BACKGROUND: In bone marrow material from patients with various leukemias we noted that samples with either a deletion on the long arm of one chromosome 7 (7q-) or a monosomy 7 had a higher telomerase activity. Considering that introduction of a chromosome 7 into a cancer cell line had been reported to eliminate telomerase activity, that 7q- is a common negative prognostic finding in cancers, and that the deleted segment (band 7q31) contains an unidentified tumor suppressor gene, we wondered if this gene might be a telomerase inhibitor. RESULTS: We found no significant difference in telomerase activity between the three groups of patient samples. In contrast to reports on tumor cell lines we observed no amplification of the telomerase genes. METHODS: We analyzed telomerase activity and copy number of the telomerase genes hTERT and hTR in frozen archival bone marrow samples from leukemia patients with a referral diagnosis of AML, and either a monosomy for chromosome 7, a deletion on the long arm of chromosome 7 (7q-), or none of these aberrations. Telomerase activity was measured with a commercially available kit, and the copy number of the telomerase genes was tested by FISH. CONCLUSIONS: We found no evidence of a telomerase inhibitor in band 7q31. The lack of telomerase gene amplification found in cell lines from solid tumors could reflect that this amplification is a property of solid tumors, not of hematological cancers

A Kallioniemi

A Zang

AI Soder

H Huang

JC Zenklusen

JW Shay

Jørn E Koch

K Nakabayashiki

L Hayflick

LA Bryce

N Serakinci

Nedime Serakinci

R Villa

S Knuutila

S Lewis

T de Lange

V Lundblad

WC Hahn

English

PubMed

Springer - Publisher Connector

BioMed CentralBMC Medical Genetics
ssOpen AcceBMC Medical Genetics 2002, 3 xResearch article
Telomerase activity in human leukemic cells with or without 
monosomy 7 or 7q-
Nedime Serakinci1 and Jørn E Koch*2
Address: 1Present Address: Institute of Human Genetics, University of Aarhus, DK-8000 Aarhus C, Denmark and 2Institute of Pathology, Aarhus 
Kommunehospital, Nørrebrogade 44, DK-8000 Aarhus C, Denmark
E-mail: Nedime Serakinci - nedi@humgen.au.dk; Jørn E Koch* - jokoc@akh.aaa.dk
*Corresponding author
Abstract
Background: In bone marrow material from patients with various leukemias we noted that
samples with either a deletion on the long arm of one chromosome 7 (7q-) or a monosomy 7 had
a higher telomerase activity. Considering that introduction of a chromosome 7 into a cancer cell
line had been reported to eliminate telomerase activity, that 7q- is a common negative prognostic
finding in cancers, and that the deleted segment (band 7q31) contains an unidentified tumor
suppressor gene, we wondered if this gene might be a telomerase inhibitor.
Results: We found no significant difference in telomerase activity between the three groups of
patient samples. In contrast to reports on tumor cell lines we observed no amplification of the
telomerase genes.
Methods: We analyzed telomerase activity and copy number of the telomerase genes hTERT and
hTR in frozen archival bone marrow samples from leukemia patients with a referral diagnosis of
AML, and either a monosomy for chromosome 7, a deletion on the long arm of chromosome 7
(7q-), or none of these aberrations. Telomerase activity was measured with a commercially
available kit, and the copy number of the telomerase genes was tested by FISH.
Conclusions: We found no evidence of a telomerase inhibitor in band 7q31. The lack of
telomerase gene amplification found in cell lines from solid tumors could reflect that this
amplification is a property of solid tumors, not of hematological cancers.
Background
Somatic cells have a limited potential for cell division, in
part due to an absence of telomerase activity in such cells
[1]. Cancer cells grow indefinitely, either depending on te-
lomerase activity or an alternative mechanism for tel-
omere elongation (ALT) [2,3]. It has long been suspected
that the absence of telomerase activity, and the resulting
limited potential for cell division, acts as an anti-oncogen-
ic mechanism in normal somatic cells. In line with this
view, recent reports demonstrated that telomerase activa-
tion was among the genetic changes needed to transform
normal cells in vitro[4], and that telomerase inhibition
caused growth arrest and cell death in cancer cell cultures
[5]. In cancer cells, telomerase activation has been de-
scribed, as well as up-regulation of preexisting telomerase
activity. However, the mechanisms for regulating of tel-
