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Conventional tumor markers are prognostic indicators in patients with head and neck squamous cell carcinoma
Cancer Letters 155 (2000) 163±168
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Conventional tumor markers are prognostic indicators in patients with head and neck squamous cell carcinoma Yuichi Kimura, Shigeharu Fujieda*, Tetsuji Takabayashi, Takeshi Tanaka, Chizuru Sugimoto, Hitoshi Saito Department of Otorhinolaryngology, Fukui Medical University, Shimoaizuki, Matsuoka-cho, Yoshida-gun, Fukui 910-1193, Japan Received 24 January 2000; received in revised form 9 March 2000; accepted 9 March 2000
Abstract We tested for squamous cell carcinoma-related antigen (SCC), carcinoembryonic antigen (CEA), ferritin, immunosuppressive acid protein (IAP) and sialic acid in the serum from 247 patients with head and neck squamous cell carcinoma prior to therapy. Signi®cant correlations were found between IAP and tumor size, lymph node metastasis, and clinical stage (P , 0:0001, P , 0:001, and P , 0:0001). Also, sialic acid and SCC were also correlated with tumor size, lymph node metastasis, and clinical stage. Moreover IAP, sialic acid and SCC were strongly associated with survival rate (P , 0:0001, P 0:0230 and P 0:0159). A multivariate Cox proportional hazard model demonstrated that being positive for IAP was an independent predictor for patients with H&NSCC (P 0:0115). The results indicate that IAP, sialic acid and SCC are useful as prognostic factors. q 2000 Elsevier Science Ireland Ltd. All rights reserved. Keywords: Tumor marker; Prognosis; Squamous cell carcinoma-related antigen; Immunosuppressive acid protein; Sialic acid; Head and neck cancer
1. Introduction An ideal tumor marker should include correlations with the stage of the cancer, detection in the early stage, and recurrence or metastasis after therapy. Also, it is important for the tumor marker to predict the prognosis of patients based on initial levels of a marker or on changing levels. Numerous studies have been conducted in attempts to identify a tumor marker in the serum for the presence of squamous cell carcinomas in head and neck lesions [1±8]. However, an ideal marker to bene®t patients with H&NSCC has not been found. The bene®t of tumor markers for patients * Corresponding author. Fax: 181-776-61-8118. E-mail address: [email protected] (S. Fujieda).
with head and neck squamous cell carcinomas (H&NSCC) is controversial. One of the most important problems in the treatment of H&NSCC is the management of metastatic spread to locoregional lymph nodes. Survival rate is often related to lymph node metastasis in H&NSCC. If a tumor marker is able to re¯ect overall survival, the data should be potentially useful in guiding additional treatment for H&NSCC. From the number of markers available, we have chosen a panel of ®ve tumor markers: immunosuppressive acid protein (IAP); sialic acid; squamous cell carcinoma-related antigen (SCC); carcinoembryonic antigen (CEA) and ferritin in patients with H&NSCC. Also, we analyzed the correlation between the patients' clinicopathological factors and serum
0304-3835/00/$ - see front matter q 2000 Elsevier Science Ireland Ltd. All rights reserved. PII: S 0304-383 5(00)00423-7
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Y. Kimura et al. / Cancer Letters 155 (2000) 163±168
levels of tumor markers. We found that IAP, out of ®ve markers, is an independent prognostic marker in patients with H&NSCC using an Cox proportional hazard model. 2. Materials and methods 2.1. Patients We studied 247 patients with H&NSCC. All patients were treated and subsequently followed at Fukui Medical University from 1983 to 1995. The primary sites were as follows: maxillary sinus, 32 cases; nasopharynx, 16 cases; oral cavity, 51 cases; oropharynx, 11 cases; hypopharynx, 24 cases; larynx, 113 cases (Table 1). We employed the International Union Against Cancer rules for head and neck cancer (tumor-node-metastasis (TNM) classi®cation) for the clinical staging of H&NSCC. Serum samples were collected from patients with H&NSCC prior to therapy. No patient had distant metastasis in the ®rst examination. Eighty percent of the patients (197 cases) were followed for 5 years. The others were followed for at least 3 years. None of the 247 cases were lost to follow up. The patients were treated with standard therapy, depending on the stage. Patients with maxillary sinus SCCs underwent tumor resection, irradiation (from 50 to 60 Gy) with chemotherapy (cisplatin, 5-¯uorouracil, and vitamin A) after surgery. Patients with nasopharyngeal SCCs received irradiation (from 60 to 70 Gy) with chemotherapy (cisplatin, 5-¯uorouracil, and vitamin A). If metastatic neck lymph node did not disappear after irradiation therapy, standard neck dissection was performed.
