Highest Dose Level Research Articles

Tailoring Radiation to the Contralateral Neck for Midline HPV-Mediated Oropharyngeal Squamous Cell Carcinoma (p16+ OPSCC)

De-intensification strategies for locally advanced p16+ OPSCC are under active investigation, including omission of contralateral neck radiation (RT) for well-lateralized T1-2N0-2a. This study evaluates the outcomes of omitting contralateral neck nodal levels for p16+ OPSCC that are not well-lateralized. A retrospective review was done of 134 consecutive patients with non-metastatic p16+ OPSCC that were not well-lateralized (< 1 cm from midline or ≥ 1 cm extension to soft palate or base of tongue) who were treated with definitive chemoradiation from 2013 to 2019. IMRT to an EQD2 6364-6735 cGy (200-220 cGy per fraction) was used to treat the primary tumor and ipsilateral or bilateral neck, with tailored nodal level coverage and dose, and with concurrent paclitaxel and carboplatin. Contralateral neck RT was tailored in 121 patients: omitted in 11, limited to level II in 9, and limited to level II and III in 101 (contralateral lower neck omitted); the remaining 13 had bilateral level II-IV neck RT. Local and regional recurrences were evaluated. Median follow up was 45.34 months. Local and/or regional recurrence was noted in 9/134 (6.7%). 3 (2%) recurred in the contralateral neck in unirradiated tissue (out-of-field). All three were salvaged. The remaining 6 recurrences (4%) were in-field; one was at the primary site and 5 were nodal (all ipsilateral). 4 of the 5 regional recurrences were in a location that was treated to EQD2 7000 cGy (all had PET avid nodal disease upfront that was included in the high dose level); the other region of recurrence was treated to an EQD2 6353 cGy (the only recurrence in a never smoker). 4 of the 5 neck recurrences underwent neck dissection; one further underwent adjuvant RT to 5940 cGy in 33 fractions and remains disease-free. 2 of those who underwent salvage neck dissection alone subsequently developed distant metastases. The majority of first recurrences were distant, 19.4% (26/134). 3-year progression free survival (PFS) was 74.41% (95% CI, 65.63-81.26). Taken together, regional recurrence rate in the contralateral, unirradiated neck was 2.5% (3/121) and ultimate locoregional control was 99% (132/134). Our tailored approach demonstrates a contralateral neck local control of 97.5% (118/121) regardless of tumor lateralization; those who recurred in unirradiated cervical lymph nodes were successfully salvaged. Limiting this analysis is moderate sample size and retrospective nature, but it raises important alternative de-escalation strategies (RT volume reduction) for p16+ OPSCC aside from dose reduction, which is under active investigation in phase II/III trials. Improved systemic therapies are needed to combat distant metastases. Prospective randomized clinical trials are needed for further evaluation of nodal omission for definitive chemoradiation of non-well-lateralized p16+ OPSCC.

Equivalent Dose Estimation in FLASH Irradiation with a Deep Learning Approach

Ultra-high dose rate (FLASH) irradiation has been reported to provide decreased normal tissue toxicity without compromising tumor control compared with conventional (CONV) irradiation. However, a comprehensive understanding of the FLASH biological effect requires precise quantification of radiobiology. The study is to explore whether deep learning (DL) can tackle the task. As a proof of concept, we investigate a DL model for estimating FLASH dose to its equivalent CONV dose. Healthy C57Bl/6 female mice underwent FLASH (200Gy/s; n = 43) or CONV (0.12Gy/s; n = 41) whole abdominal irradiation using ∼16 MeV electron beams with a dose escalation scheme of 5 groups (n = 8 or 9) at 1Gy increments: 12-16Gy FLASH, 11-15Gy CONV. 4 days post-irradiation, 9 jejunum cross-sections per mouse were H&E stained for histological analysis. Each cross-section image was processed to remove lumen background and oversampled into multiple large-scale and small-scale patches along jejunal circumference. In CONV dataset, we randomly selected the data of 32 mice (80%) for model training and the rest (20%) for model validation. A ResNet101-based DL model, pre-trained with an unsupervised contrastive learning scheme, was retrained with only CONV training set to estimate corresponding CONV dose. For comparison, a crypt counting (CC) approach was implemented by manually counting the number of regenerating crypts on each cross-section image. An exponential function of dose vs crypt number was fitted with the CONV training set and used for dose estimation on the testing set. Mean squared error (MSE) was used to assess the accuracy of DL and CC approaches in estimating dose levels in CONV irradiation. The validated DL model was applied to the FLASH set to project FLASH dose into corresponding CONV dose that results in equivalent biological response. The CONV dose estimated by DL and CC approaches and DL-estimated FLASH equivalent dose were summarized in Table 1. The DL model achieved an MSE of 0.21 Gy2 on CONV testing set compared with 0.32 Gy2 of the CC approach. FLASH equivalent dose estimated by DL model for 12, 13, 14, 15 and 16Gy were 12.16±0.40, 12.53±0.32, 12.72±0.24, 12.85±0.20 and 13.04±0.27 Sv, respectively. Our proposed DL model can accurately estimate the CONV dose based on histological images. The DL predictions of FLASH dataset demonstrate that FLASH may reduce normal tissue toxicity with a lower equivalent dose, especially at high irradiated dose levels. Our study indicates that deep learning can be potentially used to assess the equivalent dose of FLASH irradiation to normal tissue.

