Non-irradiated Lesions Research Articles

Abstract PO1-19-10: Durvalumab + datopotamab deruxtecan in patients with PD-L1 positive advanced/metastatic triple-negative breast cancer: Arm 8 of the phase 1b/2, open label, platform BEGONIA study

Abstract Background: Patients (pts) with advanced/metastatic triple-negative breast cancer (a/mTNBC) have limited treatment options and poor prognosis. Immune checkpoint inhibitors combined with chemotherapy is the standard of care for pts with PD-L1 positive tumors; still, most pts progress within a year, highlighting a need for new treatments. BEGONIA is an ongoing Simon 2-stage, multicenter, multi-arm platform study evaluating the safety and efficacy of durvalumab (D), an anti–PD-L1 monoclonal antibody, with or without paclitaxel, in combination with novel oncology therapies as first-line treatment for a/mTNBC (NCT03742102). Datopotamab deruxtecan (Dato-DXd) is an antibody-drug conjugate (ADC) consisting of a humanized anti-trophoblast cell surface antigen 2 (TROP2) IgG1 monoclonal antibody linked to a topoisomerase I inhibitor payload via a plasma stable, tumor selective, tetrapeptide-based cleavable linker. TROP2 is broadly expressed on breast and other epithelial tumors and Dato-DXd monotherapy showed antitumor activity in heavily pretreated mTNBC pts (Bardia SABCS 2022 P6-10-03). Pts treated with Dato-DXd+D in Arm 7 of BEGONIA showed a 74% objective response rate (ORR) with median 7.2 months follow-up (Schmid SABCS 2022 PD11-09). Although responses were observed in both PD-L1–high and –low-expressing tumors, most pts (87%) had PD-L1–low-expressing tumors. In Arm 8 of BEGONIA, pts determined to have PD-L1 positive tumors by pre-existing/local testing will be recruited and treated with Dato-DXd+D to evaluate efficacy and safety in a PD-L1–high population. Methods: Eligible female pts are aged ≥18 years with untreated unresectable, locally advanced or mTNBC; ≥6 months between completion of treatment for earlier-stage breast cancer and recurrence of distant disease; ≥12 months since prior taxane therapy; ECOG PS 0/1; adequate organ function; and ≥1 nonirradiated measurable lesion. For inclusion in Arm 8, a PD-L1 positive tumor as determined by local immunohistochemistry (IHC) testing is required (either pre-existing or obtained during prescreening). Exclusion criteria for Arm 8 are clinically significant corneal disease; history or suspicion of interstitial lung disease/pneumonitis; underlying pulmonary disorder; prior exposure to immune-mediated therapy; or prior treatment with an ADC containing a topoisomerase I inhibitor. Arm 8 will evaluate Dato-DXd (6 mg/kg) + D (1120 mg) given intravenously every 3 weeks until disease progression or unacceptable toxicity. Tumors will be assessed per RECIST v1.1 every 6 weeks for 48 weeks, then every 12 weeks thereafter. A safety run-in will not occur for Arm 8, as Dato-DXd+D was evaluated in Arm 7 and was tolerable with no DLTs reported. Part 1 of Arm 8 will enroll 30 pts. The primary endpoint of Part 1 is safety and tolerability. A futility analysis will be performed with an option to expand Arm 8 to Part 2, where an additional 27 pts will be enrolled if expansion criteria are met. The primary endpoint for Part 2 is ORR. Secondary endpoints include ORR (Part 1 only), testing for antidrug antibodies and treatment pharmacokinetics (Part 1 only), duration of response (DoR), progression-free survival (PFS), and overall survival (OS). ORR will be summarized with descriptive statistics and 95% Clopper-Pearson confidence intervals. Kaplan-Meier analysis will be used for DoR, PFS, and OS. All tumor response and survival endpoints will be based on investigator assessments. Additional PD-L1 testing with the VENTANA PD-L1 (SP263) Assay will be performed on pretreatment tumor samples; high expression is defined as ≥10% of the tumor area populated by PD-L1–expressing tumor or immune cells. TROP2 expression will be assessed by IHC. Enrollment is ongoing. Funding: AstraZeneca/Daiichi Sankyo Citation Format: Peter Schmid, Cynthia Ma, Robert Huisden, Ross Stewart, Katrin Heider, Petra Vuković, Neelima Denduluri. Durvalumab + datopotamab deruxtecan in patients with PD-L1 positive advanced/metastatic triple-negative breast cancer: Arm 8 of the phase 1b/2, open label, platform BEGONIA study [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO1-19-10.

Hypofractionated radiotherapy combined with lenalidomide improves systemic antitumor activity in mouse solid tumor models.

Background: Hypofractionated radiotherapy (hRT) can induce a T cell-mediated abscopal effect on non-irradiated tumor lesions, especially in combination with immune checkpoint blockade (ICB). However, clinically, this effect is still rare, and ICB-mediated adverse events are common. Lenalidomide (lena) is an anti-angiogenic and immunomodulatory drug used in the treatment of hematologic malignancies. We here investigated in solid tumor models whether lena can enhance the abscopal effect in double combination with hRT. Methods: In two syngeneic bilateral tumor models (B16-CD133 melanoma and MC38 colon carcinoma), the primary tumor was treated with hRT. Lena was given daily for 3 weeks. Besides tumor size and survival, the dependence of the antitumor effects on CD8+ cells, type-I IFN signaling, and T cell costimulation was determined with depleting or blocking antibodies. Tumor-specific CD8+ T cells were quantified, and their differentiation and effector status were characterized by multicolor flow cytometry using MHC-I tetramers and various antibodies. In addition, dendritic cell (DC)-mediated tumor antigen cross-presentation in vitro and directly ex vivo and the composition of tumor-associated vascular endothelial cells were investigated. Results: In both tumor models, the hRT/lena double combination induced a significant abscopal effect. Control of the non-irradiated secondary tumor and survival were considerably better than with the respective monotherapies. The abscopal effect was strongly dependent on CD8+ cells and associated with an increase in tumor-specific CD8+ T cells in the non-irradiated tumor and its draining lymph nodes. Additionally, we found more tumor-specific T cells with a stem-like (TCF1+ TIM3- PD1+) and a transitory (TCF1- TIM3+ CD101- PD1+) exhausted phenotype and more expressing effector molecules such as GzmB, IFNγ, and TNFα. Moreover, in the non-irradiated tumor, hRT/lena treatment also increased DCs cross-presenting a tumor model antigen. Blocking type-I IFN signaling, which is essential for cross-presentation, completely abrogated the abscopal effect. A gene expression analysis of bone marrow-derived DCs revealed that lena augmented the expression of IFN response genes and genes associated with differentiation, maturation (including CD70, CD83, and CD86), migration to lymph nodes, and T cell activation. Flow cytometry confirmed an increase in CD70+ CD83+ CD86+ DCs in both irradiated and abscopal tumors. Moreover, the hRT/lena-induced abscopal effect was diminished when these costimulatory molecules were blocked simultaneously using antibodies. In line with the enhanced infiltration by DCs and tumor-specific CD8+ T cells, including more stem-like cells, hRT/lena also increased tumor-associated high endothelial cells (TA-HECs) in the non-irradiated tumor. Conclusions: We demonstrate that lena can augment the hRT-induced abscopal effect in mouse solid tumor models in a CD8 T cell- and IFN-I-dependent manner, correlating with enhanced anti-tumor CD8 T cell immunity, DC cross-presentation, and TA-HEC numbers. Our findings may be helpful for the planning of clinical trials in (oligo)metastatic patients.

