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Ionising radiation causes secondary tumours and/or enduring cognitive deficits, especially in children. Proton radiotherapy reduces exposure of the developing brain in children but may still cause some lasting effects. Recent observations show that ultra-high dose rate radiation treatment (≥40 Gy/s), called the FLASH effect, is equally effective at tumour control but less damaging to surrounding tissue compared with conventional dose rate protons (0.03–3 Gy/s). Most studies on the FLASH effect in brain and other tissues with different radiation modalities (electron and photon radiation), show FLASH benefits in these preclinical rodent models, but the data are limited, especially for proton FLASH, including for dose, dose rate and neurochemical and neurobehavioural outcomes. Tests of neurocognitive outcomes have been limited despite clinical evidence that this is the area of greatest concern. The FLASH effect in the context of proton exposure is promising, but a more systematic and comprehensive approach to outcomes is needed.

Recent studies suggest that ultra-high dose rates of proton radiation (>40 Gy/s; FLASH) confer less toxicity to exposed healthy tissue and reduce cognitive decline compared with conventional radiation dose rates (~1 Gy/s), but further preclinical data are required to demonstrate this sparing effect. In this study, postnatal day 11 (P11) rats were treated with whole brain irradiation with protons at a total dose of 0, 5, or 8 Gy, comparing a conventional dose rate of 1 Gy/s vs. a FLASH dose rate of 100 Gy/s. Beginning on P64, rats were tested for locomotor activity, acoustic and tactile startle responses (ASR, TSR) with or without prepulses, novel object recognition (NOR; 4-object version), striatal dependent egocentric learning ([configuration A] Cincinnati water maze (CWM-A)), prefrontal dependent working memory (radial water maze (RWM)), hippocampal dependent spatial learning (Morris water maze (MWM)), amygdala dependent conditioned freezing, and the mirror image CWM [configuration B (CWM-B)]. All groups had deficits in the CWM-A procedure. Weight reductions, decreased center ambulation in the open-field, increased latency on day-1 of RWM, and deficits in CWM-B were observed in all irradiated groups, except the 5 Gy FLASH group. ASR and TSR were reduced in the 8 Gy FLASH group and day-2 latencies in the RWM were increased in the FLASH groups compared with controls. There were no effects on prepulse trials of ASR or TSR, NOR, MWM, or conditioned freezing. The results suggest striatal and prefrontal cortex are sensitive regions at P11 to proton irradiation, with reduced toxicity from FLASH at 5 Gy.

Background Intracranial germ cell tumors (GCTs) comprise 3%-5% of pediatric primary central nervous system (CNS) tumors in Western countries. Though they are related in embryonic origin to gonadal GCTs, which are considered highly treatable with cisplatin-based chemotherapy regimens, intracranial GCTs vary in malignant potential and sensitivity to radiation and chemotherapy, generally carrying a worse prognosis. Metastases of intracranial GCTs outside of the CNS are rare, indicate a poor prognosis, and their salvage treatment is not well established. Case A 15-year-old boy presented with bifocal (suprasellar and pineal) intracranial nongerminomatous germ cell tumors of mixed origin. The tumors were treated to full response with a multimodal approach of neoadjuvant chemotherapy, surgical resection, and adjuvant craniospinal proton radiation. Nine months following treatment completion, the patient presented with an enlarged cervical lymph node determined on excisional biopsy to be a recurrence of pure germinoma from the primary tumors. Salvage treatment involved high-dose chemotherapy and autologous stem cell transplantation; however, the patient denied further treatment prior to planned focal radiotherapy. Thirty months post-treatment, the patient is well with no evidence of recurrence. Conclusion This case demonstrated the successful salvage treatment of an extraneural recurrence of an intracranial GCT using surgical resection and a high-dose chemotherapy and autologous stem-cell transplantation regimen, highlighting the unique factors which led to the selection of this regimen.

