KIT Alterations Research Articles

Molecular characterization and pathogenesis of gastrointestinal stromal tumor.

Most gastrointestinal stromal tumors (GISTs) harbor activating mutations in the receptor tyrosine kinase gene KIT or platelet-derived growth factor receptor alpha (PDGFRA), and the resultant activation of downstream signals plays a pivotal role in the development of GISTs. The sites of the tyrosine kinase gene mutations are associated with the biological behavior of GISTs, including risk category, clinical outcome and drug response. Mutations in RAS signaling pathway genes, including KRAS and BRAF, have also been reported in KIT/PDGFRA wild-type GISTs, though they are rare. Neurofibromin 1 (NF1) is a tumor suppressor gene mutated in neurofibromatosis type 1. Patients with NF1 mutations are at high risk of developing GISTs. Recent findings suggest that altered expression or mutation of members of succinate dehydrogenase (SDH) heterotetramer are causally associated with GIST development through induction of aberrant DNA methylation. At present, GISTs with no alterations in KIT, PDGFRA, RAS signaling genes or SDH family genes are referred to as true wild-type GISTs. KIT and PDGFRA mutations are thought as the earliest events in GIST development, and subsequent accumulation of chromosomal aberrations and other molecular alterations are required for malignant progression. In addition, recent studies have shown that epigenetic alterations and noncoding RNAs also play key roles in the pathogenesis of GISTs.

Pharmacokinetic-driven phase I study of DCC-2618 a pan-KIT and PDGFR inhibitor in patients (pts) with gastrointestinal stromal tumor (GIST) and other solid tumors.

2515 Background: DCC-2618 is a potent switch control inhibitor of KIT and PDGFR kinases active in a broad range of mutations. GIST is an important disease to achieve a proof-of-concept due to the heterogeneity of KIT resistance mutations, which emerge on treatment with approved KIT inhibitors. Methods: This was a PK-guided dose escalation study of oral DCC-2618 (QD or BID q28 days) in advanced solid tumors. FDG-PET scans were used to assess changes in FDG uptake in GIST pts after 3 wks of therapy. Next generation sequencing (NGS) of plasma cell-free (cf) DNA was performed throughout the study to assess and quantify KIT and other molecular alterations in drug targets and potential mechanisms of resistance. Results: 38 pts were enrolled (30 GIST; 4 glioma; 1 mastocytosis, 3 other carcinoma) to 8 dose levels: BID doses: 20 (4 pts), 30 (4), 50 (5), 100 (6), 150 (6) and 200 mg (3); QD doses: 100 (5) and 150 mg (4). Safety of evaluable pts is as follows: G3 or G4 adverse effects (regardless of attribution and occurring in > 1 pt) included anemia (5), lipase increase (4), hypertension (2). Two of the G3/4 lipase increase at 100 mg BID and 200 mg BID were DLTs. All G3/4 lipase increase were asymptomatic. G1/2 AEs (considered at least possibly related to DCC-2618) and occurring in ≥15% (n > 5) of pts include fatigue (12), alopecia & lipase increase (7), weight decrease (6). Starting with 50 mg BID dose level, trough concentrations of total drug exceeded the IC90 of the least sentivitive KIT mutations. Plasma concentrations > 5μM were achieved starting at 100 mg BID and the selection of the expansion phase dose is being finalized. Of 18 pts with KIT mutant GIST assessed by FDG PET, 14 (78%) had partial metabolic response per EORTC criteria. RANO/RECIST partial responses (PRs) were reported in 3 patients (1 GBM with PDGFRA/KIT amplifications and 2 GIST with Ex 11 & 17 / Ex 11 & 18 mutations, respectively). NGS of plasma cfDNA revealed 44 KIT mutations in baseline samples from 19 of 21 pts with GIST. Conclusions: DCC-2618 is well tolerated with encouraging preliminary activity in GIST pts with a broad spectrum of mutations and prior therapies. PR was also seen in a pt with GBM with PDGFRA/KIT amplifications. Clinical trial information: NCT02571036.