Annotation of CD8+ T-cell function via ICAM-1 imaging identifies FAK inhibition as an adjuvant to augment the antitumor immunity of radiotherapy.

Background: Radiotherapy (RT) may trigger systemic antitumor immunity, manifesting as regression of non-irradiated lesions (abscopal effect). Intracellular adhesion molecule-1 (ICAM-1) is a key molecule involved in the abscopal effect of RT. However, the specific function of ICAM-1 in CD8+ T cells during antitumor immune responses remains unclear. Herein, we investigated whether noninvasive imaging of ICAM-1 can be used to annotate CD8+ T-cell function, thereby better selecting combinational therapy to enhance the antitumor immunity induced by RT. Methods: Using knockout mouse models, we investigated the role of ICAM-1 expressed on CD8+ T cells in the antitumor immunity of RT and conducted drug screening guided by ICAM-1-targeted noninvasive imaging. Results: The systemic antitumor effect of RT relies on the expression of ICAM-1 on CD8+ T cells. ICAM-1 expression is essential for CD8+ T-cell activation, proliferation, and effector function. Noninvasive annotation of the proliferation and effector function of CD8+ T cells by ICAM-1-targeted imaging identified VS-6063, a focal adhesion kinase inhibitor, as a new adjuvant to augment systemic antitumor immunity of RT in an immunologically "cold" tumor model. Mechanistically, VS-6063 overcomes the physical barriers in tumors and promotes the migration and infiltration of CD8+ T cells primed by RT into distant tumors. Conclusion: Our findings highlight that molecular imaging of ICAM-1 levels provides a dynamic readout of the proliferation and effector function of tumor-infiltrating CD8+ T cells, which facilitates the high-throughput exploitation of new combinational drugs to maximize the systemic antitumor effect of RT.

Solid tumours showing oligoprogression to immune checkpoint inhibitors have the potential for abscopal effects

PurposeGiven the uncertainty surrounding the abscopal effect (AE), it is imperative to identify promising treatment targets. In this study, we aimed to explore the incidence of AE when administering radiotherapy to patients with oligoprogressive solid tumours while they are undergoing treatment with immune checkpoint inhibitors (ICIs).Materials and methodsIn this multicentre prospective observational study, oligoprogressive disease was defined as a < 20% increase in lesions compared to > 2 months before enrolment. We enrolled patients who requested radiotherapy during the ICI rest period between 2020 and 2023. AE was considered present if ≥ 1 non-irradiated lesion decreased by ≥ 30% before the next line of systemic therapy started.ResultsTwelve patients were included in this study; the common primary lesions were in the lungs (four patients) and kidneys (three patients). AEs were observed in six (50%) patients, with a median time to onset of 4 (range 2–9) months after radiotherapy. No significant predictors of AEs were identified. Patients in the AE group had a significantly better 1-year progression-free survival (PFS) rate than those in the non-AE group (p = 0.008). Two patients from the AE group were untreated and progression-free at the last follow-up. Four (33%) patients experienced grade 2 toxicity, with two cases attributed to radiotherapy and the other two to ICI treatment. No grade 3 or higher toxicities were observed in any category.ConclusionPatients with oligoprogressive disease may be promising targets with potential for AEs. AEs can lead to improved PFS and, in rare cases, to a certain progression-free period without treatment.Secondary Irradiating solid tumours in patients with oligoprogressive disease during immune checkpoint inhibitor therapy may be a promising target with the potential for abscopal effects (AEs). AEs can lead to improved progression-free survival and, in rare cases, to a certain progression-free period without treatment.

Radiotherapy with or without immunotherapy in metastatic melanoma: efficacy and tolerability

Introduction Radiotherapy (RT) is primarily considered as a palliative treatment in patients with metastatic melanoma. However, observations suggest that when RT is combined with immune checkpoint inhibitors (ICI), it can induce an immune response leading to an anti-tumoral effect also distant from the irradiated area – a phenomenon called ‘abscopal effect’. The frequency and circumstances of abscopal effect among metastatic melanoma patients remains uncertain and further research is necessary. Material and method This retrospective study included all metastatic melanoma patients who received non-stereotactic RT in Stockholm, Sweden in 2015–2020. Patients were grouped depending on if RT was given at start of ICI (RT + ICI(start)), at ICI progression (RT + ICI(salvage)) or without ICI (RT(only)). Response rates in irradiated (RR(irradiated)) and overall response rates in non-irradiated (ORR(non-irradiated)) metastases were evaluated together with survival and toxicity in each cohort. Results In the RT + ICI(start) (n = 47), RT + ICI(salvage) (n = 41) and RT(only) (n = 55) cohorts, RR(irradiated) was 70.7%, 67.5% and 43.1% (p = 0.018) while the ORR(non-irradiated) was 36.1%, 14.8% and 0.0% (p = 0.003), and the median overall survival was 18.2, 15.0 and 7.2 months, respectively (p = 0.014). Local response to RT was in all cohorts associated with longer survival (p < 0.001). The frequency of grade ≥3 immune-related adverse events was 17.0% and 19.5% in the RT + ICI(start) and RT + ICI(salvage) cohorts. No increased frequency of RT-related adverse events was seen in the RT + ICI cohorts, compared to the RT(only) cohort. Conclusion This retrospective study showed that melanoma patients receiving RT in combination with ICI had a superior antitumoral response in both irradiated and non-irradiated lesions as compared to patients receiving only RT. Additionally, a subgroup of patients receiving RT when progressing on ICI experienced tumor regression also in non-irradiated areas.

Doxorubicin Enhances the Abscopal Effect Depending on Tumor Cell Mitochondrial DNA and cGAS/STING

