Cancer Epidemiology Biomarkers & Prevention Vol. 9, 1211-1214, November 2000
© 2000 American Association for Cancer Research
Plasma Concentrations of Reputed Tumor-associated Soluble CD44 Isoforms (v5 and v6) in Smokers Are Dose Related and Decline on Smoking Cessation
David A. Scott1,
John A. Stapleton,
Richard M. Palmer,
Ron F. Wilson,
Gay Sutherland,
Paula Y. Coward,
Gunnar Gustavsson,
Edward W. Odell and
Robin N. Poston
Dental Clinical Research [D. A. S., R. M. P., R. F. W., P. Y. C.], Oral Medicine and Pathology [E. W. O.], and Centre for Cardiovascular Biology and Medicine [R. N. P.], Guy’s, King’s and St. Thomas’ Schools of Medicine, Dentistry and Biomedical Sciences, King’s College London, London SE1 9RT, United Kingdom; Tobacco Research Section, National Addiction Centre, Institute of Psychiatry, King’s College London, London SE5 8AF, United Kingdom [J. A. S., G. S.]; and Pharmacia & Upjohn, Helsingborg S-25109, Sweden [G. G.]
 |
Abstract
|
|---|
There is some evidence to suggest that smoking may affect circulating
levels of CD44 (sCD44) molecules. Therefore, we investigated the effect
of smoking on the circulating level of sCD44 by comparing the change in
total sCD44, sCD44v5, and sCD44v6 concentrations over 1 year in a group
of people who quit smoking (n = 30) and a control
group of people who continued to smoke (n = 30).
Smoking status and compliance were monitored by analysis of plasma
cotinine and expired CO levels and also by self-reported tobacco use.
We show a dose-dependent relationship between smoke intake and baseline
plasma concentrations of reputed tumor-associated CD44 variant isoforms
(sCD44v5 and sCD44v6) in smokers (n = 60). There
was a significant decline in the level of both sCD44v5 and sCD44v6 in
quitters as compared with continuing smokers [-13.2 (95% confidence
interval, -7.6 to -18.8; P < 0.001) and -62.2
ng/ml (95% confidence interval, -33.9 to -90.6;
P < 0.001), respectively], but not in the total
sCD44 concentration. These results show that the increased
concentrations of sCD44v5 and sCD44v6 in smokers are dose related and
reversible and suggest that the attributed diagnostic specificity and
prognostic value of sCD44 molecules in malignant and inflammatory
disease may be affected by smoking status.
|
Introduction
|
|---|
Aberrant expression of variant CD44 isoforms has been identified
in certain premalignant lesions and carcinomas and has been associated
with tumor growth, metastatic potential, and poor prognosis
(1, 2, 3, 4, 5, 6, 7, 8)
. Furthermore, circulating levels of total sCD44 and
specific soluble CD44 isoforms have been shown to correlate with tumor
metastasis in some malignancies, including non-Hodgkin’s lymphoma and
breast, gastric, and colon carcinomas (9, 10, 11, 12, 13)
. The level
of soluble CD44 is also known to be higher in the body fluids of
subjects with particular inflammatory conditions, such as rheumatoid
arthritis (14
, 15)
, pouchitis and colitis
(16)
, and bronchitis (17)
. Recent evidence
suggests that tobacco smoking might lead to increased concentrations of
circulating total CD44 (sCD44) and specific tumor-associated sCD44
isoforms (18, 19, 20)
. Therefore, we examined the influence of
smoking and smoking cessation on plasma levels of total CD44 and the
reputed inflammation- and tumor-associated isoforms sCD44v6 and sCD44v5
(10
, 13
, 15
, 19, 20, 21, 22, 23, 24, 25, 26)
. Because it has been suggested that
circulating levels of sCD44v5 and sCD44v6 may have prognostic
significance in specific malignancies (13
, 22, 23, 24
, 26)
,
this issue is of primary importance.
|
Materials and Methods
|
|---|
Subjects.
Thirty subjects known to have stopped smoking completely for 52 weeks
(quitters) and a control group of 30 subjects who continued to smoke
(continuing smokers) were sampled retrospectively from the database of
a large smoking cessation treatment trial (27)
. Pulse and
blood pressure were taken at baseline. Otherwise, subjects attending
the smoking cessation clinics were not examined medically but were
apparently fit and healthy. Those with a recent history of heart or
malignant diseases were excluded. As described previously,
quitters were those who reported not smoking at all treatment
follow-ups (weeks 2, 4, 8, 12, 22, 26, and 52) and registered as
nonsmokers on analysis of expired air CO concentration (<10 ppm) at
each visit and on analysis of plasma cotinine concentration (<15
ng/ml) at 52 weeks, whereas continuing smokers were those who reported
having failed to remain abstinent from tobacco smoking throughout the
year and registered >9 ppm carbon monoxide in expired breath and a
plasma cotinine concentration of >50 ng/ml at 52 weeks
(27)
. Plasma was sampled at baseline, when all 60 subjects
were smoking normally just prior to their quit attempt, and 52 weeks
later, when 30 subjects had been abstinent from smoking for 1 year.
