VOLUME 22 䡠 NUMBER 22 䡠 NOVEMBER 15 2004

JOURNAL OF CLINICAL ONCOLOGY O R I G I N A L R E P O R T

Prevalence of KIT Expression in Human Tumors

From the Institute of Pathology,
University of Basel; Novartis Pharma
AG, Basel, Switzerland; and Diomeda
Life Sciences Inc, Rockville, MD.
Submitted October 21, 2003; accepted
July 15, 2004.
The first two authors contributed
equally to this work.
Authors’ disclosures of potential con-
flicts of interest are found at the end of
this article.
Address reprint requests to Guido
Sauter, MD, Institute of Pathology,
University of Basel, Schönbeinstrasse
40, 4031 Basel, Switzerland; e-mail:
guido.sauter@unibas.ch.
© 2004 by American Society of Clinical
Oncology
0732-183X/04/2222-4514/$20.00
DOI: 10.1200/JCO.2004.10.125
Philip Th. Went, Stephan Dirnhofer, Marcel Bundi, Martina Mirlacher, Peter Schraml, Sara Mangialaio,
Sasa Dimitrijevic, Juha Kononen, Alessandro Lugli, Ronald Simon, and Guido Sauter
A B S T R A C T
Purpose
KIT is a target for imatinib mesylate (Gleevec; Novartis Pharma, Basel, Switzerland).
Gastrointestinal stromal tumors (GISTs) express KIT and respond favorably to imatinib
therapy. To determine other tumors in which such a molecular targeted therapy might be
indicated, we investigated KIT expression in different human tumor types. Because recent
studies in GISTs suggest that KIT-activating mutations predict response to imatinib therapy, we
also sequenced a subset of positive tumors.
Materials and Methods
More than 3,000 tumors from more than 120 different tumor categories were analyzed by
immunohistochemistry in a tissue microarray format. Seven commercially available anti-KIT
antibodies were initially evaluated. The antibody A4502 (DAKO) was selected for analysis
because of a high frequency of positivity in GIST and low staining background in other
tissues. To determine the frequency of KIT mutations in various tumor types, the exons 2,
8, 9, 11, 13, and 17 (where mutations previously were reported) were sequenced in 36
tumors with strong KIT expression.
Results
KIT positivity was detected in 28 of 28 GISTs (100%), 42 of 50 seminomas (84%), 34 of 52
adenoid-cystic carcinomas (65%), 14 of 39 malignant melanomas (35%), and eight of 47
large-cell carcinomas of the lung (17%), as well as in 47 additional tumor types. KIT
mutations were found in six of 12 analyzed GISTs, but only in one of 24 other tumors.
Conclusion
The results suggest that KIT expression occurs infrequently in most tumor types and that,
with the exception of GISTs, KIT gene mutations are rare in immunohistochemically
KIT-positive tumors.

J Clin Oncol 22:4514-4522. © 2004 by American Society of Clinical Oncology

high (90% to 95%) that immunohistochem-

INTRODUCTION
KIT (CD117) is a transmembrane tyrosine
kinase that acts as a receptor for mast cell
growth factor (also known as stem cell factor
or kit ligand). It belongs to the type III fam-
ily of receptor kinases and can be detected in
several normal cell types including hemato-
poietic cells, germ cells, interstitial cell of
Cajal, ductal breast epithelium, mast cells,
and melanocytes.1-5 KIT expression has
been detected in a variety of different tumor
entities. In gastrointestinal stromal tumors
(GISTs), the frequency of KIT positivity is so
ical KIT detection is considered a prerequi-
site for the histologic diagnosis of GISTs. A
smaller fraction of KIT-positive specimens
has been described in at least 46 additional
tumor entities.1,3,4,6-10
KIT expression in malignant tumors is
of topical interest because KIT is one of the
targets of the tyrosine kinase inhibitor ima-
tinib mesylate (STI571, Gleevec). Imatinib
initially was shown to be effective in the
treatment of chronic myeloid leukemia, in
which it targets the kinase function of the
BCR/ABL fusion protein.11,12 Subsequently,