Localized radiotherapy (RT) can cause a T cell-mediated abscopal effect on non-irradiated tumor lesions, especially in combination with immune checkpoint blockade (ICB). However, this effect is still clinically rare and improvements are highly desirable. We investigated whether triple combination with a low dose of clinically approved liposomal doxorubicin (Doxil) could augment abscopal responses compared with RT+ICB, Doxil+ICB, or RT+Doxil. We used Doxil in combination with RT and αPD1 in two tumor models (B16-CD133 melanoma and MC38 colon carcinoma) with mice bearing two tumors, only one of which was irradiated. Triple therapy with RT, αPD1, and single low-dose Doxil strongly enhanced the RT-induced abscopal effect compared to all double and single treatments in both tumor models (p < 0.05, n = 5-10 mice/group). Complete cures of non-irradiated tumors were mainly observed in triple-treated mice. Triple therapy induced more cross-presenting dendritic cells (DCs) and tumor-specific CD8 T cells than RT/αPD1 and Doxil/αPD1 (p < 0.05, n = 5 mice/group), particularly in non-irradiated tumors. CD8 T cell depletion or implanting STING-deficient tumor cells abolished the abscopal effect. By using inhibitors and knockout cells, we show that doxorubicin/Doxil-induced IFNβ1 markedly depended on the cGAS/STING pathway (p < 0.05) which drives antitumor CD8 T cell responses through cross-presenting DCs. In mitochondrial DNA (mtDNA)-depleted tumor cells, doxorubicin/Doxil induced less IFNβ1 (p < 0.05), the related T cell-recruiting chemokine CXCL10 (p < 0.0001), and ATP (p < 0.0001); coincubation with mtDNA-depleted tumor cells strongly reduced IFNβ1 (p < 0.01) secretion by DCs. Implantation of mtDNA-depleted tumor cells, particularly at the non-irradiated site, substantially diminished the Doxil-enhanced abscopal effect and tumor infiltration by tumor-specific CD8 T cells (p < 0.05). Single low-dose Doxil can substantially enhance the RT-induced abscopal effect, with a strong increase in cross-presenting DCs and CD8 tumor-specific T cells particularly in abscopal tumors compared with RT/αPD1 and Doxil/αPD1. The mtDNA/cGAS/STING/IFN-I axis is important for the immunogenic doxorubicin effects. Our findings may be helpful for the planning of clinical radiochemoimmunotherapy trials in (oligo)metastatic patients.

Comparison of Two Triple Therapy Regimens for Enhancing the Abscopal Effect in Mice

Localized radiotherapy (RT) can cause a T cell-mediated abscopal effect on non-irradiated tumor lesions, especially in combination with immune checkpoint blockade (ICB). However, this effect is still clinically rare and improvements are highly desirable. We here compared two triple therapy regimens (RT+⍺PD1+⍺CTLA4 vs. RT+⍺PD1+IL2/⍺IL2 complexes (IL2c)) for their efficacy to improve systemic (abscopal) anti-tumor effects. In mice bearing bilateral subcutaneous C51 colon carcinoma tumors, the primary tumor was irradiated with two fractions of 8 Gy. ⍺PD1 and ⍺CTLA4 were given concomitantly and weekly thereafter; IL2c was given i.p. for 3 consecutive days. Besides tumor size and survival, tumor-specific CD8+ T cells were determined flow cytometrically using MHC-I tetramers and various antibodies. In addition, isolated TILs were cultured with PMA, ionomycin, and Brefeldin A in vitro to assess polyfunctionality (based on cytokine production) of CD8+ and CD4+ TILs. The abscopal effect was significantly stronger in mice treated with the ⍺CTLA4-containing triple combination (n = 13) than in mice treated with the IL2c-containing triple combination (n = 9) (p<0,05), and compared to mice treated with RT+⍺PD1 (n = 8) (p<0,05) or RT+⍺CTLA4 (n = 10) (p<0,05), respectively. The IL2c triple combination induced the abscopal effect better than RT+⍺PD1 (p<0,05). The ⍺CTLA4 triple therapy improved survival and resulted in complete cures of 8/13 mice. In mice treated with ⍺CTLA4-containing triple therapy, control of the irradiated tumor was partially dependent and that of the non-irradiated tumor was strongly dependent on T cells, primarily CD8 T cells but also CD4 T cells. With ⍺CTLA4 triple treatment (n = 9), the frequency and absolute numbers of polyfunctional TNF-⍺+IFN-γ+ AH-1 tumor-specific CD8+ T cells, IFN-γ+IL2+ AH-1 tumor-specific CD8+ T cells, TNF-⍺+IFN-γ+ CD4+ and IFN-γ+IL2+ CD4+ effector T cells were higher than with the IL2c triple combination (n = 9) (p<0,05), particularly in the non-irradiated tumors. RT+⍺PD1+⍺CTLA4 triple treatment induced more cytotoxic effector TILs than RT+⍺PD1+IL2c triple treatment, enhancing the abscopal effect and inducing a high abscopal cure rate. More experiments underway to reveal mechanisms.

Abstract PR05: Changing the radiation and immune-oncology paradigm in patients with head and neck squamous cell carcinoma (HNSCC) with the radioenhancer NBTXR3: From bench to bedside

Abstract Purpose/Objectives: Immune checkpoint inhibitors (ICIs) have revolutionized cancer treatment, yet most patients (pts) ultimately develop ICI resistance. Overcoming this resistance is a major clinical challenge. Radiation therapy (RT) has emerged as a promising combination with ICIs since in some settings it may produce an immunomodulatory effect. NBTXR3, a novel radioenhancer composed of functionalized hafnium oxide nanoparticles, is administered by a single intratumoral injection. NBTXR3 increases RT energy deposition inside tumor cells, subsequent tumor cell death, and tumor antigen release and thus in combination with ICI may overcome resistance. Materials/Methods: Pre-Clinical: Abscopal assays were conducted in immunocompetent mice. Anti-PD-1 sensitive or resistant lung tumor cell lines were injected in both legs. Intratumoral NBTXR3 injection (or vehicle) followed by RT was performed in right leg (primary) tumors only. Some mice also received anti-PD-1 injections alone or in combination with anti-LAG3 and anti-TIGIT. Tumor growth was monitored, and immune cell infiltrates analyzed by IHC. Clinical: A multicenter phase I trial [NCT01946867] is evaluating NBTXR3/RT in pts with AJCC 8th stage III-IVA or T3, T4 SCC of the oral cavity or oropharynx ineligible to cisplatin. Pts received NBTXR3 at the recommended dose (22% of baseline tumor volume) followed by RT (IMRT 70 Gy in 35 fractions). A multicenter phase I trial [NCT03589339] is evaluating NBTXR3/RT/anti-PD-1 in 3 cohorts of pts with advanced solid tumors eligible for anti-PD-1: 1) Locoregional recurrent or recurrent and metastatic HNSCC, 2) Lung metastases, or 3) Liver metastases. NBTXR3 was administered by intratumoral injection. Stereotactic Body RT (SBRT) was delivered at tumor site-specific doses per standard practice. 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. NBTXR3/RT/anti-PD-1 improved local and systemic control in mice bearing anti-PD-1 resistant tumors, produced long-term memory, and reduced spontaneous lung metastases. NBTXR3/RT/anti-PD-1/anti-LAG3/anti-TIGIT improved tumor control, extended survival and produced a robust long-term memory anti-tumor immune response. Clinically, NBTXR3 intratumoral injection was feasible and safe. Preliminary signs of efficacy observed in pts with locally advanced HNSCC treated with NBTXR3/RT will be presented. Preliminary signs of efficacy were observed in pts treated with NBTXR3/RT/anti-PD-1 including in pts resistant to anti-PD-1, as well as in remote non-irradiated lesions. Responses in pts with HNSCC will be presented. Conclusions: With demonstrated radioenhancement as well as immune system priming, NBTXR3 positions itself to be a paradigm shift catalyst in the treatment of pts with head and neck cancers. These data support further development of NBTXR3/RT alone as well as in combination with anti-PD-1 and/or other ICIs. Citation Format: Colette J. Shen, Christophe Le Tourneau, Zoltán Takacsi-Nagy, Xavier Liem, Jessica Frakes, Jiaxin Niu, Jared Weiss, Jimmy Caudell, Yun Hu, James W. Welsh, Sebastien Paris, Anais Debard, Patricia Said, Pavel Tyan, Tanguy Y. Seiwert, Zsuzsanna Papai, Omar I. Vivar, Leonard A. Farber, Ari Rosenberg. Changing the radiation and immune-oncology paradigm in patients with head and neck squamous cell carcinoma (HNSCC) with the radioenhancer NBTXR3: From bench to bedside [abstract]. In: Proceedings of the AACR-AHNS Head and Neck Cancer Conference: Innovating through Basic, Clinical, and Translational Research; 2023 Jul 7-8; Montreal, QC, Canada. Philadelphia (PA): AACR; Clin Cancer Res 2023;29(18_Suppl):Abstract nr PR05.

