The Prognostic Impact of Grading in FIGO IB and IIB Squamous Cell Cervical Carcinomas.

Background Tumor grade is one of the more controversial factors, and the data regarding its prognostic impact in squamous cell carcinoma (SCC) of the uterine cervix are controversial. Methods The histological slides of 467 surgically treated FIGO stage IB1 to IIB cervical SCC were re-examined regarding the prognostic impact of the histological tumor grade based on the degree of keratinization (conventional tumor grade) according to the WHO recommendation on recurrence-free and overall survival as well as on the prediction of pelvic lymph node involvement. Results 46.0% presented with well-differentiated tumors (G1, n = 215), 30.6% with moderate (G2, n = 143) and 23.3% with poor differentiation (G3, n = 109). The recurrence-free survival was significantly reduced in patients with poorly differentiated tumors (G1: 81.4%, G2: 70.6%, G3: 64.2%; p = 0.008). There was no impact on overall survival. Because of the lack of survival differences between G1- and G2-tumors, they were merged into low-grade tumors, and their prognostic outcome was compared to the high-grade group (G3-tumors). Based on this binary conventional grading system there was a significantly longer recurrence-free (low-grade: 77.1% vs. high-grade: 64.2%; p = 0.008) and overall survival (low-grade: 76.0% vs. high-grade: 65.1%; p = 0.031) in the low-grade group. However, both the conventional three-tiered and the binary grading systems (separating tumors into a low- and high-grade group) failed to predict pelvic lymph node involvement (p = 0.9 and 0.76, respectively). Conclusion A binary grading model for the conventional tumor grade (based on the degree of keratinization) in SCC of the uterine cervix may be suitable for the prognostic survival evaluation but failed to predict pelvic lymph node involvement.

Introduction

There are several reports in the literature on survival and disease-free intervals and on their relationship to surgical-pathological factors in cervical carcinoma of the uterus 1 , 2 , 3 , 4 . Among these factors, pelvic lymph node involvement and tumor stage are well-established prognostic parameters. The histopathological grading of cervical cancer, however, is one of the more controversial factors, especially in squamous cell carcinomas (SCC) 2 ,  5 ,  6 . Historically, grading of cervical SCCs was performed using Broderʼs system or modifications thereof based on the degree of keratinization, cytological atypia and mitotic activity 7 ,  8 . This grading system has been adopted by the WHO classification and is still valid, but it tends to be vague and unprecise 6 ,  9 . Therefore, the present study was designed to evaluate the prognostic impact of the conventional grading based on the degree of keratinization within a large cohort of SCCs. A potential correlation of the conventional tumor grade with the recurrence-free and overall survival as well as pelvic lymph node metastases was used to determine the prognostic significance of grading in squamous cell cervical carcinomas. This may help to better define the role of tumor grade in cervical SCC and its possible consequences on clinical treatment decisions.

Material and Methods

Patients

Data from 467 consecutive squamous cell cervical cancer patients, clinically staged FIGO IB1 to IIB, who underwent upfront surgery were obtained from the files of the Institute of Pathology, University Hospital of Leipzig, Germany. Patients who received neoadjuvant therapy, those with incomplete local tumor resection and non-squamous cell histology were excluded from the study. Prior to the introduction of the TMMR-technique 10 at our institution, all women were treated by a radical abdominal hysterectomy Piver type III 11 . All patients with parametrial involvement and/or lymph node metastases underwent adjuvant intracavitary radiotherapy (ICRT) and external beam chemoradiation.

Pathological examination

The pathological examination of the radical hysterectomy specimen was performed in a standardized manner 12 . All tumors were staged and classified according to the WHO and TNM-classifications 6 ,  13 .

