Microscopic processes during electron cyclotron resonance microwave nitrogen plasma-assisted molecular beam epitaxial growth of GaN/GaAs heterostructures: Experiments and kinetic modeling

A set of delta-GaNyAs1–y/GaAs strained-layer superlattices grown on GaAs (001) substrates by electron cyclotron resonance (ECR) microwave plasma-assisted molecular beam epitaxy (MBE) was characterized by ex situ high resolution X-ray diffraction (HRXRD) to determine nitrogen content y in the nitrided GaAs monolayers as a function of growth temperature T. A first order kinetic model is introduced to quantitatively explain this y(T) dependence in terms of an energetically favorable N for As anion exchange and thermally activated N-surface desorption and surface segregation processes. The nitrogen surface segregation process, with an estimated activation energy Es ~ 0.9 eV appears to be significant during the GaAs overgrowth of GaNyAs1–y layers, and is shown to be responsible for strong y(T) dependence

Intraoperative radiation therapy for early-stage breast cancer: a single-institution experience

Background: To assess outcomes and toxicity after low-energy intraoperative radiotherapy (IORT) for early-stage breast cancer (ESBC).  Materials and methods: We reviewed patients with unilateral ESBC treated with breast-conserving surgery and 50-kV IORT at our institution. Patients were prescribed 20 Gy to the surface of the spherical applicator, fitted to the surgical cavity during surgery. Patients who did not meet institutional guidelines for IORT alone on final pathology were recommended adjuvant treatment, including additional surgery and/or external-beam radiation therapy (EBRT). We analyzed ipsilateral breast tumor recurrence, overall survival, recurrence-free survival and toxicity. Results: Among 201 patients (median follow-up, 5.1 years; median age, 67 years), 88% were Her2 negative and ER positive and/or PR positive, 98% had invasive ductal carcinoma, 87% had grade 1 or 2, and 95% had clinical T1 disease. Most had pathological stage T1 (93%) N0 (95%) disease. Mean IORT applicator dose at 1-cm depth was 6.3 Gy. Post-IORT treatment included additional surgery, 10%; EBRT, 11%; adjuvant chemotherapy, 9%; and adjuvant hormonal therapy, 74%. Median total EBRT dose was 42.4 (range, 40.05-63) Gy and median dose per fraction was 2.65 Gy. At 5 years, the cumulative incidence of ipsilateral breast tumor recurrence was 2.7%, the overall survival rate was 95% with no breast cancer-related deaths, and the recurrence-free survival rate was 96%. For patients who were deemed unsuitable for postoperative IORT alone and did not receive recommended risk-adapted EBRT, the IBTR rate was 4.7% versus 1.7% (p = 0.23) for patients who were either suitable for IORT alone or unsuitable and received adjuvant EBRT. Cosmetic toxicity data was available for 83%, with 7% experiencing grade 3 breast toxicity and no grade 4–5 toxicity. Conclusions: IORT for select patients with ESBC results in acceptable outcomes in regard to ipsilateral breast tumor recurrence and toxicity

The α-emitter astatine-211 targeted to CD38 can eradicate multiple myeloma in a disseminated disease model

Minimal residual disease (MRD) has become an increasingly prevalent and important entity in multiple myeloma (MM). Despite deepening responses to frontline therapy, roughly 75% of MM patients never become MRD-negative to ≤10-5, which is concerning because MRD-negative status predicts significantly longer survival. MM is highly heterogeneous, and MRD persistence may reflect survival of isolated single cells and small clusters of treatment-resistant subclones. Virtually all MM clones are exquisitely sensitive to radiation, and the α-emitter astatine-211 (211At) deposits prodigious energy within 3 cell diameters, which is ideal for eliminating MRD if effectively targeted. CD38 is a proven MM target, and we conjugated 211At to an anti-CD38 monoclonal antibody to create an 211At-CD38 therapy. When examined in a bulky xenograft model of MM, single-dose 211At-CD38 at 15 to 45 µCi at least doubled median survival of mice relative to untreated controls (P 150 days) for 50% to 80% of mice, where all untreated mice died in 20 to 55 days (P < .0001). Treatment toxicities were transient and minimal. These data suggest that 211At-CD38 offers the potential to eliminate residual MM cell clones in low-disease-burden settings, including MRD. We are optimistic that, in a planned clinical trial, addition of 211At-CD38 to an autologous stem cell transplant (ASCT) conditioning regimen may improve ASCT outcomes for MM patients

The Alpha Emitter Astatine-211 Targeted to CD38 Can Eradicate Multiple Myeloma in Minimal Residual Disease Models

