Abstract Background: Death receptor 5 (DR5) is a member of the TNFR superfamily that initiates the extrinsic apoptotic pathway by activating caspases. CS-1008 is a humanised, monoclonal IgG1 agonistic antibody to human DR5 created by CDR grafting of the murine antibody TRA-8. The aim of this study was to investigate the impact of CS-1008 dose on biodistribution, quantitative tumor uptake and anti-tumor response in patients with mCRC. Methods: Pts with mCRC who had received at least 1 course of chemotherapy (CT) were treated with weekly IV CS-1008 infusions in 5 non-sequential cohorts (Co). Different loading doses were used on days 1 and 8 (Co 1: 0.2 and 6 mg/kg; Co 2: 1 and 6 mg/kg; Co 3: 2 and 6 mg/kg; Co 4: 4 and 4mg/kg; Co 5: 6 and 2 mg/kg), followed by a weekly CS-1008 dose of 2 mg/kg. Cycle 1 encompassed 7 wks of therapy (D1 and D36 doses trace-labeled with 111In); additional weekly CS-1008 was scheduled as 4-wk cycles. Primary endpoints were to determine (1) the influence of CS-1008 dose on initial biodistribution, pharmacokinetic (PK) and tumor uptake of 111In-CS-1008 following single infusion; (2) changes in biodistribution, PK and tumor uptake following sequential doses. Secondary endpoints were to determine (1) anti-tumor response; (2) changes in tumor metabolism; (3) serum apoptosis biomarkers and serum tumor response markers. Results: Nineteen pts with a median age of 64 years and 2-6 prior CT lines were enrolled as follows: Co 1, 2 pts; Co 2, 4 pts; Co 3, 5 pts; Co 4, 3 pts; and Co 5, 5 pts. Twelve pts showed tumor uptake of 111In-CS-1008, 3 at each of the 1, 2, 4 and 6 mg/kg D1 dose levels. 111In-CS-1008 uptake in tumor was variable: some pts showed no uptake, in others uptake was observed in all measurable lesions. Liver metastases showed poor uptake of CS-1008. No significant differences were observed in tumor uptake between D1 and D36, and no effect of dose on tumor uptake was seen. DR5 expression in archived samples did not correlate with 111In-CS-1008 uptake, nor with clinical outcome. 111In-CS-1008 biodistribution showed gradual blood pool clearance and no discernible abnormal uptake in normal tissue. CS-1008 PK was not affected by dose or repeated drug administration. At restaging, there were 8 SD, 1 PR and 10 PD. The duration of PR was 3.7 months (mos). The mean duration of SD was 4 mos (range, 2.6-6.7 mos). Among the group of pts who showed CS-1008 uptake in tumor, 58% had clinical benefit (SD or PR), compared with 28% of pts in the group with no tumor uptake. The lesions that showed CS-1008 uptake were less likely to progress even in pts with overall PD at restaging. Conclusions: 111In-CS-1008 uptake in tumor predicts SD or PR. Tumor DR5 expression, assessed by 111In-CS-1008 imaging, reveals real-time heterogeneous DR5 expression, both on a per pt and on a lesion by lesion basis, and appears to be a promising predictive imaging biomarker of clinical benefit in pts with mCRC receiving CS-1008. Citation Format: Marika Ciprotti, Niall C. Tebbutt, Fook T. Lee, Sze T. Lee, Dave C. McKee, Graeme J. O'Keefe, Sylvia J. Gong, Geoffrey Chong, Hui K. Gan, Wendie Hopkins, Bridget Chappell, Nancy Y. Guo, Fiona E. Smyth, Archie N. Tse, Mendel Jansen, Manabu Matsumura, Rira Watanabe, Robert A. Beckman, Jon Greenberg, Andrew M. Scott. A phase I imaging and pharmacodynamic trial of CS-1008 in patients (pts) with metastatic colorectal cancer (mCRC). [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1174. doi:10.1158/1538-7445.AM2013-1174
1206 Objectives: Targeting immune checkpoint proteins is changing the landscape of cancer therapy. Recently, the phase III PACIFIC trial demonstrated clear overall survival benefit in the Durvalumab treated cohort in patients with stage III non-small cell lung cancer.[1] Remarkably, treatment benefit was also seen in patients with tumours with low PD-L1 expression. However, PD-L1 characterisation was performed on single, pre-treatment tumour biopsies. PD-L1 expression can be up-regulated over the course of chemo- and radiotherapy treatment. The Immuno-PET study aims to examine the dynamics of PD-L1 expression via 89Zr-Durvalumab PET imaging before, during and after chemoradiotherapy. To support the large number of clinical productions required for this study, we have developed a fully automated protocol for the radiosynthesis of [89Zr]Zr-DFOSq-Durvalumab.