Measurement of Plasma Levels of Total sCD44, sCD44v5, and sCD44v6.
sCD44 concentrations were determined blind to smoking status at 52
weeks by ELISA [total sCD44, monoclonal antibody BU52 (BU52 recognizes
the standard/hematopoietic form of sCD44; R&D Systems, Abingdon,
United Kingdom); sCD44v5 and v6, monoclonal antibodies VFF8 and VFF18,
respectively (Bender Medsystems, Vienna, Austria)]. Each assay
was performed in duplicate. Absorbance was measured at 450 nm using a
Dynatech MR 700 automated microplate reader. The concentration of sCD44
was calculated from a calibration curve using duplicate standard
concentrations of CD44 standard, CD44v5, and CD44v6.
Statistical Evaluation.
The relation between baseline measures of smoke intake (expired CO,
plasma cotinine, and daily cigarette consumption) and concentrations of
sCD44 molecules was modeled by bivariate linear regression. The
response due to smoking cessation was measured as the change in sCD44
concentration between baseline and 1 year. The t test was
used to examine the difference in change between quitters and
continuing smokers.
|
Results
|
|---|
There were no significant differences in mean age [44 (SD =
10) and 43 (SD = 8) years], gender (15 females and 14 females),
or smoke intake at baseline [mean cigarette consumption, 26 (SD =
10) and 25 (SD = 7) cigarettes/day] between the group who
subsequently succeeded in quitting and those who failed to quit,
respectively. Pulse and blood pressure were taken at baseline, but
neither correlated with CO or other measures of smoke intake,
CD44v5/CD44v6, or cessation outcome.
The change in sCD44v5 and sCD44v6 concentrations over 1 year from
baseline in quitters and continuing smokers is presented in Fig. 1
. The reductions in both sCD44v5 and sCD44v6 levels were significant
when analyzed in the quitters alone at 52 weeks compared to baseline
(-16.2 and -79.4 ng/ml respectively; both P < 0.001)
and when comparing quitters with continuing smokers [mean difference
in change in sCD44v5 = -13.2 ng/ml (95% confidence interval,
-7.6 to -18.8); mean difference in change in sCD44v6 = -62.2
ng/ml (95% confidence interval, -33.9 to -90.6); both
P < 0.001]. The difference in the change in total
CD44 levels over 52 weeks in the quitters (-19.9 ng/ml) compared with
the change in continuing smokers (-11.5 ng/ml) was not significant
(P = 0.380).
View larger version (11K):
[in this window]
[in a new window]
|
Fig. 1. Change in (A) sCD44v5 and (B) sCD44v6
concentrations (ng/ml) over 1 year in those who stopped smoking and
those who continued to smoke. Error bars, SE of the
mean.
|
|
At baseline, there was no significant difference in sCD44, sCD44v5, or
sCD44v6 levels between those who subsequently quit and those who
continued smoking. In regression analyses at baseline, expired
CO showed a linear dose-response relationship to sCD44v5 and sCD44v6
concentration. The increase in sCD44v5 concentration was estimated to
be 0.95 ng/ml per 1 ppm expired CO [t(1,58) = 3.0;
r = 0.370; P = 0.004]. The increase in
sCD44v6 concentration was estimated to be 4.2 ng/ml per 1 ppm expired
CO [t(1,58) = 4.3; r = 0.490; P < 0.001]. Expired CO also correlated with the plasma concentrations
of CD44v5 (r = 0.27) and CD44v6 (r =
0.35) at 52 weeks in the continuing smokers. There was a significant
correlation between cotinine and CD44v6 at baseline (r = 0.32; P = 0.01), but not between cotinine and CD44v5
(r = 0.18; P = 0.16).