Clinical and pathological characteristics of gastrointestinal stromal tumor (GIST) metastatic to bone.

Our aim in this study was to describe the clinical, morphological, and molecular profile of gastrointestinal stromal tumor (GIST) metastatic to bone. We analyzed the morphological, phenotypic, and molecular characteristics of seven cases, and in addition reviewed 17 cases from literature. Sequence analysis of KIT and PDGFRA genes was possible for six cases. For the GIST cases with bone metastasis, the most common primaries were small intestine (29%), stomach (25%), and rectum (21%). Sites of bone metastases were vertebrae (11), pelvis (8), femur (8), ribs (6), humerus (5), skull (3), scapula (1), and mandible (1). The size ranged from 1.5 to 13cm (median, 3.8cm). Bone metastases without involvement of any other organ were seen in 17% of the cases and were solitary in 14 (58%). Adjacent soft tissue involvement was present in nearly half of the patients. Bone metastasis was either manifest at the time of diagnosis (28%) or occurred after a mean period of 4.7years (3months-20years). Morphologically, neoplastic cells were spindle in 67%, epithelioid in 13%, and mixed epithelioid and spindle in 20%. CD117, DOG1, and CD34 were positive in 88, 86, and 85% of the cases, respectively. KIT Exon 11 mutations were the most frequent gene alteration (78%), followed by KIT Exon 13 mutations. Of 17 of the cases with available follow-up information, 7 (41%) patients developed bone metastasis under imatinib therapy. Five patients (29%) died of disease within a mean of 17months. Bone metastases from GIST are usually found in patients with advanced disease and typically present as lytic masses with occasional soft tissue involvement. We could not identify any KIT or PDGFRA alterations predisposing to bone metastasis.

Genomic characterization of acral lentiginous melanoma: Identification of altered metabolism as a potential therapeutic target.

9524Background: Acral lentiginous melanoma (ALM) is a histologic subtype of melanoma with distinct clinical and biological features compared to common cutaneous melanoma. KIT alterations have been ...

Molecular cytology genotyping of primary and metastatic GIstromal tumors by using a custom two-gene targeted next-generation sequencing panel with therapeutic intent.

In an era of precision medicine, customized genotyping of GI stromal tumors by screening for driver mutations will become the standard of care. The fidelity of genotype concordance between paired cytology smears and surgical pathology specimens is unknown. In patients with either primary or metastatic sporadic disease, we sought to determine the frequency of KIT and PDGFRA pathogenic alterations within such specimens, imatinib sensitivity, and the concordance of pathogenic alterations between paired specimens. DNA obtained from cytology smears from 36 patients, 24 of whom had paired surgical pathology specimens, underwent targeted next-generation sequencing by using a custom panel to evaluate somatic mutations within KIT (exon 2, 9, 10, 11, 13, 14, 15, 17, 18) and PDGFRA (exon 12, 14, 15, 18) genes. Patients with KIT and PDGRFA wild-type genes completed the Qiagen Human Comprehensive Cancer GeneRead DNAseq Targeted Array V2. Genotyping revealed KIT and PDGFRA mutations in 68% and 15% of patients. The wild-type population did not harbor mutations in BRAF, RAS family, SDHB, SETD2, or NF1. Imatinib sensitivity based on the oncogenic kinase mutation prevalence was estimated to be 68%. Mutational concordance between paired cytology and surgical pathology specimens was 96%. Our data have demonstrated the ability to stratify either primary or metastatic gastrointestinal stromal tumors by mutational subtype using a targeted next-generation sequencing 2 gene mutation panel. We highlight the ability to use cytology specimens obtained via minimally invasive techniques as a surrogate to surgical specimens given the high mutational landscape concordance between paired specimens.