Methods: Automated radiolabelling of DFOSq-Durvalumab with zirconium-89 was optimized on the disposable cassette based MultiSyn module using a modified kit. The system was programmed using an Excel based step list and synthesis progression was monitored by the built-in radioactivity detectors. Purification was performed interactively by observing the characteristic radioactivity trace recorded by one of the radioactivity detectors located at the position of the PD-10 column. Activity losses were tracked using a dose calibrator and minimized by optimizing fluid transfers, reaction buffer, antibody formulation additives and pH. [89Zr]Zr-DFOSq-Durvalumab was reformulated and sterile filtered using the built-in syringe drives. Quality control was performed on the formulated product with respect to radiochemical purity (iTLC), specific activity, protein integrity (SEC-HPLC), immunoreactive fraction, and stability over time when incubated in human serum.
Results: DFOSq-Durvalumab with an average chelator-to-antibody ratio of 3.69 was used for radiolabelling experiments and the MultiSyn module was setup following the schematic in Figure 1. Reaction kinetics in succinate pH 6 were significantly faster compared to HEPES pH 7.2 with 90% conversion observed in 15 and 60 minutes, respectively. This buffer and pH change also reduced the amount of residual radioactivity in the Zr-89 isotope vial from 24% to 0.5% ± 0.2% (n=5). Further radioactivity losses were seen in the reactor vial which could not be removed by rinsing. Additives in the reaction buffer such as 1% HSA or 0.02% Tween 80 were both effective at reducing the amount of residual [89Zr]Zr-DFOSq-Durvalumab. The latter reduced losses in the reactor from 36% ± 6% (n=4) to 0.2% ± 0.0% (n=2). Interactive peak collection via the PD-10 column radiation profile (supplementary figure) allowed reproducible collection of the product fraction with low amounts of radioactivity remaining on the PD-10 column (3.0% ± 1.2%, n=3). Residual on the sterile filter was 6.2% ± 1.9% (n=3) and the remaining kit components such as manifolds, transfer syringes, and tubing accounted for 4.3% ± 1.4% (n=3) of radioactivity losses. In summary, the total process yield was improved from 13% to 74.5% ± 5.4% (n=3) and the total process time was 45 minutes. Typically, 164 MBq of product was formulated in a volume of 2.1 mL of 0.5% sodium gentisate in PBS + 0.02% Tween 80 with a specific activity of 182 MBq/mg. Radiochemical purity and immunoreactive fraction were always >99% and >93% at end-of-synthesis, respectively, and dropped to 90% and 74% after incubation in human serum for 7 days at 37°C. SEC-HPLC analysis of formulated [89Zr]Zr-DFOSq-Durvalumab showed very good antibody integrity with levels of aggregation at 3.9% ± 0.8% (n=3).
Conclusion: Fully automated production of [89Zr]Zr-DFOSq-Durvalumab for clinical use was achieved with minimal exposure to the operator. The cassette based approach allows for multiple consecutive productions on the same day which will have clinical impact considering the growing number of clinical trials investigating 89Zr-labelled antibodies.
Abstract Glioblastoma (GBM) is the most frequent and lethal primary brain neoplasm. EphA3 is a tumor restricted antigen expressed in 38–40% of GBM and 100% of the tumor vasculature. Ifabotuzumab is a non-fucosylated IgG1κ antibody targeting EphA3 receptor. A Phase 1 study of ifabotuzumab in haematological malignancies was well tolerated and clinically active. Here we report on a Phase I dose escalation and biodistribution study of ifabotuzumab in recurrent glioblastoma. DESIGN The primary objective is to determine the toxicity and recommended phase II dose of Ifabotuzumab in GBM patients (pts). Secondary objectives are to determine the biodistribution and pharmacokinetics (PK) of 89Zr-Ifabotuzumab, the frequency of EphA3 positive GBM and response rates. Eligible pts received a trace (5mg) dose of zirconium-89 labelled ifabotuzumab (89Zr-ifab) on day 1 followed by sequential PET imaging over 1 week to determine its biodistribution, frequency of in situ EphA3 expression and tumor uptake. Safety and PK assessments were undertaken. On Day 8, pts commenced weekly ifabotuzumab infusions until PD. Three cohorts are planned (3.5mg/kg, 5.25 mg/kg, 7.9 mg/kg). On day 36, pts received both 89Zr-ifab and Ifabotuzumab, to assess receptor occupancy. Response rate (RANO) and survival data were collected. RESULTS: To date, 7 of 12 pts have enrolled (6 at 3.5 mg/kg,1 at 5.25 mg/kg; Mean age: 55 years (±12.6), 4 are male). Treatment emergent AEs included infusion reactions, seizures, cerebral oedema, rash, pruritis, headaches, eye disorder. Most were considered related to study drug, seizures and infusion reactions were readily managed with increased premedications after the first occurrence. Best response in cohort 1 is SD for 23 weeks. 89Zr-ifab PET/CT scans showed rapid, specific targeting at all known tumor sites and in all pts, but no normal tissue uptake. MRI scans showed predominant T2/FLAIR changes, occasionally marked, which were consistent with treatment effect on tumor vasculature.