Correlations of CD44 variant concentration with cigarette consumption
were small and nonsignificant (CD44v5, r = 0.07;
CD44v6, r = 0.16). A positive correlation between the
plasma concentration of sCD44v5 and sCD44v6 was noted
[r = 0.870; P < 0.001]. However,
there was no correlation between several measures of smoke intake
(expired CO, plasma cotinine, and reported cigarette consumption) and
total sCD44 concentration. As expected, the correlation between CO and
cotinine at baseline was strong (r = 0.49;
P < 0.001).
|
Discussion
|
|---|
We have previously shown a small but significant elevation of
total sCD44 in smokers (18)
, but we found no
dose-dependent relationship between serum cotinine level and total CD44
concentration. In contrast, Kittl et al. (19)
reported that the mean concentration of sCD44v5 in smokers was double
that of nonsmokers. We hypothesized that the effect of smoking on sCD44
was not the result of a blanket effect of tobacco smoking on all CD44
gene products.
In the present study, there was no dose-dependent relationship noted
between any smoke intake measure (plasma cotinine, expired CO, and
daily cigarette consumption) and total sCD44 level, and no reduction in
the plasma concentration of total sCD44 was seen in quitters 1 year
after smoking cessation as compared with the level in continuing
smokers. The fact that there was also no evidence of significant
correlations between sCD44v5 or sCD44v6 levels and the number of
cigarettes smoked daily is not surprising. Self-reported consumption is
not only subject to recording error but is also an unreliable measure
of smoke intake, given wide variations in smoking styles. It should
also be noted that only those smoking 15 or more cigarettes/day were
included in the trial, which restricted the range of values for
consumption.
In contrast, by using biochemical markers as dependable indices
of tobacco smoke exposure, we have provided strong evidence that plasma
concentrations of CD44v5 and CD44v6 in smokers are dose related and
that levels of sCD44v5 and sCD44v6 significantly decline after smoking
cessation. This suggests that the elevated levels of sCD44v5 and
sCD44v6 are attributable to the smoking experience. Therefore, we
suggest that the ascribed diagnostic specificity and prognostic value
of sCD44v5 and sCD44v6 in inflammatory and malignant disease may be
altered. It has been previously reported, for example, that circulating
levels of sCD44v5 and sCD44v6 were 81 and 237 ng/ml, respectively, in
subjects with rheumatoid arthritis in comparison to 33 and 166 ng/ml,
respectively, in a control group with miscellaneous inflammatory
rheumatic diseases (15)
. Martin et al.
(10)
observed elevated levels of sCD44v5 and sCD44v6 in
subjects with node-positive breast cancer (46 and 250 ng/ml,
respectively) compared with healthy controls (24 and 154 ng/ml
respectively; Ref. 10
). Similarly, Harn et al.
(22)
observed elevated levels of sCD44v5 and sCD44v6 in
subjects with gastric carcinoma (69 and 217 ng/ml, respectively)
compared with healthy controls (25 and 148 ng/ml, respectively; Ref.
10
). Our data suggest that the effect of smoking on the
specific sCD44 variant concentration may be as great as that previously
attributed to malignancy (10
, 13
, 22)
. Therefore, the
significance of reported elevations in sCD44v5 and sCD44v6
concentration in inflammatory and malignant disease will be very
difficult to interpret if the influence of smoking and/or smoking
cessation has not been considered.
All human isoforms of the CD44 family of adhesion molecules are
encoded by a single gene. Alternate splicing of 12 of the 19 exons in
the human CD44 gene leads to the production of
multiple variant isoforms (11
, 28
, 29)
. Many CD44 isoforms
are tissue specific, but the full range of soluble variant isoform(s)
of sCD44 that are influenced by smoking remains to be clarified.
Interestingly, van Hal et al. (25)
recently
observed that most sCD44v6 molecules in the serum of patients with head
and neck squamous cell carcinoma appeared to derive from normal
epithelial cell compartments, despite the high level of expression of
the CD44v6 domain on head and neck squamous cell tumors. Thus, further
investigations into the character and source of elevated sCD44
molecules in smokers are warranted and may shed light on the relevance
of increased sCD44 levels in the context of the etiology of
cancer and other smoking-related diseases.
|
Footnotes
|
|---|
The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
1 To whom requests for reprints should be
addressed. Present address: Department of Oral Biology, Faculty of
Dentistry, The University of Manitoba, 780 Bannatyne Avenue, Winnipeg,
Manitoba, Canada R3E 0W2. Phone: (204) 789-3705; Fax:
(204) 789-3913 
Received 3/22/00;
revised 8/ 9/00;
accepted 9/ 5/00.