Long-term outcome of molecular subgroups of GIST patients treated with standard-dose imatinib in the BFR14 trial of the French Sarcoma Group

BackgroundThe added value of tumoural genomic profiles to conventional clinico-biological factors to predict progression-free survival (PFS) and overall survival (OS) was prospectively investigated in patients with advanced gastrointestinal stromal tumours (GIST) treated in the BFR14 study. MethodsOf the 434 included patients, mutational analysis was performed in 322 patients. Survival analysis was performed in patients with validated mutational status. ResultsMutational status was validated in 228 patients. We identified 196 patients with tumours harbouring 200 KIT alterations (exon 11: 173 patients, exon 9: 22 patients, exon 17: 3 patients, exon 13: 2 patients; 4 patients had double KIT mutations), 6 patients with PDGFRA mutations and 26 patients with wild-type (WT) GIST genotype. On a median follow-up of 73 months, median PFS/OS were 12.3/54.9 months for WT GIST, 12.6/55 months for KIT exon 9, and 39.4 months/not reached (69.1% at 5 years) for KIT exon 11. Tumour size, female gender, KIT exon 11 mutations and CD34 positivity were independent prognostic factors for a higher PFS. A higher OS was predicted by performance status (PS) <2, low neutrophil and normal lymphocyte counts, KIT exon 11 mutations, non-advanced tumour and female gender. KIT exon 11 mutations at codons 557–558 showed better tumour response (p=0.028) but shorter PFS (p=0.0176). ConclusionsIn GIST patients, presence of a KIT exon 11 mutation is an independent prognostic factor for PFS and OS, along with gender, PS, tumour size, lymphocyte and neutrophil counts. Subsets of exon 11 mutations are associated with significantly different response patterns and PFS.

Genetic mutations in primary malignant melanoma of the esophagus: case report and literature review

The most frequent genetic aberrations in mucosal melanoma are activating mutations of c-KIT. Primary malignant melanomas of esophagus (PMME) are uncommon entities, with aggressive biological behavior and poor prognosis. The better definition of their genotype could improve therapeutic options. We report a case of a 66 years old man with a PMME in the lower third of the esophagus. Analysis of c-kit, KRAS, NRAS and BRAF genes resulted negative for mutations. On the basis of a computerized (PuMed/Medline) bibliography search we retrieved a total of other 35 cases of PMME analyzed for genetic alterations in RAS, BRAF, and KIT. When we compared mutations frequency of PMME with those of other mucosal melanomas, it appeared that PMME are characterized by a relative higher percentage of NRAS mutations. PMME seem to show a specific pattern of genetic alterations suggesting that they could represent a distinct entity among mucosal melanomas.

Abstract 619: Identification of clinically actionable genomic alterations in the tumor and circulation of pancreatic cancer patients

Abstract Pancreatic adenocarcinoma has the worst overall mortality of any solid tumor, with only 6% of patients surviving after 5 years. To evaluate the clinical implications of genomic alterations in this low cellularity tumor type, we deeply sequenced the genomes of 134 enriched pancreatic adenocarcinomas from patients who underwent potentially curative resections. Given the low neoplastic cellularity of pancreatic cancers, we enriched for neoplastic cells either by macrodissection of primary tumors or by flow-sorting of tumor nuclei, and performed deep sequencing (high coverage) of these enriched samples using next-generation sequencing approaches. We obtained a total of &amp;gt;1Tb of sequence data, resulting in an average coverage within the target regions of &amp;gt;200-fold for each tumor analyzed by whole-exome sequencing and &amp;gt;750-fold for each tumor analyzed by targeted cancer gene sequencing. These approaches allowed us to identify sequence changes, including single base and small insertion or deletion mutations, as well as copy number alterations in &amp;gt;20,000 genes in the whole-exome analyses and in 116 specific genes in the targeted analyses. These analyses revealed that somatic mutation of chromatin remodeling genes were associated with improved progression-free and overall survival. Alterations in genes with potential clinical utility were observed in a majority of cases and included alterations of AKT1, AKT2, BRCA2, ERBB2, KIT, and PIK3CA. Non-invasive liquid biopsy analyses were performed before and after surgery to evaluate the presence of circulating tumor DNA in the plasma of 83 patients. Through these approaches, we were able to diagnose early stage pancreatic tumors in the majority of patients and to detect the presence of circulating tumor DNA prior to clinical relapse. These observations provide genetic markers of clinical outcome in pancreatic cancer and suggest new avenues for personalized therapy. Citation Format: Mark Sausen, Jillian Phallen, Vilmos Adleff, Siân Jones, Rebecca J. Leary, Karli Lytle, Sonya Parpart-Li, Derek Murphy, Michael T. Barrett, David C. Linehan, Anirban Maitra, Ralph Hruban, Daniel D. Von Hoff, Julia S. Johansen, Luis A. Diaz, Jeffrey A. Drebin, Victor E. Velculescu. Identification of clinically actionable genomic alterations in the tumor and circulation of pancreatic cancer patients. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 619. doi:10.1158/1538-7445.AM2015-619

