Examining Gastrointestinal Stromal Tumors Biology Essay

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Gastrointestinal Stromal Tumors (GIST) are rare malignancies of the gastrointestinal tract. It has been labelled as the most common mesenchymal malignancy of the GIT. [1] The progress made in this relatively recently defined entity is fascinating. The better understanding about the molecular mechanisms of the disease made it possible to incorporate successfully targeted therapy into the treatment armamentarium. FDA approval of Imatinib (IM) for the management of metastatic GIST came first in 2002. Now IM is also used as an adjuvant treatment in an effort to improve the high recurrence rates. The numerous ongoing phase I/II trials aim at further improvement of the results and also using novel Tyrosine Kinase Inhibitors (TKI) for the management of GIST.

The annual incidence of GIST in United States is estimated around ~4000-5000 per year.[1] Before 2000, GIST was estimated to account for 1-3% of all GI malignancies. Prevalence must be higher as many patients live with small GISTs often detected at autopsy. It is not clearly explained why some of these small tumors do not manifest during the lifetime. There is no published data regarding the prevalence of GIST in India. The entity is described more clearly in the recent times as majority were labelled as leiomyosarcomas in the past and since immunohistochemistry is widely available.

Before 1999, the diagnostic criteria for GIST remained controversial and somewhat confusing. GIST was initially a purely descriptive term developed in 1983 by Mazur and Clark to define intra-abdominal tumors that were definitely not carcinomas and that also failed to exhibit features of either smooth muscle or nerve cells. [10] However, pathologists recognized that there was not a completely clear differential expression of muscle or nerve antigenic markers when careful immunohistochemical analyses were performed on certain mesenchymal tumors of the gut. Several names were applied to these tumors based on the variable patterns of cell lineage markers described by a variety of pathologists across the world.

GIST were often previously diagnosed as leiomyomas or leiomyosarcomas because of their histologic resemblance to those forms of smooth muscle neoplasms. Despite this, it had long been recognized that a subset of these tumors that arose in the bowel wall had a number of peculiar histologic features and likely represented a different entity altogether.[11] These GI tract leiomyosarcomas were also noted to be exceptionally resistant to standard chemotherapy regimens compared to those arising in other anatomic sites Additional differences between GISTs and leiomyosarcomas became apparent with the application of modern immunohistochemical techniques in the 1980s. By these assays, a significant number of these tumors were noted to lack the characteristic muscle antigens that defined leiomyosarcomas located elsewhere in the body. Other terms were generated based on the fact that neural crest antigens such as neuron-specific enolase and S-100 could be demonstrated in GIST cells, which led to the terms plexosarcomas and gastrointestinal autonomic nerve tumors. [1]

Additional research in immunohistochemical analysis of GISTs in the early 1990s revealed that a significant proportion of these tumors expressed the CD34 antigen. [12] This antigen is shared between hematopoietic stem cells as well as vascular and myofibroblastic cells. It was initially hoped that CD34 might prove to be a key differentiating feature between GISTs and other spindle cell tumors of the GI tract, such as schwannomas or leiomyomas. CD34 expression characterized only approximately half of all GIST cases, and a proportion of smooth muscle and Schwann cell tumors could also express CD34. Therefore, CD34 was neither a sensitive nor a specific marker to distinguish GIST from other mesenchymal neoplasms.

In 1998, Hirota et al described the expression of c-Kit protein as an incontestable feature of GISTs, further separating them from other gastrointestinal mesenchymal tumors.[2] At present, GIST is considered to be a specific category different from true smooth muscle tumors and neurogenic tumors. c-kit belongs to the class III receptor tyrosine kinases (RTKs), together with platelet-derived growth factor receptor- α (PDGFRA), colony-stimulating factor-1 receptor (CSF-1-R), vascular endothelial growth factor receptors 1 and 2 (Flt-1 and Flk-1, respectively), Flk-2, and Flt-4. The KIT and PDGFRA genes map to chromosome 4q12.[13] The RTKs are characterized by the presence of an extracellular domain, a transmembrane domain, a juxtamembrane domain, and an intracellular domain where the 2 kinase domains are lodged. [figure ]

Activation of c-kit occurs when by the binding of stem cell factor, the receptor homodimerizes and experiences conformational transformations that lead to the activation of the kinase domains. These, in turn, transphosphorylate the tyrosine residues of the opposing homodimerized receptor, allowing its association with substrates of various kinds. Like the majority of RTKs, c-kit has been attributed many physiologic functions such as cell survival, proliferation, differentiation, adhesion, and apoptosis by signaling through the MAP kinase, PI3-kinase, and JAK/STAT pathways.[13] c-kit signaling is essential for normal erythropoiesis, lymphopoiesis, gametogenesis, and melanogenesis and for the correct development and function of mast cells. A dysfunctional activation of this RTK, therefore, has been involved in diverse neoplasias such as mastocytosis/mast cell leukemia, germ cell tumors, small cell lung carcinoma, acute myeloid leukemia, neuroblastoma, melanoma, ovarian carcinoma, and breast carcinoma, besides GISTs. [1]

