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Keywords:

  • penile carcinoma;
  • Ki-67;
  • lymph nodes

Abstract

  1. Top of page
  2. Abstract
  3. INTRODUCTION
  4. PATIENTS AND METHODS
  5. RESULTS
  6. DISCUSSION
  7. CONFLICT OF INTEREST
  8. REFERENCES

OBJECTIVE

To investigate the Ki-67 labelling index (LI) as a prognostic factor for the outcome of penile carcinoma, as in squamous cell carcinoma (SCC) of the larynx the expression of this marker correlates with histological features indicative of prognosis.

PATIENTS AND METHODS

We retrospectively analysed the records of 44 patients in whom primary SCC of the penis was treated with amputation and bilateral lymphadenectomy (pT1, in 24, pT2 in 20, pN+ in 10; G1 in 12, G2 in 28 and G3 in four). During a mean follow-up of 35.6 months, four patients had disease progression. Tumour tissue was stained immunohistochemically using the streptavidin-biotin method. The mean Ki-67 LI was defined as the percentage of total tumour cells that were Ki-67-positive. The results were compared with pathological tumour stage, grade, nodal status and clinical disease progression.

RESULTS

The mean (range) Ki-67 LI was 40.5 (6.4–93.0)%; a high mean Ki-67 LI was significantly inversely correlated with tumour differentiation (P < 0.005) and there was a tendency for a high Ki-67 LI to be associated with advanced local tumour stage, nodal metastasis and clinical disease progression, but these correlations were not statistically significant (P = 0.07, 0.07 and 0.06, respectively).

CONCLUSIONS

The Ki-67 LI is correlated with tumour grade in penile cancer, and may indicate a greater risk of nodal metastasis.


Abbreviations
SCC

squamous cell carcinoma

LI

labelling index

PCNA

proliferating cell nuclear antigen.

INTRODUCTION

  1. Top of page
  2. Abstract
  3. INTRODUCTION
  4. PATIENTS AND METHODS
  5. RESULTS
  6. DISCUSSION
  7. CONFLICT OF INTEREST
  8. REFERENCES

The prognosis for patients with squamous cell carcinoma (SCC) of the penis is strongly influenced by presence and extent of regional lymph-node metastases [1,2]; 30–60% of patients with penile carcinoma present with enlarged lymph nodes at physical examination, which can be caused by metastatic involvement or inflammatory reaction. The incidence of occult lymph-node metastases in clinically node-negative cases is 10–15%[3]. The depth of invasion, tumour grade, vascular and lymphatic involvement are the most important risk factors related to the occurrence of nodal metastases in penile carcinoma [4–8]. Inguinal lymphadenectomy is associated with a high morbidity rate of 30–68%[1,9,10]. Therefore, additional prognostic markers that can predict the presence of lymph-node involvement are useful. In the present study we investigated the relationship of Ki-67 labelling in primary penile carcinoma with relevant clinicopathological variables and survival of the patients.

PATIENTS AND METHODS

  1. Top of page
  2. Abstract
  3. INTRODUCTION
  4. PATIENTS AND METHODS
  5. RESULTS
  6. DISCUSSION
  7. CONFLICT OF INTEREST
  8. REFERENCES

We retrospectively analysed the records of 44 patients (mean age 61.4 years, range 35–89) in whom primary SCC of the penis was treated in our department by partial or total amputation and bilateral lymphadenectomy, between 1992 and 2003. The 2003 TNM system was used for staging. None of the patients had received previous treatment or had distant metastases.

Briefly, 5 µm sections of cancer tissue fixed in neutral buffered formalin were stained immunohistochemically for Ki-67 using the streptavidin-biotin method (MIB-1; Dako, Hamburg). To facilitate antigen retrieval, the sections were incubated in a microwave oven in boiling 10 mmol/L citrate buffer for 10 min. After incubation with normal horse serum (Vector Labs, Burlinghame, CA), the slides were incubated with Ki-67 antibody (1 : 100 dilution) for 1 h at 37 °C. Diaminobenzidine was used as final chromogen and haematoxylin as the nuclear counterstain. An archived case of a highly proliferating non-Hodgkin-lymphoma (type Burkitt-lymphoma) prepared as described above was used as the positive control. Slides treated the same way but omitting the specific antibody served as negative controls. Four representative regions to the tumour were selected (by D.D.) and ≥ 1000 cells scored on each slide at high magnification (×400) under light microscopy (by B.N. and D.D.). The mean Ki-67 labelling index (LI) was defined as the percentage of total tumour cell nuclei that were Ki-67-positive. The results were compared with pT stage, nodal status, grade and clinical disease progression at follow-up.

