Pulsed Electric Fields in Oncology: A Snapshot of Current Clinical Practices and Research Directions from the 4th World Congress of Electroporation.

The 4th World Congress of Electroporation (Copenhagen, 9-13 October 2022) provided a unique opportunity to convene leading experts in pulsed electric fields (PEF). PEF-based therapies harness electric fields to produce therapeutically useful effects on cancers and represent a valuable option for a variety of patients. As such, irreversible electroporation (IRE), gene electrotransfer (GET), electrochemotherapy (ECT), calcium electroporation (Ca-EP), and tumour-treating fields (TTF) are on the rise. Still, their full therapeutic potential remains underappreciated, and the field faces fragmentation, as shown by parallel maturation and differences in the stages of development and regulatory approval worldwide. This narrative review provides a glimpse of PEF-based techniques, including key mechanisms, clinical indications, and advances in therapy; finally, it offers insights into current research directions. By highlighting a common ground, the authors aim to break silos, strengthen cross-functional collaboration, and pave the way to novel possibilities for intervention. Intriguingly, beyond their peculiar mechanism of action, PEF-based therapies share technical interconnections and multifaceted biological effects (e.g., vascular, immunological) worth exploiting in combinatorial strategies.

Bleomycin electrosclerotherapy (BEST) for the treatment of vascular malformations. An International Network for Sharing Practices on Electrochemotherapy (InspECT) study group report.

BACKGROUND: Biomedical applications of electroporation are expanding out of the field of oncology into vaccination, treatment of arrhythmias and now in the treatment of vascular malformations. Bleomycin is a widely used sclerosing agent in the treatment of various vascular malformations. The application of electric pulses in addition to bleomycin enhances the effectiveness of the drug, as demonstrated by electrochemotherapy, which utilizes bleomycin in the treatment of tumors. The same principle is used in bleomycin electrosclerotherapy (BEST). The approach seems to be effective in the treatment of low-flow (venous and lymphatic) and, potentially, even high-flow (arteriovenous) malformations. Although there are only a few published reports to date, the surgical community is interested, and an increasing number of centers are applying BEST in the treatment of vascular malformations. Within the International Network for Sharing Practices on Electrochemotherapy (InspECT) consortium, a dedicated working group has been constituted to develop standard operating procedures for BEST and foster clinical trials. CONCLUSIONS: By treatment standardization and successful completion of clinical trials demonstrating the effectiveness and safety of the approach, higher quality data and better clinical outcomes may be achieved.

European e-Delphi process to define expert consensus on electrochemotherapy treatment indications, procedural aspects, and quality indicators in melanoma.

BACKGROUND: Skin metastases are an important co-morbidity in melanoma. Despite broad adoption, electrochemotherapy implementation is hindered by a lack of treatment indications, uncertainty regarding procedural aspects, and the absence of quality indicators. An expert consensus may harmonize the approach among centres and facilitate comparison with other therapies. METHODS: An interdisciplinary panel was recruited for a three-round e-Delphi survey. A literature-based 113-item questionnaire was proposed to 160 professionals from 53 European centres. Participants rated each item for relevance and degree of agreement on a five-point Likert scale, and received anonymous controlled feedback to allow revision. The items that reached concordant agreement in two successive iterations were included in the final consensus list. In the third round, quality indicator benchmarks were defined using a real-time Delphi method. RESULTS: The initial working group included 122 respondents, of whom 100 (82 per cent) completed the first round, thus qualifying for inclusion in the expert panel (49 surgeons, 29 dermatologists, 15 medical oncologists, three radiotherapists, two nurse specialists, two clinician scientists). The completion rate was 97 per cent (97 of 100) and 93 per cent (90 of 97) in the second and third rounds respectively. The final consensus list included 54 statements with benchmarks (treatment indications, (37); procedural aspects, (1); quality indicators, (16)). CONCLUSION: An expert panel achieved consensus on the use of electrochemotherapy in melanoma, with a core set of statements providing general direction to electrochemotherapy users to refine indications, align clinical practices, and promote quality assurance programmes and local audits. The residual controversial topics set future research priorities to improve patient care.

Electrochemotherapy is an effective locoregional therapy for skin metastases from melanoma, a problem faced by almost half of patients with metastatic disease. The lack of comparative studies and the heterogeneity of its clinical application among centres make it challenging to support consistent, evidence-based recommendations. To address this unmet need, a three-round online survey was conducted to establish a consensus on treatment indications, standard operating procedures, and quality indicators. In the survey, a panel of 100 European melanoma experts agreed on 56 statements that can be used to improve patient selection, homogenize treatment application, and monitor outcomes.