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 KIT in Human Tumors
significant treatment responses were also reported in pa-
tients with advanced KIT-positive GIST.13-15 It is has been
suggested that the response rate to imatinib may be partic-
ularly high in KIT-expressing tumors that also harbor acti-
vating KIT mutations.16
Previous studies investigating KIT expression in hu-
man tumors have used a variety of antibodies, staining
protocols, and scoring criteria,1,3,4,6-10,17,18 which reduce
the comparability of these studies. Thus, identification of
tumor entities that may benefit from imatinib therapy re-
quires investigation of KIT expression in a wide variety of
tumors using comparable assessment protocols. To this
end, we used a multitumor tissue microarray (TMA) con-
taining more than 3,500 paraffin-embedded tumor samples
representing more than 120 tumor types and subtypes19 to
evaluate the epidemiology of KIT expression across a di-
verse array of human tumors. In addition, selected KIT-
positive tumors were also sequenced to elucidate the
epidemiology of KIT gene mutations.
MATERIALS AND METHODS
TMA
A total of 3,911 tissue samples from the archives of the Insti-
tute of Pathology, University of Basel (Basel, Switzerland), were
assessed including 3,556 primary tumors from 134 tumor types
and subtypes, and 355 samples from 34 different normal tissues.
Tissues were fixed in formalin for variable time periods and then
embedded in paraffin. TMA construction was as described previ-
ously.19 Briefly, tissue cylinders with a diameter of 0.6 mm were
punched from representative tumor areas of a donor tissue block
using a semiautomated precision instrument and brought into
seven different recipient paraffin blocks each containing between
400 and 612 individual samples. Four-micrometer sections of
the resulting multitumor TMA blocks were transferred to an
adhesive-coated slide system (Instrumedics Inc., Hackensack, NJ).
Immunohistochemistry
Seven antibodies were initially evaluated on 120 different
tumor samples enriched with 15 GIST and 105 tumors derived
from known KIT-positive and -negative entities as well as 30
different normal tissue types. Various pretreatment protocols on
this specially constructed KIT test array were used and the follow-
ing antibodies were tested: DAKO A4502 (dilutions 1:10 to 1:600),
Novocastra NCL-CD117 (dilutions 1:10 to 1:80), NeoMarkers
MS-271 (dilutions 1:100 to 1:1,000), MBL code No. 566 (dilutions
1:200 to 1:400), Santa Cruz sc-1494 (1:10 to 1:10,000), Santa Cruz
sc-13508 (dilutions 1:10 to 1:10,000), and Santa Cruz sc-168 (di-
lutions 1:10 to 1:200,000). For all of these antibodies, endogenous
peroxidase was blocked using 0.3% hydrogen peroxidase diluted
in methanol for 30 minutes. Standard avidin biotin technique
(Vector ABC kit; Vector Laboratories, Burlingame, CA) was used
with diaminobenzidine for visualization. A preabsorption experi-
ment using CD117 peptide stock solution (Neomarkers PP1518;
NeoMarkers, Freemont, CA) was used as a negative control; this
