Estrogen and Progesterone Receptor Status in Leiomyoma and Radiofrequency Ablation Outcome

Abstract

Objective: Radiofrequency ablation (RFA) is a noninvasive therapeutic technique for treating symptomatic uterine leiomyomas. This study aimed to prospectively evaluate the relationship of estrogen receptor (ER) and progesterone receptor (PR) in leiomyoma with the volume reduction rate and improvement in symptoms following RFA. Design and Methods: Forty-nine women with symptomatic single-nodule uterine leiomyoma underwent transvaginal RFA. Fine-needle aspiration biopsy was performed 1 month before RFA, and the patients were divided into two groups based on the status of ER and PR in leiomyoma: strongly to moderately positive (group I, 32 patients) and weakly positive to negative (group II, 17 patients). The leiomyoma volume was measured by three-dimensional ultrasonography before and during the operation and in postoperative months 1, 3, 6, 9, 12, and 18. The impact of the symptoms on health-related quality of life (HRQL) was assessed on first and last follow-up visits by using the Uterine Fibroids Symptom and Quality of Life (UFS-QOL) questionnaire. Results: The preoperative diameter (6.7±1.7 vs. 6.3±1.5 cm) and leiomyoma volume (83.8±23.0 vs. 82.9±34.8 cm³) did not significantly differ between groups I and II. The final volume reduction rate in groups I and II was 48.2%±15.7% and 86.8%±7.6%, respectively. Compared to group I, group II showed significantly greater volume reduction rate, improvement in the symptom severity, and HRQL scores (p<0.01). Conclusions: The initial ER and PR immunohistochemical status is important to the outcome of leiomyoma RFA, and influences its effectiveness. (J GYNECOL SURG 27: 133)

Introduction

Uterine leiomyomas are the most common benign tumors in women of reproductive age, and affect 20%–50% of this population. The prevalence of these tumors increases with age in the reproductive period.^1,2 Leiomyoma is a hormonally responsive tumor, which contains estrogen receptor (ER) and progesterone receptor (PR), and its growth rate is dependent upon the ovarian steroid hormones.^3,4

The ER and PR status influence the rate of leiomyoma shrinkage after treatment with a gonadotropin-releasing hormone (GnRH) agonist. The status of ER and PR expression decreased by 85%, 49%, and 36% at 6 months after GnRH agonist treatment in leiomyoma patients,⁵ and the reduction rate of B_max (maximum binding density of receptor) for PRs in the myometrium correlated with the volume reduction rate of leiomyomas, which was measured using magnetic resonance imaging (MRI).⁶

A previous study suggested that GnRH agonist therapy before magnetic resonance-guided focused ultrasound improves thermoablative treatment effects.⁷ However, the relationship between the status of the ovarian hormone receptors ER and PR and thermoablative treatment was not evaluated. It was hypothesized that the initial status of ER and PR in leiomyoma could affect the treatment outcomes of thermoablative therapy. The aim of this study was to prospectively evaluate the relationship of ER and PR in leiomyoma along with the rates of volume reduction and the improvement in the symptoms after radiofrequency ablation (RFA).

Materials and Methods

Premenopausal women with symptomatic single-nodule uterine myoma were included in this study. From December 2004 to September 2006, 49 patients were enrolled. All patients said they had did not have plans for any more childbearing and declined hysterectomy or myomectomy. The exclusion criteria were as follows: previous treatment with GnRH agonists, having a myoma of >8 cm in diameter, presence of any abnormalities in cancer screening tests, abnormal coagulation test results, current pregnancy, recent pelvic/endometrial inflammatory disease, and a positive chlamydia/gonorrhea PCR test.

To confirm the pathology and steroid hormone receptor status, needle biopsy was performed at three central areas of the myoma under ultrasonographic guidance during the luteal phase 1 month before myolysis. Based on the status of the hormone receptors, the patients were divided into the following two groups: strong to moderate positive status of ER and PR (group I, 32 patients) and weakly positive to negative status of ER and PR (group II, 17 patients).

All patients were extensively counseled on the potential risks and benefits of the procedure and on the possible alternative surgical treatments. The protocol for the study was inspected and approved by the Ethics and Research Committee of the Catholic University of Korea.

The pre- and postoperative myoma volumes were measured by three-dimensional ultrasonography (Accuvix XQ, Medicine). Sonographic evaluation was repeated at postoperative months 1, 3, 6, 9, 12, and 18. In order to minimize interpersonal variation, the ultrasonograms were assessed by a single trained gynecologist.

The impact of the symptoms on health-related quality of life (HRQL) was assessed at the first and last follow-up visits by using the Uterine Fibroids Symptom and Quality of Life (UFS-QOL) questionnaire.⁸ This questionnaire contains 37 questions regarding the symptoms of uterine fibroids and their impact.

Equipment

The RF delivery system (RF Medical System, M-1004) consisted of an RF generator that operates at 400 kHz with a maximum power of 120 W and at temperatures ranging from 40°C to 99°C. The generator displays the temperature of the needle tip, tissue impedance characteristics, and procedure time. The system is connected via a needle to a 25-cm-long 18-gauge needle with an exposed tip at the distal end.

