Abstract
Aim: High [18F]-2-fluorodeoxyglucose (18F-FDG) -uptake of primary tumor, assessed by pretreatment positron emission tomography combined computed tomography (PET/CT), has indicated poor overall survival (OS) in head and neck cancer (HNC).We investigated the correlation between 18F-FDG-uptake and in vitro chemosensitivity of cisplatin using histoculture drug response assay in HNC. Patients and Methods: Twenty-eight patients were evaluated. The maximum standardized uptake value (SUVmax)and inhibition index (I.I.) cisplatin were calculated as 18F-FDG-uptake and in vitro chemosensitivity of cisplatin. Results: Each SUVmax≥10.5 or I.I.cisplatin <50 could significantly differentiate shorter survival group by OS analyses. I.I.cisplatin of patients with SUVmax ≥10.5 was significantly greater. In 19 patients with SUVmax ≥10.5, those who received treatment with cisplatin-based chemotherapy exhibited a significant correlation with longer OS. Conclusion: Cisplatin hasthe potential to improve OS for HNC patients that were predicted as shorter OS by18F-FDG-PET/CT.
Positron emission tomography with computed tomography (PET/CT) employing a glucose analogue, [18F]-2-fluorodeoxyglucose (18F-FDG), are widely used imaging procedures for accurately staging various cancers (1, 2). The semi-quantitative measurement of the maximum 18F-FDG-uptake in the primary tumor, which is assessed using 18F-FDG-PET/CT, is usually obtained according to the maximum standardized uptake value (SUVmax) (3-13). Many reports of patients with head and neck squamous cell carcinoma (SCC), non-small cell lung cancer and other cancers have demonstrated that a high SUVmax of the primary tumor is reported to predict shorter overall survival (OS) (3-12). Several investigators have suggested that patients who are predicted as poor-prognosis by high SUVmax of primary tumor need more aggressive treatment, such as chemotherapy. The correlation, however, between SUVmax and chemosensitivity, which is reported as a useful predictor of response to chemotherapy, has not been fully investigated (3, 4).
The histoculture drug response assay (HDRA), which is an in vitro chemosensitivity assay, has been reported as a useful predictor for response to chemotherapy in various cancers (14-18). We have investigated the appropriate concentration in the HDRA for cisplatin, which is the most commonly used agent for head and neck cancer (HNC) (14). Moreover, the results from the HDRA in several cancers, such as HNC, non-small lung cancer and esophageal cancer,have been significantly correlated with survival (15-17). To our best knowledge, correlation between SUVmax and HDRA of cisplatinin patients with HNC has not been investigated so far.
In the present study, we studied the correlation between SUVmax and HDRA of cisplatin in patients with HNC.
Patients and Methods
Patients. Tumor specimens from patients undergoing both pretreatment with 18F-FDG-PET/CT at the Nagoya PET Imaging Center and radical treatment for HNC at the Department of Head and Neck Surgery, Aichi Cancer Center were included in the present study. Twenty-eight tumor specimens from primary tumor sites, which were collected from radical surgery without preoperative chemotherapy or biopsy before treatment, were successfully subjected to HDRA analysis of cisplatin between August 2004 and February 2006. This study was approved by the institutional review board and all patients provided informed consent for all treatments and examinations. Sites of primary tumors were as follows: oropharynx, 9; oral cavity, 7; major salivary gland, 6; maxillary sinus, 2; hypopharynx, 2; larynx, 2. Histologic types of head and neck cancer were 21 patients with SCC and 7 patients with non-SCC (mucoepidermoid carcinoma, 2; adenoid cystic carcinoma, 1; carcinoma ex pleomorphic adenoma, 1; salivary duct carcinoma, 1; epithelial-myoepithelial carcinoma, 1; adenocarcinoma, not otherwise specified, 1). At the first visit, a routine physical examination, nasopharyngoscopy and a blood chemistry test, including blood glucose level were performed;the mean blood glucose level at the first visit was 102.8±18.0 mg/dl (mean±standard deviation (S.D.)). The clinical TNM classification of the international Union against Cancer (sixth edition) was diagnosed by the aforementioned examinations, enhanced cervical computed tomography or magnetic response imaging and 18F-FDG-PET/CT.