Patients with oral and oropharyngeal SCCs of stages I and II underwent only tumor resection, and stage III and IV patients underwent tumor resection, standard neck dissection, and irradiation (60 Gy) after surgery. The patients with hypopharyngeal SCCs underwent tumor resection, standard neck dissection, and irradiation (60 Gy) after surgery. Patients with laryngeal SCCs of stages I and II received only irradiation (from 60 to 70 Gy), and stage III and IV patients underwent total laryngectomy, standard neck dissection, and irradiation (60 Gy) after surgery. 2.2. Measurement of tumor markers Commercially available kits were used and applied according to the manufacture's instructions: radioimmunoassay with SCC (Dainabot Ltd, Tokyo, Japan), CEA (Daiichi RI, Tokyo, Japan) and ferritin (Daiichi RI). IAP values were measured by using the commercial plates which were supplied by Kayaku Antibiotics Research (Tokyo, Japan). Sialic acid was determined with a UV method (Kyokuto Pharmaceutical Industrial Company, Tokyo, Japan). A special technician measured the ®ve markers without prior clinical knowledge. The normal ranges for the ®ve markers were determined from almost 1000 healthy volunteers in Fukui Medical University. A marker value over the normal upper range was determined as a positive value. The upper limit of the normal values were as follows: IAP, 500 ng/ml; sialic acid, 65 mg/dl; SCC, 1.5 ng/ml; CEA, 2.0 ng/ml; ferritin, 220 ng/ml for men and 80 ng/ml for women. 2.3. Statistical analysis
Table 1 Site of the primary tumor in patients with untreated H&NSCC Site of lesion
Maxillary sinus Nasopharynx Oral cavity Oropharynx Hypopharynx Larynx Total
No. of cases
32 16 51 11 24 113 247
Stage I
II
III
IV
1 0 8 0 0 48 57
1 3 16 0 2 35 57
19 4 11 3 8 14 59
11 9 16 8 14 16 74
The associations between clinical characteristics and the positive rate of each tumor marker, or its values, were analyzed using the chi-square test or the Mann±Whitney U-test, respectively. The overall survival rate of patients with H&NSCC was determined and plotted according to the Kaplan±Meier test and analyzed using log-rank tests. Macintosh personal computers (Apple Computers, Cupertino, CA) with Stat View software (Abacus Concepts, Inc., Berkeley, CA) were used for all statistical analysis.
Y. Kimura et al. / Cancer Letters 155 (2000) 163±168
3. Results 3.1. Immunosuppressive acid protein Serum IAP values in 227 patients with H&NSCC were measured before treatment. The average IAP level in T4 was signi®cantly higher than that in T1 or T2 (mean ^ SD, T4: 675.4 ^ 260.5 ng/ml vs. T1: 371.8 ^ 121.2, P , 0:0001, vs. T2: 450.3 ^ 165.7, P , 0:0001, respectively). We also observed the average IAP level in patients with metastasis of neck lymph node metastasis was higher than that in those without lymph node metastasis (549.8 ^ 250.1 vs. 461.8 ^ 184.3, P , 0:005). Eighty-®ve of 227 patients (37%) were determined as positive for IAP (Fig. 1A). A positive IAP rate was positively correlated with tumor size (T classi®cation) and clinical stage (P , 0:0001, P , 0:0001, respectively). Also, the positive rate of IAP in patients with lymph node metastasis was higher than that in those without lymph node metastasis (P , 0:001). The survival rates of the IAP-positive and IAP-negative groups
165
were calculated with the Kaplan±Meier method. Although 80.5% of the IAP-negative groups reached an overall survival rate of 5 years, 53.0% of the IAPpositive groups survived for that length of time. There was a signi®cant difference in overall survival between the IAP-positive and IAP-negative groups (P , 0:0001, Fig. 2). 3.2. Sialic acid Serum sialic acid values in 210 patients with H&NSCC were measured before treatment. One hundred and ®fteen of 210 patients (55%) showed abnormal levels of sialic acid (Fig. 1B). Thirteen of 50 patients (26%) with T1 had elevated sialic acid levels. However, of 42 patients with T4 primary lesions, 35 patients (83%) had abnormal levels. The average sialic acid level in T4 was signi®cantly higher than that in T1 or in T2 (T4: 80.7 ^ 16.2 mg/dl vs. T1: 62.5 ^ 13.2, P , 0:0001, vs. T2: 68.4 ^ 11.6, P , 0:0001, respectively). Also, the average sialic acid level in patients with neck lymph node metastasis
Fig. 1. (A) Association between clinicopathological factors and IAP. The positive rate of IAP was 37% (85 cases) and the negative rate was 63% (142 cases). A positive rate of IAP was correlated with tumor size, clinical stage and lymph node metastasis. (B) Association between clinicopathological factors and sialic acid. The positive rate of sialic acid was 55% (115 cases) and the negative rate was 45% (95 cases). A positive rate of sialic acid was correlated with tumor size, clinical stage and lymph node metastasis. (C) Association between clinicopathological factors and SCC. The positive rate of SCC was 26% (60 cases) and the negative rate was 74% (173 cases). A positive rate of SCC was correlated with tumor size, clinical stage and lymph node metastasis. (D) Association between clinicopathological factors and CEA. The positive rate of CEA was 29% (65 cases) and the negative rate was 71% (160 cases). There was no correlation between CEA and tumor size, clinical stage or lymph node metastasis. (E) Association between clinicopathological factors and ferritin. The positive rate of ferritin was 10% (20 cases) and the negative rate was 90% (177 cases). There was no correlation between ferritin and tumor size, clinical stage or lymph node metastasis.
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Fig. 2. Kaplan±Meier analysis of 5-year-overall survival of patients with H&NSCC: IAP-positive and negative. Overall survival was signi®cantly lower in IAP-positive patients.
