Invasive pattern grading score designed as an independent
prognostic indicator in oral squamous cell carcinoma
Yun-Ching Chang, Shin Nieh,
1Su-Feng Chen,
2Shu-Wen Jao,
3Yu-Lu Lin
4& Earl Fu
5 Graduate Institute of Life Sciences,1Department of Pathology and Graduate Institute of Pathology, National Defence Medical Centre & Tri-Service General Hospital,2Department of Dental Hygiene, China Medical University,3Department of Surgery,4Graduate Institute of Pathology and5Department of Dentistry and Graduate Institute of Dental Sciences, National Defence Medical Centre & Tri-Service General Hospital, Taipei, Taiwan, ChinaDate of submission 5 May 2009
Accepted for publication 11 December 2009
Chang Y-C, Nieh S, Chen S-F, Jao S-W, Lin Y-L & Fu E (2010) Histopathology
Invasive pattern grading score designed as an independent prognostic indicator in oral
squamous cell carcinoma
Aims: To test the validity of an invasive pattern grading score (IPGS) developed for oral squamous cell carcinoma (OSCC) as a prognostic indicator and to elucidate the relationship between the IPGS and clinical parameters.
Methods and results: The IPGS was applied to a total of 153 cases of OSCC. There were significant correlations between IPGS and distant metastasis (P = 0.01) or recurrence (P = 0.001). However, there were no sig-nificant correlations between IPGS and gender, age, size or extent, location, status of lymph node
metasta-sis, clinical staging, or histological grading. Cases of OSCC with higher IPGS were associated with poor patient survival (P < 0.001) and higher probability of tumour recurrence (P = 0.001). Intraobserver (j = 0.74) and interobserver agreement (j = 0.67) were very satisfactory.
Conclusions: Our study confirms the validity of the IPGS, an indicator that is simple and easy to use. IPGS not only provides histological assessment of biological behaviour, but also offers an independent prognostic factor that may influence the treatment of OSCC.
Keywords: clinical parameters, independent prognostic indicator, invasive pattern grading score, oral squamous cell carcinoma
Abbreviations: IPGS, invasive pattern grading score; OSCC, oral squamous cell carcinoma; POI, pattern of invasion; TMA, tissue microarray; TNM, tumour node metastasis
Introduction
Oral squamous cell carcinoma (OSCC) is one of the most common head and neck carcinomas, and its morbidity and mortality are increasing both in Taiwan and worldwide.1 Despite substantial developments in both diagnosis and therapy in recent decades, the
prognosis of male predominant OSCC remains poor.2,3 Extensive local invasion and⁄ or frequent regional lymph node metastases are usually present even at initial diagnosis, resulting in the unpredictable prog-nosis of OSCC.4 Clinical assessment by the tumour node metastasis (TNM) system is widely and routinely used to define the extent of tumour load and thus determine treatment options for patients with OSCC.5 One of the major criticisms of the TNM system is that it ignores individual histological characteristics of tu-mours.6Therefore, many workers have devised histo-logical grading systems to predict the biohisto-logical behaviour and recommended prognostic markers for OSCC, such as cell morphometry, proliferation-associ-Address for correspondence: S Nieh and S-F Chen, Department of
Pathology and Graduate Institute of Pathology, School of Medicine, National Defence Medical Centre and Tri-Service General Hospital, No. 325, Cheng-Kung Rd., Sec.2, Neihu 114, Taipei, Taiwan, China. e-mails: [email protected] and [email protected] S.N. and S-F.C. contributed equally to this work.
pathological use. Multi-parameter prognostic models and scoring systems have been developed and refined over the last two decades based on histological variables that include nuclear pleomorphism, mitotic index, lymphocytic response, tumour growth pattern, tumour thickness, degree of keratinization, depth of invasion, and pattern of invasion (POI).10–13 The histological features of OSCC may differ widely from area to area within the same tumour due to tumour heterogeneity and is subject to inter- and intraobserver disagreement.14 There is general agreement that the most useful prognostic information can be deduced from the invasive front of the tumour, where the deepest and presumably most aggressive cells reside.5,8,15,16
In our previous publication,17 we clearly demon-strated that intense staining of Fascin was identified at the invasive front of the tumour cell nests in patients with OSCC. This phenomenon has also been described by other investigators. We further noticed that increased Fascin expression is correlated with aggres-sive behaviour and poor patient outcome in addition to higher histological grades. Therefore, we designed a modified invasive pattern grading score (IPGS) inspired by the Gleason grading system, which gives a total IPGS consisting of the sum of the two most prevalent patterns present at on the invasive front of the tumour. The aim of the current study was to test the validity of the newly defined IPGS as a reliable and independent prognostic indicator by correlating IPGS with clinical parameters.