antigen matches exactly the peptide used by DAKO to generate the
polyclonal antibody A4502.
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In normal tissues, a cell type–specific distribution of KIT
expression was recorded (⫹, weak; ⫹⫹, moderate; ⫹⫹⫹,
strong). For tumor tissues, the percentage of positive cells was
estimated and the staining intensity was semiquantitatively re-
corded as 1⫹, 2⫹, or 3⫹. For statistical analyses, the staining
results were categorized into four groups. Tumors without any
staining were considered negative. Tumors with 1⫹ staining in-
tensity in less than 60% of cells and 2⫹ intensity in less than 30%
of cells were considered weakly positive. Tumors with 1⫹ staining
intensity in ⱖ 60% of cells, 2⫹ intensity in 30% to 79%, or 3⫹
intensity in less than 30% were considered moderately positive.
Tumors with 2⫹ intensity in ⱖ 80% or 3⫹ intensity in ⱖ 30% of
cells were considered strongly positive. Only membranous and
membranous or cytoplasmic staining was considered for analysis
because cytoplasmic staining alone proved to be false-positive in
all preabsorption control experiments.
c-KIT Sequence Analysis
Thirty-six tumors (18 seminomas, 12 GISTs, two melanomas,
two small-cell lung carcinomas, one giant cell tumor of the tendon
sheath, and one lymphoepithelial carcinoma of the pharynx) showing
strongly positive KIT staining were selected for mutation analysis.
Deparaffinization of the formalin-fixed tissues and DNA extraction
were performed according to established protocols (Qiagen, Basel,
Switzerland). The exons 2, 8, 9, 11, 13, and 17 of the KIT gene, where
mutations were previously reported, were amplified using a semi-
nested polymerase chain reaction approach. All exons were se-
quenced directly using Big Dye Terminators Cycle Sequencing Ready
Reaction Kit (Applied Biosystems, Foster City, CA). Primers de-
signed for polymerase chain reaction and sequence reactions
are listed in Table 1. Sequence products were analyzed on an
ABI Prism 310 Genetic Analyzer (Applied Biosystems).
RESULTS
Antibody Evaluation
The results of our comparison of seven different anti-
bodies are listed in Table 2. The antibodies sc-1494, sc-168,
Table 1. PCR Primers for KIT Mutation Analysis
Exon Primers Used for First and Seminested PCR
2 Forward 5⬘-CATCCATCCATCCAGGAAAA-3⬘
Reverse 5⬘-ACTGCAGAAAGCCAAGCATT-3⬘
Nested reverse 5⬘-GTTGGTGCACGTGTATTTGC-3⬘
8 Forward 5⬘-GCTGAGGTTTTCCAGCACTC-3⬘
Reverse 5⬘-CAGTCCTTCCCCTCTGCAT-3⬘
Nested reverse 5⬘-CCTCTGCTCAGTTCCTGGAC-3⬘
9 Forward 5⬘-TCCTAGAGTAAGCCAGGGCTT-3⬘
Reverse 5⬘-TGGTAGACAGAGCCTAAACATCC-3⬘
Nested forward 5⬘-AGCCAGGGCTTTTGTTTTCT-3⬘
11 Forward 5⬘-CCAGAGTGCTCTAATGACTG-3⬘
Reverse 5⬘-AGCCCCTGTTTCATACTGAC-3⬘
Nested reverse 5⬘-ACTCAGCCTGTTTCTGGGAAACTC-3⬘
13 Forward 5⬘-GCTTGACATCAGTTTGCCAG-3⬘
Reverse 5⬘-AAAGGCAGCTTGGACACGGCTTTA-3⬘
Nested forward 5⬘-TGACATCAGTTTGCCAGTTG-3⬘
17 Forward 5⬘-TCCTTACTCATGGTCGGATC-3⬘
Reverse 5⬘-AAGAGACGAACTGTCAGGAC-3⬘
Nested reverse 5⬘-ACTGTCAAGCAGAGAATGGG-3⬘
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 Went et al