|
References
|
|---|
-
Bourguignon L. Y. W., Gunja-Smith Z., Iida N., Zhu H. B., Young L. J. T., Muller W. J., Cardiff R. D. CD44v(3,8–10) is involved in cytoskeleton-mediated tumor cell migration and matrix metalloproteinase (MMP-9) association in metastatic breast cancer cells. J. Cell. Physiol., 176: 206-215, 1998.[Medline]
-
Herrera-Gayol A., Jothy S. Adhesion proteins in the biology of breast cancer: contribution of CD44. Exp. Mol. Pathol., 66: 149-156, 1999.[Medline]
-
Kuo M. Y., Cheng S. J., Chen H. M., Kok S. H., Hahn L. J., Chiang C. P. Expression of CD44s, CD44v5, CD44v6 and CD44v7–8 in betel quid chewing-associated oral premalignant lesions and squamous cell carcinomas in Taiwan. J. Oral Pathol. Med., 27: 428-433, 1998.[Medline]
-
Lesley J., Hyman R., English N., Catterall J. B., Turner G. A. CD44 in inflammation and metastasis. Glycoconj. J., 14: 611-622, 1997.[Medline]
-
Okamoto I., Morisaki T., Sasaki J-I., Miyake H., Matsumoto M., Suga M., Ando M., Saya H. Molecular detection of cancer cells by competitive reverse transcription-polymerase chain reaction analysis of specific CD44 variant RNAs. J. Natl. Cancer Inst., 90: 307-315, 1998.[Abstract/Free Full Text]
-
Sasaki J. I., Tanabe K. K., Takahashi K., Okomato I., Fujimoto H., Matsumoto M., Suga M., Ando M., Saya H. Expression of CD44 splicing isoforms in lung cancers: dominant expression of CD44v8–10 in non-small cell lung carcinomas. Int. J. Oncol., 12: 525-533, 1998.[Medline]
-
Stauder R., Eisterer W., Thaler J., Gunthert U. CD44 isoforms in non-Hodgkin’s lymphoma: a new independent prognostic factor. Blood, 85: 2885-2899, 1995.[Abstract/Free Full Text]
-
Yu Q., Stamenkovic I. Localization of matrix metalloproteinase 9 to the cell surface provides a mechanism for CD44-mediated tumour invasion. Genes Dev., 13: 35-48, 1999.[Abstract/Free Full Text]
-
Guo Y. J., Liu G., Wang X., Jin D., Wu M., Ma J., Sy M. S. Potential use of soluble CD44 in serum as an indicator of tumour burden and metastasis in patients with gastric or colon cancer. Cancer Res., 65: 422-426, 1994.
-
Martin S., Jansen F., Bokelmann J., Kolb H. Soluble CD44 splice variants in metastasizing human breast cancer. Int. J. Cancer, 74: 443-445, 1997.[Medline]
-
Naot D., Sionov R. V., Ish-Shalom D. CD44: Structure, function and association with the malignant process. Adv. Cancer Res., 71: 241-319, 1997.[Medline]
-
Ristamaki R., Joensuu H., Lappalainen K., Teerenhovi L., Jalkanen S. Elevated serum CD44 level is associated with unfavourable outcome in non-Hodgkin’s lymphoma. Blood, 90: 4039-4045, 1997.[Abstract/Free Full Text]
-
Yamane N., Tsujitani S., Makino M., Maeta M., Kaibara N. Soluble CD44 variant 6 as a prognostic indicator in patients with colorectal cancer. Oncology (Basel), 56: 232-238, 1999.[Medline]
-
Haberhauer G., Kittl E. M., Skoumal M., Hubl W., Wagner E., Bayer P. M., Bauer K., Dunky A. Increased serum levels of soluble CD44-isoform v5 in rheumatic diseases are restricted to seropositive rheumatoid arthritis. Acta Med. Austriaca, 24: 23-25, 1997.[Medline]
-
Kittl E. M., Haberhauer G., Ruckser R., Selleny S., Rech-Weichselbraun I., Hinterberger W., Bauer K. Serum levels of soluble CD44 variant isoforms are elevated in rheumatoid arthritis. Rheumatol. Int., 16: 181-186, 1997.[Medline]
-
Stallmach A., van Look M., Scheiffele F., Ecker K. W., Feifel G., Duchmann R., Zeitz M. IgG, albumin and sCD44 in whole gut lavage fluid are useful clinical markers for assessing the presence and activity of pouchitis. Int. J. Colorectal Dis., 14: 35-40, 1999.[Medline]
-
Kato S., Matsubara Y., Taniguchi Abe, K., Yoshinaga R., Yamashiro S., Mukai H., Kadota J., Kawano S., Matsukura S. Evaluation of soluble CD44 in the BALF before and after treatment of DPB (diffuse panbronchitis) with macrolide antibiotics. Jpn. J. Antibiot., 51: S38-S40, 1998.