Association between Radiation Dose and Local Failure in Recurrent High-Risk Neuroblastoma

Historically, standard-of-care (SoC) for high-risk neuroblastoma recurrences is irradiation to 2160 cGy for gross or microscopic disease. Boosting gross disease to 3000-3600 cGy remains controversial. Prior reports have demonstrated worse local control with gross disease irradiated to <3000 cGy. We sought to evaluate whether higher radiation doses delivered to sites of recurrence affected local control. We identified seventy-five high-risk neuroblastoma patients aged 6 months to 26 years that completed definitive treatment between 2007 and 2021 at two large academic centers. Only patients with distant recurrences treated with radiotherapy were included for analysis. Treatments were stratified into 1800-2160 cGy or 3000-3600 cGy dose levels. Local failure after irradiation was defined as progression within the radiation field after salvage RT. The Fine-Gray competing risk model was used to identify cumulative incidence of local failure (CILF) after irradiation with competing risk of death. Thirteen patients experienced recurrence after completion of definitive treatment, representing 30 metastatic lesions. Thirteen lesions received 1800-2160 cGy and seventeen received 3000-3600 cGy. With a median follow-up of 42.0 months (IQR 30.8-60.8), local failure after irradiation occurred in 10% (3/30) of metastatic lesions. Median time to failure was 7 months. All three failures had been treated to 2160 cGy; two in the calvarium and one in the femur. There were no local failures among patients treated to 3000-3600 cGy. The 1-year and 2-year CILF for the low- versus high-dose levels were 15% versus 0% (p<0.01) and 25% versus 0% (p<0.01), respectively. Four of the 13 patients (31%) died at a median time of 12.5 months after first recurrence; all four had distant recurrences. Dose-escalated irradiation ≥3000 cGy was associated with improved local control in recurrent high-risk neuroblastoma; prospective study and validation is warranted.

Acute Antidepressant Activity Investigation of Selected African Medicinal Plants in Mice: A Preliminary Study

The therapeutic gap arising from high global incidence of depressive disorders and the efficacy and toxicity shortcomings of existing antidepressant drugs indicate a need to further search the plant kingdom with the goal of discovering novel antidepressant pharmacotherapeutic agents. Crude 50% ethanol Terminalia catappa (CETC), Tapinanthus dodoneifolius (CETD) and Bryophyllum pinnatum (CEBP) leaf extracts were investigated for their acute antidepressant activity on the forced swim (FST) and tail suspension (TST) tests. Groups of mice (n = 6) were each exposed to the tests 1 hour following oral administration of distilled water (10 ml/kg), extracts (125, 250 and 500 mg/kg body weight) and fluoxetine (20 mg/kg), using the mean immobility time as endpoint. Qualitative phytochemical analysis of these crude extracts indicated the presence of alkaloids, cardiac glycosides, saponins, phenolic compounds, tannins, steroids, carbohydrates, flavonoids, terpenoids and anthraquinones. Compared to distilled water treatments (FST, 84.16 ±3.92; TST, 82.85 ±5.84), CETC (FST, 84.17 ±4.44, 71.67 ±1.89, &amp; 44.31 ±8.23*; TST, 79.03 ±2.62, 74.10 ±3.11, &amp; 52.57 ±6.00*) and CEBP (FST, 77.02 ±2.70, 66.11 ±4.42 &amp; 49.99 ±4.20*; TST, 69.51 ±7.24, 49.73 ±11.85* &amp; 46.39 ±5.68**) treatments caused dose-dependent and significant (P &lt; 0.05) reductions in the mean immobility times on both paradigms while CETD caused dose-dependent and significant (P &lt; 0.05) reductions of this parameter only on the TST. Overall, acute antidepressant activities at the highest dose level (500 mg/kg) of all extracts approximated that of the standard antidepressant fluoxetine (20 mg/kg). The findings of this study are a justification for the widespread ethnomedicinal uses of extracts of these plants. There is a need to further characterize their antidepressant and other neuropharmacological effects. 1.

A generalisation of the method of regression calibration

There is direct evidence of risks at moderate and high levels of radiation dose for highly radiogenic cancers such as leukaemia and thyroid cancer. For many cancer sites, however, it is necessary to assess risks via extrapolation from groups exposed at moderate and high levels of dose, about which there are substantial uncertainties. Crucial to the resolution of this area of uncertainty is the modelling of the dose–response relationship and the importance of both systematic and random dosimetric errors for analyses in the various exposed groups. It is well recognised that measurement error can alter substantially the shape of this relationship and hence the derived population risk estimates. Particular attention has been devoted to the issue of shared errors, common in many datasets, and particularly important in occupational settings. We propose a modification of the regression calibration method which is particularly suited to studies in which there is a substantial amount of shared error, and in which there may also be curvature in the true dose response. This method can be used in settings where there is a mixture of Berkson and classical error. In fits to synthetic datasets in which there is substantial upward curvature in the true dose response, and varying (and sometimes substantial) amounts of classical and Berkson error, we show that the coverage probabilities of all methods for the linear coefficient alpha are near the desired level, irrespective of the magnitudes of assumed Berkson and classical error, whether shared or unshared. However, the coverage probabilities for the quadratic coefficient beta are generally too low for the unadjusted and regression calibration methods, particularly for larger magnitudes of the Berkson error, whether this is shared or unshared. In contrast Monte Carlo maximum likelihood yields coverage probabilities for beta that are uniformly too high. The extended regression calibration method yields coverage probabilities that are too low when shared and unshared Berkson errors are both large, although otherwise it performs well, and coverage is generally better than these other three methods. A notable feature is that for all methods apart from extended regression calibration the estimates of the quadratic coefficient beta are substantially upwardly biased.

Nanomicelle curcumin-induced testicular toxicity: Implications for altered mitochondrial biogenesis and mitophagy following redox imbalance

The purpose of this study was to examine the effects of nano-micelle curcumin (NMC)-induced redox imbalance on mitochondrial biogenesis and mitophagy. For this purpose, 24 mature male Wistar rats were divided into control and NMC-received groups (7.5, 15, and 30 mg/kg) groups. After 48 days, the Nrf1, Nrf2, and SOD (Cu/Zn) expression levels, as well as GSH/GSSG, NADP+ /NADPH relative balances (elements involved in redox homeostasis) were analyzed. Moreover, to explore the effect of NMC on mitochondrial biogenesis, the expression levels of Mfn1, Mfn2, OPA1, Fis1, and Drp1 were investigated. Finally, the expression levels of Parkin/PARK and PINK (genes involved in mitochondrial quality control), as well as LC3-I/II (mitophagy marker), were analyzed. Observations showed that NMC, dose-dependently, altered GSH/GSSG, NADP+ /NADPH relative balances, suppressed SOD expression and diminished its biochemical level, and repressed Nrf1 and Nrf2 expression levels. Moreover, it could change the Mfn1, Mfn2, OPA1, Fis1, and Drp1 expression pattern and stimulate the Parkin/PARK and PINK as well as LC3-I/II expression levels, dose-dependently. In conclusion, chronic and high-dose NMC is able to suppress the redox capacity by down-regulating the Nrf1 and Nrf2 expression. Finally, at high-dose levels, it is able to trigger mitophagy signaling in the testicles.