Comprehensive Genomic Profiling of Carcinoma of Unknown Primary Site: New Routes to Targeted Therapies.

For carcinoma of unknown primary site (CUP), determining the primary tumor site may be uninformative and often does not improve outcome. To discover opportunities for targeted therapies in patients with CUP not currently searched for in routine practice. Comprehensive genomic profiling on 200 CUP formalin-fixed paraffin-embedded specimens (mean, 756× coverage) using the hybrid-capture-based FoundationOne assay at academic and community oncology clinics. Presence of targetable genomic alterations (GAs) in CUP and responses to targeted therapies. There were 125 adenocarcinomas of unknown primary site (ACUPs) and 75 carcinomas of unknown primary site without features of adenocarcinoma (non-ACUPs). At least 1 GA was found in 192 (96%) of CUP specimens, with a mean (SD) of 4.2 (2.8) GAs per tumor. The most frequent GAs were in TP53 (110 [55%]), KRAS (40 [20%]), CDKN2A (37 [19%]), MYC (23 [12%]), ARID1A (21 [11%]), MCL1 (19 [10%]), PIK3CA (17 [9%]), ERBB2 (16 [8%]), PTEN (14 [7%]), EGFR (12 [6%]), SMAD4 (13 [7%]), STK11 (13 [7%]), SMARCA4 (12 [6%]), RB1 (12 [6%]), RICTOR (12 [6%]), MLL2 (12 [6%]), BRAF (11 [6%]), and BRCA2 (11 [6%]). One or more potentially targetable GAs were identified in 169 of 200 (85%) CUP specimens. Mutations or amplifications of ERBB2 were more frequent in ACUPs (13 [10%]) than in non-ACUPs (3 [4%]). Alterations of EGFR (10 [8%] vs 2 [3%]) and BRAF (8 [6%] vs 3 [4%]) were more common in ACUPs than in non-ACUPs. Strikingly, clinically relevant alterations in the receptor tyrosine kinase (RTK)/Ras signaling pathway including alterations in ALK, ARAF, BRAF, EGFR, FGFR1, FGFR2, KIT, KRAS, MAP2K1, MET, NF1, NF2, NRAS, RAF1, RET, and ROS1 were found in 90 (72%) ACUPs but in only 29 (39%) non-ACUPs (P < .001). Almost all CUP samples harbored at least 1 clinically relevant GA with potential to influence and personalize therapy. The ACUP tumors were more frequently driven by GAs in the highly druggable RTK/Ras/mitogen-activated protein kinase (MAPK) signaling pathway than the non-ACUP tumors. Comprehensive genomic profiling can identify novel treatment paradigms to address the limited options and poor prognoses of patients with CUP.