-
Scott D. A., Coward P. Y., Wilson R. F., Poston R. N., Odell E. W., Palmer R. M. Serum concentration of total soluble CD44 is elevated in smokers. Biomarkers, 5: 240-244, 2000.
-
Kittl E. M., Ruckser R., Rech-Weichselbraun I., Hinterberger W., Bauer K. Significant elevation of tumour-associated isoforms of soluble CD44 in serum of normal individuals caused by cigarette smoking. Eur. J. Clin. Chem. Clin. Biochem., 35: 81-84, 1997.[Medline]
-
Kittl E. M., Ruckser R., Selleny S., Samek V., Hofmann J., Huber K., Reiner A., Ogris E., Hinterberger W., Bauer K. Evaluation of soluble CD44 splice variant v5 in the diagnosis and follow-up in breast cancer patients. Exp. Clin. Immunogenet., 14: 264-272, 1997.[Medline]
-
Camacho F. I., Munoz C., Sanchez-Verde L., Saez A. I., Alcantara M., Rodriguez R. CD44v6 expression in inflammatory bowel disease is associated with activity detected by endoscopy and pathological features. Histopathology, 35: 144-149, 1999.[Medline]
-
Harn H. J., Ho L. I., Shyu R. Y., Yuan J. S., Lin F. G., Young T. H., Liu C. A., Tang H. S., Lee W. H. Soluble CD44 isoforms in serum as potential markers of metastatic gastric carcinoma. J. Clin. Gastroenterol., 22: 107-110, 1996.[Medline]
-
Gansauge F., Gansauge S., Rau B., Schleiblich A., Poch B., Schoenberg M. H., Beger H. G. Low serum levels of soluble CD44 variant 6 are significantly associated with poor prognosis in patients with pancreatic carcinoma. Cancer (Phila.), 80: 1733-1739, 1997.[Medline]
-
Sheen-Chen S. M., Chen W. J., Eng H. L., Sheen C. C., Chou F. F., Cheng Y. F. Evaluation of the prognostic value of serum soluble CD44 in patients with breast cancer. Cancer Invest., 17: 581-585, 1999.[Medline]
-
van Hal N. L., van Dongen G. A., Ten Brink C. B., Heider K. H., Rech-Weichselbraun I., Snow G. B., Brakenhoff R. H. Evaluation of soluble CD44v6 as a potential serum marker for head and neck squamous cell carcinoma. Clin. Cancer Res., 5: 3534-3541, 1999.[Abstract/Free Full Text]
-
Zeimet A. G., Widschwendter M., Uhl-Steidl M., Muller-Holzner E., Daxenbichler G., Marth C., Dapunt O. High serum levels of soluble CD44 variant isoform v5 are associated with favourable clinical outcome in ovarian cancer. Br. J. Cancer, 76: 1646-1651, 1997.[Medline]
-
Tonnesen P., Paoletti P., Gustavsson G., Russell M. A., Saracci R., Gulsvik A., Rijcken B., Sawe U. Higher dosage nicotine patches increase one-year smoking cessation rates: results from the European CEASE trial. Eur. Resp. J., 13: 238-246, 1999.[Abstract]
-
Ponta H., Wainwright D., Herrlich P. The CD44 protein family. Int. J. Biochem. Cell Biol., 30: 299-305, 1998.[Medline]
-
Screaton G. R., Bell M. V., Jackson D. G., Cornelis F. B., Gerth U., Bell J. I. Genomic structure of DNA encoding the lymphocyte homing receptor CD44 reveals at least 12 alternatively spliced exons. Proc. Natl. Acad. Sci. USA, 89: 12160-12164, 1992.[Abstract/Free Full Text]
This article has been cited by other articles:
|
|
|
|
 
E. J. Franzmann, E. P. Reategui, K. L. Carraway, K. L. Hamilton, D. T. Weed, and W. J. Goodwin
Salivary Soluble CD44: A Potential Molecular Marker for Head and Neck Cancer
Cancer Epidemiol. Biomarkers Prev.,
March 1, 2005;
14(3):
735 - 739.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
B.W.M. Willemse, D.S. Postma, W. Timens, and N.H.T. ten Hacken
The impact of smoking cessation on respiratory symptoms, lung function, airway hyperresponsiveness and inflammation
Eur. Respir. J.,
March 1, 2004;
23(3):
464 - 476.
[Abstract]
[Full Text]
[PDF]
|
|
|
Top
Abstract
Introduction
