Summary

Background: When exposed to radiotherapy (RT), NBTXR3 nanoparticles increase radiation dose deposition from within the cancer cells. NBTXR3 is intended for a single intratumor injection. Results from a phase II/III clinical trial in patients with locally advanced Soft Tissue Sarcoma demonstrated significant superiority and clinical benefits of NBTXR3 activated by RT compared to RT alone, and was well tolerated. NBTXR3 is currently being evaluated in several other tumors including head and neck, liver, and pancreatic cancer as a single agent or in combination with anti-PD1. Moreover, preclinical studies have demonstrated that NBTXR3 can produce a significant abscopal effect, whereas RT alone cannot. Here, we explored the impact of NBTXR3 activated by RT on CD8+ infiltrates and TcR repertoire diversity change, and the effect on the immunopeptidome of cancer cells.

Methods: CT26 (murine colorectal cancer cells) were subcutaneously injected in BALB/c mice in one flank. Then, tumors were intratumorally injected with NBTXR3 (or vehicle) and irradiated 24 hours later with 4Gy per fraction for 3 consecutive days. Tumors were collected 3 days after the last RT fraction and immune cell infiltrates were measured using immunohistochemistry (IHC) and digital pathology. For TcR repertoire sequencing, the same workflow was followed, except cells were injected in both flanks. Only right tumors received treatment, while left tumors remained untreated. For immunopeptidome analysis, in vitro cells were irradiated by 4Gy. After one day, cells were collected for isolation and sequencing of MHC I-loaded peptides.

Results: IHC analyses showed a significant increase of CD8+ T cell infiltrates in tumors of mice treated with NBTXR3+RT, while RT alone had no significant effect. In addition, NBTXR3+RT treatment was able to increase TcR repertoire diversity, both in treated and untreated tumors, compared to RT alone. Finally, analysis of immunopeptidome showed that NBTXR3+RT changed the profile of MHC-I-loaded peptides.

Conclusions: Our in vivo data indicate that NBTXR3+RT can modulate the microenvironment of treated tumors, leading to enhanced CD8+ T cell infiltration as well as modification of the TcR repertoire, both in treated and distant untreated tumors. These NBTXR3+RT-induced responses may be related to changes in the immunopeptidome of cancer cells triggered by this treatment.

Summary

Purpose/Objective(s): NBTXR3, a novel radioenhancer activated by radiotherapy (RT) demonstrated superior efficacy, as preoperative treatment in patients with locally advanced soft tissue sarcoma (LA STS) compared to RT alone. Primary endpoint of pCR rate (16% vs 8%; P = 0.044) and main secondary endpoint of R0 margin rate (16% vs 8%; P = 0.042) were met while no modification of the early RT-associated safety profile was observed, leading to market authorization. Here we report on the long-term safety, limb function and quality of life.

Materials/Methods: This phase II/III randomized (1:1), international trial included adult patients with LA STS of the extremity or trunk wall, requiring preoperative RT. Patients received either a single intratumoral injection of NBTXR3 (equivalent to 10% of tumor volume, at 53.3g/L), plus EBRT (arm A) or EBRT alone (arm B) (50 Gy in 25 fractions), followed by surgery. Here we report on safety of NBTXR3+RT which was evaluated as secondary endpoint. Data were recorded on the “all treated population” during at least a two-year follow-up. Important parameters related to HR-QoL including functional outcome were studied using the EQ-5D, RNLI, TESS and MSTS questionnaires.

Results: Patients had at least two-year follow-up and the lost to follow-up rate was very low (1.9%). RT-related SAEs were observed in 11.2% (10/89) vs 13.3% (12/90) in A vs B. Post-treatment AEs, any grade, were observed in 51.7% (46/89) vs 57.8% (52/90) and serious post-treatment AEs in 13.5% (12/89) vs 24.4% (22/90) of patients in A vs B. Long-term safety continues to demonstrate that NBTXR3 plus RT has no impact on post-surgical wound complications (24.7% vs 36.7%, A vs B). Furthermore, the evaluation of radiation late toxicities in limbs such as fibrosis, arthrosis and edema that may alter limb function showed no difference between arms (4.5% vs 7.7%, 2.2% vs 0.0% and 6.7% vs 2.2% respectively in A vs B). In addition, sequelae or chronic tissue disturbances at the former tumor localization were similar in both treatment arms, confirming that the increase of energy dose deposit and the physical presence of NBTXR3 did not impact post-treatment limb functions. Accordingly, HR-QoL evaluation yielded no difference in functional outcome. Finally, second primary cancer was observed in 1 patient in arm A and 6 patients in arm B and the intratumoral injection of NBTXR3 did not induce cancer cell seeding at the former tumor site.

