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ORIGINAL ARTICLE
Year : 2014  |  Volume : 51  |  Issue : 1  |  Page : 35-39
 

Gastrointestinal stromal tumor: Analysis of outcome and correlation with c-kit status in Indian population


1 Medical and Pediatric Oncologist, Caritas Cancer Institute, Kottayam, Kerala, India
2 Department of Medical Oncology, Cancer Institute (WIA), Adyar, Chennai, India

Date of Web Publication18-Jun-2014

Correspondence Address:
S Cyriac
Medical and Pediatric Oncologist, Caritas Cancer Institute, Kottayam, Kerala
India
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Source of Support: Technical support from Oncquest Laboratories Ltd., Conflict of Interest: None


DOI: 10.4103/0019-509X.134616

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 » Abstract 

Aims: The aim of the present study is to analyse the outcome and genotypic pattern of metastatic GIST patients which is largely unknown in India. Materials and Methods: The present study was a retrospective analysis of 24 patients of metastatic GIST. The case records were analysed for clinical profile, treatment response and prognostic factors. The archival samples were retrieved for c-kit mutation analysis in all but 5 patients for mutation analysis. Results: The median age of the study population was 56 years. At a median follow up of 29 months, the PFS was 45% at 2 years. Activating c-kit mutations were detected in 10 cases (52.6%). 80% of the mutations were located in Exon 11. Conclusions: The outcome of metastatic GIST patients has definitely improved from a virtually incurable state to a disease where median OS has reached 60 months. The genotype of Indian patients with GIST may be different from the western population which needs to be confirmed in a larger study.


Keywords: Gastrointestinal stromal tumor, Imatinib, c-kit


How to cite this article:
Cyriac S, Rajendranath R, Sagar T G. Gastrointestinal stromal tumor: Analysis of outcome and correlation with c-kit status in Indian population. Indian J Cancer 2014;51:35-9

How to cite this URL:
Cyriac S, Rajendranath R, Sagar T G. Gastrointestinal stromal tumor: Analysis of outcome and correlation with c-kit status in Indian population. Indian J Cancer [serial online] 2014 [cited 2020 Dec 2];51:35-9. Available from: https://www.indianjcancer.com/text.asp?2014/51/1/35/134616



 » Introduction Top


Gastrointestinal Stromal Tumors (GIST) are the most common mesenchymal tumors of the gastrointestinal tract. [1] The molecular hallmark of entity was discovered only 10 years back. Approximately 80-90% of the GIST exhibit activating KIT mutations. [2] The prognosis of metastatic GIST was considered dismal and hardly any patient survived beyond 1-2 years. The discovery of Imatinib and its use against Chronic Myeloid Leukemia led to more researches for targeted therapy. [3] Subsequently, the ability of Imatinib to inhibit KIT and PDGFR was noted. In two large phase III studies comparing imatinib dose levels (400 mg/day vs. 800 mg/day), the median progression-free survival (PFS) for either arm was approximately 20 months, and median overall survival (OS) was approximately 50 months. [4],[5] So the fact that the natural history of GIST has changed was clear.

Metastatic GIST management has taken a new turn in our country also after the introduction of Imatinib. Support programs like GIPAP (Glivec International Patient Assistance Program) provide free drugs to all patients with metastatic GIST. There is hardly any data about the treatment outcome of metastatic GIST in India. It would be interesting to check whether our patients behave similar to the reported series. The genotype pattern in our population also is largely unknown. This study is conducted to address both these important issues.


 » Materials and Methods Top


The present study was a retrospective analysis of 24 patients of metastatic GIST initiated on Imatinib from January 2002 to December 2007. The case records were analysed for clinical profile, treatment response and prognostic factors. Patients received treatment with imatinib at the standard dose of 400 mg/day, which was continued until there was disease progression, unacceptable toxicity, or patient refusal. Hemogram was monitored at each visit and liver/renal function tests were monitored.

The response assessment was done by Ultrasound examination in many patients and hence the definitions could not be applied on a stringent basis. A partial response was defined as >50% reduction in the maximum diameter of lesion documented in original CT scan/Ultrasound and no appearance of new lesions. A complete response was defined as total disappearance of all lesions. Stable disease patients had 25-50% reduction in size of existing lesions. Progressive disease patients had to have new lesions or >25% increase in size of existing lesions.

Dose was increased in all patients documented to have progression. Toxicities were graded as per the National Cancer Institute Canada (NCIC) criteria.

