Percutaneous irreversible electroporation of hepatocellular carcinoma: Contrast-enhanced ultrasound-findings during 1-year follow-up

Abstract

PURPOSE:

To assess the postprocedure findings after percutaneous irreversible electroporation (IRE) of hepatocellular carcinoma (HCC) in contrast-enhanced ultrasound (CEUS).

MATERIALS AND METHODS:

Percutaneous IRE was performed in a total of 22 patients with 24 HCC tumours following interdisciplinary tumour board review. The lesions were documented using CEUS before, immediately and within 24 hours after IRE. During follow-up CEUS was performed at 6 weeks and 3, 9, and 12 months after ablation. Two experienced radiologists evaluated the acquired CEUS image date in a consensus reading.

RESULTS:

Median tumour size before treatment was 13.7±4.8 mm (short axis) and 16.0±5.2 mm (long axis). All HCC lesions showed arterial hyperenhancement in CEUS. Median size of the ablation defect after ablation was 29.3±5.2 mm (short axis) and 31.6±5.6 mm (long axis). After IRE all tumours showed complete devascularization. The size of the ablation defects showed significant shrinkage and reduced peripheral enhancement in the course of follow-up. At 12 months follow-up the ablation defect size decreased to 16.7±4.3 mm (short axis) and 18.3±4.1 mm (long axis).

CONCLUSION:

CEUS showed a complete devascularization of HCC tumours after IRE. Post-intervetional peripheral enhancement returned to normal during follow-up and may represent zones of reversible damage of cellular integrity through electroporation. A significant shrinkage of the ablation defects during 12 month of follow-up was seen in all cases.

1 Introduction

There are a number of ablative technologies available for the percutaneous ablation of hepatic tumours. Most of these technologies rely on the transfer of thermal energy into the target area to cause cell death [1]. Unfortunately, there are several limitations of thermal ablation technology due to the unintended thermal damage of adjacent structures or organs [2]. In addition, complete tumour destruction in the target area can be hindered by the heat sink effect, whereby the target temperature is not or not long enough reached within the ablation zones adjacent to flowing blood (“heat sink effect”) [3]. At least in theory, irreversible electroporation (IRE) may have the potential to overcome aforementioned limitations of thermal ablation as instead of heat a number of nanosecond-long electric impulses are used to induce a breakdown of the transmembrane potential [4]. Via creation of nanopores in the cell membrane with influx and efflux of ions through the lipid bilayer cellular apoptosis instead of necrosis is induced [5].

The high diagnostic potential of contrast-enhanced ultrasound (CEUS) in relation to the detection and characterization of malignant hepatic lesions has already been demonstrated in several clinical and experimental studies [6, 7]. However, data about images of morphological findings of CEUS after IRE of HCC is scarce. Therefor, the aim of this study was to describe imaging findings of CEUS during 12-month follow-up after percutaneous ablation of HCC using irreversible electroporation.

2 Materials and methods

2.1 Study design

This retrospective study was conducted at a single institution and IRB approval was waived due to the retrospective nature of the study. A total of 22 patients (mean age 63.4±5.1 years) with 24 hepatocellular carcinomas that were treated with percutaneous IRE were included into the analysis (see Table 1). A diagnosis of HCC was made in accordance with the guidelines of the American Association fort the Study of Liver Diseases [8] based on CEUS findings in combination with CT, MRI or pathology as demanded by the guidelines and as elaborated by discussion of every case in the interdisciplinary tumour board. The indication for IRE was provided in the scope of an interdisciplinary tumour board review. Written informed consent was obtained from all patients. The study conforms the requirements set out by this journal.