Table 2. Sensitivity and Specificity of Seven Different Anti-KIT

Antibodies on a Test Array
% GISTⴱ % Other Tumorsⴱ Background Staining
Antibody (n ⫽ 8) (n ⫽ 72) in Stromal Cells

A4502 100 54 ⫺
Nr. 566 88 40 ⫺
NCL-CD117 50 42 ⫺
sc-13508 13 3 ⫺
sc-1494 88 71 ⫹⫹
MS-271 100 97 ⫹⫹
sc-168 100 100 ⫹⫹⫹

Abbreviation: GIST, gastrointestinal stromal tumor.

ⴱ
Positive tumors.

and MS-271 were not pursued further because of an unac-

ceptably high level of background staining in stromal cells
(Figs 1C to 1H). No background problem was observed for
the antibodies A4502, sc-13508, No. 566, and NCL-CD117.
Among these, A4502 was selected because the data sug-
gested the highest sensitivity for this antibody. All other
remaining antibodies (sc-13508, No. 566, NCL-CD117)
failed to stain all GISTs and yielded a lower rate of positivity
in our non-GIST test tumors. Examples of GIST that could
not be stained with sc-13508 and NCL-CD117 are shown in
Figs 1I to 1L. Overall, our comparative data suggested that
A4502 was the most sensitive and specific antibody. In
addition, A4502 had the advantage that a specific peptide
was available for negative control experiments, which
blocked specific reactions by adding a surplus of KIT anti-
gen. A4502 was therefore selected to analyze the large mul-
titumor TMA. A4502 was optimally applied at a 1:300
dilution at room temperature for 2.5 hours, after 3 minutes
of cooking under pressure in 10 mmol/L sodium citrate
buffer (pH 6.0) for antigen retrieval.
KIT Immunostaining in Normal Tissues
In normal tissues, an unequivocal membranous KIT
staining was observed in the following cell types: secretory
cells of mammary glands (⫹⫹), basal cells of skin (⫹⫹),
thymic epithelial cells (⫹⫹), mast cells (⫹⫹), interstitial
cells of Cajal (⫹⫹), and spermatogonia in testicular tubules Fig 1. Evaluation of seven different anti-KIT antibodies. (A to H) KIT
(⫹). Organs found to be completely KIT-negative included immunostaining of a gastrointestinal stromal tumor (GIST) and a malignant
cerebrum, uterine cervix, colon, endometrium, esophagus, schwannoma using the antibodies A4502 [(A) GIST; (B) malignant schwan-
noma], sc1494 [(C) GIST; (D) malignant schwannoma], MS-271 [(E) GIST; (F)
fat tissue, gall bladder, heart, kidney, liver, lung, lymph malignant schwannoma], and sc-168 [(G) GIST; (H) malignant schwannoma].
node, myometrium, oral cavity mucosa, ovary, pancreas, (I to L) Samples of GIST stained with A4502 [considered to be most
sensitive: (I) and (K)] and the antibodies (J) NCL-CD117 and (L) sc-13508. (I)
parathyroid, salivary gland, prostate, skeletal muscle, small and (J), and (K) and (L) are from identical tumors, respectively.
intestine, intestinal smooth muscle, stomach mucosa, thy-
roid, and urothelium.
KIT Immunostaining Tumors tumors with purely cytoplasmic staining showed reduction
Staining was considered true-positive if the reaction of staining after the preabsorption control, confirming false-
product was localized to the cell membrane alone or to the positive staining. At least occasional positivity was seen in 52
cell membrane and cytoplasm simultaneously. In these of 134 tumor entities (Table 3). The epidemiologically most
cases, preabsorption controls were negative. None of 142 important KIT-positive tumor entities included GIST (100%),

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 KIT in Human Tumors

Table 3. KIT-Negative Tumors Table 3. KIT-Negative Tumors (continued)

Tumor Entity No. Tumor Entity No.