Abstract Introduction Multiple myeloma (MM) is considered incurable but patients achieving minimal-residual disease (MRD) negative status following treatment have significantly better overall and progression-free survival. MM is highly heterogeneous both between and within patients, limiting the curative potential of novel agents targeting specific pathways. However all MM is highly sensitive to radiation. The α-emitter astatine-211 (211At) deposits a very large amount of energy (~100 keV/μm) within a few cell diameters (50-90 μm) resulting in irreparable double strand DNA breaks, making 211At, targeted to MM cells, particularly suited to eliminating MRD. CD38 is expressed on malignant plasma cells regardless of mutational status, and CD38 monoclonal antibodies (mAbs) constitute a proven targeted therapy for MM but do not alone eradicate disease. We proposed that 211At conjugated to an anti-CD38 mAb could effectively eliminate MM MRD, and tested this hypothesis in cellular and murine models. Methods We conjugated the anti-CD38 mAb OKT10 and an isotype matched control mAb, BHV1, to the amine-reactive labeling agent B10-NCS and labeled the final constructs with 211At. To assess in vitro cell binding we incubated each labeled construct with CD38+ cell lines, washed, and then measured cell pellet radioactivity in a gamma counter. To assess cytotoxicity we incubated CD38+ and CD38- cell lines with unlabeled or 211At-labeled OKT10-B10 for 60 hrs, then assayed viability. NOD.Cg-Rag1tm1Mom Il2rgtm1Wjl/SzJ (NRG) mice bearing H929luc or OPM-2luc MM xenografts were generated by subcutaneous (SQ) flank injection of 107 cells 7 days prior to treatment. MRD was modeled by intravenous (IV) injection of 2.5 - 5 x 105 cells 5 days prior to treatment. Radioimmunotherapy (RIT) was administered by IV injection of 7.5 - 45 µCi of 211At-OKT10-B10 or 211At-BHV1-B10. For biodistribution studies (n = 5/group) mouse tissues were harvested 24 hrs post RIT and measured in a gamma counter. For therapy studies (n = 8-10/group), all mice received syngeneic bone marrow transplant 3 days post RIT. Disease progression was assessed by tumor dimensions, luminescence imaging and survival. Results 211At-CD38 mAb selectively bound and killed CD38+ but not CD38- MM cells in vitro. In vivo, biodistribution experiments demonstrated that 211At-CD38 RIT delivered 2.4 times more radiation to MM xenografts than did control 211At-BHV1 RIT (p = 0.007), and delivered significantly higher dose to tumor than to healthy tissues including lung (p = 0.04) and kidney (p = 0.015). In murine therapy studies, 211At-CD38 RIT at 15 - 45 µCi at least doubled median survival relative to untreated controls in each of two MM SQ xenograft models (p 15 µCi 211At-CD38-RIT [p = 0.016] and all other groups [p Conclusions The efficacy of CD38 targeted 211At appears to be a function of disease distribution and malignant plasma cell access, as compellingly demonstrated by our models. Bulky tumor geometry reduces mAb penetration. In contrast, the isolated cells and small tumor clusters that define MRD are readily accessible to mAbs, creating optimal conditions for α-emitter cell kill. In an era of highly potent MM therapy, preventing relapse remains frustratingly rare. Our approach is both agnostic to high-risk cytogenetic features and offers the potential to eliminate all residual MM cell clones. These encouraging findings will be explored in a clinical trial of 211At-CD38 RIT. Download : Download high-res image (203KB) Download : Download full-size image Disclosures Orozco: Actinium Pharmaceuticals: Research Funding. Jones: Juno Therapeutics, a Celgene Company: Employment, Equity Ownership. Till: Mustang Bio: Patents & Royalties, Research Funding. Gopal: Teva: Research Funding; Spectrum: Research Funding; Janssen: Consultancy, Research Funding; BMS: Research Funding; Incyte: Consultancy; Gilead: Consultancy, Research Funding; Seattle Genetics: Consultancy, Research Funding; Brim: Consultancy; Pfizer: Research Funding; Aptevo: Consultancy; Takeda: Research Funding; Merck: Research Funding; Asana: Consultancy. Green: Juno Therapeutics: Patents & Royalties, Research Funding

Novel Bispecific Antibody Targeting CD45 and 90Y-DOTA Is Effective Therapy for Acute Myeloid Leukemia in Preclinical Murine Models