3.6 ^ 9.5 mg/dl vs. T1: 1.1 ^ 1.9, P , 0:02, vs. T2: 1:5 ^ 3:3, P , 0:03, respectively). Also, a positive rate of SCC was positively correlated with lymph node metastasis and clinical stage (P , 0:05, P , 0:0001, respectively). When the survival rates of the SCC-positive and SCC-negative groups were calculated, the overall survival rate for 5 years in the SCC-positive group was signi®cantly lower than that in the SCC negative group (76.8 vs. 59.0%) (P , 0:05, Fig. 4). 3.4. Carcinoembryonic antigen and ferritin
was higher than that in those without lymph node metastasis (74.2 ^ 14.9 vs. 68.1 ^ 14.5, P , 0:0001). The positive rate of sialic acid was also correlated with tumor size (P , 0:0001), lymph node metastasis (P , 0:05), and clinical stage (P , 0:0001) similar to IAP. The survival rates of the sialic acid-positive and sialic acid-negative groups were calculated with the Kaplan±Meier method. The associations of sialic acid with overall survival for 5 years are shown in Fig. 3. The sialic acid-positive group showed signi®cantly lower probability of overall survival than the sialic acid-negative group (67.1 vs. 80.6%) (P , 0:05).
Serum CEA values in 225 patients and serum ferritin values in 197 patients with H&NSCC were measured before treatment. Sixty-®ve patients (29%) were positive for CEA and 20 patients (10%) were positive for ferritin. There was no correlation between a positive rate for CEA and T classi®cation, lymph node metastasis, clinical stage or overall survival rate for 5 years (Fig. 1D). Neither was there were correlation between a positive rate for ferritin nor any of the clinical factors (Fig. 1E).
3.3. Squamous cell carcinoma antigen Serum SCC in 233 patients with H&NSCC was measured before treatment. The association between SCC and clinical features is shown in Fig. 1C. Only 26% of patients had elevated SCC levels. However, a positive rate of SCC was correlated with tumor size (T classi®cation). The average SCC level in T4 was signi®cantly higher than that in T1 or in T2 (T4:
Finally, we examined the prognostic value of tumor markers with multivariate analysis using Cox's proportional hazard model (Table 2). IAP was significant as an independent prognostic indicator for overall survival, as well as for lymph node metastasis. The risk ratio of death was 3.237 among patients who were IAP-positive vs. those who were IAP-negative (P 0:0115). We also examined the association between the value of tumor markers and histopatho-
Fig. 3. Kaplan±Meier analysis of 5-year-overall survival of patients with H&NSCC: sialic acid-positive and negative. Overall survival was signi®cantly lower in sialic acid-positive patients.
Fig. 4. Kaplan±Meier analysis of 5-year-overall survival of patients with H&NSCC: SCC-positive and negative. Overall survival was signi®cantly lower in SCC-positive patients.
3.5. Cox's proportional hazard model
Y. Kimura et al. / Cancer Letters 155 (2000) 163±168 Table 2 Multivariate hazard analysis Prognostic factors
Hazards 95% Cl a ratio
IAP (1 or 2) Sialic acid (1 or 2) SCC (1 or 2) Tumor size (T3/T4 or T1/T2) Lymph node metastasis (1 or 2)
3.237 0.690 0.654 2.323 3.741
a
P value
1.301±8.050 0.0115 0.275±0.690 0.4288 0.318±0.654 0.2473 1.093±4.939 0.0285 1.985±7.049 ,0.0001
CI, con®dence interval.
logical features. However, there were no correlations (data not shown). 4. Discussion In this study, we demonstrated that a positive rate for IAP, sialic acid, and SCC before treatment were correlated with large tumor size, positive lymph node metastasis, and advanced clinical stage in patients with H&NSCC. Furthermore, we found that the overall survival rate for 5 years in patients positive for IAP, sialic acid, or SCC was lower than that in those patients negative for the three tumor markers, suggesting that IAP, sialic acid, and SCC are prognostic indicators in patients with H&NSCC. The level of IAP in the serum was the most signi®cant independent prognostic indicator for overall survival in H&NSCC, as shown by Cox's proportional hazards multivariate analysis. IAP was puri®ed from the ascitic ¯uid of stomach cancer patients and has been found to be elevated in the sera of patients with colorectal cancer, ovarian cancer, and similar diseases [9]. Yamanaka et al. [8] reported that positive rates of IAP were found to be elevated as the disease stage advanced and useful parameters for monitoring the disease stage of head and neck cancer patients, which agrees with our results. IAP and sialic acid are classi®ed into the same group as the markers of in¯ammation [10,11]. The coincidental rate between the states of IAP and sialic acid (positive/positive or negative/negative) was 75.0% in H&NSCC, suggesting that the IAP level is suf®cient to be checked. Regarding the cost of the measurements, IAP was the cheapest among the ®ve tumor markers. CEA and ferritin were not associated with clinical
167