Cancer and the immune system are intimately related. Much of the bulk of tumors is comprised of stromal leukocytes with immune functions, which serve to both promote and inhibit tumor growth, invasion and metastasis. The T lymphocytes of the adaptive immune system are essential for tumor immunity, and these T cells are generated by cross-priming against tumor associated antigens. Dendritic cells (DCs) are essential in this process, serving as the cellular link between innate and adaptive immunity. As a prerequisite for priming of adaptive immune responses, DCs must take up tumor antigens, process them and present them in the context of the major histocompatibility complex (MHC). DCs also serve as sensors of innate activation signals from cancer that are necessary for their activation and effective priming of cancer specific T cells. Here we discuss the role of DCs in the sensing of cancer and in priming the adaptive response against tumors. Furthermore, we present the essential role of the Stimulator of Interferon Genes (STING) signaling pathway in producing type I interferons (IFNs) that are essential in this process.

Proton radiotherapy causes less off-target effects than X-rays but is not without effect. To reduce adverse effects of proton radiotherapy, a model of cognitive deficits from conventional proton exposure is needed. We developed a model emphasizing multiple cognitive outcomes. Adult male rats (10/group) received a single dose of 0, 11, 14, 17, or 20 Gy irradiation (the 20 Gy group was not used because 50% died). Rats were tested once/week for 5 weeks post-irradiation for activity, coordination, and startle. Cognitive assessment began 6-weeks post-irradiation with novel object recognition (NOR), egocentric learning, allocentric learning, reference memory, and proximal cue learning. Proton exposure had the largest effect on activity and prepulse inhibition of startle 1-week post-irradiation that dissipated each week. 6-weeks post-irradiation, there were no effects on NOR, however proton exposure impaired egocentric (Cincinnati water maze) and allocentric learning and caused reference memory deficits (Morris water maze), but did not affect proximal cue learning or swimming performance. Proton groups also had reduced striatal levels of the dopamine transporter, tyrosine hydroxylase, and the dopamine receptor D1, effects consistent with egocentric learning deficits. This new model will facilitate investigations of different proton dose rates and drugs to ameliorate the cognitive sequelae of proton radiotherapy.

Purpose There are sparse data defining the dose response of radiation therapy (RT) to the hypothalamus and pituitary in pediatric and young adult patients with brain tumors. We examined the correlation between RT dose to these structures and development of endocrine dysfunction in this population. Materials and Methods Dosimetric and clinical data were collected from children and young adults (< 26 years of age) with brain tumors treated with proton RT on three prospective studies (2003 to 2016). Deficiencies of growth hormone (GH), thyroid hormone, adrenocorticotropic hormone, and gonadotropins were determined clinically and serologically. Incidence of deficiency was estimated using the Kaplan-Meier method. Multivariate models were constructed accounting for radiation dose and age. Results Of 222 patients in the study, 189 were evaluable by actuarial analysis, with a median follow-up of 4.4 years (range, 0.1 to 13.3 years), with 31 patients (14%) excluded from actuarial analysis for having baseline hormone deficiency and two patients (0.9%) because of lack of follow-up. One hundred thirty patients (68.8%) with medulloblastoma were treated with craniospinal irradiation (CSI) and boost; most of the remaining patients (n = 56) received involved field RT, most commonly for ependymoma (13.8%; n = 26) and low-grade glioma (7.4%; n = 14). The 4-year actuarial rate of any hormone deficiency, growth hormone, thyroid hormone, adrenocorticotropic hormone, and gonadotropin deficiencies were 48.8%, 37.4%, 20.5%, 6.9%, and 4.1%, respectively. Age at start of RT, time interval since treatment, and median dose to the combined hypothalamus and pituitary were correlated with increased incidence of deficiency. Conclusion Median hypothalamic and pituitary radiation dose, younger age, and longer follow-up time were associated with increased rates of endocrinopathy in children and young adults treated with radiotherapy for brain tumors.