As a detailed description of the different grades is not included in the current WHO classification 6 , tumors were graded in accordance to previous studies 14 ,  15 ,  16 ,  17 . In well-differentiated tumors (G1), the tumor cell nests were composed of keratinocyte-like cells with easily visible keratinization features (layered or cytoplasmic keratin). In the poorly differentiated tumors (G3), the squamous morphology was only noticeable in a small area of the tumor, not exceeding 1% of the whole tumor. The moderately differentiated tumors (G2) showed an intermediate degree of squamous differentiation between the well- and poorly differentiated ones ( Fig. 1 a to c ).

Fig. 1

 Histologic images of different grades of squamous cell carcinoma of the uterine cervix. a  G1: well-differentiated SCC: tumor cell nests with easily visible keratinization features (layered or cytoplasmic keratin). b  G2: moderately differentiated SCC with focal keratinization within the tumor cells. c  G3: tumor cell nests with squamous features without keratinization.

The original H & E-stained slides were re-examined on low power magnification (× 25). If necessary, the infiltrating tumor cell nests were screened at intermediate power fields (× 100) for single cell keratinization and intercellular bridges. Two to six slides per case were investigated (mean 2.7 per case).

Follow-up data

Follow-up data were retrieved from the medical records. Written informed consent was obtained from all patients. Additionally, the study was approved by the Institutional Review Board.

Recurrence-free survival (RFS) was calculated from the day of diagnosis until tumor recurrence or end of follow-up. Overall survival (OS) was calculated from the day of diagnosis until death or end of follow-up. Kaplan-Meier survival curves and Log rank tests were used to analyze the survival data. Cox regression analyses were fitted to estimate the impact of grading. All statistical analyses were performed with IBM SPSS Statistics version 24.0.

Results

Grading and prognosis

A total of 467 cases were available for review. The patient characteristics are summarized in Table 1 .

Table 1  Patient characteristics (n = 467).

Follow-up (months)
Median (range)	69 (2 – 182)
Age (years)
Median (range)	41 (23 – 74)
Mean ± SD	42 ± 11
Post-surgical stage distribution
pT1b1	233 (49.9%)
pT1b2	55 (11.8%)
pT2a	49 (10.5%)
pT2b	130 (27.8%)
Pelvic lymph node involvement
No (pN0)	314 (67.2%)
Yes (pN1)	153 (32.8%)
Tumor grade
G1	215 (46.0%)
G2	143 (30.6%)
G3	109 (23.3%)
Lymphovascular space involvement
None (L0)	169 (36.2%)
Yes (L1)	298 (63.8%)
Recurrent disease
No	346 (74.1%)
Yes	121 (25.9%)

46.0% (n = 215) presented with well-differentiated tumors (G1), 30.6% (n = 143) showed moderate differentiation (G2) and 23.3% (n = 109) were poorly differentiated (G3).

The three grading groups were prognostically relevant for recurrence-free (p = 0.008; Fig. 2 a ) but not for overall survival (p = 0.089; Fig. 2 b ).

Fig. 2

 Kaplan-Meier curves for the prognostic impact of conventional tumor grading in squamous cell carcinoma of the uterine cervix using a 3-tiered grading system (please see text). a  Recurrence-free survival. b  Overall survival.

A separate analysis of well- and moderately differentiated tumors regarding recurrence-free and overall survival was performed and revealed no significant difference (p = 0.002 for 5-year recurrence-free survival and p = 0.033 for 5-year overall survival, Table 2 ). This is illustrated in Fig. 2 b where the survival curves are merging. Therefore, the well- and moderately differentiated carcinomas were merged into a low-grade group, whereas the poorly differentiated carcinomas formed the high-grade group. 358/467 patients (76.6%) were found to have low-grade, 109/467 (23.3%) high-grade tumors.

Table 2  5-year-recurrence-free (RFS) and -overall survival (OS) of the different grading groups using the conventional grading system for squamous cell carcinomas of the uterine cervix (p-values [Log rank test]).