outcomes or clinicopathological states in H&NSCC. However, the measurement of CEA has been established to be useful in patients with cancer of the aerodigestive tract [12,13]. In previous papers, CEA was often elevated in cancers of the aerodigestive system and some investigators suggested a correlation between CEA and disease stage [12,13]. Also, patients with H&NSCC often have double cancers in the aerodigestive tract. In fact, we found one patient with esophagus cancer and three with colon cancers in the 65 CEA-positive patients with H&NSCC with a high level of CEA, suggesting that CEA is a good parameter to detect other cancers in patients with H&NSCC. With respect to ferritin, some investigators have reported an increase in ferritin levels in patients with a variety of tumors, and in some studies ferritin levels in head and neck cancer patients proved a useful prognosis marker [1]. However, in this study, we concluded that ferritin was of no value as a marker to detect patients with H&NSCC. SCC was isolated from a squamous cell carcinoma of the uterine cervix [14]. This tumor marker has been investigated in patients with squamous cell carcinomas at other sites, including the esophagus, anal canal, lungs, head and neck [2,3,5±7,15±17]. Thus, SCC was found to be highly speci®c for squamous cell carcinoma. Although IAP in the serum sometimes elevates due to in¯ammation, SCC does not. When the elevation of both IAP and SCC was found in patients with H&NSCC, we speculated that they had recurrent and distant metastatic disease. We wish to stress that the combined measurement of IAP, SCC and CEA was the best combination for monitoring H&NSCC in terms of cost and effectiveness. Tumor markers have been demonstrated to be clinically helpful in detecting various human neoplasms in the last few decades. However, the role of serum tumor markers in the management of H&NSCC has attracted relatively less interest compared with that of other tumors. Since the lesions of H&NSCC are easy to see and accessible, it is easy to obtain samples for biopsy. The positive rate of each tumor marker was low in this study (10 to 55% among ®ve tumor markers). Furthermore, in patients in early stages, few patients (16 IAP cases, 41 sialic acid cases, 12 SCC cases, 26 CEA cases, eight ferritin cases) had high levels of tumor markers. These results suggested that conventional tumor markers, at least the ®ve
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items we measured, are useless to detect early H&NSCC. However, a predictable tumor marker for prognosis would bene®t the treatment for H&NSCC. Adjuvant therapy following surgical resection and induction chemotherapy followed by surgery have been proposed as the best approaches for improving results. Although much more extensive study is needed to adequately evaluate the practical applications of these procedures, the present results indicate that serum IAP, sialic acid and SCC levels may be useful monitors for determining whether to perform adjuvant chemotherapy or post-operative irradiation for H&NSCC. In particular, aggressive therapy for patients with high IAP levels prior to therapy should be considered. References [1] P.E. Maxim, R.W. Velti, Serum ferritin as a tumor marker in patients with squamous cell carcinoma of head and neck, Cancer 57 (1986) 305±311. [2] C.D.E. Eibing, J.T. Johnson, R.L. Wagner, S. Su, SCC-RIA in the diagnosis of squamous cell carcinoma of head and neck, Laryngoscope 99 (1989) 117±124. [3] W. Fischbach, T. Meyer, K. Barthel, Squamous cell carcinoma antigen in the diagnosis and treatment follow-up of oral and facial squamous cell carcinoma, Cancer 65 (1990) 1321±1324. [4] A.I. Dreyfuss, J.R. Clark, J.W. Andersen, Lipid-associated sialic acid, squamous cell carcinoma antigen, carcinoembryonic antigen, and lactic dehydrogenase levels as tumor markers in squamous cell carcinoma of head and neck, Cancer 70 (1992) 2499±2503. [5] M.B. Straka, R.L. Wagner, J.T. Johnson, K.K. Kachman, D.E. Eibling, The lack of utility of a tumor marker panel in head and neck carcinoma. Squamous cell carcinoma antigen, carcinoembryonic antigen, lipid-associated acid, and CA-125, Arch. Otolaryngol. Head Neck Surg. 118 (1992) 802±805. [6] E.K. Walther, N. Dahlmann, H.T. Gorgulla, Tumor markers in
[7]
[8] [9]
[10] [11] [12] [13] [14] [15]
[16]
[17]
the diagnosis and follow-up of head and neck cancer: role of CEA, CA 19-9, SCC, TK, and dTTpase, Head Neck 15 (1993) 230±235. C.H. Snyderman, F. D'Amico, R. Wagner, D.E. Eibling, A reappraisal of squamous cell carcinoma antigen as a tumor marker in head and neck cancer, Arch. Otolaryngol. Head Neck Surg. 121 (1995) 1294±1297. N. Yamanaka, Y. Harabuchi, T. Himi, A. Kataura, Immunosuppressive substance in the sera of head and neck cancer patients, Cancer 62 (1988) 1293±1298. K. Tamura, Y. Shibata, Y. Matsuda, N. Ishida, Isolation and characterization of an immunosuppressive acidic protein from ascitic ¯uids of cancer patients, Cancer Res. 41 (1981) 3244± 3252. R. Voigtmann, J. Pokorny, A. Meinshausen, Evaluation and limitations of the lipid-associated sialic acid test for the detection of human cancer, Cancer 64 (1989) 2279±2283. N. Stefenelli, H. Klotz, A. Engel, P. Bauer, Serum sialic acid in malignant tumors, bacterial infections and chronic liver disease, J. Cancer Res. Clin. Oncol. 109 (1985) 55±59. A.M. Steward, D. Dixson, N. Zamcheck, A. Aisenberg, Carcinoembryonic antigen in breast cancer patients: Serum levels and disease progress, Cancer 33 (1974) 1246±1252. N.A. Silverman, J.C. Alexandar Jr., P.B. Chretien, CEA levels in head and neck cancer, Cancer 37 (1976) 2204±2211. H. Kato, F. Miyauchi, H. Morioka, T. Fujino, T. Torigoe, Tumor antigen human cervical squamous cell carcinoma, Cancer 43 (1979) 585±590. H. Kato, K. Tamai, H. Morioka, M. Nagai, T. Nagaya, T. Torigoe, Tumor-antigen TA-4 in the detection of recurrence in cervical squamous cell carcinoma, Cancer 54 (1984) 1544± 1546. A.L. Picardo, A.J. Torres, M. Maestro, D. Ortega, J. GarciaAsenjo, J. Muguerza, Quantitative analysis of carcinoembryonic antigen, squamous cell carcinoma antigen, CA125 and CA50 cytosolic content in non-small cell lung cancer, Cancer 73 (1994) 2305±2311. A.L. Picardo, M. Diez, A. Torres, M. Maestro, D. Ortega, F. Hernando, Analysis of the prognostic signi®cance of cytosolic determination of CA 125 tumor-associated antigen, carcinoembryonic antigen and squamous cell carcinoma antigen in patients with nonsmall cell lung carcinoma, Cancer 77 (1996) 1066±1072.