Materials and methods
p a t i e n t a n d t i s s u e s p e c i m e n sTissue specimens of 176 patients with OSCC were retrieved from the archives of the Department of Pathology, Tri-Service General Hospital (Taipei, Tai-wan) from January 2000 to December 2002. The srudy of all specimens received ethical approval from Tri-Service General Hospital Institutional Review Board (Board No. 096-05-0008). The majority of cases were selected from radical surgical specimens of OSCC. Twenty-three cases were lost to follow-up or had insufficient clinicopathological data for analysis and were therefore excluded. Initially, a tissue microarray (TMA) of OSCC was constructed and examined for analysis of IPGS but was found to be ineffective for
invasive front. Thus, a total of 153 cases of OSCC were included in the study.
h i s t o p a t h o l o g i c a l e v a l u a t i o n
All the histopathological slides were concurrently reviewed and evaluated independently by two qualified pathologists (the first observer and the second observer) using the same type of microscope without any prior knowledge of each patient’s clinical details. When the opinions of the two evaluators differed, consensus was reached by discussion. To assess intraobserver agree-ment, the same slides were reassigned to the first observer, who was blinded to the results of the first assessment and to the outcome after an interval of 3 months.
i n v a s i o n p a t t e r n g r a d i n g s c o r e
The IPGS was based on the classification of POI originally introduced by Jakobsson et al., and further defined by Bryne et al.15,18 POI type 1 represents tumour invasion in a broad pushing manner with a smooth outline. POI type 2 represents tumour invasion with broad pushing ‘fingers’, or separate large tumour islands, with a stellate appearance. POI type 3 repre-sents invasive islands of tumour greater than 15 cells per island. POI type 4 represents invasive tumour islands smaller than 15 cells per island including cord-like and single-cell invasion. We further modified the above-mentioned POI, leading to a newly designed IPGS. A search for the deep invasive front of OSCC interfacing with the stromal tissue to signify the invasive pattern or patterns was performed and scored correspondingly under both low- and high-power field microscopy. Because of the histological variations within each tumour, the two most prevalent patterns at the invasive front of OSCC, the predominant, or primary grade and the less extensive, or secondary grade, were given a total score, the IPGS. Tertiary patterns were not considered in the IPGS. Since each invasive pattern score was assigned a number between 1 and 4, the total summed scores ranged from 2 to 8. Furthermore, in the case of a consistent invasive pattern, if only one grade was present, IPGS was gained by doubling the invasive score. Twenty percent was used as the minimal cut-off for incorporating a tumour invasion pattern into the IPGS. Comparisons and illustrations of the four main previously described
POIs of OSCC correlated with the new IPGS are shown in Figure 1.
s t a t i s t i c a l a n a l y s i s
v2 test (with adequate Yates’ correction when 2· 2 table and d.f. = 1, or with Fisher’s exact test when expected value <5 in >20% table cells) was used to measure the significance between clinical parameters and IPGS or recurrence. The log rank test was used to compare differences in survival between two groups. The prognostic significance of clinicopathological parameters and IPGS for overall survival was assessed by Cox’s regression model. Survival curves were obtained by the Kaplan–Meier method. P < 0.05 was considered to be significant. j statistics were used to investigate the reliability of the IPGS by measuring inter- and intraobserver agreement. A value of j < 0.20 was considered as poor agreement, 0.21– 0.40 as fair, 0.41–0.60 as moderate, 0.61–0.80 as good and values >0.81 as very good agreement.19 SPSS software v.13.0 (SPSS UK Ltd, Woking, UK) was used for analysis.