Treatments. Ten of thepatients underwent radical treatment with chemotherapy, while the remainder underwent radical treatment without chemotherapy. The treatment included radiotherapy with chemotherapy or induction chemotherapy and the regime of chemotherapy was cisplatin-based chemotherapy, which comprises combination of cisplatin and 5-fluorouracil (5-FU)(19). In accordance with a previous report, 28 patients were grouped by primary treatment modality: curative surgery with or without radiotherapy (surgery group, n=21) and radical radiotherapy with cisplatin-based chemotherapy (radiation group, n=7) (5). The selection of primary treatment modality depended on the histologic type and on whether patients hoped for their organ preservation. Following the completion of treatment, the patients were followed-up at our outpatient clinic. Those identified with early locoregional recurrence underwent radical salvage therapy.
HDRA and inhibition index (I.I.) cisplatin. For the tumor specimens, the in vitro chemosensitivity of cisplatin (Nippon Kayaku, Tokyo, Japan) was examined using the HDRA according to methods described previously (14, 18). The tumor specimens obtained from surgery or biopsy were immediately placed in 35 ml of RPMI 1640 (Sigma, St.Louis, MO, USA) and stored at 4°C. The specimens were washed three times with HBSS (Sigma) and aseptically cut in 10 mg fragments. Collagen sponge gel (Gel Foam; Pharmacia &Upjohn, Kalamazoo, MI, USA) was cut into 1 cm squares and placed into a 24-well microplate (Becton Dickinson Labware, Franklin Lakes, NJ, USA),which contained RPMI 1640 with 20% fetal calf serum (FCS) (Gibco, GrandIsland, NY, USA) at a concentration of 20 μg/ml for cisplatin; an amount just sufficient to contact the collagen gel. Two fragments of the cut specimens, which were placed on the collagen gel, were cultured for 7 days at 37°C in 5 % CO2 atmosphere. For the control group, the same approach was followed and specimens were cultured for 7 daysin culture medium containingonly 1640 RPMI with 20% FCS. The specimens were cultured in two or three wells each for the control and cisplatingroups. After histoculture, 100 μlHBSS containing 0.06 % collagenase type I (Sigma) and 3-(4, 5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H tetrazolium bromide (MTT) (Sigma) buffer saline solution containing 0.1 M sodium succinate were added to each well and incubated for 16 h, time after which the media were removed. For the extraction of the MTT formazan product, the cells were incubated in 0.5 ml of dimethyl sulfoxide (Sigma) for 2 h. Samples of 100 μl from each well were measured by a microplate reader (ImmunoMini NJ-2300; Nalge Nunc International, Rochester, NY, USA). The absorbance was measured at the wavelength of 540nm by using a reference wavelength of 630 nm. The efficacy of cisplatin was calculated by the following formula:I.I. (%)=(1–mean absorbance per gram of treated tumor/mean absorbance per gram of control tumor) ×100. The interval between the start of the HDRA assay and the start of therapy was 2.21±4.52 days (mean±S.D.).
18F-FDG-PET/CT. All patients were scanned using a FDG-PET/CT (DiscoveryLS:GE; Fairfield, CT, USA). The intervals between the 18F-FDG-PET/CT examination and the start of HDRA assay was 16.1±12.0 days and the interval between the 18F-FDG-PET/CT examination and the start of therapy was 17.0±11.5 days. Patients underwent intravenous injection of 18F-FDG (186-315 MBq) after fasting for at least 6 h according to procedures published previously (13).Two experienced radiologists viewed all images on a Xeleris (GE) and the SUV was calculated by the following formula: SUV=Tissue concentration (Bq/g)/{Injection dose (Bq)/body weight (g)}
The SUVmax of the primary tumor was obtained from a region of interest, which was designed as a site of abnormal accumulation on the coronal image.
