Most women with breast cancer who undergo breast-conserving surgery receive whole-breast irradiation. We examined whether the addition of regional nodal irradiation to whole-breast irradiation improved outcomes. We randomly assigned women with node-positive or high-risk node-negative breast cancer who were treated with breast-conserving surgery and adjuvant systemic therapy to undergo either whole-breast irradiation plus regional nodal irradiation (including internal mammary, supraclavicular, and axillary lymph nodes) (nodal-irradiation group) or whole-breast irradiation alone (control group). The primary outcome was overall survival. Secondary outcomes were disease-free survival, isolated locoregional disease-free survival, and distant disease-free survival. Between March 2000 and February 2007, a total of 1832 women were assigned to the nodal-irradiation group or the control group (916 women in each group). The median follow-up was 9.5 years. At the 10-year follow-up, there was no significant between-group difference in survival, with a rate of 82.8% in the nodal-irradiation group and 81.8% in the control group (hazard ratio, 0.91; 95% confidence interval [CI], 0.72 to 1.13; P=0.38). The rates of disease-free survival were 82.0% in the nodal-irradiation group and 77.0% in the control group (hazard ratio, 0.76; 95% CI, 0.61 to 0.94; P=0.01). Patients in the nodal-irradiation group had higher rates of grade 2 or greater acute pneumonitis (1.2% vs. 0.2%, P=0.01) and lymphedema (8.4% vs. 4.5%, P=0.001). Among women with node-positive or high-risk node-negative breast cancer, the addition of regional nodal irradiation to whole-breast irradiation did not improve overall survival but reduced the rate of breast-cancer recurrence. (Funded by the Canadian Cancer Society Research Institute and others; MA.20 ClinicalTrials.gov number, NCT00005957.)

Combined radiochemotherapy is the currently used therapy for locally advanced pancreatic ductal adenocarcinoma (PDAC), but normal tissue toxicity limits its application. Here we test the hypothesis that inhibition of ATR (ATM-Rad3-related) could increase the sensitivity of the cancer cells to radiation or chemotherapy without affecting normal cells. We tested VE-822, an ATR inhibitor, for in vitro and in vivo radiosensitization. Chk1 phosphorylation was used to indicate ATR activity, γH2AX and 53BP1 foci as evidence of DNA damage and Rad51 foci for homologous recombination activity. Sensitivity to radiation (XRT) and gemcitabine was measured with clonogenic assays in vitro and tumor growth delay in vivo. Murine intestinal damage was evaluated after abdominal XRT. VE-822 inhibited ATR in vitro and in vivo. VE-822 decreased maintenance of cell-cycle checkpoints, increased persistent DNA damage and decreased homologous recombination in irradiated cancer cells. VE-822 decreased survival of pancreatic cancer cells but not normal cells in response to XRT or gemcitabine. VE-822 markedly prolonged growth delay of pancreatic cancer xenografts after XRT and gemcitabine-based chemoradiation without augmenting normal cell or tissue toxicity. These findings support ATR inhibition as a promising new approach to improve the therapeutic ration of radiochemotherapy for patients with PDAC.