Objective: The Pediatric Proton/Photon Consortium Registry (PPCR) is a comprehensive data registry composed of pediatric patients treated with radiation. It was established to expedite outcomes-based research. The attributes which allow the PPCR to be a successful collaboration are reviewed. Methods and materials: Current eligibility criteria are radiotherapy patients < 22 years treated at one of the 15 US participating institutions. Detailed health and treatment data are collected about the disease presentation and treatment exposures, and annually thereafter, in REDCap (Research Electronic Data Capture). DICOM (Digital Imaging and Communications in Medicine) imaging and radiation plans are collected through MIM/MIMcloud. An optional patient-reported quality-of-life (PedsQL) study is administered at 10 sites. Results: Accrual started October 2012 with 2,775 participants enrolled as of 25 July 2019. Most patients, 62.0%, were treated for central nervous system (CNS) tumors, the most common of which are medulloblastoma (n = 349), ependymoma (n = 309), and glial/astrocytoma tumors (n = 279). The most common non-CNS diagnoses are rhabdomyosarcoma (n = 284), Ewing’s sarcoma (n = 153), and neuroblastoma (n = 130). While the majority of participants are US residents, 18.7% come from 36 other countries. Over 685 patients participate in the PedsQL study. Conclusions: The PPCR is a valuable research platform capable of answering countless research questions that will ultimately improve patient care. Centers outside of the USA are invited to participate directly or may engage with the PPCR to align data collection strategies to facilitate large-scale international research. Advances in knowledge: For investigators looking to carry out research in a large pediatric oncology cohort or interested in registry work, this paper provides an updated overview of the PPCR.

Purpose Craniospinal irradiation (CSI) is an important component of therapy for many pediatric central nervous system malignancies. Proton therapy is increasingly available and used for minimizing radiation exposure to normal tissues. The absence of an exit dose with proton therapy mandates decisions regarding coverage of the vertebral bodies (VB) in non–skeletally mature patients. Although the contents within the thecal sac represent the true clinical target volume (CTV), some physicians target the entire VB in growing children because of concerns over asymmetrical growth. This study aims to assess current practice patterns regarding VB coverage for pediatric patients undergoing CSI. Methods and Materials Pediatric radiation oncologists were identified from the Particle Therapy Co-Operative Group pediatric subcommittee membership or affiliation with US proton centers. Potential participants were contacted by e-mail with a link to an institutional review board–approved, anonymized web-based survey distributed in June 2017 with follow-up in October 2017. The survey used skip logic and included up to 11 questions regarding practice patterns. Results Thirty-three physicians responded to the survey (39%), 5 of which were excluded for lack of recent pediatric proton CSI experience. Of the 28 included responses, 23 physicians sometimes treat the entire VB and 5 physicians report always treating the entire VB. Most common responses regarding anterior CTV expansion for uncertainty were no expansion (n = 9) and 3 to 4 mm (n = 8). Most physicians modify the anterior CTV margin to protect normal structures, most commonly esophagus (n = 15), thyroid (n = 6), heart (n = 5), bowel (n = 4), and pharynx (n = 2). Conclusions Vertebral body coverage in proton CSI varies among radiation oncologists in respect to target delineation, CTV expansions, and modifications for organs at risk. These data suggest the radiation oncology community may benefit from a standardized approach to pediatric proton-based CSI.

Medulloblastoma (MB) is the most common malignant (WHO Grade IV) primary brain cancer in children, adolescents, and young adults. Radiotherapy (RT) is a mainstay of MB treatment, as it is for most childhood and adult cancers. RT dose and frequency needed to achieve efficacy in MB patients severely impacts survival outcomes and is the cause of long-term cognitive deficits. To improve on short-term side effects and long-term complications, scanning beam proton therapy is employed, when available. While this recent technological advance significantly reduces damage to surrounding healthy brain tissue, survivors continue to experience induced radiation damage, including neurocognitive sequelae. To impact survivors’ health-related quality of life and caregivers’ emotional and financial burden, it is critical to identify approaches that reduce RT dose to mitigate side-effects without impacting RT effectiveness. We are investigating targeting a unique MB electrochemical vulnerability as a means to sensitize MB tumor cells to RT. There are four MB molecular subgroups: wingless, sonic-hedgehog, Group 3, and Group 4. Our analysis of 763 MB tumor transcriptomes reveals that all Group 3 MB tumors share an enhanced expression of genes-coding for subunits of the Type-A GABA receptor (GABAAR), a chloride channel. Using patch-clamp electrophysiology, we found that GABAARs conduct Cl− in MB cells and that a brain-penetrant benzodiazepine (BZ) enhances this effect and triggers cytotoxic responses commensurate with mitochondrial depolarization. We find that BZ combined with RT, even at a sub-lethal dose, is highly effective in impairing the viability of MB tumor cells, greater than RT alone. Our BZis capable of penetrating the blood-brain barrier in minutes, is metabolically stable, and showed no toxicity in a primate model. We are investigating its suitability to be used concomitant with proton beam radiotherapy, replacing standard of care vincristine, to reduce radiation-induced brain toxicity experienced by MB patients and survivors while not decreasing RT effectiveness. Citation Format: Daniel Pomeranz Krummel, Laura Kallay, Debanjan Bhattacharya, Vaibhavkumar Gawali, Kamdem T. Donatien, Taukir Ahmed, James M. Cook, Michael Lamba, Susanne Wells, Ralph E. Vatner, Mathieu Sertorio, Dan T. Ionascu, Soma Sengupta. Targeting a unique electrochemical vulnerability in a pediatric brain tumor to potentiate proton beam radiotherapy [abstract]. In: Proceedings of the AACR Virtual Special Conference on Radiation Science and Medicine; 2021 Mar 2-3. Philadelphia (PA): AACR; Clin Cancer Res 2021;27(8_Suppl):Abstract nr PO-009.