Protective effect of carbon dots derived from scrambled Coptidis Rhizoma against ulcerative colitis in mice.

Introduction: Ulcerative colitis (UC) is a chronic and progressive inflammatory disease of the intestines. The primary symptoms, such as bloody diarrhea, can result in weight loss and significantly diminish the patient's quality of life. Despite considerable research endeavors, this disease remains incurable. The scrambled Coptidis Rhizoma (SCR) has a rich historical background in traditional Chinese medicine as a remedy for UC. Drawing from a wealth of substantial clinical practices, this study is focused on investigating the protective effects and underlying mechanisms of the active component of SCR, namely SCR-based carbon dots (SCR-CDs), in the treatment of UC. Methods: SCR-CDs were extracted and isolated from the decoction of SCR, followed by a comprehensive characterization of their morphological structure and functional groups. Subsequently, we investigated the effects of SCR-CDs on parameters such as colonic length, disease activity index, and histopathological architecture using the dextran sulfate sodium (DSS)-induced colitis mice model. Furthermore, we delved into the assessment of key aspects, including the expression of intestinal tight junction (TJ) proteins, inflammatory cytokines, oxidative stress markers, and gut microbial composition, to unravel the intricate mechanisms underpinning their therapeutic effects. Results: SCR-CDs displayed a consistent spherical morphology, featuring uniform dispersion and diameters ranging from 1.2 to 2.8nm. These SCR-CDs also exhibited a diverse array of surface chemical functional groups. Importantly, the administration of SCR-CDs, particularly at higher dosage levels, exerted a noteworthy preventive influence on colonic shortening, elevation of the disease activity index and colonic tissue impairment caused by DSS. These observed effects may be closely associated with the hygroscopic capability and hemostatic bioactivity inherent to SCR-CDs. Concurrently, the application of SCR-CDs manifested an augmenting impact on the expression of intestinal TJ proteins, concomitantly leading to a significant reduction in inflammatory cell infiltration and amelioration of oxidative stress. Additionally, SCR-CDs treatment facilitated the restoration of perturbed gut microbial composition, potentially serving as a fundamental mechanism underlying their observed protective effects. Conclusion: This study demonstrates the significant therapeutic potential of SCR-CDs in UC and provides elucidation on some of their mechanisms. Furthermore, these findings hold paramount importance in guiding innovative drug discovery for anti-UC agents.

Phase 1 trial of 4-1BB-based adoptive T-cell therapy targeting human telomerase reverse transcriptase in patients with advanced refractory solid tumors

Background aimsHuman telomerase reverse transcriptase (hTERT) is an attractive target for anti-cancer therapies. We developed an effective method for generating hTERT-specific CD8+ T cells (hTERT-induced natural T cells [TERTiNTs]) using peripheral blood mononuclear cells (PBMCs) from patients with solid cancers and investigated their feasibility and safety. MethodsThis was a single-center phase 1 trial using a 3 + 3 dose escalation design to evaluate six dose levels of TERTiNTs. PBMCs from each patient were screened using an hTERT peptide panel to select those that stimulated CD8+ T cells. The four most stimulatory peptides were used to produce autologous CD8+ T cells from patients refractory or intolerant to standard therapies. Eligible patients received a single intravenous infusion of TERTiNTs at different dose levels (4 × 108 cells/m2, 8 × 108 cells/m2 and 16 × 108 cells/m2). Pre-conditioning chemotherapy, including cyclophosphamide alone or in combination with fludarabine, was administered to induce lymphodepletion. ResultsFrom January 2014 to October 2019, a total of 24 patients with a median of three prior lines of therapy were enrolled. The most common adverse events were lymphopenia (79.2%), nausea (58.3%) and neutropenia (54.2%), mostly caused by pre-conditioning chemotherapy. The TERTiNT infusion was well tolerated, and dose-limiting toxicities were not observed. None of the patients showed objective responses. Seven patients (30.4%) achieved stable disease with a median progression-free survival of 3.9 months (range, 3.2–11.3). At the highest dose level (16 × 108 cells/m2), four of five patients showed disease stabilization. ConclusionsThe generation of TERTiNTs was feasible and safe and provided an interesting disease control rate in heavily pre-treated cancer patients.

Continuous photon energy modulation in IMRT of pancreatic cancer

Purpose: To develop a novel IMRT optimization method based on the principle of photon energy synthesis that simultaneously optimizes fluence map and beamlet energy. The method was validated on pancreatic cancers to demonstrate the benefits of the additional degree of freedom of photon energy in IMRT.Methods: Previous work has demonstrated that the effect of a photon beam of known energy can be achieved by the combination of two existing energy photons in the proper ratio. It further implied that any energy photon can be synthesized. Based on this, we propose the concept of continuous beamlet energy modulation in IMRT, or IMRT-BEM. The IMRT-BEM was modeled as the simultaneous optimization of two fluence maps, one for the low energy beam and one for the high energy beam, and it was implemented in an in-house inverse planning system. The IMRT-BEM was applied on 10 pancreatic cancer cases, where the IMRT-BEM plan was compared with single-energy IMRT plans of 6 MV (IMRT-6MV) and 15 MV photons (IMRT-15MV).Results: The IMRT-BEM plan provides a noticeable reduction to the volume irradiated at the high dose level (PTV105%) for PTV, at least 24.7% (6.4 ± 6.8 vs. 31.1 ± 18.7 (p = 0.005) and 43.8 ± 19.8 (p = 0.005) for IMRT-BEM, IMRT-6MV, and IMRT-15MV respectively). For target dose coverage, there were statistically significant improvements between the IMRT-BEM plans and the other two plans in terms of CI and HI. Compared to the IMRT-6MV plan, there were significant reductions in the Dmean of the spinal cord, liver, bowel, duodenum, and stomach. The irradiation volumes of the medium dose (V20Gy, and V40Gy) for the duodenum and bowel were reduced significantly. There were no significant differences between the IMRT-BEM and IMRT-15MV plans except for the Dmean of the spinal cord and the duodenum, the V20Gy, and V40Gy for the duodenum, and the V20Gy of the stomach.Conclusion: IMRT-BEM has certain dosimetric advantages for PTV and improves OAR sparing in pancreatic cancer, and can be effectively used in radiation treatment planning, providing another degree of freedom for planners to improve treatment plan quality.