RESULTS

  1. Top of page
  2. Abstract
  3. INTRODUCTION
  4. PATIENTS AND METHODS
  5. RESULTS
  6. DISCUSSION
  7. CONFLICT OF INTEREST
  8. REFERENCES

The pathological stage was pT1 in 24 patients and pT2 in 20; 10 had lymph-node metastases. During a mean follow-up of 35.6 months, six patients had disease progression and two were lost to follow-up. The distribution of histological grade is shown in Table 1. There was positive Ki-67 immunostaining in all cases, with a mean (range) LI of 40.5 (6.4–93)%. The proliferation rate was categorized as low (<40.5%) or high (≥40.5%), from the mean Ki-67 LI. A high LI was significantly associated with poorly differentiated tumours (P < 0.005, Fig. 1). The mean LI was higher in patients with regional lymphatic spread (51.4%) than in those with localized disease confined to the penis (37.6%), but this difference was not statistically significant (P = 0.07). There was also a tendency for high Ki-67 expression with advanced local tumour stage and disease progression, but these correlations were not statistically significant either (P = 0.07 and 0.06).

Table 1.  The relationship between mean Ki-67 LI and clinicopathological variables
VariableN tumoursKi-67 LI, %
Grade
G11220
G22846
G3 466
T stage
T12436.2
T22045.3
Lymph nodes
N03438.6
N11051.4
No progression3839.1
Disease progression 648.7
image

Figure 1. (A) Diffuse nuclear immunohistochemical staining for Ki-67 in most tumour cells of a high-grade SCC of the penis (original × 100). (B) Weakly positive immunostaining for Ki-67 in a well-differentiated penile carcinoma (original × 100).

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DISCUSSION

  1. Top of page
  2. Abstract
  3. INTRODUCTION
  4. PATIENTS AND METHODS
  5. RESULTS
  6. DISCUSSION
  7. CONFLICT OF INTEREST
  8. REFERENCES

Ki-67 is a non-histone nuclear matrix protein expressed in all cell-cycle phases except G0. An assessment of Ki-67 protein expression by immunohistochemistry is a reliable means of evaluating tumour cell proliferation [11]. To our knowledge, there are no previous reports on the correlation between the Ki-67 LI and clinicopathological variables for SCC of the penis. Studies show that for SCC of the head and neck there is a correlation between the proliferation rate determined by Ki-67 and tumour de-differentiation, nodal involvement or disease progression [12–14]. In the present study there was a significant association only between the Ki-67 LI and tumour grade, but there was a tendency towards a higher LI with tumour stage, nodal metastasis and clinical disease progression at follow-up. The lack of correlation for these factors might be because there were too few patients.

In a recent report, Martins et al.[15] examined the proliferative activity in penile carcinoma using immunostaining for proliferating cell nuclear antigen (PCNA); the PCNA LI had a significant correlation in univariate analysis with the presence of nodal metastasis. In contrast to the present findings for Ki-67 LI, there was no correlation between the PCNA LI and tumour grade.

Emerson et al.[8] found that the depth of stromal tumour invasion, measured by a computerized micrometer, and vascular invasion were predictive for cancer progression in patients with penile SCC. In two other studies the immunohistochemical overexpression of tumour-suppressor gene product p53 in penile carcinoma was correlated with disease progression [15,16].

Further investigations of molecular markers to predict tumour behaviour in penile carcinoma, and cohorts with more cases, are needed for this relatively rare tumour. Besides known prognostic factors like pT stage and grade, they might be helpful in selecting patients who would benefit from inguinal lymphadenectomy.

REFERENCES

  1. Top of page
  2. Abstract
  3. INTRODUCTION
  4. PATIENTS AND METHODS
  5. RESULTS
  6. DISCUSSION
  7. CONFLICT OF INTEREST
  8. REFERENCES
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