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Conventional tumor markers are prognostic indicators in patients with head and neck squamous cell carcinoma Yuichi Kimura, Shigeharu Fujieda*, Tetsuji Takabayashi, Takeshi Tanaka, Chizuru Sugimoto, Hitoshi Saito Department of Otorhinolaryngology, Fukui Medical University, Shimoaizuki, Matsuoka-cho, Yoshida-gun, Fukui 910-1193, Japan Received 24 January 2000; received in revised form 9 March 2000; accepted 9 March 2000
Abstract We tested for squamous cell carcinoma-related antigen (SCC), carcinoembryonic antigen (CEA), ferritin, immunosuppressive acid protein (IAP) and sialic acid in the serum from 247 patients with head and neck squamous cell carcinoma prior to therapy. Signi®cant correlations were found between IAP and tumor size, lymph node metastasis, and clinical stage (P , 0:0001, P , 0:001, and P , 0:0001). Also, sialic acid and SCC were also correlated with tumor size, lymph node metastasis, and clinical stage. Moreover IAP, sialic acid and SCC were strongly associated with survival rate (P , 0:0001, P 0:0230 and P 0:0159). A multivariate Cox proportional hazard model demonstrated that being positive for IAP was an independent predictor for patients with H&NSCC (P 0:0115). The results indicate that IAP, sialic acid and SCC are useful as prognostic factors. q 2000 Elsevier Science Ireland Ltd. All rights reserved. Keywords: Tumor marker; Prognosis; Squamous cell carcinoma-related antigen; Immunosuppressive acid protein; Sialic acid; Head and neck cancer
1. Introduction An ideal tumor marker should include correlations with the stage of the cancer, detection in the early stage, and recurrence or metastasis after therapy. Also, it is important for the tumor marker to predict the prognosis of patients based on initial levels of a marker or on changing levels. Numerous studies have been conducted in attempts to identify a tumor marker in the serum for the presence of squamous cell carcinomas in head and neck lesions [1±8]. However, an ideal marker to bene®t patients with H&NSCC has not been found. The bene®t of tumor markers for patients * Corresponding author. Fax: 181-776-61-8118. E-mail address: [email protected] (S. Fujieda).
with head and neck squamous cell carcinomas (H&NSCC) is controversial. One of the most important problems in the treatment of H&NSCC is the management of metastatic spread to locoregional lymph nodes. Survival rate is often related to lymph node metastasis in H&NSCC. If a tumor marker is able to re¯ect overall survival, the data should be potentially useful in guiding additional treatment for H&NSCC. From the number of markers available, we have chosen a panel of ®ve tumor markers: immunosuppressive acid protein (IAP); sialic acid; squamous cell carcinoma-related antigen (SCC); carcinoembryonic antigen (CEA) and ferritin in patients with H&NSCC. Also, we analyzed the correlation between the patients' clinicopathological factors and serum
0304-3835/00/$ - see front matter q 2000 Elsevier Science Ireland Ltd. All rights reserved. PII: S 0304-383 5(00)00423-7
164
Y. Kimura et al. / Cancer Letters 155 (2000) 163±168
levels of tumor markers. We found that IAP, out of ®ve markers, is an independent prognostic marker in patients with H&NSCC using an Cox proportional hazard model. 2. Materials and methods 2.1. Patients We studied 247 patients with H&NSCC. All patients were treated and subsequently followed at Fukui Medical University from 1983 to 1995. The primary sites were as follows: maxillary sinus, 32 cases; nasopharynx, 16 cases; oral cavity, 51 cases; oropharynx, 11 cases; hypopharynx, 24 cases; larynx, 113 cases (Table 1). We employed the International Union Against Cancer rules for head and neck cancer (tumor-node-metastasis (TNM) classi®cation) for the clinical staging of H&NSCC. Serum samples were collected from patients with H&NSCC prior to therapy. No patient had distant metastasis in the ®rst examination. Eighty percent of the patients (197 cases) were followed for 5 years. The others were followed for at least 3 years. None of the 247 cases were lost to follow up. The patients were treated with standard therapy, depending on the stage. Patients with maxillary sinus SCCs underwent tumor resection, irradiation (from 50 to 60 Gy) with chemotherapy (cisplatin, 5-¯uorouracil, and vitamin A) after surgery. Patients with nasopharyngeal SCCs received irradiation (from 60 to 70 Gy) with chemotherapy (cisplatin, 5-¯uorouracil, and vitamin A). If metastatic neck lymph node did not disappear after irradiation therapy, standard neck dissection was performed.