omerase activity described so far all have a quantitative ef-
Published: 29 October 2002
BMC Medical Genetics 2002, 3:11
Received: 10 July 2002
Accepted: 29 October 2002
This article is available from: http://www.biomedcentral.com/1471-2350/3/11
© 2002 Serakinci and Koch; licensee BioMed Central Ltd. This article is published in Open Access: verbatim copying and redistribution of this article are 
permitted in all media for any purpose, provided this notice is preserved along with the article's original URL.Page 1 of 6
(page number not for citation purposes)
BMC Medical Genetics 2002, 3 http://www.biomedcentral.com/1471-2350/3/11fect on the enzyme, whereas no switch closing telomerase
down has been discovered [6].
Analyzing bone marrow samples from leukaemia patients
in another study, we made the observation that samples
with either a deletion on the long arm of one chromo-
some 7 (7q-) or a full monosomy 7 displayed a signifi-
cantly higher average telomerase activity compared to
samples without these abnormalities (unpublished re-
sults). This chance observation made us wonder if chro-
mosome 7 might harbor a telomerase suppressor, the loss
of which would cause increased telomerase activity in cas-
Figure 1
CGH analysis of chromosome 7 in the 7q- samples and FISH analysis of the relevant regions on chromosomes 3, 5 and 7. A.
CGH analysis of pooled DNA from the 7q- samples hybridized against control DNA from a normal male donor. The CGH
profile for chromosome 7 is shown. A deletion is seen in the 7q31 region as indicated on the idiogram. B. Triple color hybridi-
zation for 7CEN/D7S522, hTERT, and hTR. 7CEN (orange arrows) and hTR (yellow arrows) appear in green. In the micro-
scope the hTR signal was bright and clearly visible, but in the present representation it appears weak due to the auto exposure
in the green channel being governed by the stronger 7CEN signal (compare with panel D). Both D7S522 (orange arrows) and
hTERT (red arrows) appear red in this representation. In the microscope, each of the two probes appeared in its recognizable
reddish color, but the difference could not be reproduced here. C. Dual color hybridization of hTERT (red) and chromosome
7 probes (green centromere juxtapositioned with red 7S522). While both hTERT loci are labeled, one chromosome 7 has only
a centromere signal, and not a 7q31 signal. D. Dual color hybridization of the hTR and hTERT probes showing two signals for
each probe. This was the general finding in all 65 samples. In the absence of the green centromere 7 signal the hTR probe is
clearly visible after auto exposure.Page 2 of 6
(page number not for citation purposes)
BMC Medical Genetics 2002, 3 http://www.biomedcentral.com/1471-2350/3/11es with -7 or 7q-. Looking into the literature we found that
introduction of a chromosome 7 into a telomerase posi-
tive cancer cell line eliminates the telomerase activity [7],
that 7q- is a common finding in many cancers [8,9] with
an invariably ominous prognosis, and that the commonly
deleted segment (band 7q31) contains a yet unidentified
tumor suppressor gene [10]. Based on these reports we
formed the hypothesis that the unknown tumor suppres-
sor in 7q31 might in fact be a telomerase inhibitor, per-
haps an on/off switch for telomerase activity.
Another factor that might potentially affect telomerase ac-
tivity is amplification of the telomerase genes. Such am-
plification has been reported for a number of solid tumors
and cancer cell lines [11,12], and could be speculated to
overwhelm even otherwise intact telomerase inhibition.
For the study reported here we therefore combined analy-
sis with FISH probes and measurements of telomerase ac-
tivity on all archival frozen bone marrow samples we had
available from patients with a registration as AML and cy-
togenetic finding of either -7 or 7q-. Samples without a
recognized chromosome 7 abnormality were analyzed as
control. For the FISH part of the study, probes for 7q31
were combined with probes for the two telomerase genes,
hTERT and hTR [12,13].
We found no amplification of either hTR or hTERT in any
of the tumor categories, and while FISH and CGH could
confirm the 7q31 deletions identified by G-banding, the
average telomerase activity did not differ significantly be-
tween the tumors. This result leads to two conclusions.
One is that since we found no effect on telomerase activity
of chromosome 7 aberrations in this larger patient mate-
rial, we find no evidence of a telomerase suppressor gene
in 7q31. The other conclusion is that since we found no