Adenocarcinoma of colon 47 Neurofibroma 38

Adenocarcinoma of esophagus 7 Non-Hodgkin’s lymphoma, diffuse large B cell 22
Adenocarcinoma of gall bladder 27 Non-Hodgkin’s lymphoma, others 26
Adenocarcinoma of urinary bladder 5 Opticus glioma 1
Adenocarcinoma of uterus 1 Rhabdomyosarcoma 14
Adenolymphoma of salivary gland 6 Schwannoma 43
Adenoma of adrenal gland 14 Serous carcinoma of endometrium 22
Adenomatoid tumor 10 Serous carcinoma of ovary 48
Alveolar sarcoma 1 Small-cell carcinoma of esophagus 1
Squamous cell carcinoma of larynx 44
Acute myelogenous leukemia 1
Squamous cell carcinoma of anus 4
Anaplastic carcinoma of thyroid 6
Squamous cell carcinoma of oral cavity 47
Angiomyolipoma 1
Squamous cell carcinoma of penis 39
Angiosarcoma 4
Squamous cell carcinoma of skin 41
Apocrine carcinoma of breast 3
Squamous cell carcinoma of vagina 5
Astrocytoma 41
Synovial sarcoma 3
Benign histiocytoma 30
Teratoma of testis 14
Capillary hemangioma 25
Tubular carcinoma of breast 28
Carcinoid tumor 46
Yolk sac tumor of ovary 1
Carcinoma of adrenal gland 5
Carcinosarcoma of uterus 6 Abbreviation: MALT, mucosa-associated lymphoid tissue.
Chronic myelogenous leukemia 5
Colon adenoma, mild dysplasia 40
Colon adenoma, moderate dysplasia 42
Colon adenoma, severe dysplasia 39 seminoma (84%), malignant melanoma (36%), follicular
Craniopharyngeoma 4 carcinoma of the thyroid (23%), large-cell carcinoma (17%),
Cribriform carcinoma of breast 6
and small-cell carcinoma of the lung (7%). No positive sam-
Dermatofibrosarcoma protuberans 3
Diffuse adenocarcinoma of stomach 24 ples were found in 81 tumor types (Table 4). Examples of
Endometrioid stromal sarcoma 3 KIT-positive and KIT-negative tumors are shown in Fig 2. A
Endometrioid carcinoma of endometrium 47 total of 355 (9%) of the 3,911 arrayed tissue spots were nonin-
Endometrioid carcinoma of ovary 46 formative because of missing tissue or a lack of tumor cells in
Ependymoma 10 the arrayed tissue.
Epithelioid hemangioma 1
Epithelioid sarcoma 2 Mutation Analysis
Esthesioneuroblastoma 3 Mutations in exon 11 were seen in six of 12 GISTs
Giant cell tumor of tendon sheath 31 (50%) and one of two KIT-expressing melanomas. The
Granular cell tumor 7
types of mutations are shown in Table 5. No mutations
Hemangiopericytoma 9
Hepatocellular carcinoma 40
in exons 2, 8, 9, 11, 13, and 17 were found in seminoma
Hodgkin’s lymphoma 54 (n ⫽ 18), giant-cell tumor of the tendon sheath (n ⫽ 1),
Hormone-refractory carcinoma of prostate 39 lymphoepithelial carcinoma of the pharynx (n ⫽ 1), or
Inverted papilloma of urinary bladder 1 small cell lung cancer (n ⫽ 2).
Kaposi’s sarcoma 24
Leiomyoma 62
Leiomyosarcoma 37 DISCUSSION
Lipoma 25
Liposarcoma 28
Lobular carcinoma of breast 44 After trastuzumab (Herceptin) for treatment of HER-2–
Lymphoepithelial carcinoma of pharynx 5 positive breast cancer, imatinib therapy of KIT-positive
Malignant fibrous histiocytoma 29 GISTs represents another example of a U.S. Food and Drug
Malignant schwannoma 8 Administration (FDA) –approved rationally targeted can-
MALT lymphoma 43
cer therapy requiring immunohistochemical tumor analy-
Medullary carcinoma of thyroid 9
Medulloblastoma 5
sis to identify patients most amenable to such therapy.
Merkel cell carcinoma of skin 6 However, the chronology of the development of drug
Mucinous carcinoma of breast 26 and diagnostic tools differs between trastuzumab and ima-
Mucinous carcinoma of ovary 16 tinib. Trastuzumab was specifically designed to target the
Mucoepidermoid carcinoma of salivary gland 5 HER-2 protein, and a diagnostic kit to identify potentially
responding tumors was developed early and approved by