The oncogenic potential of mutant, uncontrollably active KIT in the pathogenesis of GIST in humans was further supported by the identification of a family that exhibited an autosomal dominant inheritance pattern of GIST. Genetic analysis of this kindred revealed that they harbored a germline activating KIT mutation, similar to the mutations that were seen in sporadic cases of GIST.[14] Often, these tumors may not present clinically until the second or third decade of life, and some even present in far advanced age. KIT mutations have also been documented in very small (less than 1 cm) GISTs that were detected incidentally and that appear morphologically benign. These findings support the hypothesis that activating mutations in the KIT protooncogene represent an early transforming mechanism in GIST oncogenesis. However, since many tumors harboring this mutation can remain small for years, there must be other key signaling steps that confer an aggressive and malignant phenotype to GIST cells. These other molecular pathways remain poorly understood. Unique elements of the downstream signaling cascades in GIST are being actively elucidated, and these appear to differ from KIT signaling in hematologic neoplasia in that the STAT5 pathway is not typically activated in GIST, whereas STAT1 and STAT3 are activated at a high level. [15]

In GISTs, KIT or PDGFRA mutations cause constitutive oncogenic signaling in the absence of their ligands. The uncontrolled RTK activity results in the activation of the PI3K-AKT and MEK-MAPK pathways accompanied by relatively low level signal transducer and activation of transcription (STAT)1 and STAT3 activation, leading to alterations in cell cycle, protein translation, metabolism, and apoptosis.[13]

There is a broad spectrum of c-Kit mutations in GISTs, ranging from 30% to 90%, and c-Kit status can constitute a prognostic factor for survival. Most of the mutations are located in the juxtamembrane domain (exon 11), followed by the extracellular domain (exon 9), and seldom are in the kinase domains (exon 13 and 17). About 35% of GISTs lacking c- kit mutations have intragenic activation mutations in PDGFRA, the most common, located in exons 12, 14, and 18. Mutations of c-Kit and PDGFRA seem to be mutually exclusive oncogenic events in GISTs.[1]

Various types of mutation can be found in exon 11 including missense mutations, insertions, and deletions. Tandem repeat mutations are infrequently seen in the distal part of KIT exon 11. These changes were predominantly found in female stomach and have been proposed to be associated with a quite indolent clinical course. More controversial are GISTs with deletion mutations in and around KIT exon 11 codon 557-558. Some studies have shown an aggressive clinical course and poor prognosis, but these findings have not been confirmed by others. Loss of heterozygosity in the KIT locus has been associated with high proliferative activity and increased metastatic potential.[16,17]

GISTs with an exon 9 mutation arise most commonly in the small intestine and are frequently high-risk tumors. In the vast majority of cases, exon 9 mutations are characterized by insertion of six base pairs, a duplication of Ala and Tyr and are found in primary as well as relapsed or advanced GISTs. According to a recent study, KIT exon 13 and exon 17 mutant GISTs are slightly overrepresented among the intestinal group of GISTs, and if tumors with an exon 13 mutation occur in the stomach, they tend to be slightly larger and more aggressive than "average"gastric GISTs. The majority of KIT exon 13 and 17 mutations are substitutions and, in small intestinal GISTs, these mutations have no substantial impact on clinicopathologic features when compared to the "average" small intestinal GIST.[16]

PDGFRA mutations, identified in approximately 8% of GISTs, involve mainly (6-7%) either exon 18 (kinase activation loop) or exon 14 (ATP-binding pocket) and rarely (less than 1%) exon 12 (JM). Mutations in PDGFRA exon 14 and 18 are mostly missense mutations. The subset of GISTs with a PDGFRA mutation that is associated with a commonly benign clinical course is limited to the stomach and omentum, lack KIT expression by immunohistochemistry (IHC), and preferentially shows epithelioid morphology. GISTs with a mutation D842V in exon 18 of PDGFRA are resistant to imatinib and sunitinib.[16]

The tumor predominantly affects adults at a median age of 58 years with a slight male preponderance. GISTs occur throughout the GI tract and are most commonly seen in the stomach (60%), jejunum and ileum (30%), duodenum (5%), colorectum (4%), and rarely the esophagus and appendix. [1] Tumors lacking any association with the bowel wall are known as extra gastrointestinal stromal sarcomas and more often occur in the omentum, mesentery, or retroperitoneum.

Clinical symptoms associated with GIST include abdominal pain, fatigue, dysphagia, satiety, and obstruction. Patients may present with chronic GI bleeding which leads to anemia or acute GI bleeding which may be caused by erosion through the gastric or bowel mucosa or rupture into the abdominal cavity causing life-threatening intraperitoneal hemorrhage. Most of the symptomatic tumors were more than 5 cms in size. Small GISTs mainly present as incidental findings during endoscopy, surgery, or radiologic studies for other reasons, whereas patients with malignant GIST often present with disseminated disease. Vast majority of the metastases at presentation are intraabdominal, either to the liver, omentum and peritoneal cavity.

