Bertrand Leroux

Background: Stereotactic ablative radiotherapy (SABR) is increasingly used in the treatment of localized and metastatic renal cell carcinoma (RCC), a malignancy traditionally considered radioresistant. Beyond direct cytotoxicity, SABR may promote immunogenic cell death and modulate the tumor immune microenvironment, though the underlying mechanisms remain incompletely understood. Objectives and Methods: This study examined the immunomodulatory effects of two high-dose irradiation regimens (8 Gy and 3 × 8 Gy) in an in vitro model using two RCC cell lines (ACHN, Caki-2) and peripheral blood mononuclear cells (PBMCs) from healthy donors. Results: The 3 × 8 Gy regimen more effectively reduced tumor cell viability and proliferation, particularly in ACHN cells, suggesting differential radiosensitivity. Both regimens induced secretion of IL-6, IL-8, TGF-β, and VEGF, with levels varying by cell line and dose. Caki-2 cells exhibited a cytokine profile consistent with a pro-inflammatory and potentially immunosuppressive phenotype. Conditioned media from irradiated cells were used to stimulate PBMCs, revealing divergent responses. Media from 3 × 8 Gy-irradiated ACHN cells enhanced PBMC proliferation and increased CD8⁺ T cells and CD11c⁺ monocytes, along with IFN-γ, IL-2, and TNF-α secretion, suggesting immunostimulatory effects. Conversely, media from Caki-2 cells had minimal impact on PBMC proliferation and increased TGF-β levels. Conclusions: These results indicate that high-dose irradiation can differentially modulate immune responses in RCC cell lines, depending on tumor intrinsic properties and irradiation regimen. Further in vivo studies are warranted to validate these findings and support development of SABR immunotherapy combinations guided by predictive immune biomarkers.

Photodynamic therapy (PDT) has shown improvements in cancer treatment and in the induction of a proper anti-tumor immune response. However, current photosensitizers (PS) lack tumor specificity, resulting in reduced efficacy and side effects in patients with intraperitoneal ovarian cancer (OC). In order to target peritoneal metastases of OC, which overexpress folate receptor (FRα) in 80% of cases, we proposed a targeted PDT using a PS coupled with folic acid. Herein, we applied this targeted PDT in an in vivo mouse model of peritoneal ovarian carcinomatosis. The efficacy of the treatment was evaluated in mice without and with human peripheral blood mononuclear cell (PBMC) reconstitution. When mice were reconstituted, using a fractionized PDT protocol led to a significantly higher decrease in the tumor growth than that obtained in the non-reconstituted mice (p = 0.0469). Simultaneously, an immune response was reflected by an increase in NK cells, and both CD4+ and CD8+ T cells were activated. A promotion in cytokines IFNγ and TNFα and an inhibition in cytokines TGFβ, IL-8, and IL-10 was also noticed. Our work showed that a fractionized FRα-targeted PDT protocol is effective for the treatment of OC and goes beyond local induction of tumor cell death, with the promotion of a subsequent anti-tumor response.

Photodynamic therapy (PDT) is a two-stage treatment relying on cytotoxicity induced by photoexcitation of a nontoxic dye, called photosensitizer (PS). Using 5-aminolevulinic acid (5-ALA), the pro-drug of PS protoporphyrin IX, we investigated the impact of PDT on hepatocellular carcinoma (HCC). Optimal 5-ALA PDT dose was determined on three HCC cell lines by analyzing cell death after treatment with varying doses. HCC-patient-derived tumor hepatocytes and healthy donor liver myofibroblasts were treated with optimal 5-ALA PDT doses. The proliferation of cancer cells and healthy donor immune cells cultured with 5-ALA-PDT-treated conditioned media was analyzed. Finally, therapy efficacy on humanized SCID mice model of HCC was investigated. 5-ALA PDT induced a dose-dependent decrease in viability, with an up-to-four-fold reduction in viability of patient tumor hepatocytes. The 5-ALA PDT treated conditioned media induced immune cell clonal expansion. 5-ALA PDT has no impact on myofibroblasts in terms of viability, while their activation decreased cancer cell proliferation and reduced the tumor growth rate of the in vivo model. For the first time, 5-ALA PDT has been validated on primary patient tumor hepatocytes and donor healthy liver myofibroblasts. 5-ALA PDT may be an effective anti-HCC therapy, which might induce an anti-tumor immune response.