T-Cell Infiltration and Clonality May Identify Distinct Survival Groups in Colorectal Cancer: Development and Validation of a Prognostic Model Based on The Cancer Genome Atlas (TCGA) and Clinical Proteomic Tumor Analysis Consortium (CPTAC).

Predicting the survival outcomes of patients with colorectal cancer (CRC) remains challenging. We investigated the prognostic significance of the transcriptome and tumour-infiltrating lymphocyte T-cell receptor (TIL/Tc-TCR) repertoire and analysed TIL/Tc-TCR sequences of The Cancer Genome Atlas (TCGA) and the Clinical Proteomic Tumor Analysis Consortium (CPTAC) CRC cohorts. Using a multivariate Cox regression, we tested whether TIL/Tc-TCR repertoire, patient and tumour characteristics (stage, sidedness, total non-synonymous mutations, microsatellite instability (MSI) and transcriptional signatures) correlated with patient overall survival (OS) and designed a prognostic nomogram. A multivariate analysis (C-index = 0.75) showed that only patient age, disease stage, TIL/Tc degree of infiltration and clonality were independent prognostic factors for OS. The cut-offs for patients' allocation to TIL/Tc abundance subgroups were determined using a strategy of maximally selected rank statistics with the OptimalCutpoints R package. These were "high", "low" and "very high" (90 th percentile) TIL/Tc infiltration-stratified OS (median not reached, 67 and 44.3 months; p < 0.001); the results were validated in the CPTAC cohort. TIL/Tc clonality was prognostic (median OS in "high" vs. "low" clonality not reached and 67.3 months; p = 0.041) and independent of TIL/Tc infiltration. Whilst tumour sidedness was not prognostic, the "very highly" infiltrated tumours were prevalent among right-sided CRCs (p = 0.039) and showed distinct immunological features, with lower Th1 signature (p = 0.004), higher PD-L1 expression (p < 0.001) and likely enrichment in highly suppressory IL1R1+ Tregs (FoxP3 and IL1R1 overexpression, p < 0.001). TIL/Tc abundance and clonality are independent prognosticators in CRC and, combined with clinical variables, refine risk stratification. We identified a subset of CRCs with "very high" TIL/Tc infiltration, poor prognosis and distinct genetic and immunologic features, which may benefit from alternative therapeutic approaches. These results need validation in prospective patient cohorts.

Tumour infiltrating B cells discriminate checkpoint blockade-induced responses.

BACKGROUND: Immune cell-driven anti-cancer activity is paramount for effective responses to checkpoint inhibitors (ICB). However, the contribution of the different immune cell subsets in the circulation and within the tumour is poorly understood. MATERIALS AND METHODS: To elucidate the role of the different cell subsets in anti-tumour responses elicited by ICB, we performed single-cell analysis of the transcriptome and surface proteome of paired pre- and early on-treatment metastatic melanoma tumour biopsies and matched peripheral blood mononuclear cell samples. We next compared the survival of metastatic melanoma patients treated with ICB according to the abundance of pre-treatment tumour-infiltrating B cell clonotypes. RESULTS: We identified cell clusters associated with disease control or progression, defined differential expression of biological pathways likely involved in the immune awakening against the tumour and examined how cell-cell communication patterns between the tumour cell subsets change during treatment. Furthermore, we discovered that B cells (immunoglobulin expression and abundance of B cell clonotypes) discriminate the clinical response after ICB and propose that B cells likely contribute to anti-tumour immunity by antigen presentation through major histocompatibility complex molecules. Finally, we demonstrated that the abundance of tumour-infiltrating B cell clonotypes at baseline identifies two distinct risk groups, a finding that we confirmed in an independent cohort. CONCLUSIONS: Our exploratory translational study provides new insights on the mechanistic role of B cells in anti-melanoma immunity during treatment with ICB. Additionally, we support pre-treatment B cell tumour infiltration as a promising prognostic biomarker to be further validated as a tool for clinical risk stratification.

Electrochemotherapy for the treatment of cutaneous squamous cell carcinoma: The INSPECT experience (2008-2020).