Phase I/II clinical trial of nivolumab in combination with oligo-fractionated irradiation for unresectable advanced or recurrent gastric cancer

BackgroundAlthough immune checkpoint inhibitors (ICI) targeting for PD-1 axis is a promising approach for advanced gastric cancer (GC) patients, the response rate is still limited. Induction of synergistic effect of irradiation with ICI targeting for the PD-1 axis can be an attractive strategy. The aim of this study was to assess the effect of the combination of irradiation with anti-PD-1 therapy for advanced GC.MethodsWe conducted a single-arm, phase I/II trial in GC patients treated with a combination of nivolumab and oligo-fractionated irradiation (22.5 Gy/5 fractions/5 days) (NCT03453164). Eligible patients (n = 40) had unresectable advanced or recurrent GC which progressed after primary and secondary chemotherapy with more than one lesion. The primary endpoint is the disease control rate (DCR) of non-irradiated target lesions and the secondary endpoints are the median survival time (MST), safety, and DCR of irradiated lesions.ResultsWe observe that the DCR for the non-irradiated target as the abscopal effect is 22.5% (90% confidence interval (CI), 12.3–36.0), and the DCR for the irradiated lesion is 40.0% (90% CI, 26.9–54.2). The median survival time is 230 days (95% CI, 157–330), and grade 3 and higher adverse events (AEs) are observed in 16 patients (39 %) with no obvious additional AEs when adding irradiation.ConclusionsThe present study suggests that the combination of nivolumab with oligo-fractionated irradiation has the potential to induce a promising anti-tumor effect for advanced GC.

Adding liposomal doxorubicin enhances the abscopal effect induced by radiation/αPD1 therapy depending on tumor cell mitochondrial DNA and cGAS/STING

BackgroundLocalized radiotherapy (RT) can cause a T cell-mediated abscopal effect on non-irradiated tumor lesions, especially in combination with immune checkpoint blockade. However, this effect is still clinically rare and improvements...

Doxorubicin to enhance the abscopal effect depending on tumor cell mitochondrial DNA and cGAS/STING.

e14616 Background: Localized radiotherapy (RT) can cause a T cell-mediated abscopal effect on non-irradiated tumor lesions, especially in combination with immune checkpoint blockade (ICB). However, this effect is still clinically rare and improvements are highly desirable. We investigated whether triple combination with a low dose of clinically approved liposomal doxorubicin (Doxil) could augment abscopal responses compared with RT+ICB, Doxil+ICB, or RT+Doxil. Methods: We used Doxil in combination with RT and αPD1 in two tumor models (B16-CD133 melanoma and MC38 colon carcinoma) with mice bearing two tumors, only one of which was irradiated. Results: Triple therapy with RT, αPD1, and single low-dose Doxil strongly enhanced the RT-induced abscopal effect compared to all double and single treatments in both tumor models ( p &lt; 0.05, n = 5-10 mice/group). Complete cures of non-irradiated tumors were mainly observed in triple-treated mice. Triple therapy induced more cross-presenting dendritic cells (DCs) and tumor-specific CD8 T cells than RT/αPD1 and Doxil/αPD1 ( p &lt; 0.05, n = 5 mice/group), particularly in non-irradiated tumors. CD8 T cell depletion or implanting STING-deficient tumor cells abolished the abscopal effect. By using inhibitors and knockout cells, we show that doxorubicin/Doxil-induced IFNβ1 markedly depended on the cGAS/STING pathway ( p &lt; 0.05) which drives antitumor CD8 T cell responses through cross-presenting DCs. In mitochondrial DNA (mtDNA)-depleted tumor cells, doxorubicin/Doxil induced less IFNβ1 ( p &lt; 0.05), the related T cell-recruiting chemokine CXCL10 ( p &lt; 0.0001), and ATP ( p &lt; 0.0001); coincubation with mtDNA-depleted tumor cells strongly reduced IFNβ1 ( p &lt; 0.01) secretion by DCs. Implantation of mtDNA-depleted tumor cells, particularly at the non-irradiated site, substantially diminished the Doxil-enhanced abscopal effect and tumor infiltration by tumor-specific CD8 T cells ( p &lt; 0.05). Conclusions: Single low-dose Doxil can substantially enhance the RT-induced abscopal effect, with a strong increase in cross-presenting DCs and CD8 tumor-specific T cells particularly in abscopal tumors compared with RT/αPD1 and Doxil/αPD1. The mtDNA/cGAS/STING/IFN-I axis is important for the immunogenic doxorubicin effects. Our findings may be helpful for the planning of clinical radiochemoimmunotherapy trials in (oligo)metastatic patients.

Abstract OT2-10-03: HCRN BRE 19-433: A Multi-institutional Phase II Study to Evaluate Efficacy and Safety of TAlazoparib, Radiotherapy and Atezolizumab in gBRCA 1/2 negative Patients with PD-L1+ Metastatic Triple Negative Breast Cancer (TARA)