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 Went et al

Table 4. KIT Protein Expression in Human Tumors

KIT Immunostaining
Tumor Entity No.ⴱ Weak (No.ⴱ) Moderate (No.ⴱ) Intense (No.ⴱ) Total Positive (%)

Neural tumors
Glioblastoma multiforme 48 0 0 1 2
Meningeoma 44 0 1 0 2
Oligodendroglioma 25 2 1 0 12
Skin tumors
Skin, basalioma 41 1 1 0 4
Skin, benign appendix tumor 28 3 3 1 25
Skin, benign nevus 42 1 2 1 9
Skin, malignant melanoma 39 5 3 6 36
Hemato-lymphoid neoplasms
Thymoma 24 0 1 0 4
Soft tissue tumors
Fibrosarcoma 9 1 0 11
Glomus tumor 9 1 0 0 11
PNET 17 0 2 1 17
Male genital tumors
Prostate carcinoma, untreated 55 1 0 0 2
Testis, nonseminomatous germ cell tumor 48 2 0 0 4
Testis, seminoma 50 4 8 30 84
Breast
Breast, ductal carcinoma 49 2 1 0 6
Breast, medullary carcinoma 28 0 3 3 21
Breast, phylloides tumor 13 1 1 4 46
Female genital tumors
Ovary, Brenner tumor 9 0 0 3 33
Ovary, dysgerminoma 2 0 0 2 100
Ovary, gonadoblastoma 1 0 0 1 100
Ovarian carcinoma, other types 15 0 1 0 6
Uterus cervix, squamous cell carcinoma 38 1 1 0 5
Uterus cervix, CIN III 18 1 0 0 5
Uterus cervix, adenocarcinoma 3 0 0 1 33
Vulva, squamous cell carcinoma 42 0 0 1 2
Upper aerodigestive and respiratory tract
Lung, adenocarcinoma 48 7 0 2 18
Lung, large-cell carcinoma 47 4 0 4 17
Lung, small-cell carcinoma 46 0 0 3 6
Lung, squamous cell carcinoma 49 1 0 0 2
Malignant mesothelioma 23 1 0 0 4
Salivary gland, acinus cell carcinoma 7 1 0 0 14
Salivary gland, adenoid-cystic carcinoma 52 5 14 15 65
Salivary gland, pleomorphic adenoma 47 8 2 0 21
Gastrointestinal tract
Esophagus, squamous cell carcinoma 34 0 2 0 5
GIST 28 0 3 25 100
Pancreas, adenocarcinoma 48 0 1 0 2
Small intestine, adenocarcinoma 11 0 1 0 9
Stomach, intestinal adenocarcinoma 47 0 1 1 4
Neuroendocrine tumors
Paraganglioma 7 0 0 1 14
Parathyroid, adenoma 15 0 0 1 6
Pheochromocytoma 28 0 1 0 3
Thyroid, adenoma 39 2 9 3 35
Thyroid, follicular carcinoma 30 1 5 1 23
Thyroid, papillary carcinoma 32 0 0 1 3
(continued on next page)

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 KIT in Human Tumors

Table 4. KIT Protein Expression in Human Tumors (continued)

KIT Immunostaining
Tumor Entity No. ⴱ
Weak (No. )ⴱ
Moderate (No.ⴱ) Intense (No.ⴱ) Total Positive (%)

Urinary tract
Kidney, chromophobic carcinoma 13 2 2 2 46
Kidney, clear cell carcinoma 46 1 1 0 4
Kidney, oncocytoma 10 2 3 3 80
Kidney, papillary carcinoma 40 1 0 0 2
Urinary bladder, transitional cell 44 2 0 0 4
carcinoma, invasive (pT2-4)
Urinary bladder, transitional cell 40 1 3 2 15
carcinoma, noninvasive (pTa)
Urinary bladder, sarcomatoid carcinoma 8 1 0 0 12
Urinary bladder, squamous cell carcinoma 6 1 0 0 16