Introduction: Cutaneous squamous cell carcinoma (cSCC) is a frequent skin cancer with a high risk of recurrence characterized by tumor infiltration and, in advanced cases, a poor prognosis. ECT (electrochemotherapy) is an alternative treatment option for locally advanced or recurrent cSCC that is unsuitable for surgical resection. In this study, we aimed to evaluate the data in the InspECT (International Network for Sharing Practice on ECT) registry of the referral centers and to clarify the indications for the use of ECT as a treatment modality for cSCC. Materials and methods: Patients with primary, recurrent or locally advanced cSCC from 18 European centers were included. They underwent at least one ECT session with bleomycin between February 2008 and November 2020, which was performed following the European Standard Operating Procedures. Results: The analysis included 162 patients (mean age of 80 years; median, 1 lesion/patient). Side effects were mainly local and mild (hyperpigmentation, 11%; ulceration, 11%; suppuration, 4%). The response to treatment per patient was 62% complete and 21% partial. In the multivariate model, intravenous drug administration and small tumor size showed a significant association with a positive outcome (objective response). One-year local progression-free survival was significantly better (p<0.001) in patients with primary tumors (80% (95% C.I. 70%-90%) than in patients with locally advanced disease (49% (95% C.I. 30%-68%). Conclusion: In the present study, ECT showed antitumor activity and a favorable safety profile in patients with complex cSCC for whom there was no widely accepted standard of care. Better results were obtained in primary and small tumors (<3 cm) using intravenous bleomycin administration.

Adjuvant skin-sparing electrochemotherapy in a breast cancer patient with a prosthetic implant: 5-year follow-up outcomes.

A 55-year-old woman with previous skin-sparing mastectomy and prosthetic reconstruction for multifocal ductal carcinoma developed homolateral axillary recurrence. Following nodal dissection, partial periprosthetic capsulectomy and the overlying breast skin excision, the pathology report revealed a positive cutaneous margin. Since further breast skin excision or radiotherapy would have compromised the prosthetic implant, and the patient was adamant about avoiding any endangering intervention, the multidisciplinary recommendation included skin-directed electrochemotherapy (ECT) in the frame of a multimodal treatment strategy. The procedure lasted 20 minutes under mild general sedation and included a bolus of intravenous bleomycin followed by local application of electric pulses using a needle electrode. The postprocedural course was uneventful, except for mild dermatologic toxicity. At 5 years, the patient is disease-free with the implant in situ. This report illustrates the proof-of-concept of adjuvant skin-sparing ECT to sterilize resection margins, preserve a breast implant and highlight procedural details to avert toxicity.

Hypothesised cutaneous sites of origin of stage III melanomas with unknown primary: A multicentre study.

Based on molecular evidence that melanomas with unknown primary (MUPs) arise from the skin, we hypothesised that sites of MUPs are disproportionately on trunk and lower limbs, sites that are not readily visible to patients and clinicians. We tested this hypothesis by inferring the anatomic site of origin of MUPs from the corresponding known cutaneous sites of melanoma patients with known primary tumours (MKPs). We analysed data from three separate cohorts of patients from Brisbane, Australia (n = 236); Manchester, UK (n = 51) and Padova, Italy (n = 33), respectively, who first presented with stage III melanoma with lymph node metastases. We matched two MKP patients to each MUP patient based on lymph node dissection (LND) site, age and sex, and imputed cutaneous sites of origin of MUPs from their two matched MKPs for study countries, giving two possible sites for each MUP per centre. Overall, results showed that MUP patients were predominantly male, and trunk was the most likely origin, comprising around a third to a half of MUPs across the three cohorts. The remaining MUP inferred sites varied by country. In the Australian cohort, the legs accounted for a third of imputed sites of MUPs, while in the UK and Italian cohorts, the most frequent site was the arms followed by the legs. Our findings suggest the need for regular and thorough skin examination on trunk and limbs, especially in males, to improve early detection of cutaneous melanoma and reduce the risk of metastatic disease at the time of presentation.

Combination of Pembrolizumab with Electrochemotherapy in Cutaneous Metastases from Melanoma: A Comparative Retrospective Study from the InspECT and Slovenian Cancer Registry.

Electrochemotherapy (ECT) is an effective locoregional therapy for cutaneous melanoma metastases and has been safely combined with immune checkpoint inhibitors in preliminary experiences. Since ECT is known to induce immunogenic cell death, its combination with immune checkpoint inhibitors might be beneficial. In this study, we aimed to investigate the effectiveness of ECT on cutaneous melanoma metastases in combination with pembrolizumab. We undertook a retrospective matched cohort analysis of stage IIIC-IV melanoma patients, included in the International Network for sharing practices of ECT (InspECT) and the Slovenian Cancer Registry. We compared the outcome of patients who received the following treatments: (a) pembrolizumab alone, (b) pembrolizumab plus ECT, and (c) ECT. The groups were matched for age, sex, performance status, and size of skin metastases. The local objective response rate (ORR) was higher in the pembrolizumab-ECT group than in the pembrolizumab group (78% and 39%, p < 0.001). The 1 year local progression-free survival (LPFS) rates were 86% and 51% (p < 0.001), and the 1 year systemic PFS rates were 64% and 39%, respectively (p = 0.034). The 1 year overall survival (OS) rates were 88% and 64%, respectively (p = 0.006). Our results suggest that skin-directed therapy with ECT improves superficial tumor control in melanoma patients treated with pembrolizumab. Interestingly, we observed longer PFS and OS in the pembrolizumab-ECT group than in the pembrolizumab group. These findings warrant prospective confirmation.