Procedure

RFA of uterine myomas was performed in lithotomy position with the patient under intravenous anesthesia with profopol sodium, single bolus (2 mg/kg) and continuous injection (5 mg/kg/h). The RF needle was transvaginally inserted through the cervical canal. Under ultrasonographic guidance, the tip of the needle was centralized in the leiomyoma. Ultrasonography was performed transabdominally or transrectally. Following centralization of the needle, ablation was performed. The temperature of the needle tip was maintained at 85°C; the generator automatically adjusted the power to maintain the selected temperature. During ablation, the myoma was monitored by ultrasonography, and the duration of ablation was determined based on the increased echogenicity of the myoma. The progression of ablation was considered successful if a postechogenic shadow was observed. The myoma exhibited high echogenicity during ablation, and ablation was stopped when the area of high echogenicity reached up to 90% of the cut area of the leiomyoma. The complete ablation of a 3-cm myoma usually takes 5 minutes, whereas that of an ∼5-cm myoma takes almost 10 minutes. After the operation, the patients were monitored for 2 hours. If the patients did not demonstrate any abnormal findings (unstable vital signs, increased temperature, severe abdominal pain, or profuse vaginal bleeding) within 2 hours postoperation, they were discharged and permitted to resume daily activities.

Pathology and immunohistochemistry

The specimens from all patients were examined by a single histologist who was blinded to the study. Immunohistochemical staining for estrogen and progesterone was performed on formalin-fixed paraffin-embedded sections by using the avidin-biotin-immunoperoxidase complex method. Three samples of each leiomyoma were used for the immunohistochemical staining procedure. Mouse monoclonal antibodies to human ER (ER1D5, 1:50 dilution; Dako Corporation, Carpinteria, CA) and PR (PGR-1A6, 1:40 dilution; BioGenex, San Ramon, CA) were used as the primary antibodies. In brief, the sections were deparaffinized in xylene, rehydrated through graded alcohols, and treated with 0.01 M citrate buffer for 15 minutes (pH 6) in a microwave oven. Subsequently, the samples were incubated with normal goat serum to reduce nonspecific binding. Biotinylated goat antimouse immunoglobulin G was used as a linker. After washing, the streptavidin-biotin complex was applied and stained with diaminobenzidine. The sections were slightly counterstained with hematoxylin. Specific staining was identified by brown coloration of the nucleus. The sections were evaluated for ER and PR expression based on the percentage of stained cells: (−) negative (<10% cells stained), (+) weakly positive (11%–50% cells stained), (++) moderately positive (51%–80% the cells stained), and (+++) strongly positive (>80% cells stained).

Statistical analysis

The leiomyoma volume reduction rate was calculated as follows: (initial volume−final volume) ×100/initial volume. Statistical differences were determined using Student's t test and the ANOVA multiple comparison test. Statistical significance was set at p<0.05.

Results

The mean age of the patients was 37.8 years (SD, 4.1 years), and the mean parity was 1.5 (SD, 0.65) (Table 1). The chief preoperative complaints included menorrhagia (41.8%), dysmenorrhea (7.0%), pelvic pain (4.0%), and mixed symptoms (47.2%). The mean initial size of the dominant myoma was 6.7 cm (SD, 1.77). The operative time ranged from 20 to 40 minutes. No intraoperative complications occurred during RFA. In most patients, the staining status was identical for ER and PR. There was only a single exception to this in group I. The specimen from this patient exhibited strongly positive ER staining but moderately positive PR staining. The specimens of 3 patients from group II exhibited trace ER staining and weakly positive PR staining. However, there was no change in the grouping.

Table 1.

Patient Characteristics

Parameters	Mean (range)
Age (years)	38.7 (27–47)
Parity (number of full-term births)	1.5 (1–3)
Initial myoma diameter (cm)	6.7 (3–7.4)
Volume (cm³)	86.2 (7.9–119.2)

Initial symptoms (%)
Menorrhagia	41.8%	Pelvic pain	4%
Dysmenorrhea	7%	Mixed symptoms	47.2%

Hormone receptor status (%)
ER	Group I	Strong	23 (46.9%)	PR	Group I	Strong	22 (44.8%)
		Moderate	9 (18.3%)			Moderate	10 (20.4%)
	Group II	Weak	5 (10.2%)		Group II	Weak	8 (16.3%)
		Trace	12 (24.4%)			Trace	9 (18.0%)

ER, estrogen receptor. PR, progesterone receptor.

The preoperative diameter (6.7±1.7 vs. 6.3±1.5 cm) and volume of leiomyoma (83.8±23.0 vs. 82.9±34.8 cm³) did not significantly differ between groups I and II (Table 2). However, in group II, the diameter reduced significantly at 18 months after RF myolysis (p<0.01) and the final volume reduction rate was 48.2%±15.7% in group I and 86.8%±7.6% in group II. Group II showed a significantly greater reduction rate than group I (p<0.01).

Table 2.