Results
m o r p h o l o g i c a l d e s c r i p t i o n a n d m e a s u r e m e n t o f i p g s i n o s c c
The clinicopathological data of 153 patients with OSCC are summarized in Table 1. According to our newly modified IPGS as described in Figure 1, each case was assigned an IPGS between 2 and 8 (Figure 2). The two most frequent IPGS expressed in a total of 153 cases of OSCC were IPGS 4 (n = 45, III + I N = 2 and II + II N = 43) and IPGS 5 (n = 39, II + III N = 38 and III + II N = 1). Groups were classified as high IPGS (‡5) or low IPGS (£4) with score 4 as the threshold point. There were 77 cases of OSCC with low IPGS and 76 cases of OSCC with high IPGS (Figure 3).
c o r r e l a t i o n b e t w e e n i p g s a n d
c l i n i c o p a t h o l o g i c a l p a r a m e t e r s i n o s c c When comparing IPGS with the clinicopathological parameters, there was no significant correlation between IPGS and gender, age, size or extent of the
Epithelium
Basement membrane
Stroma
Large islands
Invasion pattern Pattern I Pattern II Pattern III Pattern IV
Small island and thin strands Individual tumour cells Morphological characteristics Newly modified IPGS
I+I II+II III+III IV+IV
IPGS = 2
I+II / II+I II+III / III+II III+IV / IV+III
IPGS = 3 IPGS = 4 IPGS = 5 IPGS = 6 IPGS = 7 IPGS = 8 Solid sheet
with a pushing border
rence (P = 0.001) of OSCC (Table 2).
i p g s u s e d a s a n i n d e p e n d e n t p r o g n o s t i c i n d i c a t o r i n o s c c
When comparing the predictor of tumour recurrence with clinicopathological parameters, histological differ-entiation and IPGS, there were statistically significant correlations between tumour recurrence and age (P = 0.036), status of lymph node metastasis (P = 0.039), distant metastasis (P < 0.001), clinical staging (P < 0.001) and IPGS (P = 0.001). However, there were no significant correlations between tumour recurrence and gender, size or extent of the tumour, and histological grading (Table 3).
Follow-up of 153 cases with OSCC from initial diagnosis to 72 months was performed and the overall survival rate estimated at 42.5%. There was a statisti-cally significant correlation between patient survival and the size or extent of the tumour (P < 0.001), lymph node metastasis (P = 0.033), distant metastasis (P < 0.001), clinical staging (P < 0.001), tumour recurrence (P < 0.001), histological grading (P = 0.002) and IPGS (P = 0.001) by the log rank test. Cox’s multivariate analysis also showed that size or extent of the tumour (P < 0.001), positive lymph node metastasis (P = 0.037), distant metastasis (P < 0.001), clinical staging (P < 0.001), tumour recurrence (P < 0.001), histological grading (P = 0.003) and IPGS (P = 0.001) were significant prognosticators (Table 4). Kaplan–Meier plot analysis demonstrated statistical significance in terms of clinical staging, tumour recur-rence, histological grading, and IPGS (Figure 4). Cases of OSCC with high IPGS were associated with strong invasive ability and the implication of poor prognosis.
i p g s a l s o r e g a r d e d a s a r e l i a b l e i n d i c a t o r i n o s c c
Interobserver agreement was calculated from the first assessment of the first observer and that of the second observer by double blind methods. The j score of interobserver agreement in the original data was 0.69, which is good agreement. To assess intraobserver agreement, the same slides were reassigned to the first observer, who was blinded to the results of the first assessment after an interval of 3 months. The j score for intraobserver agreement (j = 0.74) was slightly higher than the interobserver scores (j = 0.69), and
Clinicopathological parameters n (%) Age Median (52.34) £52 85 (55.6) >52 68 (44.4) Gender Male 129 (84.3) Female 24 (15.7) Location Tongue 115 (75.2) Buccal mucosa 36 (23.5) Gingiva 2 (1.3) T stage 1 31 (20.3) 2 50 (32.7) 3 33 (21.5) 4 39 (25.5) N stage 0 106 (69.3) 1 27 (17.6) 2 14 (9.1) 3 6 (4) M stage 0 132 (86.3) 1 21 (13.7) TNM stage I 22 (14.4) II 33 (21.6) III 36 (23.5) IV 62 (40.5) Recurrence No 106 (69.3) Yes 47 (30.7) Differentiation Well 54 (35.3) Moderate 82 (53.6) Poor 17 (11.1)
can also be categorized as good agreement. Similarly, based on the categories of high and low IPGS, the j score of interobserver agreement was 0.72, which can also be considered good agreement, and intraobserver agreement was 0.81, which can be regarded as very good agreement (Table 5).