Heritable mutations in KIT and PDGFRA have been reported in some families.[14] The penetrance of these mutations is quite high, and most affected family members will develop solitary or multiple GISTs during their life span. The mean age of onset is younger than that of sporadic GISTs without gender differences. Most of these GISTs follow a benign course, and their morphology does not differ from that of their sporadic counterparts. GISTs are part of Carney's triad (gastric GIST, paraganglioma, and pulmonary chondroma) and Carney's dyad (paraganglioma, gastric GIST), and these GISTs are KIT/PDGFRA wild-type.[18] The genetic basis for Carney's triad is not known, although it is thought to be sporadic rather than familial. In both conditions, the presence of multiple gastric GISTs is common. The clinical features of the triad include occurrence at a young age, female predilection, tumor multifocality, slow growth, frequent metastasis, lack of response to imatinib treatment, and sometimes fatal outcome. They often affect the gastric antrum, show predominant epithelioid histology and lack the classical molecular abnormalities.

Approximately 1-2% of GISTs occur in the pediatric age group, predominantly in the second decade. Pediatric GISTs are associated with a marked female predominance, are preferentially located in the stomach, and show mainly epithelioid morphology. Although these tumors consistently express KIT protein, the majority lack KIT or PDGFRA mutations. Unlike adult GISTs, these tumors quite often spread to lymph nodes. Interestingly, pediatric KIT wild-type GISTs lack the typical cytogenetic deletions seen in adult KIT-mutant GISTs and progress to malignancy without acquiring large-scale chromosomal aberrations. The difference between pediatric KIT wild-type and adult GISTs of the stomach is further demonstrated by their separate clustering by gene expression profiling, and it is very likely that these tumors are a separate clinicopathologic entity.[1,19]

In this patient group, time to tumor progression was significantly longer on sunitinib than on prior imatinib treatment, indicating that this patient group could benefit from sunitinib as first-line treatment.[20] Sometimes pediatric GISTs are associated with pulmonary chondromas and/ or paragangliomas referred to as Carney's triad. In the pediatric age group, Carney's triad should be considered in any patient with GIST, especially if patients also present with lung nodules.

GISTs present most often as well-circumscribed, highly vascular tumors associated with the stomach or the intestine.[13] On gross examination, these tumors appear fleshy pink or tan-white and may show hemorrhagic foci, central cystic degenerative changes, or necrosis. There are three principial subtypes on morphological evaluation. The spindle cell subtype of GIST, accounting for approximately 70% of cases, is composed of cells with palely eosinophilic fibrillary cytoplasm, ovoid nuclei, and ill-defined cell borders, often with a syncytial appearance, arranged in short fascicles or whorls. GIST with epithelioid cell morphology, accounting for approximately 20%, is composed of round cells with eosinophilic to clear cytoplasm arranged in sheets and nests. Finally, approximately 10% of GISTs show mixed morphology, being composed of both spindle and epithelioid cells. Variable cellularity as well as sclerotic, collagenous, or myxoid stromal changes can be seen in each subtype. Spindle cell GISTs can show nuclear palisading or a storiform growth pattern as well as prominent paranuclear vacuolation, a morphologic feature formerly proposed to be suggestive of smooth muscle origin but far more commonly seen in GISTs. Overall, GISTs are characterized as uniform and monotonous tumors.

KIT has been demonstrated to be a very specific and sensitive marker in the differential diagnosis of mesenchymal tumors in the GI tract, and around 95% of GISTs express KIT . Different KIT-staining patterns can be observed.[21] Most GISTs show strong and diffuse cytoplasmic KIT staining often associated with dot-like staining. Only in a minority of cases is an exclusively dot-like or even a membranous staining pattern observed. The extent and patterns of KIT staining do not correlate with the type of KIT mutation and have no impact on the likelihood of response to imatinib. However, GISTs showing weak or focal KIT expression and those GISTs completely negative for KIT are more likely KIT wild-type or PDGFRA mutant GISTs. Approximately, 4-5% of GISTs are KIT negative.[22] KIT-negative GISTs preferentially occur in the stomach and usually show pure epithelioid or mixed (spindle and epithelioid) cytomorphology.

Other commonly expressed but less sensitive and specific markers are CD34, h-caldesmon, and SMA.[13] CD34 is expressed in approximately 80% of gastric tumors, 50% of those in the small intestine, and 95% of GISTs in the esophagus and rectum, whereas h-caldesmon is expressed in more than two-thirds of GISTs and SMA in 30%. S-100 and cytokeratin are only infrequently expressed in GISTs. Desmin expression has been reported rarely in GISTs. KIT negativity by no means justifies denying patients therapy with TKI (imatinib or sunitinib), as some wild-type GISTs as well as some tumors with PDGFRA mutations respond to treatment with TKI.