All patients had histological confirmation with CD 117 expression by immunohistochemistry. The archival samples were retrieved for c-kit mutation analysis in all but 5 patients for mutation analysis. Informed consent was obtained from all the patients.

Tumor cells were collected from areas containing more than 60% malignant cells. DNA was extracted from the sample using Proteinase K digestion followed by silica cartridge isolation procedure. DNA was quantitated using Smartspec 3000 spectrophotometer (Bio-Rad). The quality of DNA was ascertained by resolving on a 0.8% TBE Gel. The purified sample was stored at -80°C till further processing and was then genomic DNA was amplified in two separate PCR reactions targeted at amplification of Exons 9 and 11 of C-KIT gene. The placement of the Primers was such that it ensured amplification of the complete exons. Briefly, for each reaction 30-50 ng of DNA was taken in a 0.2 ml PCR tube. To this was added, 200 nM each of Forward primer and Reverse primer and 12.5 ul of 2X Phusion Flash Master Mix (Finnzyme), in a total reaction volume of 25 ul. After an initial denaturation at 98°C for 3 min, the PCR was run for 40 cycles at conditions: 98°C for 1 min; 62°C for 1 min; 72°C for 1 min followed by a final extension at 72°C for 5 min. The amplicon was resolved on a 3% gel to ascertain the specificity of amplification. The product was purified using AxyPrep PCR cleanup Kit supplied by Axygen as per the manufacturer's instructions. Sequencing of PCR products was carried out in both directions on ABI 3130 × l Genetic Analyzer using standard chemistries. The sequences were compared against human genome sequence (NCBI accession NT_022853) using BLAST software to identify the mutations. The chromatograms were also analyzed manually using SeqScape v2.6 software.

SPSS version 13.0 (SPSS Inc) was used for statistical analysis. Log rank test was used for univariate analysis. Cofactors investigated in the analysis included age, gender, performance status, primary site of disease, previous treatment, tumor size, sites of metastases, baseline laboratory parameters, time from diagnosis to start of Imatinib, presence of bleeding at presentation, anemia and tumor genotypes. Multivariate analysis was not attempted in view of the limited patient population. C-kit genotype relationship with outcome was assessed for only 19 patients for whom the genotypic information was available. Kaplan Meier survival plot was used for estimating the progression free survival (PFS) and Overall survival (OS). PFS was measured from the first day of imatinib treatment to disease progression or death resulting from any cause and OS was measured from the first day of treatment to death resulting from any cause.


 » Results Top


The median age of the study population was 56 years (Range 26 - 76 years). The male: female ratio was 3:1. Other patient details are described in [Table 1].

Twelve patients (50%) had metastases at diagnosis. Metastases were delayed beyond 2 years of primary diagnosis in 8 patients. Surgery was offered to two cases after diagnosis of primary. Rest was managed only with Imatinib. Anemia was present in 12 (50%) of the patients. The median size of the primary tumor was 10 centimeters (cms) The median size of the metastatic tumor was 3 cms (range: 1.4 cms - 20 cms). Most of the patients had more than 2 metastatic lesions. Spindle cell subtype of GIST was present in 20 (83.3%) patients. One patient had mixed type and in other three, epitheliod variety was seen. The mitotic rate was >5/50 high power field (hpf) in 19 patients.
Table 1: Patient characteristics

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The median time to documentation of maximum initial response was 6 months. One patient had complete response. Partial response (PR) was seen in 6 (25%) patients. Stable disease was the predominant response and was seen in 10 patients. Seven patients progressed on Imatinib. However 5 of them died inspite of dose escalation. Of the remaining 19 patients, 14 had stable disease at the time of last follow up. Four patients had partial response and one maintained complete response status.

At a median follow up of 29 months, the PFS was 45% at 2 years. The OS was 78% at 2 years. Hence a significant proportion of patients responded to higher doses of Imatinib. The Kaplan Meier survival plots for PFS and OS are demonstrated in [Figure 1] and [Figure 2].
Figure 1: Kaplan Meier survival plot for PFS

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Figure 2: Kaplan Meier survival plot for OS