Table 1
Demographic data and baseline characteristics of study population

Variable Standard deviation Minimum Maximum

Patients (n) 22

Sex (male/ female) 20/2

Age (years) 63.4 5.1 55 72

Lesion size

Short axis 13.7 4.8 7 24

Long axis 16.0 5.2 10 27

Liver function

Child A 6 (27.3%)

Child B 13 (59.1%)

Child C 3 (13.6%)

Variable	Standard deviation	Minimum	Maximum
Patients (n)	22
Sex (male/ female)	20/2
Age (years)	63.4	5.1	55	72
Lesion size
Short axis	13.7	4.8	7	24
Long axis	16.0	5.2	10	27
Liver function
Child A	6 (27.3%)
Child B	13 (59.1%)
Child C	3 (13.6%)

2.2 Technical aspects of IRE ablation

In all cases, the NanoKnife System (Angiodynamics, Latham; NY) was used. All patients received general anesthesia with complete muscle relaxation. From two to six 19– gauge unipolar electrodes were used for percutaneous tumour ablation. The electrodes are placed in parallel direction via CT fluoroscopy and ultrasound guidance so that the tips of the electrodes encompass the tumour with a distance of 15–20 mm between each probe. The electrodes have an active tip of 5–20 mm and can be adjusted manually depending on the intended size of the ablation zone. The voltage needed for safe tumour ablation is determined with the aid of a standard algorithm provided by the IRE generator and depends on the intended size of the ablation zone, the number of electrodes and their distance. The generator can produce a voltage of up to 3,000 volts and a current of up to 50 amperes. After a 270 V test impulse to ensure adequate conductivity of the tissue, 90 ECG-triggered therapeutic electrical impulses are administered by the IRE generator.

2.3 Contrast-enhanced ultrasound (CEUS)

All the lesions were evaluated by CEUS immediately before ablation. CEUS was also used to monitor the ablation process and success. In a preceding study, our working group evaluated already CEUS findings immediately after IRE (Fig. 1). For this analysis, we re-evaluated CEUS findings one day after ablation, and at 3, 6, 9, and 12 months after IRE ablation (Fig. 2–6). To do this, DICOM loops were acquired with a multi-frequency probe (1-5 MHz, Logiq E9, GE Healthcare, Chalfont St. Giles, UK) following the intravenous bolus injections of a maximum of 2.4 ml sulphur hexafluoride microbubbles (SonoVue, Bracco, Italy) followed by a flush with 10 ml of 0.9% sodium chloride solution. Saving of digital image sequences took place during arterial (15–45 seconds), portal venous (60–90 s) and late venous phase (90–120 seconds) and additionally in the late phase (3–5 minutes). Colour-coded duplex sonography (CCDS) was used to rule out vascular complications (i.e. portal vein thrombosis or major vein thrombosis of adjacent or crossing veins).

Fig.1

Pre-interventional contrast-enhanced ultrasound during arterial phase: arterial hyperenhancement of a 16×18 mm measuring hepatocellular carcinoma in segment V.

Fig.2

Same patient as Fig. 1: Postinterventional contrast-enhanced ultrasound at 24 hours after irreversible electroporation showed inhomogeneous ablation defect with perifocal hyperemia.

Fig.3

Same patient as of Fig. 1 and 2: Devascularized ablation zone with beginning shrinkage at 3 months. Perifocal hyperemia is not visible anymore.

Fig.4

6 months follow-up of the same patient as Fig. 1–3: complete devascularization of the ablation zone with continued shrinkage.

Fig.5

9 months follow-up of the same patient as Fig. 1–4: Continued shrinkage of the ablation defect to 23×32 mm.

Fig.6

Same patient as Fig. 1–5. 12 months follow-up. The ablation defect after percutaneous IRE is only residually visible.

2.4 Statistical analysis

The statistical analysis was performed using SPSS V21 for Mac. The analysis of the lesions and ablation area axes was performed with the aid of a paired T-Test. A significance level of p < 0.05 was considered statistically significant.

3 Results

A total of 24 HCC tumours in 22 patients were treated with a mean tumour diameter of 13.7±4.8 mm (short axis) and 16.0±5.2 mm (long axis). The results of the ablation defect measurements are shown in Table 2. The ablation zone dimensions appeared to significantly decrease over time (p < 0.01). Figures 7 and 8 show the course of tumours’ long and short axis during follow-up. On post-interventional CEUS imaging a lack of enhancement is seen in the ablation zone surrounded by an enhancing rim surrounding the ablation defect during arterial and portal venous phase. As known from other imaging modalities the ablation zone is well demarcated from the surrounding hepatic parenchyma.