Discussion
The inevitable histological variations in OSCC can be closely related to prognosis. In addition to subjective histological interpretations of OSCC by pathologists, the major problem encountered when investigating
OSCC is its heterogeneity both between tumours, within individual tumours, and between close but biologically different oral anatomical sites.8All of these factors complicate the prognosis of OSCC, resulting in unpredictability.
Many histological prognostic models and scoring systems have been developed in previous series for predicting the biological behaviour of OSCC. Broders’ system was first established on the basis of the proportion of highly differentiated cells in 1920.20,21 Although Broders’ system was simple and widely used, it was a poor predictor for survival or metastasis.6,22 In 1973, Jakobsson et al.18 developed a multifactorial grading system which had the advantage of scoring tumour–host interactions and tumour characteristics, but eventually proved to only be useful when applied to tongue cancers.23,24Later, Anneroth et al.25 proposed a modification of Jakobsson’s system based on the assessments of six histomorphological parameters including degree of keratinization, nuclear pleomor-phism, pattern of invasion, host response and mitotic activity. Bryne et al.15 modified Anneroth’s grading system and developed a malignancy grading system focusing only on the invasive front of the tumour. This method of grading appeared to be less time-consuming than those of Jakobsson et al. and Anneroth et al.26,27 Nevertheless, this system was not sufficiently homoge-neous to allow grading parameters to be assessed individually.
IPGS = 2 (I+I) IPGS = 3 (I+II) IPGS = 4 (II+II)
IPGS = 6 (II+IV) IPGS = 6 (III+III) IPGS = 7 (III+IV) IPGS = 8 (IV+IV) IPGS = 5 (II+III)
Figure 2. Representative microscopic photographs of OSCC with assigned IPGS from 2 to 8 (H&E stain,·200).
50 Low High 4+4 4+3 3+4 3+3 2+4 3+2 2+3 3+1 2+2 2+1 1+2 1+1 Case n umbers 45 40 35 30 25 20 15 10 5 2 3 4
Invasion pattern grading scores
5 6 7 8
Figure 3. The distribution of each individual IPGS in a total of 153 patients with OSCC.
A variety of clinical and histological parameters may indeed influence patient prognosis and outcome, but independence and reliability are important and essen-tial for a chosen prognostic parameter, especially in regard to local recurrence and overall survival. The
IPGS in our study was developed from original POI to satisfy the above-mentioned independence and reliabil-ity for prognostic evaluation of OSCC. As shown in Tables 2 and 3, we have demonstrated that IPGS is an independent parameter that does not interact with and is not influenced by the majority of the important clinicopathological parameters, including size or extent of the tumour, status of lymph node metastasis, clinical or histological grading. However, due to significant correlations between IPGS and distant metastasis and tumour recurrence, as well as poor survival, IPGS is considered to be a reliable prognostic parameter. From Clinicopathological parameters Low IPGS (N = 77) High IPGS (N = 76) P-value Age £52 41 44 NS >52 36 32 Gender Male 67 62 NS Female 10 14 Location Tongue 59 56 NS Buccal mucosa 18 18 Gingiva 0 2 T stage T1 ⁄ T2 41 40 NS T3 ⁄ T4 36 36 N stage N0 54 52 NS N1 ⁄ N2 ⁄ N3 23 24 M stage 0 72 60 0.01 1 5 12 TNM stage I ⁄ II 33 22 NS III ⁄ IV 44 54 Recurrence No 63 43 0.001 Yes 14 33 Differentiation Well 29 25 NS Moderate 43 39 Poor 5 12 NS, not significant. Clinicopathological parameters No. of cases P-value No (N = 106) Yes (N = 47) Age £52 85 65 20 0.036 >52 68 41 27 Gender Male 129 87 42 NS Female 24 19 5 T stage T1 ⁄ T2 81 61 20 NS T3 ⁄ T4 72 45 27 N stage N0 106 79 27 0.039 N1 ⁄ N2 ⁄ N3 37 27 20 M stage 0 132 102 30 <0.001 1 21 4 17 TNM stage I ⁄ II 55 48 7 <0.001 III ⁄ IV 98 58 40 Differentiation Well ⁄ moderate 136 96 40 NS Poor 17 10 7 IPGS Low (24) 77 63 14 0.001 High (58) 76 43 33 NS, not significant.