Statistical analysis. The statistical analysis was carried out using the JMP software package (version 9; SAS; Cary, NC, USA). The relationship between SUVmax and I.I.cisplatin was analyzed by a simple regression analysis. Relationships between two parameters (SUVmax and I.I.cisplatin) and clinical parameters (clinical T and N classifications, clinical stage, age, sex, tumor site, treatment group, with/without cisplatin-based chemotherapy and histologic type) were analyzed using the Spearman's rank correlation and Mann-Whitney U-test. The definition of OS time was the period from pretreatment 18F-FDG-PET/CT to death or date of last contact. Applying the method described in our and others studies, the Kaplan-Meier technique was used to estimate OS rate.Various cutoff values of both SUVmax and I.I.cisplatin were tested using the log-rank test in a univariate OS analysis (3-5). The patients were divided into two groups based on the SUVmax (SUVmax ≥10.5; SUVmax <10.5) and the I.I.cisplatin (I.I.cisplatin≥50; I.I.cisplatin<50) in the univariate OS analysis as a SUVmax of 10.5 and an I.I.cisplatin 50 were found to significantly differentiate the shorter survival group from the longer survival group. The correlation between the two groups (SUVmax ≥10.5; SUVmax <10.5 or I.I.cisplatin ≥50; I.I.cisplatin< 50) on clinical parameters was compared by the chi-square test. The Mann-Whitney's test was used to estimate relationships between the two groups from SUVmax 10.5 (SUVmax ≥10.5; SUVmax < 10.5) on I.I.cisplatin and between the two groups from I.I.cisplatin 50 (I.I.cisplatin≥50; I.I.cisplatin<50) on SUVmax. In the multivariate survival analysis, we used a Cox proportional hazards model. Further study of the multivariate analysis led to adjustments for the two groups from SUVmax 10.5 (SUVmax ≥10.5; SUVmax <10.5) and the two groups from I.I.cisplatin 50 (I.I.cisplatin≥50; I.I.cisplatin<50). In 19 patients with SUVmax ≥10.5, two groups (radical treatment with cisplatin-based chemotherapy; without cisplatin-based chemotherapy) were compared by the log-rank test. A p-value of less than 0.05 was considered to be statistically significant.
Results
The SUVmax and I.I.cisplatin of the primary tumor (mean±SD) were 14.04±7.52 and 50.98±26.6, respectively. The SUVmax was significantly correlated with the I.I.cisplatin (p<0.04, R2=0.17) as shown in Figure 1.
Relationship between SUVmax and I.I.cisplatin in 28 patients with head and neck cancer. SUVmax, maximum standardized uptake value; I.I.,inhibition index.
SUVmax and clinical parameters. The relationships between SUVmax and clinical parameters (clinical T and N classifications, clinical stage, age, sex, tumor site, treatment group, with/without cisplatin-based chemotherapy and histologic type) is shown in Table I. The SUVmax of patients with SCC was significantly higher than in non-SCC (p<0.02) and that of the radiation group was closely greater than in the surgery group (p<0.01). The SUVmax of patients who received radical treatment with cisplatin-based chemotherapy was significantly higher than in patients without cisplatin-based chemotherapy (p<0.02).
I.I.cisplatin and clinical parameters. The relationships between I.I.cisplatin and clinical parameters is shown in Table II. The I.I.cisplatin of the radiation group was significantly greater than in the surgery group (p<0.01) and that of patients who received radical treatment with cisplatin-based chemotherapy was significantly higher than the one without cisplatin-based chemotherapy (p<0.03).