The T cell receptor repertoire of tumor infiltrating T cells is predictive and prognostic for cancer survival.

Tumor infiltration by T cells is paramount for effective anti-cancer immune responses. We hypothesized that the T cell receptor (TCR) repertoire of tumor infiltrating T lymphocytes could therefore be indicative of the functional state of these cells and determine disease course at different stages in cancer progression. Here we show that the diversity of the TCR of tumor infiltrating T cell at baseline is prognostic in various cancers, whereas the TCR clonality of T cell infiltrating metastatic melanoma pre-treatment is predictive for activity and efficacy of PD1 blockade immunotherapy.

Biological factors of the tumour response to electrochemotherapy: Review of the evidence and a research roadmap.

The beneficial effects of electrochemotherapy (ECT) for superficial tumours and, more recently, deep-seated malignancies in terms of local control and quality of life are widely accepted. However, the variability in responses across histotypes needs to be explored. Currently, patient selection for ECT is based on clinical factors (tumour size, histotype, and exposure to previous oncological treatments), whereas there are no biomarkers to predict the response to treatment. In this field, two major areas of investigation can be identified, i.e., tumour cell characteristics and the tumour microenvironment (vasculature, extracellular matrix, and immune infiltrate). For each of these areas, we describe the current knowledge and discuss how to foster further investigation. This review aims to provide a summary of the currently used guiding clinical factors and delineates a research roadmap for future studies to identify putative biomarkers of response to ECT. These biomarkers may allow researchers to improve ECT practice by customising treatment parameters, manipulating the tumour and its microenvironment, and exploring novel therapeutic combinations.

Outcomes of older adults aged 90 and over with cutaneous malignancies after electrochemotherapy with bleomycin: A matched cohort analysis from the InspECT registry.

BACKGROUND: With extending life expectancy, more people are diagnosed with cutaneous malignancies at advanced ages and are offered nonsurgical treatment. We assessed outcomes of the oldest-old adults after electrochemotherapy (ECT). METHODS: The International Network for Sharing Practices of ECT (InspECT) registry was queried for adults aged ≥90 years (ys) with skin cancers/cutaneous metastases of any histotype who underwent bleomycin-ECT (2006-2019). These were subanalysed with patients aged <90 ys after matching 1:2 for tumor location, number, size, histotype, and previous treatments. We assessed ECT modalities, toxicity (CTCAE), response (RECIST), and patient perception (EQ-5D). RESULTS: Sixty-one patients represented the study cohort (median 92 ys, range 92-104), 122 the control group (median 77 ys, range 23-89). Among the oldest-old, 44 patients (72%) had primary/recurrent skin cancers, 17 (28%) cutaneous metastases. Median tumour size was 15 mm (range, 5-450). The oldest-old adults underwent ECT mainly under local/regional anaesthesia (59% vs 39% p = .012). We observed no differences regarding dose and route of chemotherapy (intravenous vs intratumoral, p = .308), electrode geometry (linear vs hexagonal, p = .172) and procedural duration (18 vs 21 min, p = .378). Complete response (57.4 [95%-CI 44.1%-70.0%] vs 64.7% [95%-CI 55.6%-73.2%], p = .222) and 1-year local control (76.7% vs 81.7, p = .092) rates were comparable. Pain and skin hyperpigmentation were mild in both groups. Skin ulceration persisted longer in the oldest-old patients (4.4 vs 2.4 months, p = .008). CONCLUSIONS: The oldest-old adults with cutaneous malignancies undergo ECT most commonly under local/regional anaesthesia with safety profiles and clinical effectiveness similar to their younger counterparts, except in case of ulcerated tumors.

Electrochemotherapy for advanced cutaneous angiosarcoma: A European register-based cohort study from the International Network for Sharing Practices of electrochemotherapy (InspECT).