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Activating c-kit mutations were detected in 10 cases (52.6%). 80% of the mutations were located in Exon 11 and 20% were present in exon 9. Nine patients had a wild type of c-kit on analysis. The most common mutations described are deletions. Among all patients with exon 11 mutations, 6 had deletions. Two patients had substitution mutations. The two patients who had this type of mutations were found to have Val560 Asp and. Structurally, almost all exon 9 duplications are identical 1525_1530 dupGCCTAT leading to Ala502_Tyr503 dup at the protein level. Both of our patients with exon 9 mutations had similar finding. Interestingly both of them had intestinal GISTs. The distribution of ckit mutation in relation with site of disease and response to Imatinib has been mentioned in [Table 2]. Of all the factors analysed for overall survival, those found to be significant were presence of anemia (P = 0.006) and performance status (P = 0.02). Patients with liver metastases had a better PFS compared to other sites of metastases. The 2 year PFS was 80% vs 67% (P = 0.04). For PFS, anemia was found to be significant predictor (P = 0.02). The c-kit mutation status was also found to be significant predictor for PFS [Table 3]. 2 year PFS for patients with exon 11 mutation and wild genotype were 58% and 31% respectively (P = 0.03).

The percentage of patients responding to Imatinib (at least PR) was higher in patients with Exon 11 mutations (57.4%) compared to those with Exon 9 (0%) or those with wild type (20%). Both the patients with Exon 9 who progressed on Imatinib 400 mg were attempted 600 mg. Both patients progressed and expired later. A total of nine deaths occurred in the entire patient group. The c-kit status was unknown in 3 (33.3%) of these patients. 2 patients had either exon 11 or exon 9 mutation. However the majority of the deaths were in wild c-kit type.
Table 2: Correlation of c‑kit mutation status with outcome

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Table 3: Univariate analysis for factors predicting PFS

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Imatinib treatment was well tolerated by all patients. None of the patients required stoppage or dose reduction because of toxicity. None of them developed febrile episodes or dermatological toxicity other than hypopigmentation. Hepatic, cardiac or renal dysfunction was not reported in any patient. None of the patients had Grade 3 or Grade 4 toxicity with 400 mg per day dose. However when the dose was hiked to 600 mg per day, four patients complained of myalgia Grade 3 and increased dyspeptic symptoms.


 » Discussion Top


The discovery that GIST cells express KIT, a receptor tyrosine kinase (RTK) growth factor receptor has changed dramatically the management of GISTs. [1] KIT is not only expressed, but is mutated in 85-90% of cases leading to constitutive activation of the receptor. Exon 11 mutations are the most common, followed in frequency by exons 9, 13, and 17. [6] A subset of wild KIT tumors have mutations in platelet derived growth factor alpha (PDGFRA). The so-called wild-type (WT) GIST do not contain mutations in either KIT or PDGFRA. The development of effective targeted therapies using small molecules like imatinib and sunitinib that are tyrosine kinase inhibitors (TKI) is stemmed on the discovery of expression of KIT and the understanding that KIT or PDGFRA are constitutively activated in GIST.

The median age of our study group was 56 years which is similar to other reported studies. [7],[8] The age of presentation has not varied significantly across ethnic groups. However there was significant sex predilection with males 3 times more commonly affected than females in our group. This is more than what is mentioned in the literature. [8],[9],[10]

Stomach was the predominant site of primary disease followed by small intestine which was affected in 25% of the patients. This distribution is similar to what is described in literature. [1] Similarly we found that liver is the predominant site of, metastatic disease.

In our study, though the 2 year PFS was poor (41%), the overall survival was 78%. This should be possibly because the patients responded to higher dose of Imatinib. Blanke et al. also failed to show superiority of higher dose of Imatinib in inducing better responses. [8] After progression on standard dose imatinib, 33% of patients who crossed over to the high-dose imatinib regimen achieved either an objective response or stable disease.

Kim et al. analyzed the relationship between treatment outcome and kinase mutational status in 113 Korean patients with advanced GISTs treated with Imatinib. [7] KIT mutations were found in exon 11 (n = 92, 81.4%) and exon 9 (n = 10, 8.8%). One patient had a PDGFRA exon 18 mutation. The overall mutation rate was 91.2%. With a median follow-up of 49.0 months, the median progression-free survival (PFS) time was 42.0 months and median overall survival (OS) time was not reached. PFS and OS times did not differ significantly according to KIT genotype. They concluded that compared with previous studies in western populations, these results suggest that ethnic differences may influence the relationship between KIT genotype and clinical outcome to imatinib.