One promising marker is discovered on GIST (DOG1), also known as TMEM16A, which is a transmembrane protein recently shown to be up-regulated in GISTs by gene expression profiling.[23] Two recent studies have suggested that antibodies against DOG1 have greater sensitivity and specificity than KIT (CD117) and CD34, and that these antibodies could serve as specific immunohistochemical markers for GIST irrespective of the underlying KIT/PDGFRA mutation or KIT expression by IHC. PDGFRA alpha is a receptor tyrosine kinase closely related to KIT. Antibodies to this kinase have been proposed to be of use in the identification of KIT-negative GISTs harboring a PDGFRA mutation. However the commercially available antibodies do not show marked sensitivity for PDGFRA. [13]

Despite these developments, a subset of KIT-negative tumors remains a diagnostic challenge, at least in terms of immunohistochemical verification; and mutational analysis should be strongly considered under these circumstances.

The main differential diagnoses of spindle-cell GIST that should be considered are smooth muscle tumors, desmoids, fibromatosis, schwannoma, inflammatory myofibroblastic tumor, inflammatory fibroid polyp, and solitary fibrous tumor. Smooth muscle tumors show brightly eosinophilic cytoplasm with defined cell borders rather than the syncytial appearance typically seen in GIST. Desmin expression is relatively specific for smooth muscle tumors and rarely positive in spindle-cell GISTs. Schwannomas occurring in the gastrointestinal tract typically show a distinctive peripheral cuff of lymphocytes and express S-100 protein and GFAP. Intraabdominal desmoid fibromatosis is morphologically characterized by long sweeping fascicles of fibroblastic/ myofibroblastic spindle cells set within a collagenous matrix. Immunohistochemistry reveals nuclear beta-catenin positivity in approximately 75% of cases. Inflammatory myofibroblastic tumors mainly occur in children and young adults. Inflammatory fibroid polyp (IFP) has a collagenous or more myxoid granulation tissue-like stroma containing fibroblasts in a pattern-less array and inflammatory cells, including numerous eosinophils. Perivascular fibrosis is commonly seen. The fibroblasts usually express CD34. The differential diagnosis for epithelioid GIST includes neuroendocrine carcinoma, glomus tumor, malignant melanoma, epithelioid leiomyosarcoma, epithelioid MPNST, and clear cell sarcoma.[1,13]

The risk of metastases after resection of the primary depends on multiple risk factors. Fletcher et al proposed a risk assessment system in 2002 based on the tumor size and mitotic count. [24]A tumor size more than 5 cms and mitotic count more than 5 per 50 high power fields(hpf) predicted aggressive clinical behaviour. However it is well known that site of primary also may predict the aggressiveness of GIST. Stomach primary may have a better outcome compared to other sites. Hence site is also included in the risk assessment system after resection of the primary. This is essential to plan adjuvant treatment after resection. The genotype of the tumor is the most significant factor which will predict the response to Imatinib.

Piotr Rutkowski et al analysed 232 cases of metastatic GIST in an effort to find out the possible predictive factors for response with Imatinib.[25] The estimated 3 year PFS for the entire cohort was 54% and the median PFS was 40.3 months. The following factors significantly and negatively influenced PFS in univariate analysis: poor baseline World Health Organization (WHO) performance status ¸2 (P < 0.00001), tumor genotype indicating other than KIT exon 11 isoform (P = 0.005), baseline high neutrophils count (P < 0.00001), age <45 years at the diagnosis (P = 0.04), mitotic index >10/50 high-power fields (HPF) (P = 0.001), GIST histological type other than spindle-cell (P = 0.03), baseline low albumin level (P = 0.0005), low baseline hemoglobin level (P < 0.00001), and primary overtly malignant tumors (unresectable and/or metastatic lesions at presentation)(P = 0.05). They identified four factors negatively affecting PFS, statistically significant (P < 0.05) in multivariate analysis: baseline poor WHO performance status ¸2, high baseline neutrophils count (>5 x 109/l), tumor genotype indicating the presence of non-exon 11 KIT mutant and mitotic index >10/50 HPF.

Generally, patients with KIT exon 11 mutant GISTs are treated with 400 mg imatinib/day, and dose escalation to 800 mg/day is recommended if patients progress on 400 mg. Clinical data did not reveal a significant benefit for KIT exon 11 mutant GISTs whether treated initially with 400 or 800 mg of imatinib. However, patients with KIT exon 9 mutations have better progression-free survival if treated with imatinib 800 mg/day than 400 mg.[7,9 This observation provides the rationale for recent consensus that KIT mutation status be evaluated routinely in inoperable GISTs, and with imatinib dose escalated immediately to 800 mg/day if a KIT exon 9 mutation is found. It is likely that the genotyping of the tumor will be routine before planning treatment for metastatic GIST. The same is extrapolated to adjuvant strategy also in an effort to tailor the duration of therapy.