Modulating Natural Methane Release from Rumen Fermentation through the Use of Ficus glomerata Leaf Tannins in Murrah Buffalo (Bubalus bubalis)

Enteric fermentation is one of the largest contributors of methane release to the environment from the livestock sector. Plant bioactive compounds can modulate rumen fermentation for reduced methanogenesis and fatty acid biohydrogenation. The present study investigates the effects of tannin extract from Ficus glomerata (FG) leaves on the rumen fermentation, methanogenesis, feed digestibility and fatty acid biohydrogenation of a total mixed ration with the aim of developing a feed supplement for enhanced livestock production and product quality with lower methane emission. The tannin extract (70% aqueous acetone extract) of FG leaves in the total mixed ration (oat hay/concentrate mixture; 1:1) was studied at four graded dose regimens (0.0 (control), 0.25 mL (FG-0.25), 0.50 mL (FG-0.50) and 1.0 mL (FG-1.0) per 60 mL of buffered rumen fluid) in three replicates for each treatment in a radio-frequency-based automatic gas production system (ANKOM-RF) at 39 °C for 24 h following the standard in vitro gas production protocol. The total gas production (mL or mL/g incubated dry matter (DM)) was gradually reduced (p &lt; 0.01) at dose levels of FG-0.50 and FG-1.0; however, it remained intermediary and comparable (p &gt; 0.05) for FG-0.25 with the control and FG-0.50. Compared to the control, the methane concentration (%) in the head space gas, as well as the total methane production (mL or mL/g DM incubated, or mL/g DM digested), were found to be gradually reduced (p &lt; 0.01) with increasing doses (0.25–1.0 mL) of FG extract. The reduced (p &lt; 0.05) feed degradability at higher levels (0.50–1.0 mL) of FG extract supplementation and the comparative (p &gt; 0.05) effects with the control at a lower level of supplementation (FG-0.25) are suggestive of the dose-responsive detrimental effects of tannins on fibrolytic microbes in the rumen. However, the ammonia concentration decreased (p &lt; 0.05) in all of the incubations compared to the control. Among the volatile fatty acids, acetate remained comparable (p &gt; 0.05) with enhanced (p &lt; 0.05) propionate at a lower dose (FG-0.25); however, a dose-dependent reduction was evident at higher dose levels (FG-0.50 and FG-1.0). The production of stearic acid (C18:0), which is a product of the rumen biohydrogenation process, was reduced (p &lt; 0.05), irrespective of the concentration of the FG extract. Compared to the control, the concentration of t-vaccenic acid (C18:1), which is a precursor of conjugated linoleic acid (CLA) in animal products, was increased in all the FG-extract-supplemented groups. It may be concluded that Ficus glomerata leaf tannins can modulate rumen fermentation for reduced methanogenesis and fatty acid biohydrogenation in a total mixed ration. As a higher level of inclusion negatively affects feed digestibility, a lower dose (0.25 mL FG extract per 60 mL fermentation fluid or 4.17 mL FG extract per L of fermentation fluid) is suggested to achieve desirable effects on methane abatement (30%) and an improvement in fatty acid profiles in animal products.

Catalytic degradation of methylene blue and bactericidal action by silver and CS-doped iron oxide nanostructures: Experimental and DFT approaches

In this research, Ag (3 and 6 wt %) and carbon sphere (CS) (3 wt. %) were successfully incorporated into Fe2O3 using a co-precipitation approach. This study aimed to degrade the methylene blue (MB) dye and investigate the bactericidal effect of doping-dependent Fe2O3. The pristine sample has good stability and is less toxic however showed poor degradation potency. To enhance the catalytic activity (CA), CS was introduced as it increases Fe2O3 adsorption capacity and improves metal oxide properties. Ag was incorporated into CS-Fe2O3, which created additional active sites and might generate reactive oxygen species (ROS). XRD patterns revealed the tetragonal and monoclinic structure of Fe2O3, and crystallinity was enhanced with dopants (Ag and CS). FTIR was performed to identify vibrational and rotational modes and functional groups of samples. SAED pattern represented the polycrystalline structure of Fe2O3 and (3 & 6 wt%) Ag/CS-Fe2O3. UV–vis spectroscopy depicted that absorption decreased with the increasing amount of Ag/CS and increased band gap energy (Eg). PL spectra of doped Fe2O3 represent the reduction in recombination rate resulting in enhanced CA. EDS spectra exhibited the presence of Fe, O, Na, Au, and C, which confirmed the elemental composition of pristine and doped samples. TEM images indicate nanorods (NRs) of Fe2O3, and HRTEM provides interlayer d-spacing increased upon doping. The nanostructures exhibited excellent CA for the degradation of MB dye in an acidic medium, around 99.44%. Ag/CS-Fe2O3 (6 wt %) nanostructures have shown a significant increase in the inhibition zone (3.65 mm) against Escherichia coli (E. coli) at high-level doses. This study suggests that (3 & 6 wt%) Ag/CS-Fe2O3 nanostructures revealed superior catalytic and antibacterial activity for wastewater treatment. First-principles calculations indicate that the adsorption energies show a relatively strong interaction between MB and (3 & 6 wt%) Ag/CS-Fe2O3.

Therapeutic Vaccination against Human Papillomavirus Type 16 for the Treatment of High-Grade Anal Intraepithelial Neoplasia in HIV+ Men.