Volume Reduction Rate of Leiomyoma According to the Initial Status of ER and PR

	Myoma (mean)
	Diameter (cm)		Volume (cm ³ )
Group	Premyolysis	Postmyolysis	Premyolysis	Postmyolysis	Reduction rate (mean, %)
I	6.7±1.7	5.2±2.2	83.8±23.0	44.4±19.0	46.4±15.7
II	6.3±1.5	3.1±1.8	82.9±34.8	11.3±6.7	86.3±7.6
P-value	NS	<0.01	NS	<0.01	<0.01

ER, estrogen receptor; PR, progesterone receptor; NS, not significant.

Symptom severity and HRQL scores showed greater improvement in group II at 18 months postoperation (Table 3).

Table 3.

Symptom Severity and HRQL Scores (Mean±SD) at Baseline and at 18 Months after Radiofrequency Myolysis

	Symptom severity score		HRQL score
Group	Baseline	18 months	Baseline	18 months
I	52.7±10.1	48.1±7.7	66.7±20.2	71.3±4.2
II	49.8±7.7	21.2±4.2^*	67.7±12.9	99.4±4.2^*

HRQOL, health-related quality of life; SD, standard deviation.

p<0.05 vs. the previous assessment.

Discussion

RFA is one of the thermal ablation therapies for symptomatic leiomyoma. Thermal ablation was first introduced in the late 1980s as a hysteroscopic technique.⁹ RF energy is an alternating current with a frequency between 10 and 900 kHZ,¹⁰ and its predominant mechanism of action is the thermal damage caused by frictional heating.¹¹ RF has been used as a minimal invasive ablation therapy for renal tumor,¹² osteoid osteoma,¹³ and focal hepatic metastasis.¹⁴ Once the cell temperature exceeds 50°C, the cell membranes melt and fuse and the proteins denature, and irreversible cell death occurs.¹⁵ The leiomyoma volume reduction rates were reported to be 41.5%, 59%, and 77% at 1, 3, and 6 months after RF myolysis, respectively,¹⁰ and Fabio et al. reported median reduction rates of 68.8% and 77.9% at 6 months and 1 year after myolysis, respectively.¹⁶ Following myolysis, the leiomyoma tissue exhibited coagulative necrosis, and interstitial vessels were obliterated. The thermal effect of RF ablation was not only coagulative necrosis but also decreased ER and PR expression in the target tissue.¹⁷

Estrogen and progesterone are believed to stimulate leiomyoma growth because the incidence of leiomyoma increases after menarche and because these tumors enlarge during pregnancy and regress after menopause.^18,19 Leiomyoma has two types of ERs—ERα and ERβ—and the ERα/ERβ ratio was found to be lower in leiomyoma than in the normal uterine myometrium.²⁰ PR expression in leiomyomata is consistent throughout the menstrual cycle, whereas ER expression varies according to the menstrual phase. The latter expression is usually higher in the proliferative phase than in the secretory phase.²¹

Some investigators reported the significance of PR in the growth rate of leiomyomas. Ichimura et al. reported that the levels of PR and not ER significantly correlate with the growth of uterine leiomyoma and suggested that progesterone influences the proliferation of uterine smooth muscle cells to a substantially greater extent than does estrogen.²² The mitotic index or labeling index of Ki-67 and the proliferation indicators of uterine smooth muscle cells are higher in the secretory phase than in the proliferative phase.²¹ Moreover, Matsuo et al. found that the apoptosis-inhibiting gene product bcl-2 was upregulated by progesterone in leiomyoma cells.²³ Collectively, the results of these studies suggest that progesterone (and possibly the upregulation of PRs by estrogen) plays a significant role in cellular proliferation in leiomyomas. Because of the limitation of the current study, i.e., the small number of patients, the effect of PR alone was not evaluated.

In the current study, it was found that the initial ER and PR status of leiomyoma was very important for the outcome of myoma treatment with RF thermoablation, not only in terms of the reduction in the rate of leiomyoma volume, but also in terms of improvement of symptoms in patients. The symptom severity and HRQL scores showed greater improvement in group II wherein weakly positive to trace ER and PR expression was observed in the leiomyoma. It is believed that the symptoms in group II patients showed greater improvement than did those in group I patients because the leiomyoma shrinkage rate was higher in group II than in group I. These results suggest that the initial ER and PR status was critical to the outcome of heat therapy in leiomyoma.

Smart et al. reported that the pretreatment of leiomyomata with GnRH agonists permits the extension of magnetic resonance-guided focused ultrasound to patients with larger tumors,⁶ and GnRH agonists potentiate the thermal effects of magnetic resonance image-guided focused ultrasound therapy.²⁴ With regard to hormone receptors, the cellular proliferation index and ER and PR expression significantly decreased 6 months after GnRH agonist treatment;⁵ moreover, dilated vessels and hyalinization in leiomyoma were observed at a higher frequency after GnRH agonist treatment.

Conclusions

In conclusion, the initial immunohistochemical status of ER and PR in leiomyoma was critical for determining the effectiveness of heat therapy. Further studies are required on the correlation of GnRH pretreatment and changes in PR and ER expression with the outcomes of RFA. It is hoped that this study will serve as a proper guideline for uterine preservation treatment in leiomyoma patients.

Footnotes

Disclosure Statement

No competing financial conflicts exist.

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