Table 2
Follow-up results up to 12 months after IRE for ablation defect size, peripheral enhancement in CEUS and local recurrence

24 Hours 3 Months 6 Months 9 Months 12 Months

Short axis 29.3±5.2 24.3±5.0 21.7±5.2 18.7±4.6 16.7±4.3

Long axis 31.6±5.6 28.1±5.1 24.3±4.9 20.9±4.4 18.3±4.1

Peripheral 24 of 24 4 of 24 0 of 23 0 of 23 0 of 22

Enhancement 100% 16.7% 0% 0% 0%

Number of local recurrence (total) ^* 1 of 24 1 of 24 2 of 24 3 of 24

4.2% 4.2% 8.3% 12.5%

	24 Hours	3 Months	6 Months	9 Months	12 Months
Short axis	29.3±5.2	24.3±5.0	21.7±5.2	18.7±4.6	16.7±4.3
Long axis	31.6±5.6	28.1±5.1	24.3±4.9	20.9±4.4	18.3±4.1
Peripheral	24 of 24	4 of 24	0 of 23	0 of 23	0 of 22
Enhancement	100%	16.7%	0%	0%	0%
Number of local recurrence (total)	^*	1 of 24	1 of 24	2 of 24	3 of 24
		4.2%	4.2%	8.3%	12.5%

^*Complete ablation was achieved in 100% of treated lesions.

Fig.7

Short axis of the ablation defects after percutaneous irreversible electroporation showed significant shrinkage in the course of follow-up at 24 hours and at 3, 6, 9, and 12 months.

Fig.8

Long axis of the ablation defects after percutaneous irreversible electroporation showed significant shrinkage in the course of follow-up at 24 hours and at 3, 6, 9, and 12 months.

All ablation defects showed peripheral arterial and portalvenous hyperenhancement at 24 hours after ablation and was still visible in 4 patients at 3 months follow-up. Rim enhancement was absent in all patients and returned to normal in all patients at 6 months follow up. Treatment efficacy was not an endpoint of this retrospective analysis. However, complete ablation was achieved in 100% of tumours. There was one case of local recurrence at 3-months, one case at 9 months and one case at 12 months (total recurrence rate after 12 months was 12.5%). In addition, treatment safety was also not an endpoint of this study, but there was one case of post-interventional portal vein thrombosis as documented already in CCDS at 24 hours follow-up.

4 Discussion

Irreversible electroporation is based on the principle of electropermeabilization, in which electric pulses are used to create nanoscale defects within the phospholipide bilayer of the cell membrane. These defects are called “nanopores” and permeate the cell membrane. Under a certain electrical threshold the formation of the nanopores during electroporation is transient i.e. reversible. Above this threshold the formation remains permanent i.e. irreversible and results in cell death due to the inability to maintain homeostasis [9]. While the ablation mechanism of IRE is still not completely understood, several reports regarding safety and efficacy in the liver have been published [10 –14]. Cross sectional imaging is regularly used to assess tumour response after locally ablative tumour therapies. However, especially in patients with impaired renal function, there are relevant limitations for those imaging modalities to answer the question of local tumour response. Especially but not solely in those patients the contrast enhanced ultrasound is the most meaningful imaging modality to asses local tumour control. Therefor, the understanding of imaging appearance of HCC tumours after treatment with IRE is crucial to determine the completeness of ablation. Recently, Padia et al. reported imaging findings at MR imaging after IRE [15]. However, to the best of our knowledge, there is no literature regarding imaging findings after IRE using CEUS for follow up. Our working group already published immediate findings of CEUS imaging after IRE [16]. The present study now deals with delayed CEUS findings after IRE. Immediate findings of CEUS after IRE showed rapid and significant reduction of microvascularization in the ablation zone while major blood vessels were not affected by IRE [16]. Also during follow-up CEUS showed persistent and complete devascularization of the ablation zones. Also during follow-up, major vessels have not been affected by IRE except for one case with portal vein thrombosis, which was documented already during postinterventional imaging at 24 hours after intervention. Interestingly and in accordance with the findings of Padia et al. who retrospectively assessed IRE ablation defects using MR imaging, we found significant shrinkage of the ablation defects during follow-up [15]. Also, the type of ablation technique used can not be differentiated after one year. IRE, RFA and MWA all leave a significantly shrinked scar after successful ablation. During follow-up CEUS imaging of ablation defects are sharply demarcated from to the surrounding hepatic parenchyma. Histopathological analysis of ablation zones after IRE in patients with HCC who subsequently underwent liver transplantation also showed complete necrosis sharply demarcated from surrounding hepatic parenchyma [17]. During ablation procedures and during follow-up, CEUS adds valuable information regarding the differentiation between liver metastases and HCC as a stand-alone diagnostic procedure e.g. on the basis of parametric analysis or as fusion imaging with CT or MRI [18, 19].