O3.7 - Enriching phase I trials with molecular alterations: Interim analysis of 708 patients enrolled in the MOSCATO 01

ABSTRACT Background: Enrolling patients with ad hoc molecular alterations is one of the current challenges in early drug development. The MOSCATO 01 trial (NCT01566019) is one of the prospective trials ongoing worldwide using high throughput analysis for characterisation of the genomic alterations in cancer patients. Methods: An “on-purpose” tumor biopsy was collected from consenting patients with metastatic cancer referred for a phase I trial. DNA extracted from the tumor biopsy was analysed by comparative genomic hybridization (CGH) (if ≥ 30% tumor cells) and by hot spot mutations by Next Generation Sequencing (NGS) (if ≥ 10% tumor cells). Recently, WES and RNAseq were implemented in the MOSCATO trial. Results: Between December 2011 and November 2014, 708 patients were enrolled in the MOSCATO 01 trial, and 639 patients (90%) had undergone a tumor biopsy. Patient characteristics were as follows: median age at biopsy: 57 years (range, 18-78); main tumor types: lung (14%), Head and neck (13%), prostate and bladder (11%), breast (10%), median of 3 previous lines of treatment. Molecular profiling with CGH and NGS was performed in 510 (80%) and 565 patients (88%) respectively, with median analysis turnaround of 21 days. Clinical significant molecular alterations were found in 288 patients (45%), most frequently in PTEN/PI3K/AKT (n = 84), FGFR/FGF (n = 46), KRAS/NRAS/HRAS (n = 24), HER2/EGFR (n = 22/18) pathways. Patients with rare genomic alterations were also identified (eg MDM2, MET, AR, ALK, KIT, and ROS1 alterations among others). Therapy could be personalised in 141 patients (49%) with matched targeted therapy or dedicated phase I clinical trials. Preliminary results of WES and RNAseq as well as the response rate of patients with specific molecular alterations treated with matched targeted therapies will be updated and presented at the TAT meeting. Conclusions: Our updated results confirm in a larger cohort of patients the feasibility of molecular characterisation of cancer patients in routine clinical practice. Such approach allows the enrichment of early-phase clinical trials with specific and rare genomic alterations

Constitutive activation of the ERK pathway in melanoma and skin melanocytes in Grey horses.

BackgroundConstitutive activation of the ERK pathway, occurring in the vast majority of melanocytic neoplasms, has a pivotal role in melanoma development. Different mechanisms underlie this activation in different tumour settings. The Grey phenotype in horses, caused by a 4.6 kb duplication in intron 6 of Syntaxin 17 (STX17), is associated with a very high incidence of cutaneous melanoma, but the molecular mechanism behind the melanomagenesis remains unknown. Here, we investigated the involvement of the ERK pathway in melanoma development in Grey horses.MethodsGrey horse melanoma tumours, cell lines and normal skin melanocytes were analyzed with help of indirect immunofluorescence and immunoblotting for the expression of phospho-ERK1/2 in comparison to that in non-grey horse and human counterparts. The mutational status of BRAF, RAS, GNAQ, GNA11 and KIT genes in Grey horse melanomas was determined by direct sequencing. The effect of RAS, RAF and PI3K/AKT pathways on the activation of the ERK signaling in Grey horse melanoma cells was investigated with help of specific inhibitors and immunoblotting. Individual roles of RAF and RAS kinases on the ERK activation were examined using si-RNA based approach and immunoblotting.ResultsWe found that the ERK pathway is constitutively activated in Grey horse melanoma tumours and cell lines in the absence of somatic activating mutations in BRAF, RAS, GNAQ, GNA11 and KIT genes or alterations in the expression of the main components of the pathway. The pathway is mitogenic and is mediated by BRAF, CRAF and KRAS kinases. Importantly, we found high activation of the ERK pathway also in epidermal melanocytes, suggesting a general predisposition to melanomagenesis in these horses.ConclusionsThese findings demonstrate that the presence of the intronic 4.6 kb duplication in STX17 is strongly associated with constitutive activation of the ERK pathway in melanocytic cells in Grey horses in the absence of somatic mutations commonly linked to the activation of this pathway during melanomagenesis. These findings are consistent with the universal importance of the ERK pathway in melanomagenesis and may have valuable implications for human melanoma research.Electronic supplementary materialThe online version of this article (doi:10.1186/1471-2407-14-857) contains supplementary material, which is available to authorized users.