Anal cancer is increasing in HIV+ men who have sex with men (MSM). Treatment options for its precursor, high-grade anal intraepithelial neoplasia (HGAIN), are suboptimal. In this phase I to II dose-finding study, we assessed the safety and efficacy of the human papillomavirus type 16 (HPV16) synthetic long peptide vaccine (SLP-HPV-01) in HIV+ MSM with HPV16-positive HGAIN. Four dosage schedules (1-5-10; 5-10-20; 10-20-40; and 40-40-40-40 μg) of SLP-HPV-01 were administered intradermally with a 3-week interval in 10 patients per dose level (DL). In each dose group, 5 patients also received 1 μg/kg pegylated IFNα-2b subcutaneously. Primary endpoints were safety and regression of HGAIN at 3, 6, and 12 months. Eighty-one of 134 screened patients (60%) had HPV16-negative HGAIN lesions, leaving 53 eligible patients. Thirteen patients were excluded, leaving 40 men. The vaccine was well tolerated. One patient developed a generalized rash. The highest dosage level induced the strongest immune responses. There was no indication for stronger reactivity in the IFNα groups. Up to 18 months of follow-up, 8/38 intention-to-treat patients had a complete clinical and histologic response and one had a partial response (in total 9/38, 23.7%). At the highest dosage level, the clinical response was 4/10 (40%). Stronger immune responses were detected among clinical responders. The highest DL is safe, immunogenic, and associated with clinical responses to HPV16-induced lesions. However, as the majority of HGAIN is caused by the other HPV types, further studies should aim at pan-HPV vaccination to prevent or treat HGAIN.

Immunogenicity, safety, and preliminary efficacy evaluation of OVX836, a nucleoprotein-based universal influenza A vaccine candidate: a randomised, double-blind, placebo-controlled, phase 2a trial

OVX836, a recombinant vaccine containing the nucleoprotein of the influenza A virus A/WSN/1933 (H1N1) and the oligomerisation domain OVX313, has displayed a good safety profile and elicited dose-dependent humoral and cellular immune responses at 90 μg or 180 μg (intramuscularly) in previous clinical trials. The aim of this study was to explore higher doses, since no maximum tolerated dose had been reached. In this phase 2a, randomised, double-blind, placebo-controlled study, we recruited 137 healthy adults aged 18-55 years in a single centre in Belgium. Participants were randomly assigned (interactive web response system; block size=4) using SAS (version 9.4) to receive one single intramuscular administration of OVX836 influenza vaccine at three doses (180 μg [n=33], 300 μg [n=35], and 480 μg [n=36]) or placebo (n=33). The two primary endpoints were the safety and the cell-mediated immune response to OVX836 at the three doses in terms of change of nucleoprotein-specific IFNγ spot forming cell (SFC) frequencies in the peripheral blood mononuclear cell (PBMC) population, measured by IFNγ ELISpot, at day 8 versus pre-injection baseline (day 1). The population used for the safety analysis is the modified intention-to-treat cohort. The population used for the immunogenicity analysis is the per-protocol cohort. This trial is registered with ClinicalTrials.gov, NCT05060887, and EudraCT, 2021-002535-39. Participants were recruited between Nov 15, 2021, and Feb 1, 2022. OVX836 had a favourable safety profile up to 480 μg without reaching the maximum tolerated dose, and showed a good safety profile at all doses with mild local and systemic reactogenicity. 7 days after vaccination, although no significant differences were observed between the doses, OVX836 increased the frequency of nucleoprotein-specific IFNγ SFCs per million PBMCs from days 1 to 8 (primary endpoint): by 124 SFCs per 106 PMBCs (95% CI 67 to 180; p=0·002) at 180 μg; by 202 SFCs per 106 PMBCs (95% CI 138 to 267; p<0·0001) at 300 μg; by 223 SFCs per 106 PMBCs (95% CI 147 to 299; p<0·0001) at 480 μg; and decreased by 1 SFCs per 106 PMBCs (95% CI -24 to 22] in the placebo group (Kruskal-Wallis test p<0·0001 followed by Mann-Whitney's tests; per-protocol cohort). Dose-dependent and polyfunctional nucleoprotein-specific CD4 T-cell responses were observed, and CD8 T-cell responses were elicited at 300 μg and 480 μg (secondary endpoints). OVX836 appears to be a safe and well tolerated candidate vaccine that elicits humoral and cellular nucleoprotein-specific immune responses (including CD8 T cells at the highest dose levels) and showed a preliminary signal of protection against influenza. Therefore, OVX836 is a promising vaccine candidate for universal influenza A prevention, that warrants further trials. OSIVAX, Bpifrance, Wallonia Region, and the EUs Horizon 2020 Research and Innovation Program.

A NOVEL IMMUNOSTIMULATORY PD-L1/OX40 TETRAVALENT BISPECIFIC ANTIBODY FOR CANCER IMMUNOTHERAPY

Abstract Single agent immune checkpoint therapy has shown substantial and durable clinical activity in many tumor types; however, only a fraction of the patients could benefit from this approach. To improve beyond the anti-PD-1/PD-L1 treatment options, bispecific antibodies (BsAb) that combines PD-L1 blockade and conditional co-stimulatory receptor activation simultaneously in one molecule have been developed and demonstrated superior anti-tumor activity in pre-clinical models. However, many of these PD-L1 based BsAb faced challenge in clinical development due to insufficient activity or unexpected toxicity. Here, we demonstrated that OX40 might be a more suitable partner for PD-L1 based BsAb design than other agonistic targets (CD27 and 4-1BB, etc.) currently in clinical studies. A novel Fc silenced tetravalent PD-L1/OX40 (EMB-09) BsAb targeting optimal OX40 binding epitope has been developed based on EpimAb’s proprietary FIT-Ig® technology. Results showed that EMB-09 maintained the parental mAb binding characteristic and retained the functional properties of each parental mAb including OX40 agonistic as well as PD-L1/PD1 inhibitory pathways. In addition, EMB-09 induced OX40 activation only in the context of PD-L1 engagement. Concurrent PD-L1/PD-1 blockade and OX40 co-stimulation by EMB-09 led to synergistic activation of T cell in vitro and exerted superior anti-tumor activity in mouse tumor models compared to anti-PD-L1 mAb. The underlining mechanism was extensively analyzed, which indicated an increased CD8+ tumor-infiltrating T-cells (TIL) as well as enhanced CD8 TIL activation status upon EMB-09 treatment. Additionally, EMB-09 was well tolerated in cynomolgus monkeys at high dose levels with a favorable safety and PK profile in a GLP-TOX study. In conclusion, as a PD-L1/OX40 BsAb with a novel biology mechanism, EMB-09 demonstrated a markedly improved anti-tumor activity compared to anti-PD-L1 mAb. The first-in-human clinal study of EMB-09 has been initiated (NCT05263180).