Response evaluation after starting TKI often can be challenging in GIST. CT scans changes do not necessarily keep up with the more dynamic changes in PET scans, and underestimate the degree of benefit. patients with RECIST stable disease on imatinib fared just as well in terms of PFS as those who had an overt response to treatment. patients with RECIST stable disease on imatinib fared just as well in terms of PFS as those who had an overt response to treatment. A number of problems arise when applying RECIST to GIST. Necrosis of tumor masses in the liver is frequent, and thus the sum-of-maximum-diameters criterion is often not met in responding patients. Also, other lesions that are isodense with liver may become less dense and appear to be new lesions, ie, RECIST progression, when in fact it is just the opposite that is true. Occasional patients will have growth of lesions with increasing edematous tumor degeneration over time before what ultimately becomes disease that decreases in size. In addition, new tumor nodules can occur within preexisting lesions without changing the absolute size of the original lesion but likely representing the occurrence and progression of resistant GIST clones. However since patients with stable disease have similar outcome like that of partial remission, people have questioned usage of other evaluation systems.[26]

One of the most impressive aspects of GIST diagnostic imaging is the use of 18F-fluorodeoxyglucose (18FDG) positron emission tomography (PET) to add functional imaging data that are complementary to the information obtained by conventional anatomic imaging. Choi and colleagues at M.D. Anderson Cancer Center evaluated clinical outcome by PET response and by RECIST, showing that PET provided a superior means to follow patients for clinical benefit. [27] Although CT or MRI scanning can assess the size of GIST lesions quite accurately, the functional imaging of GISTs with 18FDG-PET can give additional information that can assist clinicians in the management of GIST patients. The actual mechanisms responsible for the high-level avidity of GISTs for the 18FDG tracer used most commonly in PET imaging are not yet known; however, it is likely that there is a direct connection between signaling through the overactive KIT RTK and glucose transport proteins 45a.1 and 45a.2. In this way, one could explain the very rapid changes in PET imaging associated with inhibition of KIT signaling by pharmacologic means. [28,29]

Large GISTs can demonstrate centers with predominately cystic or low attenuation characteristics noted on CT or MRI scans. It is clear by 18FDG-PET scans that the internal mass of large GIST lesions can often be viewed as metabolically quiescent. This is likely due to the endogenous necrosis of very large lesions in their central portions; although GIST lesions can be very vascular, the internal portion can nonetheless represent a confluent mass of necrotic material, with the more viable aspects of the GIST pushing out toward the edges of the lesion. In addition, occasionally metastatic GIST lesions in the omentum can be subtle and easy to overlook on CT scans, because small lesions can blend into the folds of the bowel walls and be difficult for even the most experienced radiologist to detect. 18FDG-PET imaging can detect lesions at least 1 cm in size without difficulty, because neither the normal bowel nor omentum takes up the 18FDG tracer with excess avidity.

Metastases can quite often occur 10-15 years after initial surgery, and therefore long-term follow-up is required. Metastases develop primarily in the abdominal cavity and liver, rarely in the soft tissue and skin, and exceptionally rarely in lymph nodes or in the lung.[1] Clinically, it is essential to differentiate metastatic GIST from multifocal GISTs observed in patients with germline KIT or PDGFRA mutations, in patients with neurofibromatosis 1 and multiple sporadic GISTs, mainly occurring in the proximal stomach. The pathogenesis of multiple sporadic GISTs is poorly understood; however, these GISTs have been shown to harbor different KIT mutations in separate individual lesions from the same patient. Generally speaking, the clinical history, clinical presentation, morphology, mitotic activity and, in rare cases, mutational analysis should allow exact precise classification.

Unresectable or metastatic GIST are considered incurable and is conventionally thought to be chemoresistant. The mechanism for resistance may be partly explained by increased levels of p-glycoprotein and multidrug resistance protein.[1] Till Imatinib was discovered to be active against GIST, there were not much options for these patients. Joensuu et al first reported the use of imatinib in a patient with a recurrent, metastatic GIST, a 50-year-old female whose tumor demonstrated staining for CD117. She exhibited a response to 400 mg of imatinib daily that was sustained for 11 months at the time of the case report publication. [30]

This paved for further phase II trials in United States and Europe. Demetri et al studied 147 patients with GIST randomised to 400 mg or 800 mg per day of Imatinib. The partial response rates reported was 54% and the median duration of response was not reached at 24 weeks.[31] This was encouraging in a disease which was considered as a death warrant all these years. FDA approved the drug for use in metastatic GIST in 2002 itself. A Phase I study by EORTC also demonstrated activity including partial responses.[32]

In 2001, two open-label, controlled, multicenter, randomized phase III studies were started-the first led by the European Organization for Research and Treatment of Cancer (EORTC), which included the Australasian Gastro-Intestinal Trials Group and the Italian Sarcoma Group (n=946), and the second led by the Southwest Oncology Group (SWOG), which included the Cancer and Leukemia Group B, the Eastern Cooperative Oncology Group (ECOG), and the National Cancer Institute of Canada (n=746).[7,34] These studies were designed to compare 400 mg/day versus 800 mg/day of imatinib in patients with advanced inoperable or metastatic GISTs. The studies were planned simultaneously with similar protocols, and a combined analysis was prospectively defined and agreed to by both groups. Both protocols allowed patients randomized to the 400-mg/day imatinib arm who experienced progressive disease (PD) to crossover to the higher, 800 mg/day, dose of imatinib.