Phase Ii Study of Nilotinib in Melanoma Harboring Kit Alterations Following Progression or Intolerance to Prior Kit Inhibition

Phase Ii Study of Nilotinib in Melanoma Harboring Kit Alterations Following Progression or Intolerance to Prior Kit Inhibition

Guidelines for biomarker testing in metastatic melanoma: a National Consensus of the Spanish Society of Pathology and the Spanish Society of Medical Oncology

This consensus statement, conceived as a joint initiative of the Spanish Society of Pathology (SEAP) and the Spanish Society of Medical Oncology (SEOM), makes diagnostic and treatment recommendations for the management of patients with advanced or metastatic melanoma based on the current scientific evidence on biomarker use. This document thus provides an opportunity to improve healthcare efficiency and resource use, which will benefit these patients. Based on the data available so far, this expert group recommends routinely testing patients with metastatic melanoma for BRAF mutation status, as the result affects the subsequent therapeutic management of these patients. The analysis of genetic alterations in KIT may be reasonable in patients with primary tumours in acral or mucosal sites or on chronically sun-exposed skin, in an advanced condition, but not in patients with other types of melanomas. This panel believes that testing for other genetic alterations, such as NRAS mutation status in patients not carrying BRAF mutations, GNAQ/GNA11 mutational analysis or genetic alterations in PTEN, is not currently indicated as routine clinical practice, because the results do not influence treatment planning in these patients at the present time. Other important issues addressed in this document are the organisational requirements and quality controls needed for proper testing of these biomarkers, and the legal implications to be borne in mind.

Histological specificity of alterations and expression of KIT and KITLG in non‐small cell lung carcinoma

Characterization of molecules within important oncogenetic pathways may have future implications for development of therapies and biomarkers in lung cancer. One such target is the tyrosine kinase receptor KIT (c-KIT). We evaluated alterations and expression of KIT and its ligand, KITLG (also known as SCF), in 72 clinical lung tumor specimens of different histologies. Gene copy number, mRNA expression levels, and protein expression were assayed using array-based comparative genomic hybridization, real-time quantitative reverse transcription PCR and immunohistochemistry, respectively. For validation, we investigated copy number alterations and mRNA expression in external microarray data sets of 1,600 and 555 primary lung tumors, respectively. Positivity for KIT staining was most common in large cell neuroendocrine carcinoma (LCNEC) which also showed the highest KIT mRNA expression levels whereas expression was lowest in squamous cell carcinoma (SqCC). KIT mRNA expression levels were higher in KIT immunopositive samples, but expression was not affected by KIT copy numbers. Copy number gains of KIT were significantly more frequent in SqCC compared with adenocarcinoma in our own series and in the 1,600-sample data set. Immunopositivity for both KIT and KITLG in the same tumor was rare except in LCNEC. Our results highlight an increased KIT mRNA expression and frequent KIT immunopositivity in LCNEC but point out a poor correlation between KIT copy numbers and expression in SqCC, perhaps reflecting the existence of a protective mechanism against KIT alterations in this subgroup.