BackgroundA research priority in finding a cure for HIV is to establish methods to accurately locate and quantify where and how HIV persists in people living with HIV (PLWH) receiving suppressive antiretroviral therapy (ART). Infusing copper-64 (64Cu) radiolabelled broadly neutralising antibodies targeting HIV envelope (Env) with CT scan and positron emission tomography (PET) identified HIV Env in tissues in SIV infected non-human primates . We aimed to determine if a similar approach was effective in people living with HIV (PLWH).MethodsUnmodified 3BNC117 was compared with 3BNC117 bound to the chelator MeCOSar and 64Cu (64Cu-3BNC117) in vitro to assess binding and neutralization. In a clinical trial 64Cu-3BNC117 was infused into HIV uninfected (Group 1), HIV infected and viremic (viral load, VL >1000 c/mL; Group 2) and HIV infected aviremic (VL <20 c/mL; Group 3) participants using two dosing strategies: high protein (3mg/kg unlabeled 3BNC117 combined with <5mg 64Cu-3BNC117) and trace (<5mg 64Cu-3BNC117 only). All participants were screened for 3BNC117 sensitivity from virus obtained from viral outgrowth. Magnetic resonance imaging (MRI)/PET and pharmacokinetic assessments (ELISA for serum 3BNC117 concentrations and gamma counting for 64Cu) were performed 1, 24- and 48-hours post dosing. The trial (clincialtrials.gov NCT03063788) primary endpoint was comparison of PET standard uptake values (SUVs) in regions of interest (e.g lymph node groups and gastrointestinal tract).FindingsComparison of unmodified and modified 3BNC117 in vitro demonstrated no difference in HIV binding or neutralisation. 17 individuals were enrolled of which 12 were dosed including Group 1 (n=4, 2 high protein, 2 trace dose), Group 2 (n=6, 2 high protein, 4 trace) and Group 3 (n=2, trace only). HIV+ participants had a mean CD4 of 574 cells/microL and mean age 43 years. There were no drug related adverse effects and no differences in tissue uptake in regions of interest (e.g lymph node gut, pharynx) between the 3 groups. In the high protein dosing group, serum concentrations of 3BNC117 and gamma counts were highly correlated demonstrating that 64Cu-3BNC117 remained intact in vivo.InterpretationIn PLWH on or off ART, the intervention of infusing 64Cu-3BNC117 and MRI/PET imaging over 48 hours, was unable to detect HIV-1 env expression in vivo. Future studies should investigate alternative radiolabels such as zirconium which have a longer half-life in vivo.FundingFunded by the Alfred Foundation, The Australian Centre for HIV and Hepatitis Virology Research with additional support from the Division of AIDS, National Institute of Allergy and Infectious Disease, US National Institutes of Health (USAI126611). JHM and SRL are supported by the Australian National Health and Medical Research Council.
Abstract Overview: Glioblastoma (GBM) is the most frequent and lethal primary brain neoplasm, with only 10% of patients surviving 5 years (1). EphA3 is a tumor restricted antigen expressed in 38-40% of GBM and 100% of the tumor vasculature (2, 3). Ifabotuzumab is a non-fucosylated IgG1K antibody targeting EphA3 receptor (4). A Phase I study of ifabotuzumab in haematological malignancies showed it was well tolerated and clinically active (5). Here we report on a Phase I dose escalation and biodistribution study of ifabotuzumab in recurrent GBM. Study Design: The primary objective is to determine the toxicity and recommended phase II dose of Ifabotuzumab in GBM patients (pts). Secondary objectives are to determine the biodistribution and pharmacokinetics (PK) of 89Zr-Ifabotuzumab, the frequency of EphA3 positive GBM and response rates. Eligible pts had measurable tumors received a trace (5mg) dose of zirconium-89 labelled ifabotuzumab (89Zr-ifab) on day 1 followed by sequential PET imaging over 1 week to determine its biodistribution, frequency of in situ EphA3 expression and quantitative tumor uptake. Safety assessments and PK sampling was also undertaken. On Day 8, pts commenced weekly Ifabotuzumab infusions over 2 hours. Three cohorts are planned (3.5mg/kg, 5.25 mg/kg, 7.9 mg/kg). On day 36, pts received both 89Zr-ifab and Ifabotuzumab, allowing assessment of receptor occupancy. Response rate (RANO) and survival data were collected. Pts then continued on Ifabotuzumab until progression. Results: To date, 4 of 12 planned pts have enrolled. Their mean age is 53 years (±16.3) and 3 are male. Treatment emergent adverse events included infusion reactions in 4 patients, seizures in 3 pts, cerebral oedema in 1, rash in 1, headaches in 1, eye disorder in 1. Most were considered related to study drug except seizure in 2 pts, headaches and eye disorder. Seizures and infusion reactions were readily managed with increased premedications after the first occurrence. The best response was stable disease for 23 weeks. 