Eligible patients in both studies were required to have a biopsy-proven diagnosis of GIST that was CD117 positive and distantly metastatic or unresectable. Patients must not have received chemotherapy, biologic therapy, or any other investigational drug for any reason within 28 days prior to registration. Patients must not have had major surgery within 14 days prior to registration. Patients had to have an ECOG performance status score of 0-3 and adequate hematologic, hepatic, renal, and cardiac function. Patients with known brain metastases or uncontrolled medical disease were ineligible.

The endpoint of the EORTC study was progression-free survival (PFS), whereas the SWOG study evaluated overall survival (OS), with PFS as a secondary efficacy variable. In both studies the evaluation of disease progression was based on the Response Evaluation Criteria in Solid Tumors [3]. Patients were randomized on a 1:1 basis, stratified by performance status (World Health Organization score of 0-2 versus 3) and measurable versus nonmeasurable disease. In both studies, patients in the 400-mg/day arm were allowed to cross over to 800 mg/day of imatinib upon documented disease progression as long as they continued to meet the eligibility criteria required for study entry and in the absence of safety concerns. A pre planned combined analysis of the two studies was done. The objectives of this combined analysis were to assess the efficacy and safety of the two imatinib doses, to assess the efficacy and safety of a dose increase from 400 mg/day to 800 mg/day after progression (crossover subset), and to explore the impact of mutation status on efficacy in each of the two dose groups.

PFS was defined as the time from the randomization date to the date of the first documented disease progression or death, whichever came first. The time was censored at the last date of disease evaluation for patients who had not experienced disease progression or death. The OS time was defined as the time from randomization to death (for any reason). Patients still alive and on study were considered as censored at the last date on study. The best overall response was assessed locally. For the EORTC study, the best response was either a complete response (CR), a partial response (PR), stable disease (SD), or PD. For the SWOG study, the best response was either a CR, a PR, an unconfirmed CR, an unconfirmed PR, stable/no response, increasing disease, or inadequate assessment.

A total of 946 patients in total with metastatic and/or unresectable GISTs were enrolled in the EORTC study by 56 institutions from 13 countries. In the PFS analysis, there were 332 patients with disease progression in the 400-mg/day imatinib group and 324 patients with disease progression in the 800-mg/day imatinib group. The median PFS times were 19.8 and 24.0 months, in the 400-mg/day and 800-mg/day arms, respectively. In the OS analysis, there were 208 deaths among patients in the 400-mg/day imatinib group and 206 deaths among patients in the 800-mg/day imatinib group. The median survival time was 45 months in both the 400 mg/day and 800 mg/day arms. The CR and PR rates in the 400-mg/day imatinib and 800-mg/day imatinib groups were 5.3% and 5.9%, respectively, and 44.6% and 48.8%, respectively. These differences were not statistically significant (p= .0637).

The SWOG study randomized 746 patients with metastatic or unresectable GISTs. Of these, 694 were considered evaluable for analysis. The patient demographics and baseline characteristics were well balanced between the two treatment doses. The median follow-up time was 44.2 months (range, 0 - 64 months). About 20% of all patients were still on treatment at the cutoff date. The median OS times were 55.1 months and 51.3 months in the 400-mg/day and 800-mg/ day dose groups, respectively (p= .5819). The median PFS times were 17.6 and 19.7 months, in the 400-mg/day and 800-mg/day arms, respectively. The CR and PR rates in the 400-mg/day imatinib and 800-mg/day imatinib groups were 5.2% and 3.7%, respectively, and 48.1% and 49.0%, respectively (p= .2826). There were 1,640 patients in the combined EORTC- SWOG dataset. The median PFS time was longer by 4.3 months in patients receiving 800 mg/day of imatinib compared with patients receiving 400 mg/day of imatinib - 23.2 months versus 18.9 months. No difference was observed between the two dose groups with respect to OS (p = .9775). The median OS time was 48.8 months in both dose groups. The two dose groups were also nearly identical in terms of the best overall response. Overall, 5.1% of patients achieved a confirmed CR and 47.5% achieved a confirmed PR.

In the combined EORTC-SWOG analysis, the median duration of drug exposure was 24 months for both imatinib treatment groups. About one third of the patients from either dose group were treated for 36 months. The majority of imatinib-treated patients experienced adverse reactions at some time. The most frequently reported adverse reactions were edema, fatigue, nausea, abdominal pain, diarrhea, rash, vomiting, myalgia, anemia, and anorexia. Most reactions were of mild-to-moderate severity. The drug was discontinued for adverse reactions in a total of 89 patients (5.4%). Overall, the incidence of all grades of adverse reactions and the incidence of severe adverse reactions (Common Terminology Criteria grade > 3) were similar between the two treatment arms except for edema, which was reported more frequently in the 800 mg/day group. There were five grade 5 adverse events in patients receiving 400 mg/day of imatinib and 10 in patients receiving 800 mg/day of

imatinib. Three deaths, all in patients receiving 800 mg/day of imatinib, were considered by the investigator to be related to imatinib treatment, including liver dysfunction in one patient, cardiac arrhythmia in one patient, and tumor hemorrhagic necrosis in one patient.