During immediate post interventional imaging we found a hyperenhancing rim around ablation defects during the arterial phase, which might represent a perifocal zone of reversible electroporation. The identification of areas with reversible electroporation might be useful for optimizing therapy regimes, e.g. when using certain chemotherapies that require enhanced localized drug delivery as they cannot cross cellular plasma membranes. In situations of reversible electroporation the cellular uptake of drugs may be increased [20]. This rim of perifocal hyperenhancement was seen at immediate follow-up imaging at 24 hours after ablation and returned to normal during follow-up and is not visible anymore at 3 months (20 of 24 patients) or 6 months (24 of 24) of follow-up imaging. In this context, it is crucial to understand that the perifocal rim of hyperenhancement does not represent an area of tumour ablation and must not be added to the safety margin of 1 cm after tumour ablation.

However, the results of our study have to be regarded with prudence, as all the results are subject to all the biases inherent to retrospective analyses. This analysis was performed in a very small group of patients. The small group of patients is a result of the fact, that the majority of patients can be safely treated with thermal ablation techniques. Also, all our results are not based on histologic analyses and we do not know the degree to which liver cirrhosis did alter the appearance of the ablation zones after IRE.

In summary, the results of our study showed that the initial rim enhancement of IRE ablation zone returned to normal during follow-up. These zones might represent zones of reversible cellular damage due to electroporation and must not lead to overestimation of ablation zones. IRE causes a complete devascularization within its ablation defects. We observed significant shrinkage of the ablation zones after IRE, which might be result of scar formation.

References

Lencioni

, Crocetti

. Local-regional treatment of hepatocellular carcinoma. Radiology. 2012;262(1):43–58.

Kim

, Rhim

, Park

, Kim

, Choi

, et al. Percutaneous radiofrequency ablation of hepatocellular carcinomas adjacent to the gallbladder with internally cooled electrodes: assessment of safety and therapeutic efficacy. Korean J Radiol. 2009;10(4):366–76.

DSK

, Raman

, Limanond

, Aziz

, Economou

, Busuttil

, et al. Influence of large peritumoral vessels on outcome of radiofrequency ablation of liver tumors. J Vasc Interv Radiol JVIR. 2003;14(10):1267–74.

Lee

, Wong

, Prikhodko

, Perez

, Tran

, Loh

, et al. Electron microscopic demonstration and evaluation of irreversible electroporation-induced nanopores on hepatocyte membranes. J Vasc Interv Radiol JVIR. 2012;23(1):107–13.

Kim

H-B

, Sung

C-K

, Baik

, Moon

K-W

, Kim

H-S

, Yi

J-H

, et al. Changes of apoptosis in tumor tissues with time after irreversible electroporation. Biochem Biophys Res Commun. 2013;435(4):651–6.