Yeh et al. assessed 54 GIST patients in Taiwan. [9] The mutation rate for KIT was 90.7%, and included 40 patients with KIT exon 11 mutations and nine patients with KIT exon 9 mutations. Although response rates to imatinib tended to be higher in patients with KIT exon 11 mutations (57.5%) or no kinase mutations (60.0%) than in those with KIT exon 9 mutations (22.2%), the OS of these three subsets did not differ. Similar to the above study, OS did not differ in our population also. The three mentioned studies have been contrasted against our study in [Table 4].
Table 4: Comparison of present study with similar studies

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In a nation-wide study in Iceland, Tryggvason G et al. evaluated all 55 GIST patients diagnosed between 1990 and 2004. [10] Mutations were found in 52 tumors representing a 92.9% mutational rate which is higher than in our study. Most of the mutations were found in c-kit exon 11 (76.8%), followed by c-kit exon 9 (10.7%) as expected. PDGFRA mutations were only found in three tumors. No correlation of mutation type with biologic behavior was found. This study again contrasts with our patient group in that the c-kit mutation rate is lower and the same predicted PFS.

Heinrich et al. analysed mutations of c-kit or PDGFR in 127 patients in a Phase II study. [11] Activating mutations of KIT or PDGFRA were found in 112 (88.2%) and six (4.7%) GISTs respectively. Exon 11 mutations were seen in 85 patients. Patients whose tumors contained exon 11 KIT mutations had a longer event-free and overall survival than those whose tumors expressed either exon 9 KIT mutations or had no detectable kinase mutation.

In the era on Imatinib therapy for GIST, various prognostic factors are described in literature for GIST. In our patient population, anemia and wild type c-kit mutation status predicted poor PFS. Patients with liver metastases had a better PFS compared to other sites of metastases. Heinrich et al. suggested that exon 11, poor performance status, Imatinib dose and unknown mutational status predicted outcome. [11] Zhu et al. also suggested that liver metastases is not a adverse prognostic factor in Imatinib era. [12] Kim et al. found that poor performance status is the predominant factor which predicted poor outcome. [7] Poor performance status, low albumin, male sex and high neutrophil count were significantly associated with worse outcome in a study by Blanke et al.[4]

It was interesting to note the type of mutations in our patient population. We have evaluated only for the two most common mutations, exon 11 and exon 9 mutations. Deletions are the most common mutations seen in GIST and almost always occur in exon 11. Of 8 patients who had exon 11 mutations, all except two had deletions. The most common deletion described is 1690_1695delTGGAAG (Trp557_Lys558del) which was seen in 5 of our patients. This particular mutation is associated with more malignant behavior especially in gastric GISTs. [13],[14] Single nucleotide substitutions are next most common mutation and are associated with more indolent behavior. The most common missense mutations identified in GISTs are Val559Asp, Val560Asp, Trp557Arg, Val559Ala, Val559Gly and Leu576 Pro. The two patients who had this type of mutations were found to have Val560 Asp and Trp557Arg.

Exon 9 mutations are well known for the occurrence in intestinal GIST (upto 90%) and also to the response to higher doses of GIST. We had two patients with exon 9 mutations. Duplications are the third most common KIT mutations in GISTs. This is more commonly seen with exon 9 mutations. Structurally, almost all exon 9 duplications are identical 1525_1530dupGCCTAT leading to Ala502_Tyr503dup at the protein level. Both of our patients with exon 9 mutations had similar finding. Interestingly both of them had intestinal GISTs.

The 2 year PFS in patients with exon 11 mutation was 58% in our study. However the same was only 16% in those patients who had a wild type of c-kit. This suggests that a mere sampling error cannot explain the poor outcome of those patients with wild type c-kit. Hence the genotype of our patient population may be truly different. Nearing 50% of our study population had wild type of mutations. This is very high compared to the existing literature. Hence this needs to be confirmed in a larger patient population. We did not have the facility to perform tests for rare mutations. The rare mutations like exon 13, exon 17, PDGFRA etc., cannot explain this low incidence of mutations. We have already initiated genotyping of all newly diagnosed tumors at our centre.

The study had some limitations. This study assessed the outcome of the patients in a retrospective manner. Hence the criticisms for a retrospective analysis are applicable for this study. The sample size for the present study was small. The genotype of c-kit mutation status was assessed on a Trucut biopsy or endoscopic biopsy in most of the patients as it was done on archival samples. We would prefer in our future studies to get enough specimen upfront for c-kit analysis. Second line agents are not freely available in our country and hence majority of the patients could not receive drugs like Sunitinib or new drugs like Nilotinib. Hence salvage was not possible in many patients.


 » Conclusions Top


The present study demonstrated the feasibility of performing c-kit mutation analysis in India. The outcome of metastatic GIST patients has definitely improved from a virtually incurable state to a disease where median OS has reached 60 months. The genotype of Indian patients with GIST may be different from the western population which needs to be confirmed in a larger study.