BACKGROUND About one-third of children with primary CNS MMGCT experience incomplete responses to initial induction chemotherapy prior to irradiation, many of whom will subsequently relapse. Such high-risk patients are variably defined as having initial alpha-fetoprotein (AFP) elevations exceeding 1,000ng/mL, predominant histopathologies of malignant non-germinomatous GCT and incomplete responses to induction chemotherapy. Drugs targeting GCT-specific molecular markers have been identified for non-germinomatous GCT elements but have yet to be incorporated into prospective clinical trials. Four children with clearly identified HR-MMGCT characteristics have been treated on an innovative pilot regimen incorporating intensified chemotherapy and molecularly targeted agents, with avoidance or minimization of irradiation. METHODS Four children (two with pure suprasellar embryonal carcinoma (EC) - one with Down syndrome and the other with pre-diagnosis cognitive dysfunction; one with initial serum AFP exceeding 7,000ng/mL and yolk sac tumor (YST)+EC+Teratoma pathology; one with initial serum AFP exceeding 1,000ng/mL) were treated with 3 cycles of “standard” induction chemotherapy (ACNS1123), followed by 1–3 transplant cycles (thiotepa/carboplatin) each with complete radiographic and tumor marker responses. Two children with pure EC subsequently received six cycles of brentuximab-vedotin without irradiation and remain disease-free off therapy for 2–4 years. One child with YST+EC+Teratoma has subsequently received reduced dose craniospinal irradiation and pineal region boost, and will receive oral everolimus, erlotinib, palbocyclib and intravenous brentuximab-vedotin. The fourth child with YST+MT will commence everolimus, erlotinib and palbocyclib without irradiation. CONCLUSION This treatment strategy for HR-MMGCT patients provides preliminary tolerance and response data justifying extension to a multi-center trial.

Advances in radiation delivery technologies and immunotherapy have improved effective cancer treatments and long-term outcomes. Experimental and clinical trials have demonstrated the benefit of a combination of radiation therapy and immunotherapy for tumor eradication. Despite precise radiation dose delivery that is achievable by particle therapy and benefits from reactivating the antitumor immune response, resistance to both therapeutic strategies is frequently observed in patients. Understanding the biological origins of such resistance will create new opportunities for improved cancer treatment. Cancer metabolism and especially a high rate of aerobic glycolysis leading to overproduction and release of lactate is one such biological process favoring tumor progression and treatment resistance. Because of their known protumor effects, aerobic glycolysis and lactate production are potential targets for increased efficacy of radiation alone or in combination with immunotherapy. In the following review, we present an overview of the interplay of cancer cell lactate metabolism with the tumor microenvironment and immune cells. We discuss how a deeper understanding and careful modulation of lactate metabolism and radiation therapy might exploit this interplay for improved therapeutic outcome.