evidence of amplification of telomerase genes in AML, up-
regulation of telomerase activity in this type of cancer
might be different from up-regulation of telomerase activ-
ity in solid tumors.
Methods
Patient material selection
G-banding results were retrieved from the files of the de-
partment. These data had originally been obtained in the
course of routine diagnosis. Samples with a registration as
AML and a cytogenetic diagnosis of -7 or 7q- by G-band-
Figure 2
ELISA readings for telomerase activity in the three categories of patient samples.
0
0.5
1
1.5
2
2.5
3
0 1 2 3 4
GROUPS
E
L
IS
A
 R
ea
d
in
g
s
 group 1, no-del7q
 group 2, del7q
 group 3, Monosomy 7 Page 3 of 6
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BMC Medical Genetics 2002, 3 http://www.biomedcentral.com/1471-2350/3/11ing were selected. Samples in which frozen cells were also
available for telomerase activity determination were in-
cluded in the study. This gave us about 20 samples in each
of the two categories, so we added an additional 20 AML
cases without any of the abnormalities for control purpos-
es. In all three groups of patients some of the cases select-
ed eventually had to be left out for technical reasons, such
an insufficient amount of material, so the actual numbers
of samples studied became 17, 17, and 19, respectively.
There was no significant difference in age or sex distribu-
tion between the patient groups with -7, 7q- or no aberra-
tion on chromosome 7. There was also no overall
difference in the karyotype complexity between the test
and the control groups, beyond the presence or absence of
chromosome 7 aberrations.
Probes for FISH and CGH
Chromosome 7CEN/D7S522 and 7CEN7D7S486 probes
were commercially available (Vysis, Inc. Downers Grove,
IL 60515, USA, AH diagnostics, Denmark). They include
a centromere 7 probe labeled with Spectrum Green and a
locus specific probe in 7q31 labeled with Spectrum Or-
ange. FISH hybridization and post-hybridization washes
were performed as specified by the manufacturer.
The probe for the reverse transcriptase component of tel-
omerase (hTERT, band 5p15.33) was a BAC clone, and the
probe for the RNA template component (hTR gene, band
3q26) was a P1 clone. Both were kindly provided by
Nicole Keith (Glasgow, UK [12,13]). They were labeled by
standard nick translation.
Figure 3
Correlation between ELISA readings for telomerase activity and the frequency of 7q31 FISH signals in individual samples from
the 7q- patients.
0,00
0,50
1,00
1,50
2,00
2,50
3,00
1,0 1,5 2,0
Frequency of 7q31 FISH signals
E
L
IS
A
 R
ea
d
in
g
sPage 4 of 6
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BMC Medical Genetics 2002, 3 http://www.biomedcentral.com/1471-2350/3/11CGH analysis [14] of pooled DNA to determine the com-
mon deleted region in the 7q- group was performed es-
sentially as described previously by Knuutila [15]. In brief,
"test" DNA from the patients and reference DNA from a
healthy male donor was isolated with Wizard Genomic
DNA Purification Kit (Promega) and labeled by standard
nick translation. The labels used were fluorescein-12-
dUTP and tetramehylrhodamine-5-dUTP (Roche), and
both pooled patient DNA and reference DNA was labeled
in either color, so that green test could by hybridized
against red reference and red test against green reference.
FISH and CGH analysis
When surplus methanol acetic acid (standard Carnoy fix-
ative) fixed cells from the original chromosome analysis
were available these were used, otherwise freshly thawed
cells were treated with 75 mM KCl and fixed in Carnoy fix-
ative. Prior to FISH or CGH, slides were "aged" at room
temperature for 3–5 days, and denatured in 1×Tth buffer
(10 mM Tris HCL, pH 8.9 (25°C); 0.1 m KCl; 1.5 mM
MgCl2; 50 mg/ml bovine serum albumin; and 0.05% v/v
Tween 20) at 93°C for 2.5 min. Hybridizations were per-
formed as previously described [16].
Images were captured from a standard fluorescence micro-
scope (Lieca DMRB) with a SenSys CCD camera using
auto-exposure time and controlled by the "SmartCapture
VP" software (Digital Scientific Ltd.). For CGH 10 met-
aphases from each reaction was analyzed with the Quips
software (Vysis). Thresholds for gains and losses were set
at 1.20 and 0.80, respectively.
Assay for telomerase activity
Telomerase activity was measured with the commercially
available ELISA-based kit from Roche, and performed ac-
cording to the manual included with the kit. To ensure re-
producibility and that the values obtained could be
compared all samples were analysed at least twice and in