BACKGROUND: Cutaneous angiosarcoma (cAS) is a highly aggressive malignancy that challenges the radicality of surgical treatment. Electrochemotherapy (ECT), a skin-directed treatment based on cytotoxic chemotherapy combined with local electric pulses, may be an intraoperative adjunct and a new opportunity in the therapeutic strategy. This cohort study reports the experience with ECT as an option. METHODS: Data on patients with locally-advanced/metastatic cAS who underwent ECT between October 2013 and October 2018 at eight European centres were prospectively submitted to the InspECT (International network for sharing practices of ECT) register. Patients received therapy according to the European Standard Operating Procedures of ECT (ESOPE). Treatment feasibility was assessed based on tumour coverage with electrodes and recorded tissue current; treatment toxicity and tumour response were graded according to CTCAE v5.0 and RECIST v1.1 criteria, respectively; patient-reported outcomes (PRO) were evaluated using a visual analogue score (VAS) for pain, acceptance of retreatment and the EQ-5D questionnaire. RESULTS: We enrolled 20 patients with advanced cAS in the scalp/face (n = 7), breast/trunk (n = 10) or limbs (n = 3). Target tumours (n = 51) had a median size of 2.3 cm (range, 1-20). We administered 24 ECT courses using 1-4 cm treatment safety margin around tumours. In five patients, ECT was combined/sequenced with surgery. Median tissue current was 3 A (range, 1.5-10), tumour margins coverage rate was 75% (15/20 patients). The objective response rate (ORR) was 80% (complete, 40%). Grade-3 toxicity included skin ulceration (15%) and pain (10%), with no significant change of PRO scores. Bleeding control was achieved in 13/14 patients with ulcerated tumours. With a median overall survival of 12.5 months, the local progression-free survival (LPFS) was 10.9 months. CONCLUSION: ECT produces sustained response rate with minimal side effects and should be considered an option for advanced cAS. Palliative benefits include patient tolerability, local haemostasis and durable local control. Definition of optimal timing, treatment safety margins and combination with surgery need further investigation.

Electrochemotherapy of superficial tumors - Current status:: Basic principles, operating procedures, shared indications, and emerging applications.

Treatment of superficial tumors with electrochemotherapy (ECT) has shown a steep rise over the past decade and indications range from skin cancers to locally advanced or metastatic neoplasms. Based on reversible electroporation, which is a physical method to achieve transient tumor cell membrane permeabilization by means of short electric pulses, ECT increases cellular uptake of bleomycin and cisplatin and their cytotoxicity by 8,000- and 80-fold, respectively. Standard operating procedures were established in 2006 and updated in 2018. Ease of administration, patient tolerability, efficacy across histotypes, and repeatability are peculiar advantages, which make standard ECT (ie, ECT using fixed-geometry electrodes) a reliable option for controlling superficial tumor growth locally and preventing their morbidity. Consolidated indications include superficial metastatic melanoma, breast cancer, head and neck skin tumors, nonmelanoma skin cancers, and Kaposi sarcoma. In well-selected patients with oropharyngeal cancers, ECT ensures appreciable symptom control. Emerging applications include skin metastases from visceral or hematological malignancies, vulvar cancer, and some noncancerous skin lesions (keloids and capillary vascular malformations). Repeatability and integration with other oncologic therapies allow for consolidation of response and sustained tumor control. In this review, we present the basic principles of ECT, recently updated operating procedures, anesthesiological management, and provide a synthesis of the efficacy of standard ECT across histotypes.

Electrochemotherapy - Emerging applications technical advances, new indications, combined approaches, and multi-institutional collaboration.

The treatment of tumors with electrochemotherapy (ECT) has surged over the past decade. Thanks to the transient cell membrane permeabilization induced by the short electric pulses used by ECT, cancer cells are exposed to otherwise poorly permeant chemotherapy agents, with consequent increased cytotoxicity. The codification of the procedure in 2006 led to a broad diffusion of the procedure, mainly in Europe, and since then, the progressive clinical experience, together with the emerging technologies, have extended the range of its application. Herein, we review the key advances in the ECT field since the European Standard Operating Procedures on ECT (ESOPE) 2006 guidelines and discuss the emerging clinical data on the new ECT indications. First, technical developments have improved ECT equipment, with custom electrode probes and dedicated tools supporting individual treatment planning in anatomically challenging tumors. Second, the feasibility and short-term efficacy of ECT has been established in deep-seated tumors, including bone metastases, liver malignancies, and pancreatic and prostate cancers (long-needle variable electrode geometry ECT), and gastrointestinal tumors (endoscopic ECT). Moreover, pioneering studies indicate lung and brain tumors as suitable future targets. A further advance relates to new combination strategies with immunotherapy, gene electro transfer (GET), calcium EP, and radiotherapy. Finally and fourth, cross-institutional collaborative groups have been established to refine procedural guidelines, promote clinical research, and explore new indications.