The cytotoxicity and tumor-targeting properties of the anti-HER2/neu monoclonal antibody trastuzumab modified with peptides (CGYGPKKKRKVGG) harboring the nuclear localization sequence ([NLS] italicized) of simian virus 40 large T-antigen and radiolabeled with ¹¹¹In were evaluated. Methods: Trastuzumab was derivatized with sulfosuccinimidyl-4-(N-maleimidomethyl)cyclohexane-1-carboxylate (sulfo-SMCC) for reaction with NLS-peptides and labeled with ¹¹¹In using diethylenetriaminepentaacetic acid (DTPA). The immunoreactivity of ¹¹¹In-NLS-trastuzumab was determined by its ability to displace the binding of trastuzumab to SK-BR-3 human breast cancer (BC) cells. Cellular uptake and nuclear localization were evaluated in SK-BR-3, MDA-MB-361, and MDA-MB-231 BC cells, expressing high, intermediate, or very low levels of HER2/neu, respectively, by cell fractionation and confocal microscopy. Biodistribution and nuclear uptake were compared in athymic mice bearing MDA-MB-361 xenografts. The cytotoxicity of ¹¹¹In-trastuzumab and ¹¹¹In-NLS-trastuzumab was studied by clonogenic assays, and DNA damage was assessed by probing for phosphorylated histone H2AX (γH2AX) foci. Results: The dissociation constant for binding of ¹¹¹In-NLS-trastuzumab to SK-BR-3 cells was reduced <3-fold compared with that of ¹¹¹In-trastuzumab, demonstrating relatively preserved receptor-binding affinity. The receptor-mediated internalization of ¹¹¹In-trastuzumab in SK-BR-3, MDA-MB-361, and MDA-MB-231 cells increased significantly from 7.2% ± 0.9%, 1.3% ± 0.1%, and 0.2% ± 0.05% to 14.4% ± 1.8%, 6.3% ± 0.2%, and 0.9% ± 0.2% for ¹¹¹In-NLS-trastuzumab harboring 6 NLS-peptides, respectively. NLS-trastuzumab localized in the nuclei of BC cells, whereas unmodified trastuzumab remained surface-bound. Conjugation of ¹¹¹In-trastuzumab to NLS-peptides did not affect its tissue biodistribution but promoted specific nuclear uptake in MDA-MB-361 xenografts (2.4–2.9 %ID/g [percentage injected dose per gram] for ¹¹¹In-NLS-trastuzumab and 1.1 %ID/g for ¹¹¹In-trastuzumab). ¹¹¹In-NLS-trastuzumab was 5- and 2-fold more potent at killing SK-BR-3 and MDA-MB-361 cells than ¹¹¹In-trastuzumab, respectively, whereas toxicity toward MDA-MB-231 cells was minimal. ¹¹¹In-NLS-trastuzumab was 6-fold more effective at killing SK-BR-3 cells than unlabeled trastuzumab. Formation of γH2AX foci occurred in a greater proportion of BC cells after incubation with ¹¹¹In-NLS-trastuzumab compared with ¹¹¹In-trastuzumab or unlabeled trastuzumab. Conclusion: NLS-peptides routed ¹¹¹In-trastuzumab to the nucleus of HER2/neu-positive human BC cells, rendering the radiopharmaceutical lethal to the cells through the emission of nanometer−micrometer range Auger electrons. The greater cytotoxic potency of ¹¹¹In-NLS-trastuzumab compared with unlabeled trastuzumab in vitro and its favorable tumor-targeting properties in vivo suggest that it could be an effective targeted radiotherapeutic agent for HER2/neu-amplified BC in humans.

Machine learning approaches to problem-solving are growing rapidly within healthcare, and radiation oncology is no exception. With the burgeoning interest in machine learning comes the significant risk of misaligned expectations as to what it can and cannot accomplish. This paper evaluates the role of machine learning and the problems it solves within the context of current clinical challenges in radiation oncology. The role of learning algorithms within the workflow for external beam radiation therapy are surveyed, considering simulation imaging, multimodal fusion, image segmentation, treatment planning, quality assurance, and treatment delivery and adaptation. For each aspect, the clinical challenges faced, the learning algorithms proposed, and the successes and limitations of various approaches are analyzed. It is observed that machine learning has largely thrived on reproducibly mimicking conventional human-driven solutions with more efficiency and consistency. On the other hand, since algorithms are generally trained using expert opinion as ground truth, machine learning is of limited utility where problems or ground truths are not well-defined, or if suitable measures of correctness are not available. As a result, machines may excel at replicating, automating and standardizing human behaviour on manual chores, meanwhile the conceptual clinical challenges relating to definition, evaluation, and judgement remain in the realm of human intelligence and insight.

The intracellular environment hosts a large number of cancer- and other disease-relevant human proteins. Targeting these with internalized antibodies would allow therapeutic modulation of hitherto undruggable pathways, such as those mediated by protein–protein interactions. However, one of the major obstacles in intracellular targeting is the entrapment of biomacromolecules in the endosome. Here we report an approach to delivering antibodies and antibody fragments into the cytosol and nucleus of cells using trimeric cell-penetrating peptides (CPPs). Four trimers, based on linear and cyclic sequences of the archetypal CPP Tat, are significantly more potent than monomers and can be tuned to function by direct interaction with the plasma membrane or escape from vesicle-like bodies. These studies identify a tricyclic Tat construct that enables intracellular delivery of functional immunoglobulin-G antibodies and Fab fragments that bind intracellular targets in the cytosol and nuclei of live cells at effective concentrations as low as 1 μM.