different combinations of samples.
Results and Discussion
In the 7q- patient material we were able to confirm the
presence of a common deleted region in 7q31 by CGH
with pooled patient DNA (Fig 1A). Similarly, FISH with
probes D7S486 and D7S522 confirmed that one copy of
genes in that chromosome band had been lost in a frac-
tion of cells from all the 7q- patients (Fig 1B,1C and Table
1). The control probes labeled 7q- and control cells to the
same extent (Fig 1B,1C,1D and Table 1), confirming that
Table I: Results of triple color FISH reactions with probes for 7q31, hTERT and hTR given as the average number of signals per cell in 
each sample tested.
3q (hTERC) 3q (hTERC) 5p (hTERT) 5p (hTERT) 7q (D7S486) 7q (D7S486) 7q (D7S522 7q (D7S522)
7q deletion no 7q delet 7q deletion no 7q delet 7q deletion no 7q delet 7q deletion no 7q delet
1,83 1,95 1,95 1,98 1,41 1,95 1,40 1,97
1,92 1,96 1,80 1,96 1,83 1,94 1,81 1,95
1,86 1,92 1,93 1,88 1,38 1,96 1,52 1,93
1,82 1,88 1,85 1,80 1,85 1,97 1,79 1,97
1,96 1,94 1,84 1,97 1,48 1,96 1,42 1,96
1,88 1,88 1,82 1,94 1,42 1,94 1,40 1,94
1,86 1,94 1,78 1,94 1,68 1,98 1,66 1,98
1,94 1,93 1,90 1,94 1,62 1,95 1,40 1,95
1,87 1,82 1,95 1,89 1,79 2,00 1,75 2,00
1,52 1,70 1,83 1,84 1,81 1,97 1,80 1,97
1,89 1,95 1,91 1,99 1,19 1,96 1,44 1,96
1,92 1,97 1,93 2,00 1,74 1,98 1,73 1,98
1,90 1,93 1,82 1,93 1,49 1,96 1,49 1,96
1,81 1,95 1,90 1,84 1,57 1,91 1,55 1,91
1,85 1,85 1,85 1,97 1,78 1,97 1,82 1,97
1,65 1,94 1,82 1,92 1,42 1,95 1,38 1,94
1,45 1,82 1,88 1,81 1,66 1,97 1,69 1,95
Signal frequencies were generally in the 1.8–2.0 range, except that the 7q- samples showed the expected reduction in signal frequency with the 
7q31 probes. For the hTR and hTERT probes the frequencies never exceeded 2.0, nor did we observe signals that were enhanced in intensity. In 
fact, some samples instead showed deletion of one of the genes in a fraction of cells.Page 5 of 6
(page number not for citation purposes)
BMC Medical Genetics 2002, 3 http://www.biomedcentral.com/1471-2350/3/11the missing 7q31 signals were not absent due to an arti-
fact. Surprisingly, no significant difference in telomerase
activity was observed between the -7, the 7q-, and the con-
trol samples (Fig 2). Likewise, there was no correlation be-
tween telomerase activity and the fraction of cells with 7q-
in a sample (Fig 3). Thus, we found no evidence that the
tumor suppressor in band 7q31 is involved in regulating
telomerase activity.
It has been reported that the hTERT gene is amplified in a
sizeable fraction of cancers and cancer cell lines [11,12].
The inclusion of FISH probes for hTERT and hTR in this
study provided data on the possible similar telomerase
gene amplification in AML. None of the 53 samples tested
showed such amplification (Fig 1D and Table 1I), arguing
strongly against it being a common phenomenon in AML.
Hematological cancers differ from most other cancers in
that they originate from cells with a low level of natural te-
lomerase activity. Acquiring the necessary telomerase ac-
tivity for cancer growth in leukemia thus only requires up-
regulation of telomerase activity, whereas others cancers
need to overcome a complete telomerase inhibition, pos-
sibly by amplifying telomerase genes to overwhelm tel-
omerase repressors. As an illustration of this difference
between hematological cancers and other cancers, the ALT
mechanism for telomere elongation is largely confined to
the latter group. It is therefore tempting to speculate that
telomerase up-regulation in hematological cancers and in
solid tumor may develop along different pathways.
Competing interests
None declared.
Authors' contributions
NS identified the samples, carried out the experimental
work, compiled the data, performed the statistical analy-
sis, produced the illustrations, and participated in the ed-
iting of the manuscript. JK conceived of and designed the
study, supervised the experimental work and the data
compilation, and drafted and edited the manuscript.
Acknowledgements
NS was employed on a grant to JK from the Danish Medical Research 
Council. The study was largely performed on equipment obtained on in-
strumentation grants to JK from The John and Birthe Meyer Foundation.
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Telomerase activity in human leukemic cells with or without monosomy 7 or 7q-

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Telomerase activity in human leukemic cells with or without monosomy 7 or 7q-

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