Conclusion: These results demonstrate that the use of NBTXR3 did not change the late onset toxicity profile of EBRT, nor modified its bystander effect. Taken together, the long-term safety data presented here, and the previously published efficacy data reinforce the favorable benefit-risk ratio of the use of NBTXR3 in patients with LA STS. NCT02379845

Summary

Purpose/Objective(s): Immune checkpoint inhibitors (ICI) can improve outcomes in patients who respond to treatment, however most patients exhibit resistance. Overcoming this resistance is the main challenge in immune-oncology and recent studies suggest radiotherapy (RT) may improve ICI response rates. NBTXR3, composed of functionalized hafnium oxide nanoparticles, is injected intratumorally and activated by RT. NBTXR3 increases RT energy deposit inside tumor cells and subsequent tumor cell death, without adding toxicity to healthy tissues. Here we present evidence that NBTXR3 activated by RT primes the immune system, producing an anti-tumor immune response, including activation of the cGAS-STING pathway, that overcomes anti-PD-1 resistance both in murine models and patients.

/Methods: Abscopal assays were conducted in immunocompetent mice. Anti-PD-1 sensitive or resistant lung tumor cell lines were injected in both flanks. Intratumoral injection of NBTXR3 (or vehicle) followed by RT was performed in right flank (primary) tumors only. Some mice also received anti-PD-1 injections. Tumor growth was monitored, and tumor immune cell infiltrates analyzed by immunohistochemistry (IHC). Separately, in the phase II/III randomized Act.in.Sarc [NCT02379845] trial patients with locally advanced soft tissue sarcoma (STS) received either NBTXR3+RT or RT alone followed by tumor resection. Pre- and post-treatment tumor samples from patients in both groups were analyzed by IHC and Digital Pathology for immune biomarkers. The safety and efficacy of NBTXR3 plus stereotactic body radiotherapy (SBRT) in combination with anti-PD-1 is being evaluated in three cohorts of patients with advanced cancers in the Phase I 1100 [NCT03589339] trial.

Results: Pre-clinical studies demonstrated that NBTXR3+RT induces an immune response not observed with RT alone and enhances systemic control. IHC showed significant increase of CD8+ T-cell infiltrates in both NBTXR3+RT treated and untreated tumors compared to RT alone. Increased CD8+ T-cell and decreased FOXP3+ Treg density (pre- vs post-treatment) was also observed in tumors from STS patients treated with NBTXR3+RT. Furthermore, NBTXR3+RT in combination with anti-PD-1 improved local and systemic control in mice bearing anti-PD-1 resistant lung tumors, produced long-term memory, and reduced spontaneous lung metastases. Preliminary efficacy data from the 1100 trial showed tumor regression in 8/9 patients. Of note, tumor regression was observed in 6/7 patients who had progressed on prior anti-PD-1.

Conclusion: The clinical efficacy of NBTXR3+RT has been demonstrated as a single agent in STS. Here we demonstrate that it overcomes resistance to anti-PD-1 treatment mechanisms in mice and led to tumor regression in patients having progressed on anti-PD-1 therapy. These results highlight the potential of NBTXR3+RT to positively impact the immuno-oncology field.

Summary

Purpose or Objective: NBTXR3, a novel radioenhancer activated by radiotherapy (RT) demonstrated superior efficacy, as preoperative treatment in patients with locally advanced soft tissue sarcoma (LA STS) compared to RT alone. Primary endpoint of pCR rate (16% vs 8%; p=0.044) and main secondary endpoint of R0 margin rate (16% vs 8%; p=0.042) were met (Bonvalot et al. Lancet Oncol. 2019) while no modification of the early RT-associated safety profile was observed, leading to market authorization. Here we report on the long-term safety, limb function and quality of life.