89Zr-ifab PET/CT scans showed rapid, specific targeting at all known tumor sites and in all pts, but no normal tissue uptake. MRI scans showed predominant T2/FLAIR changes, occasionally marked, which were consistent with treatment effect on tumor vasculature. The mean ± SD (n=4) PK parameters for first infusion 89Zr-ifab were T½α= 10.63 ± 3.14 hr, T½β = 106.85 ± 40.60 hr, V1 = 3.88 ± 0.88 L, CL= 94.11 ± 36.54 mL/hr. Conclusions: 89Zr-Ifabotuzumab demonstrates sensitive, specific and reproducible targeting of the tumor microenvironment in GBM patients. The imaging changes suggest modulation of the tumor vasculature. Enrolment is on-going. References: 1. Stupp R, et al. Lancet Oncology 10:459-66, 2009 2. Day BW, et al. Cancer Cell 23:238-48, 2013 3. Vail ME, et al. Cancer Research 74:4470-81, 2014 4. Tomasevic N, et al. Growth Factors 32:223-35, 2014 5. Swords RT, et al. Leukemia Research 50:123-131, 2016 Citation Format: Hui Gan, Lawrence Cher, Po Inglis, Zarnie Lwin, Eddie Lau, Christian Wichmann, Uwe Ackermann, Nicole Coombs, Kirsten Remen, Nancy Guo, Sze Ting Lee, Sylvia Gong, Jodie B. Palmer, Kunthi Pathmaraj, Graeme O'Keefe, Fiona Scott, Bryan W. Day, Andrew W. Boyd, Paul Thomas, Cameron Durrant, Andrew M. Scott. Preliminary findings of a Phase I safety and bioimaging trial of KB004 (ifabotuzumab) in patients with glioblastoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr CT063.
Alpha particle therapy with an actinium-225 labelled antibody for carbonic anhydrase IX leads to a highly significant therapeutic response in a mouse xenograft model.
An entry from the Cambridge Structural Database, the world’s repository for small molecule crystal structures. The entry contains experimental data from a crystal diffraction study. The deposited dataset for this entry is freely available from the CCDC and typically includes 3D coordinates, cell parameters, space group, experimental conditions and quality measures.
Abstract Purpose ATG-101, a bispecific antibody that simultaneously targets the immune checkpoint PD-L1 and the costimulatory receptor 4-1BB, activates exhausted T cells upon PD-L1 crosslinking. Previous studies demonstrated promising anti-tumour efficacy of ATG-101 in preclinical models. Here, we labelled ATG-101 with 89 Zr to confirm its tumour targeting effect and tissue biodistribution in a preclinical model. We also evaluated the use of immuno-PET to study tumour uptake of ATG-101 in vivo. Methods ATG-101, anti-PD-L1, and an isotype control were conjugated with p -SCN-Deferoxamine (Df). The Df-conjugated antibodies were radiolabelled with 89 Zr, and their radiochemical purity, immunoreactivity, and serum stability were assessed. We conducted PET/MRI and biodistribution studies on [ 89 Zr]Zr-Df-ATG-101 in BALB/c nude mice bearing PD-L1-expressing MDA-MB-231 breast cancer xenografts for up to 10 days after intravenous administration of [ 89 Zr]Zr-labelled antibodies. The specificity of [ 89 Zr]Zr-Df-ATG-101 was evaluated through a competition study with unlabelled ATG-101 and anti-PD-L1 antibodies. Results The Df-conjugation and [ 89 Zr]Zr -radiolabelling did not affect the target binding of ATG-101. Biodistribution and imaging studies demonstrated biological similarity of [ 89 Zr]Zr-Df-ATG-101 and [ 89 Zr]Zr-Df-anti-PD-L1. Tumour uptake of [ 89 Zr]Zr-Df-ATG-101 was clearly visualised using small-animal PET imaging up to 7 days post-injection. Competition studies confirmed the specificity of PD-L1 targeting in vivo. Conclusion [ 89 Zr]Zr-Df-ATG-101 in vivo distribution is dependent on PD-L1 expression in the MDA-MB-231 xenograft model. Immuno-PET with [ 89 Zr]Zr-Df-ATG-101 provides real-time information about ATG-101 distribution and tumour uptake in vivo. Our data support the use of [ 89 Zr]Zr-Df-ATG-101 to assess tumour and tissue uptake of ATG-101.
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