117 mutation with a BRAF V600E mutation. The last patient harbored a NRAS exon 2 (gly12asp) mutation with the GGT/GAT, V600G mutation in the BRAF exon 15 codon. Consequently, concomitant KRAS and BRAF mutations are very rare. Although it is known that the survival of concomitant RAS/BRAF mutation carriers is generally poor, we have shown that survival of concomitant RAS/BRAF mutation carriers is variable.


Keywords: BRAF mutation, colorectal cancer, RAS mutation


How to cite this article:
Ates O, Yalcin S. Concomitant RAS and BRAF mutation in colorectal cancer - A report of 7 cases. Indian J Cancer 2019;56:176-9

How to cite this URL:
Ates O, Yalcin S. Concomitant RAS and BRAF mutation in colorectal cancer - A report of 7 cases. Indian J Cancer [serial online] 2019 [cited 2019 May 23];56:176-9. Available from: http://www.indianjcancer.com/text.asp?2019/56/2/176/257558





 » Introduction Top


The prognosis of metastatic (m) colorectal carcinoma (CRC) is poor. The median overall survival (OS) has been extended up to 30 months with new targeted drugs. Target treatment has addressed two pathways, the antiepidermal growth factor receptor (EGFR) and vascular endothelial growth factor pathways. Vascular endotelial growth factor-A (VEGFA) is secreted by hypoxic tumor cells, especially in Kirsten rat sarcoma viral oncogene homolog (KRAS) mutant patients. Bevacizumab is a monoclonal antibody that blocks VEGFA in the plasma. The other targeted therapeutic agents are cetuximab and panitumumab, which are monoclonal antibodies against EGFR on tumor cells.[1] Ligand binding to the extracellular domain leads to allosteric activation via receptor dimerization and tyrosine kinase transphosphorylation, thus activating the RAS/MAPK pathway (RAS-RAF-MAPK) and the phosphor inositide 3-kinase (PI3K) pathway (PI3K-phosphatase and tensin homologue-Akt).[2] The antibodies which are active with wild-type KRAS in mCRC inhibit EGFR on the tumor cell with the intracellular downstream pathways of RAS-RAF-MAPK and blockage of PI3K-PTEN-AKT. But the benefit of EGFR blockade could recently not be shown for some KRAS wild-type mCRC patients.[3],[4] The reason may be the fact that there were mutations in BRAF, PI3K, and neuroblastoma RAS (NRAS).[2],[5] BRAF is a downstream molecule of EGFR and the prognosis of BRAF mutant mCRC is poor.[6],[7],[8],[9],[10],[11] The BRAF mutation in mCRC is seen in 5–10% of cases more commonly in proximal colon and in female patients;[12] it is known that the BRAF mutation is only seen in patients with RAS wild-type CRC.[10] BRAF and RAS are not mutually exclusive.

In the present case series, we evaluated the association of RAS and BRAF mutations with clinicopathological features in CRC patients in the Turkish population and their potential roles as prognostic biomarkers.


 » Case Series Top


We reviewed a total of 600 samples tested in the clinical laboratory for RAS and BRAF mutations from 2009 to 2015. We elucidated concomitant KRAS and RAF mutations in seven patients (1%). We identified these cases randomly. Written consent was given by the patients for their information to use for research. Mutations were analyzed by pathologists experienced in gastrointestinal tumors. DNA was extracted from formalin-fixed paraffin-embedded tumor tissue and the mutation status of KRAS, NRAS (exons 2, 3, 4), and BRAF mutation (exon 15 V600, V597) was assessed using a polymerase chain reaction (PCR)-enzyme-linked mini sequence assay-based DNA sequencing method.[13] For detection of mutations, the protocol supplied by the manufacturer of PCR products was applied using the ABI PRISM SNaPshot Multiplex Kit (Applied Biosystems).