Materials and Methods: This phase II/III randomized (1:1), international trial included adult patients with LA STS of the extremity or trunk wall, requiring preoperative RT (NCT02379845). Patients received either a single intratumoral injection of NBTXR3 (equivalent to 10% of tumor volume, at 53.3g/L), plus EBRT (arm A) or EBRT alone (arm B) (50 Gy in 25 fractions), followed by surgery. Here we report on safety of NBTXR3+RT which was evaluated as secondary endpoint. Data were recorded on the “all treated population” during at least a two-year follow-up. Important parameters related to HR-QoL including functional outcome were studied using the EQ-5D, RNLI, TESS and MSTS questionnaires.

Results: Patients had at least two-year follow-up and the lost to follow-up rate was very low (1.9%). RT-related SAEs were observed in 11.2% (10/89) vs 13.3% (12/90) in A vs B. Post-treatment AEs, any grade, were observed in 51.7% (46/89) vs 57.8% (52/90) and serious post-treatment AEs in 13.5% (12/89) vs 24.4% (22/90) of patients in A vs B. Long-term safety continues to demonstrate that NBTXR3 plus RT has no impact on post-surgical wound complications (24.7% vs 36.7%, A vs B). Furthermore, the evaluation of radiation late toxicities in limbs such as fibrosis, arthrosis and edema that may alter limb function showed no difference between arms (4.5% vs 7.7%, 2.2% vs 0.0% and 6.7% vs 2.2% respectively in A vs B). In addition, sequelae or chronic tissue disturbances at the former tumor localization were similar in both treatment arms, confirming that the increase of energy dose deposit and the physical presence of NBTXR3 did not impact post-treatment limb functions. Accordingly, HR-QoL evaluation yielded no difference in functional outcome. Finally, second primary cancer was observed in 1 patient in arm A and 6 patients in arm B and the intratumoral injection of NBTXR3 did not induce cancer cell seeding at the former tumor site.

Conclusion: These results demonstrate that the use of NBTXR3 did not change the late onset toxicity profile of EBRT, nor modified its bystander effect. Taken together, the long-term safety data presented here, and the previously published efficacy data reinforce the favorable benefit-risk ratio of the use of NBTXR3 in patients with LA STS.

Summary

Background: NBTXR3, a novel radioenhancer activated by radiotherapy (RT) demonstrated superior efficacy, as preoperative treatment, in patients with locally advanced soft tissue sarcoma (LA STS), compared to RT alone. Primary endpoint of pCR rate was 16% vs 8% (p=0.044) and R0 margin rate was 77% vs 64% (p=0.042) (Bonvalot et al. Lancet Oncol. 2019). No modification of the early safety profile of RT was observed, leading to market authorization. Here we report on the long-term safety, limb function and quality of life.

Methods: This phase II/III randomized (1:1), international trial included adult patients with LA STS of the extremity or trunk wall, requiring preoperative RT (NCT02379845). Patients were treated with either a single intratumoral injection of NBTXR3 (volume equivalent to 10% of tumor volume, at 53.3g/L) plus EBRT (arm A), or EBRT alone (arm B) (50 Gy in 25 fractions), followed by surgery. The primary and main secondary efficacy endpoints were previously reported. Safety of NBTXR3+RT, as preoperative treatment, was evaluated as secondary endpoint. We present the safety analyses done in the “all treated population”, with data recorded during at least a two-year follow-up. Important parameters related to HR-QoL, including functional outcome were studied using the EQ-5D, RNLI, TESS and MSTS questionnaires.