Another Option in Our KIT of Effective Therapies for Advanced Melanoma

Another Option in Our KIT of Effective Therapies for Advanced Melanoma

Targeting Activated KIT Signaling for Melanoma Therapy

Melanomas are composed of several biologically distinct categories of neoplasms that differ in their clinical and histologic presentations, cell of origin, age of onset, ethnic variation, pathogenetic role of ultraviolet radiation, predisposing germline alterations, type of genomic instability, pattern of metastasis, and patterns of somatic mutation. The latter contain a set of recurrent gain-of-function mutations in the genes NRAS, BRAF, KIT, GNAQ, and GNA11 that tend to occur in a mutually exclusive pattern early during progression. KIT is a receptor tyrosine kinase that binds to stem-cell factor (SCF), activating a series of downstream effector pathways (Fig 1). KIT is of critical importance in melanocyte development. It is expressed in early melanoblasts, which receive positional cues from SCF-expressing epithelia, while they migrate dorsoventrally through the superficial mesenchyme to reach their destination in the epidermis and hair follicles. Loss-of-function mutations reducing KIT signaling lead to distinctive pigment phenotypes, such as piebaldism, caused by an absence of melanocytes in the skin and/or hair follicles in frontomedial regions, such as the scalp or abdomen. Genetic alterations of KIT, including somatic gain-of-function mutations and copy number increases of wild-type KIT, occur in a minority of melanomas on skin with cumulative sun-induced damage (ie, CSD melanomas); on non–hair-bearing skin, such as palms, soles, or nail apparatus (ie, acral melanomas); and on the mucosal membrane (ie, mucosal melanomas). These melanomas are typically not associated with melanocytic nevi, and melanocytic nevi presenting on acral skin do not seem to harbor KIT mutations. The absence of a

Abstract 3039: Protein kinase C-δ mediated KIT recycling in colon cancers.

Abstract Abnormal signals through the receptor tyrosine kinase (RTK) moieties are important in colon cancer both for tumorigenesis and drug targets. Wild type KIT, one of the RTKs, had been reported to be highly expressed in some colon cancers, however little is known about the functional aspects and roles of stem cell factor (SCF) dependent activation in colon cancers. We characterized colorectal cancers with KIT expression and analyzed the biological alterations of KIT after stimulation of SCF and its impact on the downstream signals. We identified wild type KIT is expressed in some colon cancer cell lines, activated by SCF, and subsequently activated AKT and ERK signaling pathways. We also demonstrated that KIT expression was gradually decreased after SCF stimulation due to lysosomal degradation of activated KIT. We found PKC activation rescues KIT by inhibiting ubiquitination of KIT after SCF treatment. Among the various types of PKCs, PKC-δ directly bound to wild type KIT and rescued SCF-KIT complexs. Additionally, we demonstrated that KIT and PKC-δ are concomitantly activated in the colon cancer tissues with wild type KIT expression, and the expression of KIT is correlated with poor survival. Based on these findings we concluded that downstream signal activation after SCF-KIT binding is continuously accomplished by the PKC-δ activation and subsequent KIT recycling. These signal pathways induced by SCF and PKC-δ mediated KIT recycling play important roles in colon cancer progression. Citation Format: Mi Sun Park, Won Kyu Kim, Meiying Song, Hoguen Kim. Protein kinase C-δ mediated KIT recycling in colon cancers. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3039. doi:10.1158/1538-7445.AM2013-3039

Lack of Correlation Between Immunohistochemical Expression of CKIT and KIT Mutations in Atypical Acral Nevi

Given the correlation between KIT mutations and immunohistochemical expression of CKIT in acral melanoma, our aim was to confirm the utility of CKIT detection as a screening tool for KIT genotyping in atypical acral nevi and to ascertain the frequency of KIT mutations in the same. Immunohistochemical staining for CKIT was performed and staining criteria were the following: negative = <10%, 1 = 11%-49%, and 2 = >50% of cells. Intensity grading was as follows: negative = 0, weak = 1, moderate = 2, and strong = 3. Genomic amplification was performed on KIT exons commonly mutated in acral melanomas (11, 13, and 17) from atypical acral nevi (23) ranging in severity from mild (9), moderate (10), and severe (4). The control group included acral nevi without atypia (19). For purposes of statistical analyses, cases with 11% or more staining of cells were compared with negative cases and cases with a staining intensity of 1 or higher were compared with the negatives. Immunohistochemical analyses revealed the following: positive staining with an intensity 1 or more in 18 of 22 (82%) of cases with atypia (5 mild; 9 moderate and 4 severe) and in 13 of 17 (76%) nevi without atypia with no statistically significant differences between both groups. Genomic analyses of exon regions revealed no abnormalities in "hotspots" frequently associated with point mutations in acral melanomas. Our findings indicate a lack of correlation between immunohistochemical expression of CKIT and KIT mutations in atypical acral nevi. Atypical acral nevi do not exhibit genetic alterations in KIT associated with acral melanomas.