Patients with oral and oropharyngeal SCCs of stages I and II underwent only tumor resection, and stage III and IV patients underwent tumor resection, standard neck dissection, and irradiation (60 Gy) after surgery. The patients with hypopharyngeal SCCs underwent tumor resection, standard neck dissection, and irradiation (60 Gy) after surgery. Patients with laryngeal SCCs of stages I and II received only irradiation (from 60 to 70 Gy), and stage III and IV patients underwent total laryngectomy, standard neck dissection, and irradiation (60 Gy) after surgery. 2.2. Measurement of tumor markers Commercially available kits were used and applied according to the manufacture's instructions: radioimmunoassay with SCC (Dainabot Ltd, Tokyo, Japan), CEA (Daiichi RI, Tokyo, Japan) and ferritin (Daiichi RI). IAP values were measured by using the commercial plates which were supplied by Kayaku Antibiotics Research (Tokyo, Japan). Sialic acid was determined with a UV method (Kyokuto Pharmaceutical Industrial Company, Tokyo, Japan). A special technician measured the ®ve markers without prior clinical knowledge. The normal ranges for the ®ve markers were determined from almost 1000 healthy volunteers in Fukui Medical University. A marker value over the normal upper range was determined as a positive value. The upper limit of the normal values were as follows: IAP, 500 ng/ml; sialic acid, 65 mg/dl; SCC, 1.5 ng/ml; CEA, 2.0 ng/ml; ferritin, 220 ng/ml for men and 80 ng/ml for women. 2.3. Statistical analysis
Table 1 Site of the primary tumor in patients with untreated H&NSCC Site of lesion
Maxillary sinus Nasopharynx Oral cavity Oropharynx Hypopharynx Larynx Total
No. of cases
32 16 51 11 24 113 247
Stage I
II
III
IV
1 0 8 0 0 48 57
1 3 16 0 2 35 57
19 4 11 3 8 14 59
11 9 16 8 14 16 74
The associations between clinical characteristics and the positive rate of each tumor marker, or its values, were analyzed using the chi-square test or the Mann±Whitney U-test, respectively. The overall survival rate of patients with H&NSCC was determined and plotted according to the Kaplan±Meier test and analyzed using log-rank tests. Macintosh personal computers (Apple Computers, Cupertino, CA) with Stat View software (Abacus Concepts, Inc., Berkeley, CA) were used for all statistical analysis.
Y. Kimura et al. / Cancer Letters 155 (2000) 163±168
3. Results 3.1. Immunosuppressive acid protein Serum IAP values in 227 patients with H&NSCC were measured before treatment. The average IAP level in T4 was signi®cantly higher than that in T1 or T2 (mean ^ SD, T4: 675.4 ^ 260.5 ng/ml vs. T1: 371.8 ^ 121.2, P , 0:0001, vs. T2: 450.3 ^ 165.7, P , 0:0001, respectively). We also observed the average IAP level in patients with metastasis of neck lymph node metastasis was higher than that in those without lymph node metastasis (549.8 ^ 250.1 vs. 461.8 ^ 184.3, P , 0:005). Eighty-®ve of 227 patients (37%) were determined as positive for IAP (Fig. 1A). A positive IAP rate was positively correlated with tumor size (T classi®cation) and clinical stage (P , 0:0001, P , 0:0001, respectively). Also, the positive rate of IAP in patients with lymph node metastasis was higher than that in those without lymph node metastasis (P , 0:001). The survival rates of the IAP-positive and IAP-negative groups
165
were calculated with the Kaplan±Meier method. Although 80.5% of the IAP-negative groups reached an overall survival rate of 5 years, 53.0% of the IAPpositive groups survived for that length of time. There was a signi®cant difference in overall survival between the IAP-positive and IAP-negative groups (P , 0:0001, Fig. 2). 3.2. Sialic acid Serum sialic acid values in 210 patients with H&NSCC were measured before treatment. One hundred and ®fteen of 210 patients (55%) showed abnormal levels of sialic acid (Fig. 1B). Thirteen of 50 patients (26%) with T1 had elevated sialic acid levels. However, of 42 patients with T4 primary lesions, 35 patients (83%) had abnormal levels. The average sialic acid level in T4 was signi®cantly higher than that in T1 or in T2 (T4: 80.7 ^ 16.2 mg/dl vs. T1: 62.5 ^ 13.2, P , 0:0001, vs. T2: 68.4 ^ 11.6, P , 0:0001, respectively). Also, the average sialic acid level in patients with neck lymph node metastasis
Fig. 1. (A) Association between clinicopathological factors and IAP. The positive rate of IAP was 37% (85 cases) and the negative rate was 63% (142 cases). A positive rate of IAP was correlated with tumor size, clinical stage and lymph node metastasis. (B) Association between clinicopathological factors and sialic acid. The positive rate of sialic acid was 55% (115 cases) and the negative rate was 45% (95 cases). A positive rate of sialic acid was correlated with tumor size, clinical stage and lymph node metastasis. (C) Association between clinicopathological factors and SCC. The positive rate of SCC was 26% (60 cases) and the negative rate was 74% (173 cases). A positive rate of SCC was correlated with tumor size, clinical stage and lymph node metastasis. (D) Association between clinicopathological factors and CEA. The positive rate of CEA was 29% (65 cases) and the negative rate was 71% (160 cases). There was no correlation between CEA and tumor size, clinical stage or lymph node metastasis. (E) Association between clinicopathological factors and ferritin. The positive rate of ferritin was 10% (20 cases) and the negative rate was 90% (177 cases). There was no correlation between ferritin and tumor size, clinical stage or lymph node metastasis.