Abbreviations: PNET, primitive neuroectodermal tumor; CIN, cervical intraepithelial neoplasia; GIST, gastrointestinal stromal tumor.
ⴱ
Sum of arrayed spots.

the FDA simultaneously with the drug in September 1998.20
Imatinib was initially designed to inhibit the BCR/ABL
fusion protein in chronic myeloid leukemia and was ap-
proved for this application by the FDA in May 2001.21 Only
later was it discovered that imatinib is also effective against
KIT-positive GISTs.14 More than 25 clinical studies are now
investigating the effect of imatinib on KIT-positive tumors
of various origins (http://www.clinicaltrials.gov). Despite
of these activities, generally accepted guidelines for identi-
fication of KIT-positive tumors are still lacking. Previous
studies have therefore used a wide variety of different KIT
antibodies, protocols, and scoring systems to identify
KIT-positive tumors. These studies often yielded discrepant
results.1-4 For example, in small-cell lung cancer22,23 the
frequency of KIT positivity ranged from 36%3 to 91%1 and
in malignant melanoma from 0%4 to 20%.7 Because of the
potential importance of assessing KIT expression for treat-
ment decisions, a reliable procedure to identify KIT overex-
pression is needed.
The TMA approach is ideally suited for development
and comparison of immunohistochemical assays. Hun-
dreds of tumors can be immunostained simultaneously
under highly standardized conditions on one TMA section.
The use of consecutive sections of a TMA block in combi-
nation with the small diameter of each arrayed tissue sample
limits each comparison to a small tissue area with a minimal
likelihood of genetic or tissue processing heterogeneity.
Our initial comparison of seven commercially available
antibodies showed considerable variance in staining inten-
sity and signal-to-noise ratio. The A4502 antibody was se-
lected because it had the highest sensitivity and produced
minimal background in other tissues. A4502 recognizes an
intracellular component of the transmembranous protein,
963 to 976 amino acids to the C-terminal. It was therefore
not surprising that our preabsorption experiments always
confirmed membranous or membranous and cytoplasmic
staining as specific, whereas tumors with pure cytoplasmic
KIT positivity proved to be false-positive.
Multitumor TMAs containing samples from a wide
variety of tumor entities are optimally suited to identify
those samples with frequent alterations of a specific gene.24
This holds true even though the absolute number of posi-
tive cases detected in a TMA study may be somewhat lower
than the true number of tumors due to regional heteroge-
neity of immunostaining. This presumed weakness of the
TMA approach, however, is apparently compensated for by
the perfect standardization of staining. All tumors of a TMA
study can be stained under absolutely identical reaction
conditions. Previous TMA studies have shown that repre-
sentative information is obtained despite the small size of
arrayed tissue per tumor.25-28 On a multitumor TMA, the
most valuable information is the relative frequency of mo-
lecular parameters across all tumor types, which results in a
rank order of potentially affected tumors. The results of this
study confirm that KIT expression is a consistent finding in
GIST and, as reported in previous studies, relatively fre-
quent in melanoma, seminoma, and adenoid-cystic carci-
noma.1,3 An additional 48 tumor types and subtypes with
varying frequencies of KIT expression were also identified
by our TMA approach.
The development of a KIT immunostaining protocol,
which is well supported by experimental data, does not
automatically solve the problem of immunohistochemical
KIT detection in clinical praxis. The experience with HER-2
testing of tumors potentially suited for trastuzumab therapy
has highlighted several inherent difficulties of immunohis-
tochemical testing. Immunohistochemistry has been shown to
be highly dependent on preanalytical handling (especially fix-
ation), and significant interlaboratory variations have been
reported despite the availability of high-quality FDA-approved
immunohistochemical test kits.29-31 These difficulties are seen
especially in low-throughput laboratories.32,33 In any case,

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 Went et al

Fig 2. Examples of immunohistochemical KIT staining in various tumors using the antibody A4502: (A) positive gastrointestinal stromal tumor; (B) positive
seminoma; (C) positive malignant melanoma; (D) KIT-negative malignant melanoma showing mast cells as positive internal controls.