The optimal duration of imatinib therapy for patients with metastatic GIST remains somewhat uncertain, but most experts consider kinase inhibition as lifelong therapy for advanced disease. Studies in which patients have interrupted imatinib dosing have reported that disease progression often follows shortly after the imatinib is stopped.[35]. Therefore, for GIST patients who achieve any measure of disease control, continued dosing with imatinib as long as the disease is not progressive appears to be the optimal course of management.

Primary resistance has been observed with all genotypic subtypes of GISTs; however, the tumors that are most likely to show primary resistance include those that are KIT and PDGFRA wild type, those that have a KIT exon 9 mutation, and those that have a PDGFRA D842V substitution.[33] The latter can be explained by intrinsic biochemical resistance of the D842V mutation to imatinib.[36] In patients with KIT exon 9-mutant tumors, inadequate dosing may account for some of the primary resistance observed. It appears that exon 9 mutations generate a kinase conformation that is less amenable to imatinib binding. In patients lacking identifiable PDGFRA or KIT mutations, one potential mechanism for resistance is a mutation in an alternate signaling pathway. Recently, several groups have identified BRAF exon 15 activating mutations in wild-type GISTs from both imatinib-naive and -resistant patients. [37]

Acquired kinase mutations are now recognized as the most common mechanism of secondary imatinib resistance. The resistance may manifest in a number ways, including growth of a nodule within a pre-existing, clinically quiescent lesion, the development of one or more new nodules, or widespread expansion of lesions throughout the liver or abdominal cavity.[38]

Unlike primary resistance, delayed imatinib resistance is associated most often with the expansion of tumor clones with secondary KIT or PDGFRA mutations.[39] Analysis of tumors of patients who progressed on the phase II B2222 imatinib trial revealed that 67% of patients with secondary resistance had tumor clones with one or more secondary kinase mutations. All secondary KIT kinase mutations were found in tumors with an underlying primary KIT mutation, and the only secondary PDGFRA mutation identified arose in a PDGFRA-mutant GIST. The secondary KIT mutations involved either the ATP binding pocket of the kinase domain (exons 13 and 14) or the kinase activation loop (exons 17 and 18; Fig. 1). No secondary mutations were identified in post-imatinib samples that lacked a primary mutation (wild-type GISTs).[40]

Imatinib currently remains the standard first-line treatment option for patients with unresectable and metastatic GISTs, especially those harboring an exon 11 mutation. However, accumulating evidence suggests that sunitinib could be effective as a first-line treatment for GISTs harboring KIT exon 9 mutation and for KIT/PDGFRA wild-type GISTs (including pediatric GISTs).[9] Sunitinib is effective against secondary imatinib-resistance mutations in the ATP-binding pocket. However, the substantial heterogeneity of resistance mutations highlights the therapeutic challenges involved in salvaging patients, especially after clinical progression on TKI monotherapies. With regard to treatment approaches, the role of newer generation KIT and PDGFRA kinase inhibitors (e.g., nilotinib, dasatinib, etc.) remains to be determined in GIST patients who are multiply resistant, i.e., after imatinib and sunitinib treatment, to TKIs. Nilotinib has been shown to be effective in advanced imatinib- and sunitinib-resistant GISTs. Using nilotinib 400 mg twice a day, the median progression-free survival and the median overall survival were 12 and 34 weeks, respectively.[41] In vitro data, using cell lines expressing imatinib resistant PDGFRA (D842V) mutants, suggest that dasatinib, a dual SRC/ABL kinase inhibitor, and IPI-504, a heat shock protein 90 inhibitor, may be a therapeutic option for patients with a GIST harboring the PDGRA (D842V9) mutation.[42]

The standard approach in the case of tumour progression on 400 mg is to increase the imatinib dose to 800 mg daily, with the possible exception of insensitive mutations. Dose escalation may be useful in the case of a KIT exon 9 mutated GIST, possibly in the case of changes in drug pharmacokinetics over time, or perhaps in the case of some molecular alterations. False progression on imaging should be ruled out, due to the response patterns. Also patient non-compliance should be ruled out as a possible cause of tumour progression, as well as drug interactions with concomitant medications. In the case of progression or intolerance on imatinib, second-line standard treatment is sunitinib. After failing on sunitinib, the patient with metastatic GIST should be considered for participation in a clinical trial of new therapies or new combinations.

It is needless to mention about the importance of compliance to treatment in order to sustain the response. Close monitoring of tumour response should be continued throughout treatment, since the risk of secondary progression persists over time.[43] Retrospective data suggest that suboptimal plasma levels of imatinib are associated with a worse outcome. Further studies are needed.