The tailless complex polypeptide-1 ring complex (TRiC) is a eukaryotic heat shock protein 60 (hsp60) molecule that has been shown to bind N-terminally extended precursors of OVA-derived SIINFEKL in vivo. Binding of peptides to TRiC was shown to be essential for their presentation on MHC class I. We demonstrate in this study that purified TRiC binds antigenic peptides in vitro as well; however, such binding is not restricted to N-terminally extended peptides, suggesting that the results obtained in vivo reflect the availability of peptides in vivo rather than structural constraints of TRiC-peptide binding. Immunization of mice with noncovalent complexes of peptides (derived from OVA or β-galactosidase) and TRiC results in cross-priming of CD8+ T lymphocytes specific for Kb/SIINFEKL or Ld/TPHPARIGL. Mechanistic dissection of this phenomenon shows that TRiC binds APC, and TRiC-chaperoned peptides are processed within the APC and presented on their MHC class I. Immunogenicity of TRiC purified from OVA- or β-galactosidase–expressing cells, that is, of endogenously generated TRiC-peptide complexes, was investigated, and such preparations were observed not to be immunogenic. Consistent with this observation, SIINFEKL or its precursors were not observed to be associated with TRiC purified from cells expressing a fusion GFP-OVA protein. In contrast, immunization with TRiC purified from a tumor elicited specific protection against a challenge with that tumor. These results are interpreted with respect to the cell biological properties of TRiC and suggest that in vivo, TRiC binds a limited proportion of peptides derived from a limited set of intracellular proteins.

The peptide-binding property of MHC is central to adaptive immunological functions. A similar property of heat shock proteins (HSPs) hsp70 and hsp90 has been implicated in Ag presentation by MHC and in cross-priming. The peptide-binding pocket of hsp70 has been characterized structurally and functionally and a peptide-binding site in gp96 (of hsp90 family) has been defined. Nonetheless, questions persist whether the specific immunogenicity of HSP preparations derives from the peptides chaperoned by the HSPs or by proteins contaminating the HSP preparations. Because absolute purity of a protein preparation is a metaphysical concept, other approaches are necessary to address the question. In this study, we demonstrate that the specific immunogenicity of gp96 preparations isolated from cells expressing β-galactosidase derives from the MHC I epitope precursors associated with the gp96 and not from contaminating β-galactosidase protein nor unassociated fragments derived from it. Although the observations here are limited to a single HSP and antigenic peptides chaperoned by it, they can be extended broadly.

Introduction: We expanded upon our previous experience using involved-field fractionated radiotherapy (IFRT) as an alternative to whole brain radiotherapy (WBRT) or stereotactic radiosurgery (SRS) for patients with surgically resected brain metastases.Material and Methods: All patients with single brain metastases who underwent surgical resection followed by IFRT at our institution from 2006-2013 were evaluated. Local recurrence-free survival, distant failure-free survival and overall survival were determined. Analyses were performed associating clinical variables with local recurrence and distant failure. Salvage approaches and toxicity of treatment for each patient were also assessed.Results: Median follow-up was 19.1 months. Fifty-six patients were treated with a median dose of 40.05 Gy/15 fractions with IFRT to the resection cavity. Local recurrence-free survival was 91.4%, distant failure-free survival was 68.4%, and overall survival was 77.7% at 12 months. No variables were associated with increased local recurrence, however melanoma histopathology and infratentorial location were associated with distant failure on multivariate analysis. Local recurrences were salvaged in 5/8 patients, and distant failures were salvaged in 24/29 patients. Two patients developed radionecrosis.Conclusions: Adjuvant IFRT is feasible and safe for well-selected patients with surgically resected single brain metastases. Acceptable rates of local control and salvage of distal intracranial recurrences continue to be achieved with continued follow-up.