Effects of Dietary Vitamin D3 Over-Supplementation on Broiler Chickens' Health; Clinicopathological and Immunohistochemical Characteristics

Introduction: Vitamin D3 is used as a supplement in the feeds of livestock, pets, and human infants. However, the presence of excessive Vitamin D3 has been shown to cause toxicity in humans and animals. This study investigated the clinicopathological aspects of Vitamin D3 toxicity in broiler chickens. Materials and methods: The median lethal dose (LD50) of Vitamin D3 was estimated by the up-and-down method. To determine long term (21 days) toxic effects of oral Vitamin D3 supplementation, 90 (14-day-old) IBL-80 unsexed chicks were randomly divided into three groups as group A (control, received basal diet), B (basal diet + Vitamin D3 at 16.67 mg/kg body weight daily), and C (basal diet + Vitamin D3 at 33.33 mg/kg body weight daily). Results: The findings indicated that broiler chickens tolerated a single oral dose of Vitamin D3 up to 550 mg/kg body weight (22,000,000 IU/kg) without mortality. The results of long-term (21 days) oral supplementation of divided doses of Vitamin D3 in broiler chickens (groups B and C) showed progressive emaciation, elevated hemoglobin, hypercalcemia, hypophosphatemia, and increased alkaline phosphatase activity. At necropsy, pale liver and kidneys, congestion and hardening of lungs, mild congestion in the brain, and soft bones were observed in Vitamin D3 treated chicks (groups B and C). Microscopically, degeneration and metastatic calcification in lung parenchyma and peribronchiolar epithelium, coagulative necrosis and calcification in kidneys, and calcification with fibroplasia in proventriculus was detected. Lungs and kidneys showed a significant difference in calcification score between groups B and C. Broiler chickens from Vitamin D3 treated groups (B and C) showed strong immunohistochemical expression of Calbindin D28K in the intestine and kidneys but weak expression in the lungs. Conclusion: This study demonstrates that broiler chicks can tolerate very high levels of a single oral dose of Vitamin D3. Toxic effects of prolonged exposure to Vitamin D3 are due to over-expression of Calbindin D28k in the intestine and kidneys, disturbing the calcium and phosphorous homeostasis and leading to metastatic calcification of vital organs. This study supports that prolonged over-supplementation of Vitamin D3 causes toxic effects; hence, appropriate dietary Vitamin D3 supplementation limits should be set.

#4447 EFFECT OF RENAL IMPAIRMENT ON IPTACOPAN PHARMACOKINETICS

Abstract Background and Aims Iptacopan (LNP023) is a first-in-class, oral, proximal complement inhibitor that specifically binds to Factor B and inhibits the alternative complement pathway (AP). Current Phase III studies of iptacopan focus on diseases associated with AP activation, such as paroxysmal nocturnal hemoglobinuria (PNH), C3 glomerulonephritis (C3G), IgA nephropathy (IgAN), and atypical hemolytic uremic syndrome. The liver is the primary route of iptacopan elimination. Based on a previous healthy volunteer pharmacokinetic (PK) study, the contribution of the renal route of iptacopan elimination was measured to be low, approximately 14%. A population PK (PopPK) analysis was done to further quantify the effect of renal impairment on iptacopan PK. Method A PopPK dataset pool was created by pooling across 6 Phase II/III studies in PNH, C3G, and IgAN indications. The pool comprised patient data on the two highest dose levels (100 and 200 mg bid) and included 2439 datapoints in 234 unique patients. Factors (baseline characteristics) that could possibly affect exposure, body weight, age, gender, ethnicity, and eGFR (mL/min/1.73 m2) at baseline were investigated. The range of eGFR was 27.4 to 142.8 mL/min/1.73 m2. Age, ethnicity, and body weight were included as the significant covariates in the final model. Results The median eGFR was 87.5 mL/min/1.73 m2. As shown in the table below, the PopPK model detected a significant (p = 4.3 × 10−11) but modest effect of eGFR on simulated mean AUC0-24. At an eGFR of 34.3, AUC0-24 is expected to increase by only 38%, compared with median eGFR. Changes of eGFR of ≤15% with respect to median eGFR were observed with eGFR range between 59.7 and 132.0 mL/min /1.73 m2. Conclusion Consistent with previously observed primarily hepatic clearance of iptacopan in preclinical and clinical studies, renal impairment down to an eGFR of 34.3 did not have a clinically meaningful effect on iptacopan PK. Currently, iptacopan PK in patients with eGFR &amp;lt;30 mL/min/1.73 m2 is being explored. These results support the use of iptacopan in patients with mild and moderate renal impairment.

Systemic VSV-IFNβ-NIS oncolytic virotherapy in patients with relapsed refractory T-cell lymphoma.