 
 » References Top

1.Demetri GD. Gastrointestinal stromal tumors. In: DeVita VT Jr, Hellman S, Rosenberg SA, editors. Cancer: Principles and Practice of Oncology, Philadelphia: Lippincott, Williams, and Wilkins; 2005. p. 1050-60.  Back to cited text no. 1
    
2.Rubin BP, Singer S, Tsao C, Duensing A, Lux ML, Ruiz R, et al. KIT activation is a ubiquitous feature of gastrointestinal stromal tumors. Cancer Res 2001;61:8118-27.  Back to cited text no. 2
    
3.Druker BJ, Tamura S, Buchdunger E, Ohno S, Segal GM, Fanning S, et al. Effects of a selective inhibitor of the Abl tyrosine kinase on the growth of Bcr-Abl positive cells. Nat Med 1996;2:561-70.  Back to cited text no. 3
    
4.Blanke CD, Rankin C, Demetri GD, Ryan CW, von Mehren M, Benjamin RS, et al. Phase III randomized, intergroup trial assessing imatinib mesylate at two dose levels in patients with unresectable or metastatic gastrointestinal stromal tumors expressing the kit receptor tyrosine kinase: S0033. J Clin Oncol 2008;26:626-40.  Back to cited text no. 4
    
5.Zalcberg JR, Verweij J, Casali PG, Le Cesne A, Reichardt P, Blay JY, et al. Outcome of patients with advanced gastro-intestinal stromal tumours crossing over to a daily imatinib dose of 800 mg after progression on 400 mg. Eur J Cancer 2005;41:1751-60.  Back to cited text no. 5
    
6.Lux ML, Rubin BP, Biase TL, Chen Cl, Maclure T, Demetri G, et al. KIT extracellular and kinase domain mutations in gastrointestinal stromal tumors. Am J Pathol 2000;156:791-800.  Back to cited text no. 6
    
7.Demetri GD, von Mehren M, Blanke CD, Abbeele AD, Eisenberg B, Roberts PJ, et al. Efficacy and safety of imatinib mesylate in advanced gastrointestinal stromal tumors. N Engl J Med 2002;347:472-7.  Back to cited text no. 7
    
8.Kim TW, Ryu M-H, Lee H, Sym SJ, Lee J-L, Chang HN, et al. Kinase Mutations and Efficacy of Imatinib in Korean Patients with Advanced Gastrointestinal Stromal Tumors. The Oncologist 2009;14:540-7.  Back to cited text no. 8
    
9.Yeh CN, Chen TW, Lee HL, Liu YY, Chao TC, Hwang TL, et al. Kinase Mutations and Imatinib Mesylate Response for 64 Taiwanese with Advanced GIST: Preliminary Experience from Chang Gung Memorial Hospital. Ann Surg Oncol 2007;14:1123-8.  Back to cited text no. 9
    
10.Trygvasson G, Hilmarsdottir B, Gunnarson GH, Jonsson JJ, Jonasson JG, Magnusson MK, et al. Tyrosine kinase mutations in gastrointestinal stromal tumors in a nation-wide study in Iceland. APMIS 2010;118: 648-56.  Back to cited text no. 10
    
11.Heinrich MC, Corless CL, Demetri GD, Blanke CD, von Mehren M, Joensuu H, et al. Kinase mutations and imatinib response in patients with metastatic gastrointestinal stromal tumor. J Clin Oncol 2003;21:4342-9.  Back to cited text no. 11
    
12.Zhu J, Yang Y, Zhou L, Jiang M, Hou M. A long term follow-up of the imatinib mesylate treatment for the patients with recurrent gastrointestinal stromal tumor (GIST): The liver metastasis and the outcome. BMC Cancer 2010;10:199-205.  Back to cited text no. 12
    
13.J Lasota, M Miettinen. Clinical significance of oncogenic KIT and PDGFRA mutations in gastrointestinal stromal tumours. Histopathology 2008;53:245-66.  Back to cited text no. 13
    
14.Eva W, Inge L, Volkmar H, Iris N, Nicola S, Erhad B, et al. Deletion of Trp - 557 and Lys - 558 in the juxtamembrane domain of the c-kit protooncogene is associated with metastatic behaviour of gastrointestinal stromal tumors. Int J Cancer 2003;106:887-95.  Back to cited text no. 14
    


    Figures

  [Figure 1], [Figure 2]
 
 
    Tables

  [Table 1], [Table 2], [Table 3], [Table 4]

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