Survival analysis. At the end of this study, the mean±SD follow-up period amongall patients, 17 patients who died (60.7% vs. all) and 11 patients found to be alive (39.3 %) was 48.2±32.4 months, 70.7±32.3 months and 33.6±23.3 months, respectively. Among the total patient population, the 3-year, 4-year and 5-year OS rates were 54.2%, 50.3% and 46.1%, respectively. Applying the method described previously in our and other studies (3-5), various SUVmax and I.I.cisplatin cut-off values were tested using the log-rank test in the OS analysis. The cut-off values with the lowest p-values were used in these analyses: SUVmax=10.5 and I.I.cisplatin=50 (Figure 2). It was shown that each of the SUVmax of ≥10.5 (p<0.02) or I.I.cisplatin <50 (p<0.04) could be used to significantly differentiate the shorter survival group by the log-rank test (Figure 3). No significant correlation between the two groups (SUVmax ≥10.5; SUVmax <10.5) on clinical parameters is shown in Table III. The correlation between the two groups (I.I.cisplatin ≥50; I.I.cisplatin <50) on clinical parameters is shown in Table IV. Patients with I.I.cisplatin ≥50 were more frequently found in the radiation group (p<0.03) and radical treatment with cisplatin-based chemotherapy (p<0.05) than patients with I.I.cisplatin <50. A significant correlation between the two groups from SUVmax 10.5 (SUVmax ≥10.5; SUVmax <10.5) on I.I.cisplatin and no correlation between the two groups from I.I.cisplatin 50 (I.I.cisplatin ≥10.5; I.I.cisplatin <10.5) on SUVmax is shown in Figure 4. The I.I.cisplatin of patients with SUVmax ≥10.5 was significantly greater than that of patients with SUVmax <10.5 (p<0.03).
Relationship between SUVmax and clinical parameters (n=28).
Relationship between I.I.cispatin and clinical parameters (n=28).
Multivariate survival analysis. We performed multivariate analysis with adjustments for the two groups from SUVmax 10.5 (SUVmax ≥10.5; SUVmax <10.5) and the two groups from I.I.cisplatin 50 (I.I.cisplatin≥50; I.I.cisplatin <50) for OS. SUVmax ≥10.5 (p<0.01) and I.I.cisplatin <50 (p<0.01) proved to be significantly shorter survival factors. Multivariate analysis for OS is shown in Table V.
Results with/without cisplatin-based chemotherapy. In 19 patients with SUVmax ≥10.5, those who received radical treatment with cisplatin-based chemotherapy exhibited a significant correlation with longer OS than those who received radical treatment without cisplatin-based chemotherapy (p<0.05) (Figure 5).
Relationships between SUVmax (<10.5/≥10.5) and clinical parameters (n=28).
Discussion
In the present study, we showed for the first time that the I.I.cisplatin of patients with SUVmax ≥10.5 in HNC was significantly greater than that of patients with SUVmax <10.5 and that both SUVmax ≥10.5 (p<0.01) and I.I.cisplatin < 50 (p<0.01) proved to be significantly shorter survival factors in multivariate analysis.
Although 18F-FDG-PET and 18F-FDG-PET/CT are important imaging procedures for the diagnosis and staging of many cancers, their full potential has yet to be established (3-5). Many investigators have demonstrated in head and neck SCC, as well as lung cancer, including various types of carcinoma, such SCC and adenocarcinoma, that higher SUVmax is correlated with shorter OS (3-9). Recently, a review and two meta-analyses of patients with head and neck SCC demonstrated that an increased SUVmax indicates a poor OS (10-12). Moreover, in our previous reports, a high SUVmax of the primary tumor was found to be associated with a shorter OS in patients with both oral SCC and pharyngeal SCC (5, 6). Our results, demonstrating a significant association between patients with SUVmax of ≥10.5 and poor OS, are in agreement with the findings of these previous studies (3-12).
p-Values of log-rank test for overall survival using different cutoff levels for SUVmax (A) and I.I.cisplatin (B) for 28 patients with head and neck cancer. SUVmax, maximum standardized uptake value; I.I.,inhibition index.
