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

Background: The majority of cancer patients are resistant to immune therapy; only around 15% respond to immune checkpoint inhibitors (ICI). Thus, strategies able to increase ICI response are of great interest. Recent work suggests radiotherapy (RT) can act as an immunomodulator to increase the proportion of ICI responders and improve clinical outcomes. However, RT dose and ultimate efficacy are limited by toxicity related to exposure of healthy tissues. NBTXR3 is a first-in-class radioenhancer administered by intratumoral injection, designed at the nanoscale to increase RT energy dose deposition within the tumor. The result is increased radiation-dependent tumor cell killing, without increasing radiation exposure of healthy tissues. Preclinical and early clinical data suggest NBTXR3 activated by RT can increase the anti-tumor immune response, producing both local and systemic (abscopal) effects. We hypothesize that NBTXR3 activated by RT, in combination with anti-PD-1 therapy (R3/RT/PD-1), will act synergistically to maximize the local RT effect while also producing a systemic response sufficient to increase the proportion of ICI responders or convert ICI non-responders to responders.

Trial Design: NANORAY-1100 [NCT03589339] is a multicenter, open-label, phase 1 study to evaluate safety and tolerability of R3/RT/PD-1 in three cohorts: (1) Locoregional recurrent or recurrent and metastatic head and neck squamous cell carcinoma (HNSCC) amenable to re-irradiation of the HN field, (2) Lung metastases from any primary cancer eligible for anti-PD-1, or (3) Liver metastases from any primary cancer eligible for anti-PD-1. Approximately two-thirds of each cohort will be composed of anti-PD-1 non-responders. NBTXR3 injection volume is based on a percentage of gross tumor volume (GTV) determined by central review. The primary objective is to determine R3/RT/PD-1 RP2D. Secondary objectives are to evaluate anti-tumor response (objective response rate; ORR) of R3/RT/PD-1, safety and feasibility of NBTXR3 injection, and NBTXR3 body kinetic profile. Exploratory objectives will assess biomarkers of R3/RT/PD-1 response, including PD-L1 status by IHC, mRNA and cytokine immune marker profiling.

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

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].

Summary

The enclosed abstract was presented at the 13th Journées cancéropole Grand Sud-Ouest at Poitiers. The abstract Hafnium oxide nanoparticles as an emergent promising treatment for solid tumors describes how hafnium oxide nanoparticles were designed at the nanoscale in the form of crystalline 50nm-particles to efficiently absorb ionizing radiation and increase the radiation dose deposited – “hot spots” of energy deposit – from within the tumor cells for efficient cell killing.

Hafnium oxide nanoparticles (NBTXR3), administered as a single intratumoral injection and activated by radiotherapy, is currently evaluated in a phase II/III clinical trial in soft tissue sarcoma (STS) [NCT02379845], and in phase I/II clinical trials for head and neck [NCT01946867], prostate [NCT02805894], liver [NCT02721056; NCT02721056] and rectum cancers [NCT02465593]. So far, patients treated with NBTXR3 received radiotherapy as planned and showed a very good local safety profile.

Summary

Immunotherapy Workshop: Bethesda, MD, June 15-16, 2017

Radiotherapy (RT) has proven its ability to function like an in-situ vaccine, showing potential for successful combination with immunotherapeutic agents. Hafnium oxide nanoparticle (HfO2-NP), undergoing clinical trials for enhancing RT, was designed as high electron density material at the nanoscale. HfO2-NPs are taken up by cancer cells and, when exposed to RT, locally increase the radiation dose deposit, triggering more cancer cells death when compared to RT. We hypothesized that HfO2-NP+RT could trigger an enhanced immune response when compared to RT, both in preclinical and clinical settings.

Summary

Background: NBTXR3 are functionalized hafnium oxide nanoparticles, undergoing seven clinical trials for enhancing radiation therapy (RT). The high electron density of the nanoparticles, when exposed to radiotherapy (NBTXR3 + RT), allow absorption/deposition of a high radiation dose within the cancer cells to physically kill the cells, and possibly improve outcome. Besides, NBTXR3 + RT has shown subsequent ability to enhance immunogenic cell death and immune response in preclinics. We hypothesized that NBTXR3 + RT could trigger an enhanced immune response when compared to RT in patients with STS.

Methods: Tumor tissues pre- (biopsy) and/or post-treatment (resection) were collected from patients (pts) with locally advanced STS, who received either NBTXR3 as intratumor injection and RT (14 pts) or RT (12 pts), as preoperative treatment (NCT02379845). Immunohistochemistry and Digital Pathology for immune biomarkers and for Immunoscore (CD3/CD8) were analyzed. Gene expression profiling and pre-optimized immune-gene signatures called Immunosign were also used.

Results: A significant increase of T cells (CD3+, CD8+) and a marked increase of CD103+ immune cell infiltration post- vs pre-treatment were observed for NBTXR3 + RT (P< 0.01), while no differences were seen for RT. Post-treatment, an increased Immunoscore (CD3 + CD8 cell densities) was observed for NBTXR3 + RT compared to RT (P < 0.07). Consistently, the up-regulation of pan immune genes expression and specifically expression of adaptive immunity genes between pre- and post-treatment, was pronounced for NBTXR3 + RT when compared to RT. Functional analysis of genes up-regulated in NBTXR3 + RT showed an enrichment of cytokine activity (IL7, IFNA, IL16, IL11, IFNG), adaptive immunity (RAG1, GZMA, TAP1, TAP2, TBX21, STAT4, IFNG, LCK, LTK, CD37, CD22) and T cell receptor signaling pathway (CD28, CTLA4, CD274, BTLA, TIGIT, CD40LG, CD5, CD3E, ZAP70).

Conclusions: NBTXR3 + RT induces a specific adaptive immune pattern. As such, it may contribute to convert “cold” tumor into “hot” tumor and be effectively combined with immunotherapeutic agents across oncology. These data warrant more tissue samples evaluation to reinforce these findings.