Schellhaas

, Waldner

, Görtz

, Vitali

, Kielisch

, Pfeifer

, et al. Diagnostic accuracy and interobserver variability of Dynamic Vascular Pattern (DVP) in primary liver malignancies - A simple semiquantitative tool for the analysis of contrast enhancement patterns. Clin Hemorheol Microcirc. 2017;66(4):317–31.

Wildner

, Schellhaas

, Strack

, Goertz

, Pfeifer

, Fießler

, et al. Differentiation of malignant liver tumors by software-based perfusion quantification with dynamic contrast-enhanced ultrasound (DCEUS). Clin Hemorheol Microcirc. 2018. 10.3233/CH-180378.

Marrero

, Kulik

, Sirlin

, Zhu

, Finn

, Abecassis

, et al. Diagnosis, staging, and management of hepatocellular carcinoma: 2018 practice guidance by the american association for the study of liver diseases. Hepatol Baltim Md. 2018;68(2):723–50.

Chang

, Reese

. Changes in membrane structure induced by electroporation as revealed by rapid-freezing electron microscopy. Biophys J. 1990;58(1):1–12.

10.

Cannon

, Ellis

, Hayes

, Narayanan

, Martin

RCG

. Safety and early efficacy of irreversible electroporation for hepatic tumors in proximity to vital structures. J Surg Oncol. 2013;107(5):544–9.

11.

Kingham

, Karkar

, D’Angelica

, Allen

, Dematteo

, Getrajdman

, et al. Ablation of perivascular hepatic malignant tumors with irreversible electroporation. J Am Coll Surg. 2012;215(3):379–87.

12.

Martin

RCG

, Philips

, Ellis

, Hayes

, Bagla

. Irreversible electroporation of unresectable soft tissue tumors with vascular invasion: effective palliation. BMC Cancer. 2014;14:540.

13.

Philips

, Hays

, Martin

RCG

. Irreversible electroporation ablation (IRE) of unresectable soft tissue tumors: Learning curve evaluation in the first 150 patients treated. PloS One. 2013;8(11):e76260.

14.

Thomson

, Cheung

, Ellis

, Federman

, Kavnoudias

, Loader-Oliver

, et al. Investigation of the safety of irreversible electroporation in humans. J Vasc Interv Radiol JVIR. 2011;22(5):611–21.

15.

Padia

, Johnson

, Yeung

, Park

, Hippe

, Kogut

. Irreversible electroporation in patients with hepatocellular carcinoma: Immediate versus delayed findings at MR imaging. Radiology. 2016;278(1):285–94.

16.

Wiggermann

, Zeman

, Niessen

, Agha

, Trabold

, Stroszczynski

, et al. Percutaneous irreversible electroporation (IRE) of hepatic malignant tumours: contrast-enhanced ultrasound (CEUS) findings. Clin Hemorheol Microcirc. 2012;52(2–4):417–27.

17.

Cheng

, Bhattacharya

, Yeh

, Padia

. Irreversible electroporation can effectively ablate hepatocellular carcinoma to complete pathologic necrosis. J Vasc Interv Radiol JVIR. 2015;26(8):1184–8.

18.

Yue

W-W

, Wang

, Xu

H-X

, Sun

L-P

, Guo

L-H

, Bo

X-W

, et al. Parametric imaging with contrast-enhanced ultrasound for differentiating hepatocellular carcinoma from metastatic liver cancer. Clin Hemorheol Microcirc. 2016;64(2):177–88.

19.

X-W

, Xu

H-X

, Wang

, Guo

L-H

, Sun

L-P

, Li

X-L

, et al. Fusion imaging of contrast-enhanced ultrasound and contrast-enhanced CT or MRI before radiofrequency ablation for liver cancers. Br J Radiol. 2016;89(1067):20160379.

20.

Mir

, Orlowski

, Belehradek

, Paoletti

. Electrochemotherapy potentiation of antitumour effect of bleomycin by local electric pulses. Eur J Cancer Oxf Engl 1990. 1991;27(1):68–72.