Results: Patients had at least two-year follow-up and the lost to follow-up rate was very low (1.9%). RT-related SAEs were observed in 11.2% (10/89) vs 13.3% (12/90) in A vs B. Post-treatment AEs, any grade, were observed in 51.7% (46/89) vs 57.8% (52/90) and serious post-treatment AEs in 13.5% (12/89) vs 24.4% (22/90) of patients in A vs B. Second primary cancer was observed in 1 patient in arm A and 6 patients in arm B. Long-term safety continues to demonstrate that NBTXR3 plus RT has no impact on post-surgical wound complications (24.7% vs 36.7%, A vs B). Furthermore, the evaluation of radiation late toxicities in limbs such as fibrosis (4.5% vs 7.7%), arthrosis (2.2% vs 0.0%) and edema (6.7% vs 2.2%) that may alter limb function showed no difference between arms. Accordingly, HR-QoL evaluation yielded no difference in functional outcome. In addition, the intratumoral injection of NBTXR3 did not induce cancer cell seeding at the former tumor site. Finally, sequelae or chronic tissue disturbances at the former tumor localization were similar in both treatment arms, confirming that the increase of energy dose deposit and the physical presence of NBTXR3 did not impact post-treatment limb functions.

Conclusions: The long-term safety results demonstrate that the addition of NBTXR3 to EBRT neither added toxicity nor modified the bystander effect of RT. The results presented here associated with the efficacy data reported previously reinforce the favorable benefit-risk ratio of the use of NBTXR3 in patients with LA STS. Clinical trial information: NCT02379845.

Summary

Background: Locally advanced soft tissue sarcoma (STS) management may include neoadjuvant or adjuvant treatment by radiotherapy (RT), chemotherapy (CT) or chemoradiotherapy (CRT) followed by wide surgical excision. While pathological complete response (pCR) to preoperative treatment is prognostic for survival in osteosarcomas, its significance for STS is unclear. We aimed to evaluate the prognostic significance of pCR to pre-operative treatment on 3-year disease-free survival (3y-DFS) in STS patients.

Methods: This is an observational, retrospective, international, study of adult patients with primary non-metastatic STS of the extremities and trunk wall, any grade, diagnosed between 2008 and 2012, treated with at least neoadjuvant treatment and surgical resection and observed for a minimum of 3 years after diagnosis. The primary objective was to evaluate the effect of pCR. (≤5% viable tumor cells or ≥95% necrosis/fibrosis) on 3y-DFS. Effect on local recurrence-free survival (LRFS), distant recurrence-free survival (MFS) overall survival (OS) at 3 years was also analyzed. Statistical univariate analysis utilized chi-square independence test and odds ratio confidence interval (CI) estimate, multivariate analysis was performed using LASSO.

Results: A total of 330 patients (median age 56 years old, range:19–95) treated by preoperative RT (67%), CT (15%) or CRT (18%) followed by surgery were included. pCR was achieved in 74/330 (22%) of patients, of which 56/74 (76%) had received RT. 3-yr DFS was observed in 76% of patients with pCR vs 61% without pCR (p < 0.001). Multivariate analysis showed that pCR is statistically associated with better MFS (95% CI, 1.054–3.417; p = 0.033), LRFS (95% CI, 1.226–5.916; p = 0.014), DFS (95% CI, 1.165–4.040; p = 0.015) and OS at 3 years (95% CI, 1.072–5.210; p = 0.033).

Conclusions: In a wide, heterogeneous STS population we showed that pCR to preoperative treatment is prognostic for survival.

Summary

Purpose: First in class hafnium oxide nanoparticles (NBTXR3) activated by radiotherapy (RT) increase radiation dose deposit within cancer cells compared to RT alone. Given that RT can prime an anti-tumor immune response we hypothesized that this response could be enhanced by NBTXR3+RT in both animals and humans.

Method & Materials: Different abscopal assays in mice were conducted. Immunocompetent mice were injected in both flanks with murine tumor cells. Intratumoral injection of NBTXR3 (or vehicle) was performed in right flank tumors, followed by RT of right flank tumors only. Tumor growth was followed and immune cell infiltrates were analyzed by immunohistochemistry (IHC). Some mice received anti-PD-1 injections and tumor growth was monitored. Pts with locally advanced soft tissue sarcoma (STS) [NCT02379845] received either NBTXR3+RT or RT alone. Pts pre- and post-treatment tumor tissues were analyzed by IHC and Digital Pathology for immune biomarkers.