7530 Background: Oncolytic virotherapy is a novel immunomodulatory therapeutic approach currently under study for relapsed refractory (R/R) hematologic malignancies. The engineered Vesicular Stomatitis Virus VSV-IFNβ-NIS used in this study encodes interferon beta (IFNβ) and sodium iodine symporter (NIS). Virally-encoded IFNβ serves as an index of viral proliferation and enhances host anti-tumor immunity. Herein, we report the updated results of the phase 1 clinical trial NCT03017820 of a one-time administration of systemic VSV-IFNβ-NIS among patients (pts) with R/R multiple myeloma (MM), T cell Lymphoma (TCL) and Acute myeloid Leukemia (AML) with particular focus on the TCL cohort treated at the highest dose level (DL4) without Ruxolitinib. Methods: VSV-IFNβ-NIS was administered as a single IV dose following a classical 3+3 phase I trial, starting at 5e9 TCID50 DL1 through 1.7e11 TCID50 DL4. A dose expansion cohort of 8 additional R/R TCL pts was added at DL4 in view of an initial signal of activity. The primary objective was to determine the maximum tolerated dose of VSV-IFNβ-NIS as a single agent. Secondary objectives were determination of safety profile and preliminary efficacy of VSV-IFNβ-NIS. Adverse events (AEs) are reported based on CTCAE V4; cytokine release syndrome (CRS) grading was based on Lee (Blood 2014) criteria. Results: To date, a total of 43 pts received VSV-IFNβ-NIS: MM (N=21), TCL (N=21) and AML (N=1). In the TCL cohort: at DL4 Group A (no Ruxolitinib), 12 pts were registered, but only 11 pts were treated (mycosis fungoides (MF) N=3; ALK- ALCL N=1; PTCL-NOS N=2; AITL N=4; Nodal PTCL, TFH N=2; ages 33-79). These pts received a median of 3 lines of therapy (range: 1-5). No dose limiting toxicities were observed. Grades 3 and higher AEs were hematologic with transient lymphopenia (63.6%), neutropenia (36.4%) being the most common. The non-hematologic AEs observed included unique toxicities such as CRS grades 1-2 (81.8%); the transient transaminitis grade 1-2 (36.4%) and grade 3 (27.3%) along with thrombocytopenia grade 1-2 (63.6%) and grade 3 (18.2%) were a reflection IFNβ toxicity. All the AEs observed were short lived. Responses were seen in pts with PTCL. At DL4, in pts with systemic PTCL (N=9), there were 5 responses including 3 complete responses (CR) with two long-lasting at 24 months, an ongoing 15 month CR, and a 5 month CR, and 2 partial responses (PR) lasting 5 months in one pts and 6 months in the other. Conclusions: These updated results of single agent VSV-IFNβ-NIS which include pts with PTCL treated at the highest dose level and a dose expansion cohort demonstrates not only the safety of VSV-IFNβ-NIS but also a strong efficacy signal in pts with PTCL with impressive durable responses. Two expansion cohorts at DL4 have been added (20 additional PTCL, 10 B-cell lymphoma pts) to obtain additional data on tolerability and efficacy and biomarkers of response in these pts. Clinical trial information: NCT03017820 .

Preliminary results of a phase I dose escalation study of IMM2510, a PD-L1 and VEGF bispecific fusion protein, in patients with advanced tumors.

2535 Background: It is well established that anti-PD-1/PD-L1 and anti-VEGF therapies have excellent synergic effects. Previous clinical trials showed that immunotherapy using anti-PD-1/PD-L1 combined with various VEGF/VERFR targeting agents achieved better efficacy for advanced tumors. IMM2510 is a novel bispecific fusion protein targeting PD-L1 and VEGFs with good efficacy and safety profile verified by our preclinical study. Here we report the preliminary data of an ongoing phase I dose escalation clinical trial of IMM2510 in human. Methods: A phase I, multicenter, open labelled, dose escalation study, was designed to evaluate the safety, efficacy, pharmacokinetics (PK) and pharmacodynamics (PD) of IMM2510. Patients (pts) with advanced solid tumors who had undergone more than two prior lines of PD-1/PD-L1 or antiangiogenic inhibitor therapies were excluded. IMM2510 was administered intravenously Q2W by using accelerated titration (0.007 mg/kg, 0.03 mg/kg) followed by 3+3 dose escalation design (0.1 mg/kg, 0.3 mg/kg, 1.0 mg/kg, 3.0 mg/kg, 6.0 mg/kg, and 10.0 mg/kg) to evaluate the safety and to determine recommended phase II dose (RP2D). Anti-tumor activity was evaluated based on RECIST v1.1. Results: As of Dec 15, 2022, 22 pts had received IMM2510 at 8 dose levels (0.007 mg/kg-10.0 mg/kg), the median age was 55.5 yrs, the median prior line of therapy was 3, ranging 1-12. No DLT was observed. The most common TRAEs (≥ 20%) were infusion related reaction (IRR) (63.6%), diarrhea (22.7%), anemia (22.7%). 4 pts experienced G3 TRAEs, including IRR (4.5%), diarrhea (4.5%), pain in extremity (4.5%), C-reactive protein increased (4.5%), procalcitonin increased (4.5%) and lymphocyte count decreased (4.5%). No G4/G5 TRAE occurred. MTD had not been reached. Based on 17 evaluable pts, 1 confirmed PR was observed after 24-week of treatment at 3.0 mg/kg in an ongoing patient with lung squamous cell carcinoma who was IO resistant to prior therapies. Additionally, SDs were seen in 4 patients (1 ovarian serous carcinoma at 0.1 mg/kg, 1 cervical squamous cell carcinoma at 3.0 mg/kg, 1 malignant pleural mesothelioma at 6.0 mg/kg, and 1 breast invasive ductal carcinoma at 10.0 mg/kg). After the cut-off date, another NSCLC patient at 10.0 mg/kg achieved PR after 9-week of treatment and was ongoing on the study. Preliminary PK analysis demonstrated that T1/2 of IMM2510 was around 6 days at the high dose levels. PD data showed that at doses of 3.0 mg/kg or greater, the circulating VEGF-A was completely blocked after first administration till the end of treatment. Conclusions: These data indicated that IMM2510 is well tolerated at the dose levels evaluated, and preliminary anti-tumor efficacy in solid tumors is encouraging. The phase I/II is ongoing. Clinical trial information: chictr20211203 .

First-in-human phase 1 trial of the safety, tolerability, pharmacokinetics, and preliminary anti-tumor activity of WSD0922-Fu: Initial report from dose escalation cohort.