Histone deacetylase inhibitors (HDACI) alone or in combination with imatinib show inhibition of cell proliferation, KIT activity and expression as well as activation of downstream pathways in KIT-positive cell lines, providing preclinical evidence that HDACI may expand the treatment options in KIT-positive GISTs.[44] IGFR inhibitors in combination with imatinib have been proposed as a treatment option mainly for wild-type GISTs which tend to be less responsive to imatinib-based therapies. The rationale for this treatment is based on detected amplification of IGF1R and protein overexpression predominantly in WT and pediatric GISTs. [45]

The frequent occurrence of secondary mutations led some investigators to think about combining drugs in an effort to target multiple steps in the pathogenesis. Some of the drugs inhibit both c-kit and PDGFRA. These include Dasatinib, Sorafenib, Motesanib etc. The feasibility of combining these drugs with Imatinib has been tried in Phase I/II trials with moderate success. The toxicity profile was reasonable. Another interesting way of interfering with the c-kit pathway is to target the downstream signalling pathways like mTOR. Phase I/II studies demonstrated the excellent activity of the combination. However it remains to be seen which subset of patients will be benefitted by this interesting combination.

Surgery is the mainstay of curative therapy in primary GIST and has traditionally played a palliative role in the advanced disease setting. In the era of targeted therapy, the role for surgery as a part of multimodality management of advanced GISTs has been looked at in small patient series and retrospective studies. One of the rationales for resecting metastases is to eliminate tumors from which drug-resistant clones might develop. The Radiation Therapy Oncology Group studied the role of preoperative imatinib followed by surgery in a phase 2 study in patients with primary locally advanced disease or with recurrent/metastatic disease. Patients with locally advanced disease received 2 years of postoperative imatinib and those with metastatic disease were continued on imatinib until progression. At 2 years, patients with locally advanced disease and metastatic disease had a PFS of 82% (95% CI 68-97) and 73% (95% CI 54-91), respectively, which are encouraging results, suggesting a benefit to surgical debulking in advanced disease.[46]

Surgical management of residual disease can be discussed in two settings. One is in the setting of progressive disease after Imatinib. Most of these patients have secondary mutations. Considering the limited options of second line agents, surgery may have a role especially when second line agents are not available for the patient. surgery of limited progression, such as the 'nodule within a mass', has been associated with a progression-free interval in the same range as for second-line treatment with sunitinib. Therefore, it may be a palliative option in the individual patient with a limited progression. Non-surgical procedures (local treatment, such as ablations, etc.) may be selected. More controversial is the role of surgery in the setting of a responding patient with residual disease. With PFS reaching 2 years in most of the studies, people have questioned its role. There is no randomised data which has addressed this issue. An ongoing EORTC trial (EORTC 62063) may throw more light on this controversial issue. Till then, this may be used only for selected patients. It is essential to continue Imatinib lifelong postoperatively.

Based on its activity in the metastatic setting, imatinib was tested as an adjuvant treatment after complete resection of primary GISTs in a randomized phase III, double-blind, placebo-controlled, multicenter trial (Z9001). [47] Following complete gross resection of a primary KIT protein-expressing GIST 3 cm or larger in size, patients were randomly assigned to take 1 year of 400 mg imatinib or placebo daily. Accrual was stopped early, because the trial results crossed the interim analysis efficacy boundary for recurrence-free survival. At a median follow-up of 19.7 months, 8% of patients in the imatinib group and 20% in the placebo group had experienced tumor recurrence (or died). Imatinib significantly improved recurrence-free survival compared with placebo (98% compared with 83% at 1 year; p < 0.0001). Based on these results, the U.S. Food and Drug Administration (FDA) approved imatinib as an adjuvant treatment for GISTs.

Although this study clearly demonstrates that empiric adjuvant imatinib reduces the rates of early recurrence, it is not yet clear whether this strategy improves overall survival over a strategy of watchful waiting. Given the strong effect of genotype on imatinib response in the metastatic setting, we hypothesize that tumor genotype will influence the efficacy of imatinib in the adjuvant setting. The recurrence rate in the imatinib group was noted to increase appreciably around 18 months after surgery, raising the concern that 1 year of therapy may be inadequate for patients at high risk for recurrence. Analysis of the correlation of tumor genotype and mitotic index-as well as other clinicopathological factors-is underway. These data may define a potential role for genotyping of primary tumors and optimization of postsurgical therapy and/or surveillance strategies.

It is critical to emphasize the importance of multidisciplinary management in the care of GIST patients. For optimal management of metastatic disease, medical oncologists, surgeons, radiologists, and nuclear medicine imaging experts must all collaborate closely to determine the best course of action for any given patient. This important message has been emphasized in the Task Force Report on GIST Clinical Practice Guidelines of the National Comprehensive Cancer Network.96 For example, disease that is initially judged as unresectable may become amenable to surgical excision after a major response induced by imatinib therapy. Most centers recommend surgical resection for such patients because it is feared that residual GIST may develop secondary mutations that could result in clinical resistance to imatinib and progression of disease. However, the role of surgery as an adjunct to imatinib therapy for patients for metastatic GIST remains unclear.