Results: Animal studies demonstrated that NBTXR3+RT induces an immune response which was not observed with RT alone. IHC showed significantly more CD8+ cells present in NBTXR3+RT treated and untreated tumors. Furthermore, NBTXR3+RT improved the effect of anti-PD-1. Similarly, increased CD8+ T cell infiltration pre- vs post-treatment was observed in tumor tissues from STS pts treated with NBTXR3+RT. An increase in biomarkers, including CD8, following NBTXR3+RT was also observed by IHC in tumor samples from STS pts compared to RT alone.

Conclusion: These results demonstrate that NBTXR3+RT induces a specific adaptive immune profile in both mice and STS pts. NBTXR3+RT also improved response to anti-PD-1 in mice, opening the potential for combination with immunotherapeutic agents in humans. We have therefore sought to investigate the safety and systemic effect of NBTXR3 activated by stereotactic ablative radiotherapy (SABR) in combination with anti-PD-1 in pts with locoregionally recurrent or metastatic (lung or liver) head and neck squamous cell carcinoma, as well as in metastatic non-small cell lung cancer and liver metastasis pts [NCT03589339].

Clinical Relevance & Application: The results of this study highlight the potential of NBTXR3 to be used in combination with immune checkpoint inhibitors in order to improve patient outcomes.

Summary

Purpose/Objective(s): Hafnium oxide nanoparticles (NBTXR3) activated by radiotherapy (RT) increase radiation dose deposit within cancer cells compared to RT alone. Currently 7 clinical trials are underway to evaluate NBTXR3+RT. To date, no dose limiting toxicities (DLTs) have been observed. Given that RT can prime an anti-tumor immune response we hypothesized that this response could be enhanced by NBTXR3+RT in both animals and humans.

Materials/Methods: Immunocompetent mice were injected in both flanks with CT26 cells. An intratumoral injection of NBTXR3 (or vehicle) was performed in right flank tumors, followed by RT (3x4Gy). Tumor growth was followed, and animals sacrificed when tumors reached 800mm3. Alternatively, tumors were collected 3 days after last RT fraction and immune cell infiltrates analyzed by immunohistochemistry (IHC). Pts with locally advanced soft tissue sarcoma (STS) [NCT02379845] received
either NBTXR3+RT or RT alone. Pre- and post-treatment tumor tissues (biopsy and tumor resection respectively) from pts were analyzed by IHC
and Digital Pathology for immune biomarkers (>16 pts per arm).

Results: Animal studies demonstrated that NBTXR3+RT can induce an immune response which was not observed with RT alone. IHC analyses showed that significantly more CD8+ cells were present in NBTXR3+RT treated and untreated tumors, compared to tumors from mice treated with RT alone. Similarly, increased CD8+ T cell infiltration pre- vs post-treatment was observed in tumor tissues from STS pts treated with NBTXR3+RT. An increase in biomarkers, including CD8 and PD1, following NBTXR3 +RT was also observed by IHC in tumor samples from STS pts compared to RT alone.

Conclusion: These results demonstrate that NBTXR3+RT induces a specific adaptive immune profile in both mice and STS pts. As such, it may convert immunologically “cold” tumors into “hot” tumors, opening the potential for combination with immunotherapeutic agents. We have therefore sought to investigate the safety and systemic effect of NBTXR3 activated by stereotactic ablative radiotherapy (SABR) in combination with anti-PD-1 antibody in pts with locoregionally recurrent or metastatic (to lung or liver) head and neck squamous cell carcinoma (HNSCC), as well as in metastatic non-small cell lung cancer (NSCLC) and liver metastasis patients [NCT03589339].

Summary

Purpose/Objective(s): Local interventional treatments of cancers include interventional radiology and radiotherapy (RT). NBTXR3, hafnium oxide nanoparticles, is deeply associated to both. Given as a single local administration it increases energy dose deposit inside tumor cells only when activated by ionizing radiation. Various interventional treatments have been used to treat cancers such as liver, lung, bone. Because entirely new therapies such as NBTXR3 are being introduced, implementation of interventional approaches is continuously growing.