	5-year recurrence-free survival	p-values
G1	80.4% (95% CI: 74.7 – 86.0%)	p = 0.089 (G1 vs. G2)	p = 0.002 (G1 vs. G3)
G2	74.2% (95% CI: 66.9 – 81.5%)
G3	64.4% (95% CI: 54.9 – 73.8%)	p = 0.162 (G2 vs. G3)
		global: p = 0.008
	5-year overall survival	p-values
G1	76.4% (95% CI: 70.0 – 82.8%)	p = 0.603 (G1 vs. G2)	p = 0.033 (G1 vs. G3)
G2	76.4% (95% CI: 69.3 – 83.5%)
G3	67.8% (95% CI: 58.6 – 77.0%)	p = 0.121 (G2 vs. G3)
		global: p = 0.089

As illustrated in Fig. 3 , there was a significant difference between low- and high-grade tumors regarding the recurrence-free and overall survival, respectively. The 5-year recurrence-free survival for G1/G2 versus G3 tumors was 77.9% (95% CI 73.4 – 82.3%) versus 64.4% for the G3 tumors (95% CI 54.9 – 73.8%; p = 0.008). G1/G2 tumors showed a 5-year overall survival of 76.9% (95% CI 72.3 – 81.5%) versus 67.8% for the G3 tumors (95% CI 58.6 – 77.0%; p = 0.031). The survival rates are shown in Table 3 .

Fig. 3

 Kaplan-Meier curves for the prognostic impact of conventional tumor grading in squamous cell carcinoma of the uterine cervix using a 2-tiered grading system (please see text). a  Recurrence-free survival. b  Overall survival.

Table 3  5-year-recurrence-free (RFS) and -overall survival (OS) of the different grading groups using the conventional grading system for squamous cell carcinomas of the uterine cervix (p-values [Log rank test]).

	5-year recurrence-free survival	p-values
G1/G2	77.9% (95% CI: 73.4 – 82.3%)	p = 0.008
G3	64.4% (95% CI: 54.9 – 73.8%)
	5-year overall survival	p-values
G1/G2	76.9% (95% CI: 72.3 – 81.5%)	p = 0.031
G3	67.8% (95% CI: 58.6 – 77.0%)

Grading and lymph node involvement

The frequency of pelvic lymph node involvement for the different grading systems used is presented in Table 5 .

Table 5  Pelvic lymph node involvement within different grading groups.

Conventional grading
	three-tired grading system			binary grading system
	G1(n = 215)	G2(n = 143)	G3(n = 109)	low-grade(n = 358)	high-grade(n = 109)
pN0	67.4%	67.8%	66.1%	67.6%	66.1%
pN1	32.6%	32.2%	33.9%	32.4%	33.9%
p-value	p = 0.953			p = 0.764

The conventional three-tiered grading system failed to predict pelvic lymph node involvement. The odds ratios for G1 versus G2 (0.98 [95% CI: 0.62 – 1.54]) and G2 versus G3 (1.08 [95% CI: 0.64 – 1.84]) were not statistically different. Also, the binary grading was unable to predict pelvic lymph node involvement (OR 1.07 [95% CI: 0.68 – 1.7]; p = 0.76).

Discussion

Clinical and postsurgical tumor stage as well as lymph node status are the most powerful predictors of outcome in cervical cancer 2 ,  3 ,  4 . The data regarding the prognostic impact of the tumor grade in squamous cell cancers (SCC) of the uterine cervix are controversial, and several approaches using different morphologic variables have been applied 15 .

Historically, cervical SCCs were graded using Broderʼs system 7 or modifications thereof based on the degree of keratinization 8 . This particular grading system of cervical SCCs is currently mentioned in the last two editions of the WHO classification 6 ,  9 and is referred to as the conventional grading in order to separate it from other grading systems. In an earlier GOG-study, the three-year disease-free interval correlated significantly with the conventional tumor grade (G1: 90.6%, G2: 86.0% and for G3: 76.1%; p = 0.001) 14 . In a small study of 97 patients, grading had an impact on overall survival 18 . Other studies including those with multivariate approaches showed no prognostic significance of conventional grading 1 ,  16 . In the present study, the conventional grading failed to show a prognostic impact on overall survival. There were no differences between well-differentiated (G1) and moderately differentiated (G2) tumors in regards to recurrence-free and overall survival ( Fig. 2 and Table 2 ). After merging G1 and G2 tumors into low-grade tumors, the two-tiered conventional grading of cervical SCCs showed a prognostic impact both on recurrence-free and overall survival ( Fig. 3 and Tables 2 and 3 ).