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Fig. 2. Kaplan±Meier analysis of 5-year-overall survival of patients with H&NSCC: IAP-positive and negative. Overall survival was signi®cantly lower in IAP-positive patients.
3.6 ^ 9.5 mg/dl vs. T1: 1.1 ^ 1.9, P , 0:02, vs. T2: 1:5 ^ 3:3, P , 0:03, respectively). Also, a positive rate of SCC was positively correlated with lymph node metastasis and clinical stage (P , 0:05, P , 0:0001, respectively). When the survival rates of the SCC-positive and SCC-negative groups were calculated, the overall survival rate for 5 years in the SCC-positive group was signi®cantly lower than that in the SCC negative group (76.8 vs. 59.0%) (P , 0:05, Fig. 4). 3.4. Carcinoembryonic antigen and ferritin
was higher than that in those without lymph node metastasis (74.2 ^ 14.9 vs. 68.1 ^ 14.5, P , 0:0001). The positive rate of sialic acid was also correlated with tumor size (P , 0:0001), lymph node metastasis (P , 0:05), and clinical stage (P , 0:0001) similar to IAP. The survival rates of the sialic acid-positive and sialic acid-negative groups were calculated with the Kaplan±Meier method. The associations of sialic acid with overall survival for 5 years are shown in Fig. 3. The sialic acid-positive group showed signi®cantly lower probability of overall survival than the sialic acid-negative group (67.1 vs. 80.6%) (P , 0:05).
Serum CEA values in 225 patients and serum ferritin values in 197 patients with H&NSCC were measured before treatment. Sixty-®ve patients (29%) were positive for CEA and 20 patients (10%) were positive for ferritin. There was no correlation between a positive rate for CEA and T classi®cation, lymph node metastasis, clinical stage or overall survival rate for 5 years (Fig. 1D). Neither was there were correlation between a positive rate for ferritin nor any of the clinical factors (Fig. 1E).
3.3. Squamous cell carcinoma antigen Serum SCC in 233 patients with H&NSCC was measured before treatment. The association between SCC and clinical features is shown in Fig. 1C. Only 26% of patients had elevated SCC levels. However, a positive rate of SCC was correlated with tumor size (T classi®cation). The average SCC level in T4 was signi®cantly higher than that in T1 or in T2 (T4:
Finally, we examined the prognostic value of tumor markers with multivariate analysis using Cox's proportional hazard model (Table 2). IAP was significant as an independent prognostic indicator for overall survival, as well as for lymph node metastasis. The risk ratio of death was 3.237 among patients who were IAP-positive vs. those who were IAP-negative (P 0:0115). We also examined the association between the value of tumor markers and histopatho-
Fig. 3. Kaplan±Meier analysis of 5-year-overall survival of patients with H&NSCC: sialic acid-positive and negative. Overall survival was signi®cantly lower in sialic acid-positive patients.
Fig. 4. Kaplan±Meier analysis of 5-year-overall survival of patients with H&NSCC: SCC-positive and negative. Overall survival was signi®cantly lower in SCC-positive patients.
3.5. Cox's proportional hazard model
Y. Kimura et al. / Cancer Letters 155 (2000) 163±168 Table 2 Multivariate hazard analysis Prognostic factors
Hazards 95% Cl a ratio
IAP (1 or 2) Sialic acid (1 or 2) SCC (1 or 2) Tumor size (T3/T4 or T1/T2) Lymph node metastasis (1 or 2)
3.237 0.690 0.654 2.323 3.741
a
P value
1.301±8.050 0.0115 0.275±0.690 0.4288 0.318±0.654 0.2473 1.093±4.939 0.0285 1.985±7.049 ,0.0001
CI, con®dence interval.