Abstract Background Although immunotherapy (IO) in combination with chemotherapy has improved progression free and overall survival in patients with PD-L1+ metastatic triple negative breast cancer (mTNBC), prognosis remains poor. A potential therapeutic strategy to restore sensitivity to IO in patients with progressive disease is to introduce agents that re-sensitize the immune system to IO leading to a more tumor-specific and less toxic treatment in the second-line setting or beyond. Both talazoparib, a PARP inhibitor, and radiation (XRT) independently increase PD-L1 expression on the tumor cell surface resulting in heightened sensitivity to IO agents like atezolizumab, a PD-L1 inhibitor. Although a local treatment, XRT has the ability to produce an abscopal effect resulting in systemic shrinkage of non-irradiated tumors outside/distant to the XRT field, a phenomenon observed in patients receiving concurrent IO. While talazoparib is standard treatment in patients with gBRCA1 and 2 mutations, it is also a potent radiosensitizing agent that suppresses homologous recombination and PARP-1-dependent nonhomologous end joining (NHEJ) repair while promoting error-prone alt-NHEJ. When combined with IO, talazoparib can amplify immune responses by generating immunogenic neo-antigens independent of gBRCA1/2 status. We, therefore, hypothesize that the combination of talazoparib, XRT, and atezolizumab will re-sensitize mTNBC tumors to IO and promote a durable tumor-specific response that spares patients from toxicities associated with traditional chemotherapy regimens. Methods This is a Phase II multi-institutional study designed to assess efficacy and safety of talazoparib, high dose XRT, and atezolizumab given in the second-, third-, or fourth-line settings to patients with mTNBC that is PD-L1 positive. A total of 23 patients with mTNBC who do not carry gBRCA pathogenic variants will be enrolled. All patients will be treated with induction talazoparib of 1mg PO daily starting Day 1 of a 28-day cycle. Patients will then receive 8 Gy x 3 fractions to 1-4 metastatic lesions QOD beginning Day 12, 13, or 14. Atezolizumab will be given intravenously (840 mg)) on Day 15 of the 1st cycle and then on Day 1 and Day 15 of the subsequent cycles. Talazoparib and atezolizumab treatment will continue until progression or severe toxicity. The primary endpoint is objective response rate (ORR) in non-irradiated lesions 8 weeks after the first dose of atezolizumab. Key inclusion criteria include biopsy proven mTNBC (ER&amp;lt; 10%, PR&amp;lt; 10%, Her2-) with at least 2 extracranial metastatic lesions. Patients must have at least 1 extracranial metastatic lesion amenable to high dose radiotherapy and at least one additional extracranial lesion of measurable disease by Response Evaluation Criteria in Solid Tumors (RECIST v1.1) that will not receive radiotherapy. Tumors must be PD-L1 positive as defined as &amp;gt;1% on IHC using the SP142 Ventana Assay. Key exclusion criteria include patients with germline BRCA pathogenic variants, more than three previous lines of chemotherapy treatment in the advanced setting with or without IO, and breast cancer progression within the first 3 months of previous IO treatment for non-metastatic or metastatic breast cancer. Sample size was determined using Simon’s 2-stage Minimax design to detect a 20% increase in ORR. The null hypothesis that the true response rate among gBRCA1/2 negative patients of 10% will be tested against a one-sided alternative. Inflammatory cytokines, circulating B cells, and ctDNA will be collected for correlative analysis. Enrollment began in April 2021. The study is managed by the Hoosier Cancer Research Network and is open to accrual at Emory University and University of Alabama at Birmingham. Clinical trial information: NCT04690855 Citation Format: Mylin A. Torres, Kevin Kalinsky, Erica Stringer-Reasor, Ahmed Elkhanany, Jolinta Lin, David M. Schuster, Sarah Friend, Jeffrey Switchenko, Manali Bhave. HCRN BRE 19-433: A Multi-institutional Phase II Study to Evaluate Efficacy and Safety of TAlazoparib, Radiotherapy and Atezolizumab in gBRCA 1/2 negative Patients with PD-L1+ Metastatic Triple Negative Breast Cancer (TARA) [abstract]. In: Proceedings of the 2022 San Antonio Breast Cancer Symposium; 2022 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2023;83(5 Suppl):Abstract nr OT2-10-03.

Combination treatment of intratumoral vidutolimod (CMP-001), radiosurgery, nivolumab, and ipilimumab for metastatic colorectal carcinoma.

123 Background: Microsatellite stable (MSS) or mismatch repair proficient (MMR-P) metastatic colorectal cancer is refractory to immune checkpoint inhibition. We hypothesized that a combination of intratumoral TLR9 agonist, radiosurgery and dual PD-1 and CTLA-4 blockade would induce a local focus of immune stimulation, evoking a systemic immune response. Methods: In this single institution investigator-initiated phase I study, patients with MSS / MMR-P metastatic colorectal cancer were treated with a priming dose of subcutaneous (s.c.) vidutolimod, three intratumoral injections of vidutolimod, and radiosurgery to a metastasis, combined with nivolumab 3mg/kg every 2 weeks and ipilimumab 1 mg/kg every 6 weeks. Following conclusion of intratumoral therapy, s.c. injections of vidutolimod commenced. Efficacy endpoints were based upon a non-irradiated and non-injected lesion. Cytokine levels were measured at baseline and at 7 (± 2) weeks. Patients were placed in 4 cohorts: (1) Safety run-in without radiosurgery (No RT); (2) Radiosurgery prior to intratumoral therapy (Early Liver RT Slow). An amendment was made due to early patient fall-out due to progressive disease; (3) Radiosurgery prior to intratumoral therapy with a condensed timeline (Early Liver RT Quick). An additional amendment was made due to unacceptable liver toxicity; (4) Radiosurgery to extrahepatic lesion following completion of intratumoral therapy (Late Extrahepatic RT). ClinicalTrials.gov Identifier NCT03507699. Results: A total of 19 patients were accrued (No RT, n=2; Early Liver RT Slow, n=7; Early Liver RT Quick, n=4; Late Extrahepatic RT, n=6). Median age was 59 years (range 40-71), 68% were male, median number of previous systemic treatments was 3 (range 2 – 5). None of the patients responded, aside from one patient in cohort #4 who achieved complete response, and was subsequently found to have a high tumor mutational burden (TMB) 79 mutations / Mb. Grade 3 liver toxicity, including elevated transaminases and hyperbilirubinemia, was reported in 0%, 0%, 75% and 17% in cohorts 1-4, respectively. Reactions following intratumoral injections, typically within 12 hours, included fever (68%), tachycardia (21%), chills (63%) and hypotension (47%). There was an increase in systemic CXCL10 levels at 7 (± 2) weeks compared to baseline, with a median of 407 versus 78 pg/ml, respectively, p&lt;0.01. Conclusions: Intratumoral liver injection of vidutolimod was associated with acute systemic symptoms attributed to cytokine release. The juxtaposition of liver irradiation and intratumoral vidutolimod injection to the same lesion was associated with unacceptable hepatic toxicity. The combination of intratumoral vidutolimod, radiosurgery, nivolumab and ipilimumab was not found to be efficacious in MSS / MMR-P colon cancer. Clinical trial information: NCT03507699 .

Abstract C012: Dissecting the reorganization of pancreatic tumor microenvironments after radiation and immunotherapy reveals insights into immunotherapy resistance