Median diagnosis age was 58 (52-61) years. Five of these patients were males. Three patients had right colon carcinoma, three patients had rectal carcinomas, and one patient had left colon carcinoma [Table 1]. All of the patients harbored concomitant RAS and BRAF mutations. The KRAS codon 13 mutation (gly13asp) was seen in three patients with BRAF variations in exon 15 (p. leu597val, c.1789C>G (CTA>GTA). Two patients had a KRAS mutation in codon 12, one of them harbored GGT/GTT, gly12val and the other patient harbored gly12asp mutation with a variation of in the BRAF exon 15 codon [Table 1]. The BRAF V600E mutation was seen only in the sixth patient, who harbored the KRAS codon 117 mutation. The last patient harbored a NRAS exon 2 (gly12asp) mutation with the GGT/GAT, V600G mutation in the BRAF exon 15 codon. Most of patients had metastasis, except for one patient (patient 2). The median OS was 47 months.
Table 1: Demographics and clinical features of the patients

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 » Discussion Top


In the present study, we have identified seven cases who harbored concomitant RAS/BRAF mutations. Most of our patients (five of them) were operated. Median OS is 47 months.

Over the last few decades, research into CRC genomics and epigenetics has significantly advanced. Three distinct pathways abnormalities have been described for the development of CRC: chromosomal instability (CIN), microsatellite instability (MSI), and CIMP pathways.[14] CIN, which would be loss or gain of a whole chromosome, occurs as a result of cumulative mutations of oncogenes and tumor suppressor genes. The second model is MSI, which is seen only in 15% of CRC, causing mutations via DNA mismatch repair genes (MMR). Defective MMR can develop from inherited germline mutations in MMR genes (MLH1, MSH2, MSH6, PMS2) associated with Lynch syndrome. Half of patients with sporadic defective MMR carry a BRAF oncogene mutation. Proximal tumors, BRAF mutations, mucinous and poorly differentiated adenocarcinoma in female patients are prone to MSI,[15],[16],[17],[18] whereas distal tumors, which are more common in men than women, are characterized by loss of heterozygosity and mutation of TP53.

KRAS is an oncogene that encodes a protein with membrane bound GTP. KRAS mutations occur in exon 2 (codon 12 and 13), exon 3 (codon 59 and 61), and exon 4 (codon117, 146) in CRC and activate the MAPK pathway.[19],[20] KRAS mutations are seen in many parts of the colon at various rates (35–45%). In addition, other RAS mutations, NRAS exon 2-3-4 or KRAS exon 3-4, are seen in 10% of wild-type cases of KRAS exon 2. In a trial in the Turkish population, the KRAS mutation frequency was 35% and 30% in recto sigmoid and colon cancer, respectively.[21] In the TURKRAS study, a KRAS mutation was detected in 45% of cases.[22] There is a significantly higher KRAS mutation frequency in female patients compared to male patients. G12D and G12V mutations are the most common mutations, with rates of 28% and 25.9%, respectively. The most common KRAS mutation was the codon 12 (gly12Ala) mutation in that trial.[21] In our study, most patients harbored the codon 13 mutation (gly13asp). The codon 13 mutation is rare. In a far eastern study, where 676 patients were evaluated[16] KRAS codon 12 and 13 mutations were observed in 25.7 and 6.8% of cases, respectively. In another study evaluating 84 patients in total, mutations of KRAS codon 12 and 13 were found in 28.5% and 3% of cases, respectively.[21]

The BRAF gene is localized in 7q34 and consists of 18 exons. Usually, exon 15 and 11 mutations are observed. The BRAF mutation is more commonly seen in serrated and hyperplastic polyps, especially in proximal colon and in female patients.[12] The BRAF mutation in mCRC is seen in 5–10% of cases. It is known that the BRAF mutation is only seen in patients with RAS wild-type CRC.[10] BRAF and RAS are not mutually exclusive. The BRAF mutation is rarely seen in KRAS mutant CRC patients.[17],[19],[23],[24] Shen et al. determined the association of these mutations in 11 patients in their studies.[16] BRAF exon 15 and exon 11 mutations were seen in seven and four patients, respectively. In our study, patients had BRAF mutation at exon 15 [Table 1]. In another trial by Mao et al., the rate of KRAS, BRAF, and NRAS was 43%, 25%, and 8%, respectively.[17] They found that six patients harbored concomitant KRAS and BRAF mutations.

BRAF mutations are seen commonly in the right colon and rarely seen in the left colon and rectum (2%).[25] In our study, we defined concomitant KRAS and BRAF mutations, in which three patients had right colon cancer, three patients had left colon, and two had rectal cancer. The BRAF mutation is often seen in women and is associated with a poor prognosis. Patients with the BRAF V600 mutation usually respond poorly to standard therapy, with median PFS 2.5 months after second-line treatment and median survival of 12 months. The OS of patients with KRAS and BRAF mutations is generally poor.[26] But the survival of some patients was longer than in the literature in our study.