3109 Background: WSD0922-Fu is an oral, central nervous system (CNS)-penetrant, small molecule, ATP non-competitive, reversible EGFR inhibitor which potently inhibits EGFR aberrations specific to non-small cell lung cancer (NSCLC) and high-grade astrocytoma (HGA). We report initial results of a first-in-human phase 1 dose-escalation trial (NCT04197934) in patients with EGFR aberrant recurrent HGA and NSCLC. Methods: Adult patients with either recurrent EGFR mutant NSCLC with CNS metastases or recurrent EGFR mutant and/or EGFR amplified HGA were enrolled. WSD0922-Fu was orally administered, and dose escalated in cohorts based on a standard 3+3 design to determine the maximum tolerated dose (MTD). Toxicities were graded per CTCAE v5.0 and preliminary efficacy was based on modified RANO criteria or RECIST 1.1 as appropriate. Results: 25 patients (17 HGA, 8 NSCLC) were enrolled and treated in six cohorts (40 mg once daily to 320 mg twice daily, 28-day cycles). The median age was 57 years (range 39-75 years). 24% were female and 36% were ECOG 0. For patients with HGA, all patients were previously treated with radiation and temozolomide; the median number of prior lines of systemic therapy was 1 (range 1-4). For patients with NSCLC, all patients were previously treated with an EGFR inhibitor (prior osimertinib 88%); the median number of prior lines of systemic therapy was 1.5 (range 1-6). The median duration of exposure to WSD0922-Fu among all patients was 54 days (range 2-258 days). The most common (≥25%) treatment-related adverse events included fatigue (68%), acneiform rash (68%), diarrhea (52%), nausea (48%) and pruritis (28%). The frequency of severe AEs was dose dependent. The most common Grade 3+ AEs (≥10%) included lymphopenia (16%), acneiform rash (12%), headache (12%) and diarrhea (12%). The MTD was defined as 160 mg twice daily. At Cycle 1 Day 15 for patients treated at this dose level, the mean terminal elimination half-life (%CV) in the plasma was 31.6 h (102%) (median 14.5 h; range 6.2-78.7 h). The mean Cmax, Tmax, oral clearance at steady state, and AUC (0-12 h) (%CV) were 4513 ng/mL (50%), 2.0 hr (69%), 6.9 L/hr/m2 (42%), and 28084 h*ng/mL (52%), respectively. Oral clearance at steady state was inversely related with dose, suggesting either inhibition or saturation of clearance processes. Among patients treated at MTD or higher dose levels and with available response data, 100% (5/5) of patients with NSCLC demonstrated clinical benefit (SD + PR) while 60% of patients had partial responses (intracranial and extracranial). Furthermore, prolonged stable disease (9 cycles) was observed in a patient with EGFRvIII mutant GBM. Conclusions: The MTD for WSD0922-Fu monotherapy is 160 mg BID. Preliminary efficacy was observed in patients with osimertinib-resistant NSCLC and EGFRvIII mutant HGA. Enrollment to expansion cohorts is ongoing. Clinical trial information: NCT04197934 .

Tazemetostat, a selective EZH2 inhibitor, with pembrolizumab as treatment of anti-PD-1 resistant head and neck squamous-cell carcinoma (HNSCC): A phase 1-2 trial.

6022 Background: EZH2 is the catalytic subunit of PRC2, which catalyzes H3K27me3 of target gene promotors and silences transcription of several cancer-related genes. EZH2 overexpression is common in HNSCC and correlated with methylation of several tumor suppressor genes. EZH2 siRNA downregulated EZH2 mRNA and protein and reduced colony formation in an EZH2-expressing HNSCC cell line. EZH2 inhibitors decreased H3K27me3 in HPV + and - cell lines. Tazemetostat, an EZH2 inhibitor, upregulated expression of HLA class I molecules in anti-PD-1-resistant HNSCC cell lines and mouse models, and increased antigen-specific CD8+ T cell proliferation, IFN-γ production, CXCL10 expression, and tumor cytotoxicity. In an anti-PD-1-resistant HNSCC model, tazemetostat and an anti-PD-1 suppressed tumor growth. The primary aims of this phase 1-2 trial were to establish 1) the recommended phase 2 dose (RP2D) of tazemetostat given with pembrolizumab and 2) the objective response rate of this regimen among patients (pts) with anti-PD-1 resistant, PD-L1 positive HNSCC. We report the results of Phase 1. Methods: In phase 1, eligible pts had recurrent/metastatic (RM) HNSCC, adequate performance status and organ function, and ability to swallow study drug. A 3+3 dose-escalation phase 1 design was used to assess 3 dose-levels of tazemetostat (400, 600, and 800 mg orally twice daily) with pembrolizumab (200 mg IV). Cycle 1 was 35 days, with tazemetostat given on days 1-35 and pembrolizumab on day 15. Subsequent cycles were 21 days, with tazemetostat given on days 1-21 and pembrolizumab on day 1. Dose-limiting toxicity (DLT), assessed during cycle 1, was defined as grade 4 neutropenia/thrombocytopenia, grade 3 febrile neutropenia, or study drug-related grades 3-4 non-hematologic adverse events (AEs). Pts must have completed cycle 1 to be evaluable for DLT assessment; otherwise, an equal number of additional pts were enrolled. The RP2D was defined as the highest dose level in which 0 of the first 3 or &lt;1 of 6 pts experienced a DLT. Results: 12 pts enrolled to phase 1: 3 each on the 400 and 600 mg dose-levels of tazemetostat and 6 on the 800 mg dose-level. Key tumor characteristics included primary site (larynx: 4, oral cavity: 3, oropharynx: 3, nasopharynx: 1, cutaneous: 1), HPV status (+: 1, -: 11), and number of lines of prior systemic treatment for RM disease (1-2: 6, &gt;3: 6). Nine pts were evaluable for DLT; 3 pts on the 800 mg dose-level did not complete cycle 1 and were not evaluable for DLT. DLT events and grade 4-5 AEs did not occur. Grade 3 AEs that occurred after cycle 1 included anemia (2 pts, both at 800 mg dose) and fatigue (1, at 400 mg dose). Across all cycles, the most common AEs included anemia (10 pts), fatigue (9), rash (7), and myalgia (4). Conclusions: Among pts with RM-HNSCC, the RP2D of tazemetostat was 800 mg twice daily when given with pembrolizumab. Enrollment to phase 2 is ongoing. Clinical trial information: NCT04624113 .