Nilotinib in patients with metastatic melanoma harboring KIT gene aberration

Patients with metastatic melanoma have a poor prognosis and few treatment options are available. We evaluated the anti-tumor activity and safety of nilotinib, a KIT inhibitor, in patients with metastatic melanoma harboring KIT alterations, either mutations or amplifications. This study was open-label, single center, prospective phase II clinical trial. Between October 2009 and April 2011, 11 patients with metastatic melanoma harboring KIT gene mutations or KIT gene amplifications were enrolled in the first stage of phase II study and nilotinib was administered orally at a dose of 400 mg twice a day until disease progression or intolerable toxicities. The primary endpoint was response rate and secondary endpoints were safety, progression-free survival (PFS) and overall survival (OS). Of 11 patients, 9 patients were evaluable for treatment response. Of 9 patients, three patients had KIT mutations in exon 11, Leu576Pro, Val559Ala and Lys558Arg; and 6 patients had KIT amplifications > 50 copies compared to control DNA. Two patients achieved partial response (22.2%) and 5 patients achieved stable disease (55.6%). In two patients who responded to nilotinib, both had KIT mutations and showed durable response for 8.4 months and 10.0+ months. Of note, one patient with KIT amplification had stable disease with response for 6 months. A decrease in tumor size from baseline was observed in four patients (44.4%). Nilotinib 800 mg/d was very well tolerated with grade 1 nausea and grade 1 dry-eye being the most common adverse events. We have decided to publish the preliminary results because anti-tumor activity of nilotinib was promising in KIT mutated patients. Although our results are preliminary, nilotinib had very favorable toxicity profile with durable response in metastatic melanoma patients with KIT mutations. The anti-tumor activity of nilotinib in melanoma patients with KIT amplification is yet to be determined in future studies. Currently, phase II nilotinib trial is ongoing in Korea as multi-center study.

KIT Pathway Alterations in Mucosal Melanomas of the Vulva and Other Sites

A significant proportion of mucosal melanomas contain alterations in KIT. The aim of this study was to characterize the pattern of KIT, NRAS, and BRAF mutations in mucosal melanomas at specific sites and to assess activation of the KIT downstream RAF/MEK/extracellular signal-regulated kinase (ERK) and phosphoinositide 3-kinase (PI3K)/AKT pathways in mucosal melanoma specimens. Seventy-one primary mucosal melanomas from various sites were studied. Mutation analysis was done by DNA sequencing. Expression of KIT, phosphorylated (p)-ERK, and p-AKT was evaluated by immunohistochemistry. KIT mutations were detected in 35% (8 of 23) of vulvar, 9% (2 of 22) of anorectal, 7% (1 of 14) of nasal cavity, and 20% (1 of 5) of penile melanomas. No KIT mutations were found in 7 vaginal melanomas. The difference in KIT mutation frequency between vulvar and nonvulvar cases was statistically significant (P = 0.014). The overall frequencies of NRAS and BRAF mutations were 10% and 6%, respectively. Notably, vaginal melanomas showed a NRAS mutation rate of 43%. KIT gene amplification (≥4 copies), as assessed by quantitative real-time PCR, was observed in 19% of cases. KIT expression was associated with KIT mutation status (P < 0.001) and was more common in vulvar than nonvulvar tumors (P = 0.016). Expression of p-ERK and p-AKT was observed in 42% and 59% of tumors, respectively, and occurred irrespective of KIT/NRAS/BRAF mutation status. NRAS mutation was associated with worse overall survival in univariate analysis. Results show that KIT mutations are more common in vulvar melanomas than other types of mucosal melanomas and that both the RAF/MEK/ERK and PI3K/AKT pathways are activated in mucosal melanoma specimens.