Abstract Immune checkpoint blockade (ICB) has revolutionized the treatment of many cancers but has been ineffective for the treatment of microsatellite stable (MSS) PDAC. The lack of efficacy of immunotherapies in PDAC is due to: 1) a desmoplastic tumor microenvironment (TME); 2) the presence of suppressive cells, including myeloid derived suppressor cells and regulatory T cells; and 3) the lack of antigen-presenting dendritic cells (DCs) that are important in priming an effective immune response to generate functionally effective tumor antigen-specific T cells. We recently completed a pilot study of dual ICB (Ipilumamab and Nivolumab) with radiation therapy (SBRT 8Gy for 3 fractions) in a cohort of 25 metastatic PDAC patients that had progressed on conventional chemotherapy; this combination conferred an impressive 18% ORR and 29% disease control rate measured on non-irradiated lesions (historical 0% ORR with ICB in PDAC). This led to a phase 2 study in 30 metastatic PDAC patients using this dual modality treatment paradigm. To understand the role of radiation and ICB in altering the PDAC tumor microenvironment we performed single-cell RNA-sequencing and TCR-sequencing (&amp;gt;180k cells), and single-nucleus RNA-sequencing (&amp;gt;300k cells) on 36 tumor biopsies (23 pre-treatment, 13 paired on-treatment between day 10 and 21) from patients undergoing treatment in our phase 2 study. Tumor tissue was taken from distinct tissue sites, including primary tumors in the pancreas, and liver and abdominal wall metastases. We identified distinct tumor cell state distributions within different tissues, and a redistribution of cells from basal/mesenchymal states to classical states after radiation. We identified several state-specific interferon stimulated gene programs thus cataloging distinct responses of epithelial cells with different transcriptional states. Importantly, we found a redistribution of T cells states towards proliferating and exhausted T cells with unique clonality after radiation. Additionally, the myeloid compartment after radiation was enriched for C1QC+ and MHCII+ macrophage subsets, as well as infiltrating CD16/CD16 monocytes and CD14 monocytes, each showing induction of unique sets of interferon stimulated genes (ISGs). We next sought to better understand immunotherapy resistance mechanisms within these PDAC patients despite finding strong ISG induction in several subsets. We analyzed covarying gene programs and identified multicellular communities of cells before and after radiation that underlie interaction networks associated with radiation. Together our data provides the most comprehensive single-cell atlas of paired biopsies to study tumor and immune cell states in the context of radiation and ICB response. Citation Format: Arnav Mehta, Aparna Parikh, Milan Parikh, Ryan Park, Moshe Sade-Feldman, Lynn Bi, Nicole Carzo, Tarin M. Grillo, Islam Baiev, Olanike Asupoto, Irena Gushterova, Tom LaSalle, Anna Gonye, Emily Blaum, Sebastien Vigneau, Ronan Chaligne, Ana Lako, Thomas Lila, David Nelson, Caroline Porter, Orr Ashenberg, Karthik Jagadesh, William L. Hwang, Christopher Smillie, David P. Ryan, David T. Ting, Theodore Hong, Dana Pe'er, Nir Hacohen. Dissecting the reorganization of pancreatic tumor microenvironments after radiation and immunotherapy reveals insights into immunotherapy resistance [abstract]. In: Proceedings of the AACR Special Conference on Pancreatic Cancer; 2022 Sep 13-16; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2022;82(22 Suppl):Abstract nr C012.

90Y-Labeled Gold Nanoparticle Depot (NPD) Combined with Anti-PD-L1 Antibodies Strongly Inhibits the Growth of 4T1 Tumors in Immunocompetent Mice and Induces an Abscopal Effect on a Distant Non-Irradiated Tumor.

The effectiveness and normal tissue toxicity of a novel nanoparticle depot (NPD) brachytherapy seed incorporating gold nanoparticles (AuNPs) labeled with β-particle emitting, 90Y (termed a "radiation nanomedicine"), were studied for the treatment of 4T1 triple-negative murine mammary carcinoma tumors in Balb/c mice and for inducing an abscopal effect on a distant non-irradiated tumor alone or combined with anti-PD-L1 immune checkpoint antibodies. Balb/c mice with two subcutaneous 4T1 tumors─a primary tumor and a distant secondary tumor were implanted intratumorally (i.t.) in the primary tumor with NPD incorporating 3.5 MBq of 90Y-AuNPs (1 × 1014 AuNPs) or unlabeled AuNPs, alone or combined with systemically administered anti-PD-L1 antibodies (200 μg i.p. three times/week for 2 weeks) or received anti-PD-L1 antibodies alone or no treatment. The primary tumor was strongly growth-inhibited over 14 d by NPD incorporating 90Y-AuNPs but only very modestly inhibited by NPD incorporating unlabeled AuNPs. Anti-PD-L1 antibodies alone were ineffective, and combining anti-PD-L1 antibodies with NPD incorporating 90Y-AuNPs did not further inhibit the growth of the primary tumor. Secondary tumor growth was inhibited by treatment of the primary tumor with NPD incorporating 90Y-AuNPs, and growth inhibition was enhanced by anti-PD-L1 antibodies. Treatment of the primary tumor with NPD incorporating unlabeled AuNPs or anti-PD-L1 antibodies alone had no effect on secondary tumor growth. Biodistribution studies showed high uptake of 90Y in the primary tumor [516-810% implanted dose/g (%ID/g)] but very low uptake in the secondary tumor (0.033-0.16% ID/g) and in normal tissues (<0.5% ID/g) except for kidneys (5-8% ID/g). Very high radiation absorbed doses were estimated for the primary tumor (472 Gy) but very low doses in the secondary tumor (0.13 Gy). There was highdose-heterogeneity in the primary tumor with doses as high as 9964 Gy in close proximity to the NPD, decreasing rapidly with distance from the NPD. Normal organ doses were low (<1 Gy) except for kidneys (4 Gy). No normal tissue toxicity was observed, but white blood cell counts (WBC) decreased in tumor-bearing mice treated with NPD incorporating 90Y-AuNPs. Decreased WBC counts were interpreted as tumor response and not toxicity since these were higher than that in healthy non-tumor-bearing mice, and there was a direct association between WBC counts and 4T1 tumor burden. We conclude that implantation of NPD incorporating 90Y-AuNPs into a primary 4T1 tumor in Balb/c mice strongly inhibited tumor growth and combined with anti-PD-L1 antibodies induced an abscopal effect on a distant secondary tumor. This radiation nanomedicine is promising for the local treatment of triple-negative breast cancer tumors in patients, and these therapeutic effects may extend to non-irradiated lesions, especially when combined with checkpoint immunotherapy.

CAMRESBRT: Randomized Phase II Trial of Camrelizumab with Stereotactic Body Radiotherapy vs. Camrelizumab Alone in Recurrent or Metastatic Head and Neck Squamous Cell Carcinoma

<h3>Purpose/Objective(s)</h3> Immune checkpoint inhibitor (ICI) therapy has significantly expanded the therapeutic armamentarium for recurrent or metastatic (R/M) head and neck squamous cell carcinoma (HNSCC). Radiation stimulation of abscopal responses, combined with ICI, has been examined in single-arm trials with various tumor types, but randomized evaluation of nivolumab with stereotactic body radiotherapy (SBRT) in R/M HNSCC was frustrated. We aimed to test whether anti–PD-1 therapy, Camrelizumab, may act synergistically with SBRT to improve response through the abscopal effect. <h3>Materials/Methods</h3> We conducted a single-center, randomized, phase II trial. Patients older than 18 years of age with R/M HNSCC who have not been treated with ICIs, regardless of PD-L1 status, are eligible for this study. This trial planned to enroll approximately 70 patients. Patients were randomly assigned (1:1), stratified by human papillomavirus status and PD-L1 status, to camrelizumab versus camrelizumab plus SBRT. Patients have at least two metastatic lesions: one that could be safely irradiated and one measurable by RECIST version 1.1. Drug: camrelizumab (200mg intravenously every 2 weeks). Radiation: SBRT (27 Gy over 3 fractions given every other day) to single lesion to start by study day (study day 1 is day of first dose of camrelizumab). The primary end point was objective response rate (ORR) in nonirradiated lesions, which was assessed by RECIST 1.1. The secondary objectives were safety and tolerability (adverse events of grade 3-4 or higher, rate of long-term adverse events), 6-month and 1-year disease control rate (DCR), 6-month and 1-year progression free survival (PFS), overall survival (OS). <h3>Results</h3> As of January 2022, 31.4% of the planned 70 subjects have been enrolled. Median age was 54.5 years; 17 patients were male. 14 patients received camrelizumab alone and 8 patients received camrelizumab plus SBRT. The PD-L1 status was known in 14 of 18 patients, including 1 patient with PD-L1 CPS (combined positive score) less than 1, 3 patients more than 20, and 10 patients between 1-20. ClinicalTrials.gov Identifier: NCT04830267. <h3>Conclusion</h3> Clinical trial in progress.