Photodynamic therapy (PDT) appears to be a promising strategy in biomedical applications. However, the complexity of its parameters prevents wide acceptance. This work presents and characterizes a novel optical device based on knitted light-emitting fabrics and dedicated to in vitro PDT involving low irradiance over a long illumination period. Technical characterization of this device, called CELL-LEF, is performed. A cytotoxic study of 5-ALA-mediated PDT on human cancer cell lines is provided as a proof of concept. The target of delivering an irradiance of 1 mW/cm² over 750 cm² is achieved (mean: 0.99 mW/cm²; standard deviation: 0.13 mW/cm²). The device can maintain a stable temperature with the mean thermal distribution of 35.1 °C (min: 30.7 °C; max: 38.4 °C). In vitro outcomes show that 5-ALA PDT using CELL-LEF consistently and effectively induced a decrease in tumor cell viability: Almost all the HepG2 cells died after 80 min of illumination, while less than 60% of U87 cell viability remained. CELL-LEF is suitable for in vitro PDT involving low irradiance over a long illumination period.

Often discovered at an advanced stage, ovarian cancer progresses to peritoneal carcinoma, which corresponds to the invasion of the serosa by multiple tumor implants. The current treatment is based on the combination of chemotherapy and tumor cytoreduction surgery. Despite the progress and standardization of surgical techniques combined with effective chemotherapy, post-treatment recurrences affect more than 60% of women in remission. Photodynamic therapy (PDT) has been particularly indicated for the treatment of superficial lesions on large surfaces and appears to be a relevant candidate for the treatment of microscopic intraperitoneal lesions and non-visible lesions. However, the impact of this therapy on immune cells remains unclear. Hence, the objective of this study is to validate the efficacy of a new photosensitizer [pyropheophorbide a-polyethylene glycol-folic acid (PS)] on human ovarian cancer cells and to assess the impact of the secretome of PDT-treated cells on human peripheral blood mononuclear cells (PBMC). We show that PS, upon illumination, can induce cell death of different ovarian tumor cells. Furthermore, PDT using this new PS seems to favor activation of the immune response by inducing the secretion of effective cytokines and inhibiting the pro-inflammatory and immunosuppressive ones, as well as releasing extracellular vesicles (EVs) prone to activating immune cells. Finally, we show that PDT can activate CD4+ and CD8+ T cells, resulting in a potential immunostimulating process. The results of this pilot study therefore indicate that PS-PDT treatment may not only be effective in rapidly and directly destroying target tumor cells but also promote the activation of an effective immune response; notably, by EVs. These data thus open up good prospects for the treatment of micrometastases of intraperitoneal ovarian carcinosis which are currently inoperable.

To date, pancreatic adenocarcinoma (ADKP) is a devastating disease for which the incidence rate is close to the mortality rate. The survival rate has evolved only 2–5% in 45 years, highlighting the failure of current therapies. Otherwise, the use of photodynamic therapy (PDT), based on the use of an adapted photosensitizer (PS) has already proved its worth and has prompted a growing interest in the field of oncology. We have developed a new photosensitizer (PS-FOL/PS2), protected by a recently published patent (WO2019 016397-A1, 24 January 2019). This photosensitizer is associated with an addressing molecule (folic acid) targeting the folate receptor 1 (FOLR1) with a high affinity. Folate binds to FOLR1, in a specific way, expressed in 100% of ADKP or over-expressed in 30% of cases. The first objective of this study is to evaluate the effectiveness of this PS2-PDT in four ADKP cell lines: Capan-1, Capan-2, MiapaCa-2, and Panc-1. For this purpose, we first evaluated the gene and protein expression of FOLR1 on four ADKP cell lines. Subsequently, we evaluated PS2’s efficacy in our cell lines and we assessed the impact of PDT on the secretome of cancer cells and its impact on the immune system. Finally, we evaluate the PDT efficacy on a humanized SCID mouse model of pancreatic cancer. In a very interesting way, we observed a significant increase in the proliferation of activated-human PBMC when cultured with conditioned media of ADKP cancer cells subjected to PDT. Furthermore, to evaluate in vivo the impact of this new PS, we analyzed the tumor growth in a humanized SCID mice model of pancreatic cancer. Four conditions were tested: Untreated, mice (nontreated), mice with PS (PS2), mice subjected to illumination (Light only), and mice subjected to illumination in the presence of PS (PDT). We noticed that the mice subjected to PDT presented a strong decrease in the growth of the tumor over time after illumination. Our investigations have not only suggested that PS2-PDT is an effective therapy in the treatment of PDAC but also that it activates the immune system and could be considered as a real adjuvant for anti-cancer vaccination. Thus, this new study provides new treatment options for patients in a therapeutic impasse and will provide a new arsenal in the fight against PDAC.