Materials/Methods: NBTXR3 is being evaluated in soft tissue sarcoma (STS, extremity, trunk wall) [NCT02379845], head and neck (HN) [NCT01946867, NCT02901483], prostate [NCT02805894], liver [NCT02721056] and rectal cancers [NCT02465593]. NBTXR3 injected volume is a percentage of baseline tumor volume, and therefore heterogeneous. Image guidance allowed for accurate injection. Standard catheters, needles, and syringes were used for preparation and injection. Importantly, percutaneous needle positioning was done within the region to be irradiated to control potential seeding of cancer cells. NBTXR3 was then activated by IMRT (STS, HN), EBRT or combination brachytherapy/EBRT boost (prostate), SBRT (liver), IMRT or IMAT (rectum).

Results: Thus far, NBTXR3 has been administered to 171 patients by intratumoral/lesional, and intraprostate injections depending on indication. NBTXR3 injections have been demonstrated safe and very well tolerated. Local infection, ulceration or massive tumor necrosis were never observed. This has been confirmed by adequate application of treatment schedules, fitting planned irradiation onset 1 to 5 days post-injection. Importantly, grade 1 ecchymosis and hematoma at puncture site (needle entry) observed in few cases always resolved spontaneously and did not impact dosimetry. Indeed, change of tumor/lesion/prostate volume resolved when water (NBTXR3 vehicle) was drained via lymphatic system. So far, inflammatory response to injection procedure itself was mild. Concerning AEs, grade 3 pain was observed in conscious patients under local anesthesia with STS close to joints (limited extensibility), and in needle shift in injection within a subcapsular liver tumor.

Conclusion: Across 7 clinical trials involving tumors in extremity, trunk wall, liver, rectum, prostate and HN, NBTXR3 injection was well tolerated and demonstrated a very good safety profile. The savoir faire of interventional radiology for local treatment of cancers supported implementation of injection procedures with specific parameters according to anatomy. Intratumoral/lesional or intraprostate injection ensures optimum bioavailability at site of irradiation, protecting patients from systemic toxicity. Future clinical research will involve other anatomical sites such as lymph nodes and lung lesions [NCT03589339].

Summary

Background: Radiotherapy (RT) can prime an anti-tumor immune response. Unfortunately, this response rarely generates total tumor destruction and abscopal effect. When activated by RT, intratumorally (IT) administered hafnium oxide nanoparticles (NBTXR3) locally increase radiation dose deposit and tumor cell death compared to RT alone. We hypothesized that NBTXR3 + RT could enhance the anti-tumor immune response, both in mice and humans.

Methods: Murine CT26 cells were injected in both flanks of immunocompetent mice. When tumor volume reached 50-120mm3, NBTXR3 (or vehicle) was injected IT in right flank tumors only, then irradiated (3x4Gy). Mice were sacrificed when tumors reached 800mm3. Alternatively, tumors were collected 3 days after last RT fraction and immune cell infiltrates analyzed by immunohistochemistry (IHC). Patients (pts) with locally advanced Soft Tissue Sarcoma (STS) (NCT02379845) received NBTXR3 + RT or RT alone. Pre- and post-treatment (biopsy and resection, respectively) tumor tissues from pts were analyzed by IHC and Digital Pathology for immune biomarkers ( > 16 pts per arm).

Results: In mice, IHC analyses showed an increase of CD8+ T cells infiltrates in both flanks of mice treated with NBTXR3+RT, while this was not observed in animals treated with RT alone. Furthermore, ICH analysis of post- vs pre-treatment samples from STS pts showed a marked increase of CD8+ and PD1 biomarkers for pts treated with NBTXR3 + RT, while no differences were seen for pts treated with RT alone.

Conclusions: NBTXR3 + RT markedly changes the tumor immune profile in a similar manner in mice and pts with STS. We hypothesize that this adaptive immune response could help convert a local tumor microenvironment to a “hot” phenotype and thus improve the efficacy of immune checkpoint inhibitors. These results led us to investigate the safety and systemic effect of NBTXR3 activated by stereotactic ablative RT (SABR) in combination with anti-PD-1 antibody in pts with locoregionally recurrent or metastatic (to lung or liver) Head and Neck squamous cell carcinoma HNSCC, as well as in metastatic non-small cell lung cancer (NSCLC) and liver metastasis patients [NCT03589339].