(A) SUVmax ≥10.5 and (B) I.I.cisplatin <50 were significantly correlated with shorter overall survival in 28 patients with head and neck cancer by log-rank test. SUVmax, maximum standardized uptake value; I.I.,inhibition index.
The HDRA, an in vitro chemosensitivity assay used in several cancers, has been reported as a useful predictor for response to chemotherapy and survival (14-17). Singh et al. reported that in a group of 41 head and neck SCC, as assessed by HDRA of 5-fluorouracil and cisplatin,thepatients who were chemosensitive had a significantly better 2-year cause-specific survival than those of chemoresistant background (15). Jung et al. found thatin a group of 104 epithelial ovarian cancer patients, including various types of histology, such as clear cell carcinoma, papillary serous adenocarcinoma and undifferentiated carcinoma, as assessed by HDRA of carboplatin, the patients who were chemosensitive to carboplatin exhibited a significantly longer progression-free survival than those who were resistant (16). Our results, demonstrating a significant association between patients with I.I.cisplatin <50 and poor OS are in line with the findings of these previous studies (15-17).
The Mann-Whitney's test was used to estimate relationships (A) between the two groups from SUVmax 10.5 (SUVmax ≥10.5; SUVmax <10.5) on I.I.cisplatin and (B) between the two groups from I.I.cisplatin 50 (I.I.cisplatin ≥50; I.I.cisplatin <50) on SUVmax. SUVmax, maximum standardized uptake value; I.I.,inhibition index.
Many researchers have investigated various characteristics, such as chemosensitivity and useful prognostic parameters in many types of cancers and reported the appropriate concentration in the HDRA for cisplatin, which is the most commonly used agent for HNC (1-18). Moreover, the results from the HDRA in several cancers, such as HNC, non-small lung cancer and esophageal cancer, were significantly correlated with survival (15-17). However, to our best knowledge, the correlation between SUVmax and HDRA of cisplatin has not been investigated thus far.Our present work showed, for the first time,that SUVmax ≥10.5 (p<0.01) and I.I.cisplatin <50 (p<0.01) proved to be significantly shorter survival factors by multivariate analysis for OS. In the present study, the I.I.cisplatin of patients with SUVmax ≥10.5, who were significantly correlated with poor OS, was significantly greater than that of patients with SUVmax <10.5. This finding suggests that cisplatin has the potential to improve OS for HNC patients with SUVmax ≥10.5, who were predicted as shorter OS by 18F-FDG-PET/CT.
Limitations of the present study include the relatively small number of subjects and various pathological type of carcinoma. Thus, in the future, analysis of larger cohorts and one pathological type of carcinoma should yield more statistically accurate results with, hopefully, applicable potential.
Conclusion
We revealed that the I.I.cisplatin of HNC patients with SUVmax ≥10.5 was significantly greater than that of patients with SUVmax <10.5 and that SUVmax ≥10.5 and I.I.CDDP <50 (p<0.01) proved to be significantly shorter survival factors by multivariate analysis for OS. Since patients who received radical treatment with cisplatin-based chemotherapy were significantly correlated with longer OS than those without in a group of 19 patients with SUVmax ≥10.5, cisplatin exhibited a potential trait to improve OS for HNC patients who were predicted as shorter OS by pretreatment with 18F-FDG-PET/CT.
In 19 patients with SUVmax ≥10.5, the patients who received radical treatment with chemotherapy were significantly correlated with longer overall survival.
Relationships between I.I. cisplatin (<50/≥50) and clinical parameters (n=28).
Multivariate overall survival analysis* of overall survival.
Acknowledgements
The Authors are grateful to the technical staff for the PET/CT operation.
This study was supported by JSPS KAKENHI Grant Number 24791821.
Footnotes
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Conflicts of Interest
The Authors have no conflicts of interests.
- Received October 1, 2014.
- Revision received November 28, 2014.
- Accepted December 4, 2014.
- Copyright© 2015 International Institute of Anticancer Research (Dr. John G. Delinassios), All rights reserved
References
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