Consequently, concomitant KRAS and BRAF mutations are very rare. Although it is known that the survival of concomitant RAS/BRAF mutation carriers is generally poor, we have shown that survival of concomitant RAS/BRAF mutation carriers is variable.

Financial support and sponsorship

Nil.

Conflicts of interest

There are no conflicts of interest.



 
 » References Top

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Allegra CJ, Jessup JM, Somerfield MR, Hamilton SR, Hammond EH, Hayes DF, et al. American Society of Clinical Oncology provisional clinical opinion: Testing for KRAS gene mutations in patients with metastatic colorectal carcinoma to predict response to anti–epidermal growth factor receptor monoclonal antibody therapy. J Clin Oncol 2009;27:2091-6.  Back to cited text no. 1
    
2.
Laurent-Puig P, Cayre A, Manceau G, Buc E, Bachet JB, Lecomte T, et al. Analysis of PTEN, BRAF, and EGFR status in determining benefit from cetuximab therapy in wild-type KRAS metastatic colon cancer. J Clin Oncol 2009;27:5924-30.  Back to cited text no. 2
    
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Karapetis CS, Khambata-Ford S, Jonker DJ, O'Callaghan CJ, Tu D, Tebbutt NC, et al. K-ras mutations and benefit from cetuximab in advanced colorectal cancer. N Engl J Med 2008;359:1757-65.  Back to cited text no. 3
    
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Douillard J-Y, Siena S, Cassidy J, Tabernero J, Burkes R, Barugel M, et al. Randomized, phase III trial of panitumumab with infusional fluorouracil, leucovorin, and oxaliplatin (FOLFOX4) versus FOLFOX4 alone as first-line treatment in patients with previously untreated metastatic colorectal cancer: The PRIME study. J Clin Oncol2010;28:4697-705.  Back to cited text no. 4
    
5.
Sartore-Bianchi A, Di Nicolantonio F, Nichelatti M, Molinari F, De Dosso S, Saletti P, et al. Multi-determinants analysis of molecular alterations for predicting clinical benefit to EGFR-targeted monoclonal antibodies in colorectal cancer. PloS One2009;4:e7287.  Back to cited text no. 5
    
6.
Vaughn CP, ZoBell SD, Furtado LV, Baker CL, Samowitz WS. Frequency of KRAS, BRAF, and NRAS mutations in colorectal cancer. Genes Chromosomes Cancer2011;50:307-12.  Back to cited text no. 6
    
7.
Douillard J-Y, Oliner KS, Siena S, Tabernero J, Burkes R, Barugel M, et al. Panitumumab–FOLFOX4 treatment and RAS mutations in colorectal cancer. N Engl J Med2013;369:1023-34.  Back to cited text no. 7
    
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Van Cutsem E, Köhne C-H, Láng I, Folprecht G, Nowacki MP, Cascinu S, et al. Cetuximab plus irinotecan, fluorouracil, and leucovorin as first-line treatment for metastatic colorectal cancer: Updated analysis of overall survival according to tumor KRAS and BRAF mutation status. J Clin Oncol2011;29:2011-9.  Back to cited text no. 8
    
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Sartore-Bianchi A, Martini M, Molinari F, Veronese S, Nichelatti M, Artale S, et al. PIK3CA mutations in colorectal cancer are associated with clinical resistance to EGFR-targeted monoclonal antibodies. Cancer Res2009;69:1851-7.  Back to cited text no. 9
    
10.
Karapetis CS, Jonker D, Daneshmand M, Hanson JE, O'Callaghan CJ, Marginean C, et al. PIK3CA, BRAF, and PTEN status and benefit from cetuximab in the treatment of advanced colorectal cancer—results from NCIC CTG/AGITG CO. 17. Clin Cancer Res2014;20:744-53.  Back to cited text no. 10
    
11.
Yokota T, Ura T, Shibata N, Takahari D, Shitara K, Nomura M, et al. BRAF mutation is a powerful prognostic factor in advanced and recurrent colorectal cancer. Br J Cancer 2011;104:856-62.  Back to cited text no. 11
    