using immunohistochemistry for predictive tumor analysis
will necessitate rigorous quality-assurance steps and highly
skilled certified laboratories. It is also noteworthy that immu-
nohistochemistry testing may be even more difficult and per-
haps less relevant for targets other than HER-2. For example,
initial studies using anti– epidermal growth factor receptor
drugs have shown little influence of the epidermal growth
factor receptor expression level (as detected by immunohisto-
chemistry) on response to therapy.34,35 Other targets such
as CD20, as a target for rituximab, are expressed in all

Table 5. KIT Mutations

Tumor Type Type of Mutation Exon Codon

GIST 2 deletions (11 and 1 bp) 11 552-555 and 557

GIST Deletion and transversion 11 570-577 and 578 tyr3phe
GIST Deletion (9 bp) 11 557-560
GIST 2 transversions 11 558 lys3glu; 559 val3asp
GIST Insertion (42 bp) 11 586 ⬍ 573-586 ⬎ 587
GIST Deletion (6 bp) 11 557-558
Melanoma Transition 11 576 leu3pro

NOTE. A total of seven mutations were found in 36 sequenced tumors.

Abbreviation: GIST, gastrointestinal stromal tumor.

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 KIT in Human Tumors

tumors of a certain type, making testing unnecessary
once a definite diagnosis is established.36
Recent data have indeed suggested that response to
imatinib may not be driven primarily by the KIT expression
level. Studies have shown that not all KIT-expressing tu-
mors will benefit from imatinib therapy.37 They indicate
that the response rate may depend on the presence of KIT
mutations in the tumor and potentially also on the location
and type of mutation.38,39 These data suggest that tumors
with exon 11 mutations respond better than those tu-
mors with exon 9 mutations, whereas the response rate is
minimal in tumors without mutations.16 Our sequencing
analysis revealed 50% exon 11 mutations in GISTs, which is
in the range of previous studies.40 The results of sequencing
analyses are more controversial in seminomas. In this study,
we failed to find mutations in 18 examined seminomas,
despite a comprehensive analysis of exons 2, 8, 9, 11, 13, and
17. Previous studies analyzing a total of 108 pure semino-
mas and dysgerminomas have found 1% exon 11 and 20%
exon 17 mutations.38,41-43 The only tumor type other than
GIST that showed a KIT mutation in our study was mela-
noma. However, our finding of an exon 11 mutation in one
of two melanomas analyzed seems to be an overestimate
of the true mutation prevalence. In a follow-up study, we
were unable to detect KIT mutations in 10 additional
KIT-expressing melanomas.44 Overall, the prevalence
of KIT mutations in tumors other than GISTs and per-
haps seminoma seems to be low. Recent studies investi-
gating 10 breast,45 10 ovarian,46 and 26 small-cell lung
cancers22 failed to find any mutations.
Our study gives a comprehensive overview of KIT ex-
pression in a diverse set of human tumors. With the excep-
tion of seminoma and melanoma, immunohistochemical
KIT-positivity was below 30% in frequently occurring tu-
mor entities. Together with the low prevalence of mutations
in KIT-expressing tumors, this suggests a relatively low
proportion of patients who might benefit from imatinib
therapy subsequent to KIT activation. However, KIT pro-
tein is not the only target of imatinib. In addition to chronic
myelogenous leukemia, responses to imatinib are known to
occur in dermatofibrosarcoma protuberans,47-49 chronic
myelomonocytic leukemia,50 and hypereosinophilic syn-
drome,51 in which other tyrosine kinases are inhibited
by imatinib.
■ ■ ■
Acknowledgment
We thank Y. Knecht and M. Kaspar for skillful techni-
cal assistance in laboratory work, and J. Schwegler for the
photographic montage.
Authors’ Disclosures of Potential
Conflicts of Interest
The authors indicated no potential conflicts of interest.

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