In the present study, both the conventional three-tiered and the binary grading system failed to predict pelvic lymph node involvement ( Table 5 ).

In multivariate analyses including pelvic lymph node status, grading and the post-surgical tumor stage, the binary grading system was prognostically significant for recurrence-free as well as overall survival ( Table 4 ).

Table 4  Cox-regression analyses for recurrence-free and overall survival.

	HR	p-values
Recurrence-free survival
Pelvic lymph node involvement
No (pN0)	ref
Yes (pN1)	2.7 (95% CI: 1.8 – 3.9)	p < 0.001
Histological tumor grade
Low-grade (G1 + G2)	ref
High-grade (G3)	1.8 (95% CI: 1.2 – 2.6)	p = 0.003
Post-surgical stage
pT1b1	ref
pT1b2	2.2 (95% CI: 1.2 – 3.5)	p = 0.013
pT2a	1.4 (95% CI: 0.7 – 2.8)	p = 0.373
pT2b	2.4 (95% CI: 1.5 – 3.7)	p < 0.001
Overall survival
Pelvic lymph node involvement
No (pN0)	ref
Yes (pN1)	2.8 (95% CI: 1.9 – 4.1)	p < 0.001
Histological tumor grade
Low-grade (G1 + G2)	ref
High-grade (G3)	1.7 (95% CI: 1.1 – 2.4)	p = 0.011
Post-surgical stage
pT1b1	ref
pT1b2	1.6 (95% CI: 0.9 – 3.0)	p = 0.107
pT2a	1.7 (95% CI: 0.9 – 3.3)	p = 0.102
pT2b	2.7 (95% CI: 1.7 – 4.1)	p < 0.001

The advantage of the binary grading system may be the easier morphological categorization using only two rather than three different categories as well as the better reproducibility. While the conventional three-tiered grading system failed to predict prognosis, the binary system was useful in predicting both a reduced recurrence-free and overall survival. One limitation of the present study may be that it includes a variety of different stages of cervical carcinoma which may impact the results. The inclusion of different stages may also be responsible for the failure to predict pelvic lymph node involvement within the different grading groups.

Many attempts regarding the definition of grading systems on SCCs of the cervix uteri have been made 15 . Within that context, an invasive front grading was suggested 19 . This type of grading was first introduced in head and neck SCCs 20 and evaluates the degree of keratinization, nuclear pleomorphism, pattern of invasion and peritumoral host response at the infiltrative edge of SCCs. In cervical cancer, however, data on this grading system are very limited. Very recently, tumor cell budding (evaluating tumor growth by cell nest size) was studied in cervical SCC 21 . The tumor budding as a variant of the grading of a malignancy was previously validated in esophageal, lung and oral SCC.

Furthermore, studies that discuss tumor grade as a prognostic factor give no detailed description of the grading system that was applied 22 ,  23 . At this point, there is no well-accepted and widely used grading system for cervical SCCs 24 ,  25 , nor has one been recommended by the current WHO classification 6 . Because of these limitations, tumor grading is not listed as a required but rather a recommended feature in the most recent recommendations of the International Collaboration on Cancer Reporting 5 for cervical cancer. The German national guidelines for the diagnosis and treatment of cervical carcinoma point out that grading should not be used as a single factor for adjuvant treatment decision 26 .

Finally, additional studies are required to define standardized and reproducible criteria for grading with stage-by-stage analyses and a multivariate approach.

Conclusions

A binary grading model for the conventional tumor grade (based on the degree of keratinization) in SCC of the uterine cervix may be suitable for the prognostic survival evaluation but failed to predict pelvic lymph node involvement.