12.
Nagasaka T, Koi M, Kloor M, Gebert J, Vilkin A, Nishida N, et al. Mutations in both KRAS and BRAF may contribute to the methylator phenotype in colon cancer. Gastroenterology 2008;134:1950-60.e1951.  Back to cited text no. 12
    
13.
Lurkin I, Stoehr R, Hurst CD, van Tilborg AA, Knowles MA, Hartmann A, et al. Two multiplex assays that simultaneously identify 22 possible mutation sites in the KRAS, BRAF, NRAS and PIK3CA genes. PLoS One2010;5:e8802.  Back to cited text no. 13
    
14.
Worthley DL, Leggett BA. Colorectal cancer: Molecular features and clinical opportunities. Clin Biochem Rev2010;31:31-8.  Back to cited text no. 14
    
15.
Gray R, Barnwell J, McConkey C, Williams N, Kerr D. QUASAR: A randomised study of adjuvant chemotherapy versus observation including 3239 colorectal cancer patients QUASAR Collaborative Group. Lancet2007;370:2020-9.  Back to cited text no. 15
    
16.
Shen Y, Wang J, Han X, Yang H, Wang S, Lin D, et al. Effectors of epidermal growth factor receptor pathway: The genetic profiling of KRAS, BRAF, PIK3CA, NRAS mutations in colorectal cancer characteristics and personalized medicine. PLoS One 2013;8:e81628.  Back to cited text no. 16
    
17.
Mao C, Zhou J, Yang Z, Huang Y, Wu X, Shen H, et al. KRAS, BRAF and PIK3CA mutations and the loss of PTEN expression in Chinese patients with colorectal cancer. PloS One 2012;7:e36653.  Back to cited text no. 17
    
18.
Sahin IH, Kazmi SM, Yorio JT, Bhadkamkar NA, Kee BK, Garrett CR. Rare though not mutually exclusive: A report of three cases of concomitant KRAS and BRAF mutation and a review of the literature. J Cancer2013;4:320.  Back to cited text no. 18
    
19.
Scheffzek K, Ahmadian MR, Kabsch W, Wiesmüller L, Lautwein A, Schmitz F, et al. The Ras-RasGAP complex: Structural basis for GTPase activation and its loss in oncogenic Ras mutants. Science1997;277:333-9.  Back to cited text no. 19
    
20.
Trahey M, McCormick F. A cytoplasmic protein stimulates normal N-ras p21 GTPase, but does not affect oncogenic mutants. Science 1987;238:542-5.  Back to cited text no. 20
    
21.
Baskin Y, Dagdeviren YK, Calibasi G, Canda AE, Sarioglu S, Ellidokuz H, et al. KRAS mutation profile differences between rectosigmoid localized adenocarcinomas and colon adenocarcinomas. J Gastrointest Oncol2014;5:265-9.  Back to cited text no. 21
    
22.
Gumus M, Dane F, Karabulut B, Uygun K, Aydin K, Orhan B, et al. Results of observational study to determine K-ras mutation rates in 2458 patients with metastatic colorectal cancer in Turkey (TURKRAS Study). Am Soc Clin Oncol 2015. doi: 10.1200/jco. 2015.33.15_suppl.e14516.  Back to cited text no. 22
    
23.
Oliveira C, Velho S, Moutinho C, Ferreira A, Preto A, Domingo E, et al. KRAS and BRAF oncogenic mutations in MSS colorectal carcinoma progression. Oncogene2007;26:158-63.  Back to cited text no. 23
    
24.
Li HT, Lu YY, An YX, Wang X, Zhao QC. KRAS, BRAF and PIK3CA mutations in human colorectal cancer: Relationship with metastatic colorectal cancer. Oncol Rep2011;25:1691-7.  Back to cited text no. 24
    
25.
Hutchins G, Southward K, Handley K, Magill L, Beaumont C, Stahlschmidt J, et al. Value of mismatch repair, KRAS, and BRAF mutations in predicting recurrence and benefits from chemotherapy in colorectal cancer. J Clin Oncol2011;29:1261-70.  Back to cited text no. 25
    
26.
Roth AD, Tejpar S, Delorenzi M, Yan P, Fiocca R, Klingbiel D, et al. Prognostic role of KRAS and BRAF in stage II and III resected colon cancer: Results of the translational study on the PETACC-3, EORTC 40993, SAKK 60-00 trial. J Clin Oncol2009;28:466-74.  Back to cited text no. 26
    



 
 
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