Combination of SABR With Anti-PD-1 in Oligoprogressive Non-Small Cell Lung Cancer and Melanoma: Results of a Prospective Multicenter Observational Study

The percentage of patients with metastatic non-small cell lung cancer (NSCLC) and melanoma who benefit from anti-programmed cell death protein 1 (anti-PD-1) is low owing to resistance mechanisms. SABR has a role in oligoprogressive disease and can improve responses to anti-PD-1. This multicenter prospective observational study aimed to determine whether concomitant anti-PD-1 and SABR to oligoprogressive sites enhance tumor response in metastatic NSCLC and melanoma. Patients with metastatic NSCLC or melanoma in progression to anti-PD-1 but continuing the same line owing to clinical benefit were referred for palliative SABR. All patients received concomitant pembrolizumab or nivolumab and SABR to 1 to 5 lesions, maintaining anti-PD-1 until further progression, unacceptable toxicity, or medical/patient decision. Objective response rate-complete responses and partial responses-was evaluated during all follow-up according to Response Evaluation Criteria in Solid Tumors 1.1. The abscopal response was evaluated 8 weeks after SABR as a ≥30% reduction in 1 to 2 predefined nonirradiated lesions. Of the 61 patients enrolled, 50 could be analyzed. With a median follow-up of 32.8 months, objective response rate was 42% (30% complete responses and 12% partial responses). Median progression-free survival was 14.2 months (95% confidence interval, 6.9-29 months). Median overall survival since SABR was 37.4 months (95% confidence interval, 22.9 months-not reached). Abscopal response was 65%, evaluated in 40 patients who fulfilled the criteria. Combined anti-PD-1 and SABR in oligoprogressive metastatic NSCLC or melanoma can achieve high rates of response and extend the clinical benefit of immunotherapy by delaying further progression and a new systemic therapy. This approach should be assessed in larger randomized trials.

Promising outlook with sugemalimab in non-small-cell lung cancer.

Anti-PD-L1 (durvalumab) immunotherapy after chemotherapy with external beam radiotherapy (ie, the PACIFIC regimen) is standard care for patients with unresectable stage III non-small-cell lung cancer (NSCLC). In the PACIFIC trial, 1 Faivre-Finn C Vicente D Kurata T et al. Four-year survival with durvalumab after chemoradiotherapy in stage III NSCLC—an update from the PACIFIC trial. J Thorac Oncol. 2021; 16: 860-867 Summary Full Text Full Text PDF PubMed Scopus (153) Google Scholar this regimen showed durable progression-free survival, with the estimated number of randomly assigned patients who remained alive at 4 years being 50% in the group that received 12 months of anti-PD-L1 consolidation therapy using durvalumab following chemotherapy plus concurrent radiotherapy versus 36% in patients who received placebo alone. Preclinical studies have shown that radiotherapy can boost anti-tumour immune response and overcome resistance to current immunotherapies. Clinical studies have shown improved responsiveness to pembrolizumab (anti-PD-1 antibody) in metastatic NSCLC, and to durvalumab (anti-PD-L1 antibody) in operable NSCLC. In the PEMBRO-RT and MDACC trials, 2 Theelen W Chen D Verma V et al. Pembrolizumab with or without radiotherapy for metastatic non-small-cell lung cancer: a pooled analysis of two randomised trials. Lancet Respir Med. 2021; 9: 467-475 Summary Full Text Full Text PDF PubMed Scopus (98) Google Scholar the best abscopal response (for non-irradiated lesions) was 20% with pembrolizumab versus 42% with pembrolizumab plus radiotherapy (odds ratio 2·96, 95% CI 1·42–6·20; p=0·0039). Median progression-free survival was 4·4 months (95% CI 2·9–5·9) with pembrolizumab alone versus 9·0 months (6·8–11·2) with pembrolizumab plus radiotherapy (hazard ratio [HR] 0·67, 95% CI 0·45–0·99; p=0·045). 2 Theelen W Chen D Verma V et al. Pembrolizumab with or without radiotherapy for metastatic non-small-cell lung cancer: a pooled analysis of two randomised trials. Lancet Respir Med. 2021; 9: 467-475 Summary Full Text Full Text PDF PubMed Scopus (98) Google Scholar In a phase 2 trial comparing neoadjuvant durvalumab alone versus neoadjuvant durvalumab plus stereotactic radiotherapy in patients with resectable early-stage NSCLC (clinical stages I–IIIA), a major pathological response was observed in 7% of the durvalumab alone group versus 53% of the durvalumab plus radiotherapy group. 3 Altorki NK McGraw TE Borczuk AC et al. Neoadjuvant durvalumab with or without stereotactic body radiotherapy in patients with early-stage non-small-cell lung cancer: a single-centre, randomised phase 2 trial. Lancet Oncol. 2021; 22: 824-835 Summary Full Text Full Text PDF PubMed Scopus (59) Google Scholar Sugemalimab versus placebo, in combination with platinum-based chemotherapy, as first-line treatment of metastatic non-small-cell lung cancer (GEMSTONE-302): interim and final analyses of a double-blind, randomised, phase 3 clinical trialSugemalimab plus chemotherapy showed a statistically significant and clinically meaningful progression-free survival improvement compared with placebo plus chemotherapy, in patients with previously untreated squamous and non-squamous metastatic NSCLC, regardless of PD-L1 expression, and could be a newfirst-line treatment option for both squamous and non-squamous metastatic NSCLC. Full-Text PDF Sugemalimab versus placebo after concurrent or sequential chemoradiotherapy in patients with locally advanced, unresectable, stage III non-small-cell lung cancer in China (GEMSTONE-301): interim results of a randomised, double-blind, multicentre, phase 3 trialSugemalimab after definitive concurrent or sequential chemoradiotherapy could be an effective consolidation therapy for patients with stage III NSCLC whose disease has not progressed after sequential or concurrent chemoradiotherapy. Longer follow-up is needed to confirm this conclusion. Full-Text PDF

Histopathologic and MR Imaging Appearance of Spontaneous and Radiation-Induced Necrosis in Uveal Melanomas: Initial Results.