Abstract Background : Acute myeloid leukemia (AML) is a heterogeneous disease with a variety of chromosomal and molecular aberrations, rendering AML a challenging disease to cure with targeted agents. Despite the genetic variability, AML uniformly responds to radiation therapy. We have previously shown therapeutic efficacy targeting CD45+ AML using the beta-emitter 90Y and a streptavidin-biotin (SA) pretargeted radioimmunotherapy (PRIT) system. Theoretical concerns, such as endogenous biotin in humans, may limit translation of this approach to the clinic. To circumvent such hurdles, we developed bispecific antibody constructs targeting both CD45 and 90Y-labeled-DOTA-biotin, and evaluated this approach in murine models of AML. Methods : We constructed a pair of plasmids using the pfuse system that code for heavy and light chains for antibody bispecific to murine CD45 (30F11) and Y-DOTA (C825). Plasmid pairs were co-transfected into HEK 293T cells for production and purificaiton of the bispecific antibody. Binding to CD45 and 90Y-labeled-DOTA-biotin was confirmed, after which biodistribution studies with 30F11-Fc-C825 were performed in a disseminated murine leukemia model. B6SJLF1 mice (n=5 per group) were injected intravenously with 105 SJL murine leukemia cells, followed 2 days later with 0.67 nmol 30F11-Fc-C825, then 5 μg DOTA-Y-dextran clearing agent 23 hr later to clear any unbound bispecific construct from circulation. Mice were then injected with 50 µCi 90Y-DOTA-biotin IP 24 hr after the bispecific antibody infusion. Tissues were harvested at 6 and 24 hr after radiolabeled DOTA-biotin, and then analyzed for radioactivity to calculate the percent injected dose per gram of tissue (% ID/g). Results : Target organs (spleen and bone marrow) from leukemia bearing mice treated with 30F11-Fc-C825 had 9.0 ± 1.5 and 8.1 ± 1.2% ID/g, respectively, 24 hr post injection. Non-target organs such as kidneys and lungs each had To investigate this approach in a clinically relevant human leukemia model, an anti-human CD45 (BC8) scFv fusion protein was similarly engineered onto a high-affinity anti-Y-DOTA (C825) scFv fusion protein. This bispecific construct (BC8-Fc-C825) was expressed and purified from CHO cells. Binding to human CD45 and 90Y-labeled-DOTA-biotin was confirmed by flow cytometry. In biodistribution studies, female athymic mice (n=5 per group) were injected subcutaneously with 107 human AML cells (HEL). Mice were then injected intravenously with 1.4 nmol BC8-Fc-C825, BC8-SA, or non-targeting CC49-Fc-C825 as the first step before receiving clearing agent. Radiolabeled 90Y-DOTA-biotin (50 µCi) was injected IP 24 hr after bispecific delivery. HEL tumors from mice treated with BC8-Fc-C825 showed peak uptake at 24 hr post injection with 7.75 ± 0.02% ID/g, with minimal uptake in non-target organs [e.g., 0.5 ± 0.3% ID/g in kidneys]. This compared favorably with HEL tumors from mice treated with BC8-SA as the first step, which had peak uptake of 6.7 ± 0.01% ID/g 24 hr post-injection. Therapy studies took athymic mice (n=10 per group) that had been given subcutaneous HEL xenografts, before receiving BC8-Fc-C825, BC8-SA, or non-binding control CC49-Fc-C825. Treated mice were given clearing agent, followed by 800 to 1500 µCi of 90Y-labeled DOTA-biotin. Mice bearing HEL tumors treated with BC8-Fc-C825 followed by 1400 µCi of 90Y-DOTA-biotin, resulted in 6 of 10 mice surviving 170 days post-injection, whereas untreated controls and HEL-bearing mice treated with non-targeting negative control CC49-Fc-C825 and 1400 µCi of 90Y-DOTA-biotin required euthanasia due to tumor size by day 26 and 32, respectively. Conclusion : Our bispecific constructs both showed selectivity for CD45+ targets and 90Y-DOTA. In two leukemia models, targeting of 90Y-DOTA to target hematopoietic tissues translated into survival benefits. Given the therapeutic efficacy, bispecific constructs with the capacity to pretarget CD45 and capture 90Y-DOTA should be added to the armamentarium of PRIT approaches. Download : Download high-res image (106KB) Download : Download full-size image Disclosures Orozco: Actinium Pharmaceuticals: Other: Research Funding to Institution for sponsored Clinical Trials. Till: Mustang Bio: Patents & Royalties, Research Funding; Genentech: Research Funding. Gopal: Seattle Genetics: Consultancy, Research Funding. Orcutt: inviCRO: Employment, Equity Ownership. Pagel: Actinium Pharmaceuticals: Membership on an entity's Board of Directors or advisory committees. Press: Roche: Honoraria, Research Funding; BMS: Honoraria; Bayer: Consultancy