Background Radiotherapy (RT) can induce T cell-mediated antitumor immune responses by multiple mechanisms. However, due to the strongly immunosuppressive tumor microenvironment that hinders T cell function, these responses are ineffective and tumors usually escape. The common gamma-chain cytokines interleukin ( IL)-2 and IL-15 promote the proliferation of activated T cells and, therefore, are prime agents for immunotherapy strategies aimed at sustaining anti-tumor T cell responses. IL2 at high doses is effective at inducing responses in a subset of cancer patients. However, the benefits of IL-2 are partially offset by serious toxicity and by regulatory T cell (Treg) stimulation. IL-15, on the other hand, has shown low toxicity and lacks Treg stimulatory activity, making it an attractive candidate for testing in combination with RT. Here we tested the hypothesis that IL-15 administration strengthens the pro-immunogenic effect of hypofractionated RT and contributes to inducing effective anti-tumor responses. Methods The poorly immunogenic TSA mouse breast cancer cells were implanted s.c. in syngeneic BALB/c mice on day 0. When tumors became palpable, mice were randomly assigned to one of 4 treatment groups (N = 4-5mice/ group): control, RT, IL-15 or RT+IL-15. RT was delivered exclusively to the primary tumor in 8 Gy fractions on days 13, 14 and 15. IL-15 (5 μg/mouse) was administered i.p. or s.c. peritumorally daily for 10 days starting on day 12. Mice were followed for tumor growth or regression. In a separate experiment, mice were euthanized on day 22 to characterize tumor-infiltrating lymphocytes (TILs). Results Tumor growth delay was seen when IL-15 was given peritumorally but not i.p. (p < 0.05 compared to control). Improved tumor regression was seen when RT was combined with IL-15 (p < 0.05), suggesting that IL15 can potentiate T cell responses elicited by RT. Consistently, analysis of TILs showed a marked increase in CD8+ T cells expressing the activation marker CD137 (35.3% in RT+IL-15 versus 5.90% In control, p < 0.05) while the increase was modest with each monotherapy (18.8% in RT, 20.7% in IL-15, p < 0.05 compared to control). In addition, we found a significant increase in the ratio of effector CD4+ T-cells to Tregs (2.5 in RT +IL-15 versus 0.78 in control, p < 0.05) whereas monotherapy had no effect (1.14 in RT, 0.96 in IL-15). Conclusions Overall these results support the hypothesis that IL-15 is a valuable partner for combination treatments with local RT. We are currently elucidating the mechanisms involved in pre-clinical models in preparation for future testing in patients.

The immune system has the ability to recognize and specifically reject tumors, and tumors only become clinically apparent once they have evaded immune destruction by creating an immunosuppressive tumor microenvironment. Radiotherapy can cause immunogenic tumor cell death resulting in cross-priming of tumor specific T-cells, acting as an in situ tumor vaccine; however, radiotherapy alone rarely induces effective anti-tumor immunity resulting in systemic tumor rejection. Immunotherapy can complement radiotherapy to help overcome tumor induced immune suppression, as demonstrated in pre-clinical tumor models. Here we provide the rationale for combinations of different immunotherapies and radiotherapy, and review the pre-clinical and emerging clinical evidence for these combinations in the treatment of cancer.

The discrepancy between the in vitro and in vivo response to radiation is readily explained by the fact that tumors do not exist independently of the host organism; cancer cells grow in the context of a complex microenvironment composed of stromal cells, vasculature, and elements of the immune system. As the antitumor effect of radiotherapy depends in part on the immune system, and myeloid-derived cells in the tumor microenvironment modulate the immune response to tumors, it follows that understanding the effect of radiation on myeloid cells in the tumor is likely to be essential for comprehending the antitumor effects of radiotherapy. In this review, we describe the phenotype and function of these myeloid-derived cells, and stress the complexity of studying this important cell compartment owing to its intrinsic plasticity. With regard to the response to radiation of myeloid cells in the tumor, evidence has emerged demonstrating that it is both model and dose dependent. Deciphering the effects of myeloid-derived cells in tumors, particularly in irradiated tumors, is key for attempting to pharmacologically modulate their actions in the clinic as part of cancer therapy.

ligands, PDL-1/2, were upregulated by RT on TSA cells and tumor-infiltrating myeloid cells, suggesting PD-1’s interaction with its ligands may limit RT-activated antitumor T-cell activity. Consistent with this hypothesis, RMP1-14 alone had no effect on tumor growth; RT delayed growth (p < 0.01), but only 1/6 mice showed tumor regression, whereas all mice receiving RT+RMP114 completely rejected tumors by day 25. In spleen, RMP1-14 had no effect on AH1-specific CD8 + T-cells