Simple SummaryUveal melanomas may undergo necrosis, both spontaneously or following radiotherapy. Nowadays radiotherapy is the preferred treatment, whereas enucleation of the eye is used in selected cases. In order to differentiate the effects of radiotherapy from spontaneous degenerative changes in uveal melanomas, we compared the appearance of necrosis, both from a histopathological point of view and from the perspective of MR imaging, in two groups of patients with uveal melanoma: a group who had undergone previous proton beam radiotherapy (secondary enucleation); a control group who had undergone enucleation without any previous radiotherapy treatment (primary enucleation). Irradiated and nonirradiated uveal melanomas differ on the basis of the histological appearance, the MR imaging appearance and the distribution of necrosis. We hope that the findings we observed could be extended to all patients with uveal melanomas treated with radiotherapy, and may enhance the accuracy of radiologists in evaluating MR examinations after radiotherapy.Necrosis in uveal melanomas can be spontaneous or induced by radiotherapy. The purpose of our study was to compare the histopathologic and MRI findings of radiation-induced necrosis of a group of proton beam-irradiated uveal melanomas with those of spontaneous necrosis of a control group of patients undergoing primary enucleation. 11 uveal melanomas who had undergone proton beam radiotherapy, MRI and secondary enucleation, and a control group of 15 untreated uveal melanomas who had undergone MRI and primary enucleation were retrospectively identified. Within the irradiated and nonirradiated group, 7 and 6 eyes with histological evidence of necrosis respectively, were furtherly selected for the final analysis; the appearance of necrosis was assessed at histopathologic examination and MRI. Irradiated melanomas showed a higher degree of necrosis as compared with nonirradiated tumors. Irradiated and nonirradiated lesions differed based on the appearance and distribution of necrosis. Irradiated tumors showed large necrotic foci, sharply demarcated from the viable neoplastic tissue; nonirradiated tumors demonstrated small, distinct foci of necrosis. Radiation-induced necrosis, more pigmented than surrounding viable tumor, displayed high signal intensity on T1-weighted and low signal intensity on T2-weighted images. The hemorrhagic/coagulative necrosis, more prevalent in nonirradiated tumors (4 out of 6 vs. 1 out of 7 cases), appeared hyperintense on T2-weighted and hypointense on T1-weighted images. Our study boosts the capability to recognize radiation-induced alterations in uveal melanomas at MRI and may improve the accuracy of radiologists in the evaluation of follow-up MR examination after radiotherapy.

Therapeutic and Immunologic Responses Elicited By in Situ Vaccination with CpG, Ibrutinib, and Low-Dose Radiation

Introduction: In situ vaccination aims to induce an immune response locally at one tumor site that propagates systemically to all tumor sites. This approach can be effective in indolent lymphoma (Brody et al., JCO 2010, Frank et al., Cancer Discov 2018, Hammerich et al., Nat Med 2019). We designed a novel clinical strategy combining in situ vaccination with systemic ibrutinib, a kinase inhibitor that modulates B and T cells. Our preclinical work had shown that combined intratumoral CpG injection and systemic ibrutinib administration was curative of systemic disease in a mouse lymphoma model, an effect that was T cell dependent (Sagiv-Barfi, Blood, 2015). Here we report the results and correlative data from the Phase I/II clinical trial testing this combination along with local low-dose radiation in adults with recurrent low-grade B cell lymphoma (NCT02927964).Methods: Enrolled patients received intratumoral injections of CpG (SD-101, 3 mg) weekly for 5 doses and local radiation (4Gy in two fractions) to the same site. Daily oral ibrutinib (560mg) began after the second intratumoral injection. Revised Lugano criteria (Cheson et al., JCO, 2014) were used to assess overall radiographic responses to therapy. Distal responses were assessed by excluding the injected site and measuring only non-injected sites. Fine needle aspirates (FNAs) were obtained from CpG-injected and non-injected nodal tumor sites pre- and post-treatment and analyzed by flow cytometry and droplet-based single-cell RNA sequencing (scRNAseq).Results: Among the twenty patients treated on study, median age was 64, 55% were male, and all but one had a diagnosis of follicular lymphoma. All patients were previously treated with an average of 2 lines of therapy, and half had previously received chemotherapy. Adverse events (AEs) were consistent with known effects of ibrutinib (including diarrhea and rash) and of CpG (including fever and flu-like reactions). No drug-related grade 4, serious, or unexpected AEs were observed. As anticipated, all patients experienced tumor reduction at the locally treated site (median 84% reduction). Remarkably, all patients experienced some tumor reduction at non-injected non-irradiated index lesions (median 45%, range 13-100%), suggesting the generation of systemic immune responses (Figure 1A). By Cheson criteria, ten patients achieved an objective response, including one complete response (ORR 50%). Despite an overall improvement in tumor burden, three patients had new or progressing non-index lesions and scored as progressive disease. Treatment induced an expansion of naïve and effector memory T cells and reductions in T follicular helper (Tfh) and activated regulatory T cells (Tregs) at the injected site. T cells with high expression of transcripts related to oxidative phosphorylation (Toxphos) increased preferentially in patients with subsequent clinical tumor reduction (Figure 1B), implicating T cell metabolism in successful generation of immune responses. Analysis of single cell T cell receptor (TCR) sequencing data revealed>300 clones that were comprised of at least 2 cells at each timepoint and which expanded or contracted at least two-fold during treatment. Expanding clones were more likely than contracting clones to be activated or memory T cells and less likely than contracting clones to be Tfh or Tregs (Figure 1C-D). Clone dynamics were often similar at the two sampled tumor sites, reflecting systemic immune responses. Finally, in vitro assays showed treatment-induced expansion of tumor-specific T cells in the peripheral blood of all 6 evaluable patients.Conclusion: The combination of oral ibrutinib, intratumoral CpG, and local low-dose radiation is safe and can generate systemic antitumor immune responses and systemic tumor shrinkage in low-grade B cell lymphoma. [Display omitted] DisclosuresShree: Gilead: Other: Spouse's employment. Khodadoust: CRISPR Therapeutics, Nutcracker Therapeutics: Research Funding; Myeloid Therapeutics: Membership on an entity's Board of Directors or advisory committees; Alexion, AstraZeneca Rare Disease: Other: Study investigator. Frank: Kite-Gilead: Membership on an entity's Board of Directors or advisory committees; Adaptive Biotechnologies: Research Funding; Allogene Therapeutics: Research Funding. Beygi: Kite/Gilead: Current Employment. Levy: GigaGen: Membership on an entity's Board of Directors or advisory committees; Teneobio: Membership on an entity's Board of Directors or advisory committees; Nurix: Membership on an entity's Board of Directors or advisory committees; Dragonfly: Membership on an entity's Board of Directors or advisory committees; Apexigen: Membership on an entity's Board of Directors or advisory committees; Viracta: Membership on an entity's Board of Directors or advisory committees; Spotlight: Membership on an entity's Board of Directors or advisory committees; Immunocore: Membership on an entity's Board of Directors or advisory committees; Walking Fish: Membership on an entity's Board of Directors or advisory committees; Kira: Membership on an entity's Board of Directors or advisory committees; Abintus Bio: Membership on an entity's Board of Directors or advisory committees; Khloris: Membership on an entity's Board of Directors or advisory committees; Virsti: Membership on an entity's Board of Directors or advisory committees; BiolineRx: Membership on an entity's Board of Directors or advisory committees; BeiGene: Membership on an entity's Board of Directors or advisory committees; Quadriga: Membership on an entity's Board of Directors or advisory committees.