logical features. However, there were no correlations (data not shown). 4. Discussion In this study, we demonstrated that a positive rate for IAP, sialic acid, and SCC before treatment were correlated with large tumor size, positive lymph node metastasis, and advanced clinical stage in patients with H&NSCC. Furthermore, we found that the overall survival rate for 5 years in patients positive for IAP, sialic acid, or SCC was lower than that in those patients negative for the three tumor markers, suggesting that IAP, sialic acid, and SCC are prognostic indicators in patients with H&NSCC. The level of IAP in the serum was the most signi®cant independent prognostic indicator for overall survival in H&NSCC, as shown by Cox's proportional hazards multivariate analysis. IAP was puri®ed from the ascitic ¯uid of stomach cancer patients and has been found to be elevated in the sera of patients with colorectal cancer, ovarian cancer, and similar diseases [9]. Yamanaka et al. [8] reported that positive rates of IAP were found to be elevated as the disease stage advanced and useful parameters for monitoring the disease stage of head and neck cancer patients, which agrees with our results. IAP and sialic acid are classi®ed into the same group as the markers of in¯ammation [10,11]. The coincidental rate between the states of IAP and sialic acid (positive/positive or negative/negative) was 75.0% in H&NSCC, suggesting that the IAP level is suf®cient to be checked. Regarding the cost of the measurements, IAP was the cheapest among the ®ve tumor markers. CEA and ferritin were not associated with clinical
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outcomes or clinicopathological states in H&NSCC. However, the measurement of CEA has been established to be useful in patients with cancer of the aerodigestive tract [12,13]. In previous papers, CEA was often elevated in cancers of the aerodigestive system and some investigators suggested a correlation between CEA and disease stage [12,13]. Also, patients with H&NSCC often have double cancers in the aerodigestive tract. In fact, we found one patient with esophagus cancer and three with colon cancers in the 65 CEA-positive patients with H&NSCC with a high level of CEA, suggesting that CEA is a good parameter to detect other cancers in patients with H&NSCC. With respect to ferritin, some investigators have reported an increase in ferritin levels in patients with a variety of tumors, and in some studies ferritin levels in head and neck cancer patients proved a useful prognosis marker [1]. However, in this study, we concluded that ferritin was of no value as a marker to detect patients with H&NSCC. SCC was isolated from a squamous cell carcinoma of the uterine cervix [14]. This tumor marker has been investigated in patients with squamous cell carcinomas at other sites, including the esophagus, anal canal, lungs, head and neck [2,3,5±7,15±17]. Thus, SCC was found to be highly speci®c for squamous cell carcinoma. Although IAP in the serum sometimes elevates due to in¯ammation, SCC does not. When the elevation of both IAP and SCC was found in patients with H&NSCC, we speculated that they had recurrent and distant metastatic disease. We wish to stress that the combined measurement of IAP, SCC and CEA was the best combination for monitoring H&NSCC in terms of cost and effectiveness. Tumor markers have been demonstrated to be clinically helpful in detecting various human neoplasms in the last few decades. However, the role of serum tumor markers in the management of H&NSCC has attracted relatively less interest compared with that of other tumors. Since the lesions of H&NSCC are easy to see and accessible, it is easy to obtain samples for biopsy. The positive rate of each tumor marker was low in this study (10 to 55% among ®ve tumor markers). Furthermore, in patients in early stages, few patients (16 IAP cases, 41 sialic acid cases, 12 SCC cases, 26 CEA cases, eight ferritin cases) had high levels of tumor markers. These results suggested that conventional tumor markers, at least the ®ve
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items we measured, are useless to detect early H&NSCC. However, a predictable tumor marker for prognosis would bene®t the treatment for H&NSCC. Adjuvant therapy following surgical resection and induction chemotherapy followed by surgery have been proposed as the best approaches for improving results. Although much more extensive study is needed to adequately evaluate the practical applications of these procedures, the present results indicate that serum IAP, sialic acid and SCC levels may be useful monitors for determining whether to perform adjuvant chemotherapy or post-operative irradiation for H&NSCC. In particular, aggressive therapy for patients with high IAP levels prior to therapy should be considered. References [1] P.E. Maxim, R.W. Velti, Serum ferritin as a tumor marker in patients with squamous cell carcinoma of head and neck, Cancer 57 (1986) 305±311. [2] C.D.E. Eibing, J.T. Johnson, R.L. Wagner, S. Su, SCC-RIA in the diagnosis of squamous cell carcinoma of head and neck, Laryngoscope 99 (1989) 117±124. [3] W. Fischbach, T. Meyer, K. Barthel, Squamous cell carcinoma antigen in the diagnosis and treatment follow-up of oral and facial squamous cell carcinoma, Cancer 65 (1990) 1321±1324. [4] A.I. Dreyfuss, J.R. Clark, J.W. Andersen, Lipid-associated sialic acid, squamous cell carcinoma antigen, carcinoembryonic antigen, and lactic dehydrogenase levels as tumor markers in squamous cell carcinoma of head and neck, Cancer 70 (1992) 2499±2503. [5] M.B. Straka, R.L. Wagner, J.T. Johnson, K.K. Kachman, D.E. Eibling, The lack of utility of a tumor marker panel in head and neck carcinoma. Squamous cell carcinoma antigen, carcinoembryonic antigen, lipid-associated acid, and CA-125, Arch. Otolaryngol. Head Neck Surg. 118 (1992) 802±805. [6] E.K. Walther, N. Dahlmann, H.T. Gorgulla, Tumor markers in
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