Table 4. Statistical correlation of clinical parameters, differentiation and invasive pattern grading score (IPGS) with survival
Variables Cut-off levels No. of cases Log-rank P-value Cox regression P-value
Age £52 versus >52 85 ⁄ 68 0.513 0.518
Gender Male versus female 129 ⁄ 24 0.714 0.717
T stage T1 ⁄ T2 versus T3 ⁄ T4 81 ⁄ 72 <0.001 <0.001
N stage N0 versus N1 ⁄ N2 ⁄ N3 106 ⁄ 47 0.033 0.037
M stage M0 versus M1 132 ⁄ 21 <0.001 <0.001
TNM stage I ⁄ II versus III ⁄ IV 55 ⁄ 98 <0.001 <0.001
Recurrence No versus yes 106 ⁄ 47 <0.001 <0.001
Differentiation Well ⁄ moderate versus poor 136 ⁄ 17 0.002 0.003
IPGS Low versus high 77 ⁄ 76 <0.001 0.001
1.0
P < 0.001 P = 0.002
P < 0.001 P = 0.001
Stage 1 and stage 2 (n = 55)
No recurrence (n = 106) IPGS 2~4 (n = 77) IPGS 5~8 (n = 76) Recurrence (n = 47) Well to moderately differentiated (n = 136) Poorly differentiated (n = 17) Stage 3 and stage 4 (n = 98)
0.8 0.6 0.4 0.2 0.0 0.00 20.00 40.00 60.00
Months after diagnosis
80.00
0.00 20.00 40.00 60.00
Months after diagnosis
80.00
0.00 20.00 40.00 60.00
Months after diagnosis
80.00
0.00 20.00 40.00 60.00
Months after diagnosis
80.00 TNM stage Stage 1/2 Stage 3/4 Differentiation Poor Well/moderated IPGS High Low Recurrence No Yes Cum sur viv al 1.0 0.8 0.6 0.4 0.2 0.0 Cum sur v iv al 1.0 0.8 0.6 0.4 0.2 0.0 Cum sur viv al 1.0 0.8 0.6 0.4 0.2 0.0 Cum sur v iv al
Figure 4. Statistical differences were found by comparisons of cumulative survival in terms of the clinical stage, histological grading, tumor recurrence, and IPGS in 153 patients with OSCC by Kaplan–Meier analysis.
the practical standpoint, IPGS is simple and easy to use by calculating and scoring the most representative invasion pattern or patterns. Based on our data, cases of OSCC with low IPGS were associated with limited invasion and a favourable prognosis, including infre-quent recurrence and relatively long survival, whereas cases of OSCC with high IPGS were associated with increased invasion and poor overall survival, a propensity to tumour recurrence and even distant metastasis. Thus, IPGS might thus influence the strategy for treatment.
As mentioned above, IPGS was derived from four types of POI. Each POI has its own morphological features and definition. Cases of verrucous carcinoma were not included in the study due to their predictable favourable outcomes and the monotonous histological presentation of pattern I without other invasion patterns.28,29 Besides these four types of invasion pattern, a fifth pattern was introduced by Brandwein-Gensler et al.30 by defining a separate and infiltrative invasion pattern as tumour satellites of any size with ‡1 mm distance of intervening normal tissue at the tumour–host interface. Although this type of POI can be seen in some cases or in some areas of the tumour, we believe that this widely dispersed pattern is rare, unpredictable and unmeasurable, as well as too rare to calculate. Furthermore, pattern 5 does not seem to be well established and accepted in the field of head and neck cancer and therefore we decided to exclude it from our study.
Based on our data, thej scores indicated good inter-and intraobserver variability in assessment of IPGS. Compared with the study performed by Sawair et al.,5 our data using IPGS derived by adding up the scores of the two most prevalent patterns at the invasive front of OSCC resulted in a more reproducible and reliable statistical j score. To the best of our knowledge, the IPGS is reported for the first time and is a reliable and independent prognostic factor that may influence the treatment plan. This is an initial study and the assessment of more cases of OSCC is required to support
Acknowledgements
This study was partly supported by the National Science Council, Republic of China, Grant No. NSC98-2314-B-016-019-MY3 (1⁄ 3); Tri-Service General Hospital, Grant No. TSGH-C98-10-S02 and Ministry of National Defence, Grant No. DOD98-43.
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