Bone Organ Research Articles

Ameloblastin as a putative marker of specific bone compartments

Ameloblastin (AMBN), a member of the enamel matrix protein family, has been recently identified as integral part of the skeleton beyond the enamel. However, the specific role of endogenous AMBN in bone tissue is not fully elucidated. This study aims at investigating mRNA expression of AMBN in wild-type mice in different bone sites from early embryonic to adult stages. AMBN mRNA expression started at pre-dental stages in mouse embryos (E10.5) in both head and body parts. Using laser capture microdissection on 3-day-old mice, we showed an unambiguous mRNA expression of AMBN in extra-dental tissue (mandible bone). Screening of AMBN mRNA expression in adult mice (15-week-old) revealed that mRNA expression of AMBN varied according to the bone site; a higher mRNA levels in mandibular and frontal bone compartments were observed when compared to tibia and occipital bones. These results strongly suggest that AMBN expression may be regulated in a site-specific manner and identify AMBN as a putative in vivo marker of the site-specific fingerprint of bone organs.

PHASE II TRIAL OF THE PHOSPHATIDYINOSITOL-3 KINASE (PI3K) INHIBITOR BUPARLISIB (BKM120) IN RECURRENT GLIOBLASTOMA CONDUCTED BY THE IVY FOUNDATION EARLY PHASE CLINICAL TRIALS CONSORTIUM

BACKGROUND: The PI3K pathway is activated in most GBMs and represents a potential therapeutic target. Buparlisib is an oral, pan-Class I PI3K inhibitor that enters the brain at therapeutic concentrations and inhibits the growth of U87 and human glioma tumor sphere models. METHODS: The Ivy Consortium conducted a phase II study of buparlisib in recurrent GBM patients with activation of the PI3K pathway (mutation, homozygous deletion or loss of IHC of PTEN, PIK3CA or PIK3RI mutations, or detectable pAKT). Additional eligibility criteria included radiologic progression, 1st or 2nd relapse, >18 yrs, KPS >60, adequate bone marrow and organ function, no prior bevacizumab or enzyme-inducing antiepileptic drugs. Patients received buparlisib 100mg daily. The study consisted of 2 concurrent cohorts. In Cohort 1 (up to 15 patients) buparlisib was given for 8-12 days prior to surgery for recurrent disease. Patients underwent FDG PET, pharmacokinetic studies, and tumor was obtained for drug concentrations and pharmacodynamic effects. In Cohort 2, 50 patients with unresectable GBM received buparlisib with a primary endpoint of PFS6. RESULTS: 13 patients were enrolled into cohort 1, 50 into cohort 2. Treatment was fairly well-tolerated with no grade 4 toxicities. Grade 3 toxicities were asymptomatic lipase elevation (6), rash (4), hyperglycemia (3), fatigue (4), elevated ALT/AST (2), and 1 each with depression, anxiety, hypophosphatemia, thrombocytopenia and lymphopenia. Analysis of tumor specimens from Cohort 1 showed reduction of pAKT by IHC in 4/6 (67%) of evaluable patients. The combined cohorts showed minimal efficacy with median PFS of 1.8 months and median OS of 10.9 months. Best response was stable disease. Only 1 patient in each cohort achieved PFS6. Of the first 40 patients who underwent exome sequencing, there were 4 PIK3CA (10%), 2 PIK3R1 (5%) and 13 PTEN (33%) mutations. Patients with PTEN loss and or PI3K mutations paradoxically had a worse outcome. CONCLUSIONS: Buparlisib is well-tolerated in patients with recurrent GBM and achieves adequate tumor concentration to inhibit pAKTS473. However, single agent efficacy was minimal. Final correlation of tumor genotype with outcome will be presented. SECONDARY CATEGORY: n/a.

Antitumor agent cabozantinib decreases RANKL expression in osteoblastic cells and inhibits osteoclastogenesis and PTHrP-stimulated bone resorption.

Cabozantinib, an inhibitor of vascular endothelial growth factor and hepatocyte growth factor signaling, decreases bone lesions in patients with prostate cancer. To determine direct effects of cabozantinib on bone, resorption in neonatal mouse bone organ culture and on gene expression, proliferation, and phenotypic markers in osteoblast and osteoclast cell lines were examined. Cabozantinib, 0.3 and 3 µM, prevented PTHrP-stimulated calcium release from neonatal mouse calvaria. Since the effect on resorption could reflect effects on osteoblasts to prevent osteoclast activation, or direct inhibition of osteoclasts, responses in osteoblastic and osteoclast precursor cell lines were examined. Twenty-four-hour treatment of osteoblastic MC3T3-E1 cells with 3 µM cabozantinib decreased expression of receptor activator of NFkB ligand (RANKL) and alkaline phosphatase. Forty-eight-hour treatment of MC3T3-E1 cells with 3 µM cabozantinib inhibited cell proliferation and decreased MTT activity. Effects on alkaline phosphatase activity were biphasic, with small stimulatory effects at concentrations below 3 µM. When RAW 264.7 osteoclast precursor cells differentiated with 20 ng/ml RANKL were co-treated for 24 h with 3 µM cabozantinib, expression of RANK, TRAP, cathepsin K, alpha v or beta 3 integrin, or NFATc1 were unaffected. Five-day treatment of RANKL-treated RAW 264.7 cells with 3 µM cabozantinib decreased TRAP and MTT activity. The results suggest that the osteoblast could be the initial target, with subsequent direct and indirect effects on osteoclastogenesis leading to decreased resorption. The multiple effects of cabozantinib on the cell microenvironment of bone are consistent with its effectiveness in reducing lesions from prostate cancer metastases.

Prognosis of non-small cell lung cancer patients with bone oligometastases treated concurrently with thoracic three-dimensional radiotherapy and chemotherapy.

BackgroundTo evaluate the efficacy of three-dimensional radiotherapy for non-small cell lung cancer (NSCLC) patients with bone metastases.MethodsClinical data for 95 NSCLC patients with bone metastases were collected and prognostic factors were analyzed. All patients received radiation to their thoracic primary tumor and ≥2 cycles of chemotherapy.ResultsOf these 95 patients, 47 patients had only bone metastases and 48 had both bone metastases and other organ metastases. Univariate analysis showed that factors that statistically significantly contributed to patients having longer overall survival (OS) included receiving a radiation dose to the primary tumor ≥63 Gy, responding to treatment and receiving ≥4 cycles of chemotherapy (p = 0.001, p = 0.037 and p = 0.009, respectively). A radiation dose to the primary tumor ≥63 Gy remained significant for patients with bone metastases only as well as those with bone and other organ metastases when they were analyzed separately (p = 0.045 and p = 0.012, respectively). For patients with bone metastases only, those with T1-2 tumors had longer OS than those with T3-4 (p = 0.048); and patients who received ≥4 cycles chemotherapy compared with those who received <4 cycles had similar OS (p = 0.385). On multivariate analysis, only a radiation dose ≥63 Gy (p = 0.028) and having only bone metastases (p = 0.006) were independent prognostic factors for better OS.ConclusionsA radiation dose to the primary tumor ≥63 Gy and having only bone metastases were associated with better OS in NSCLC patients with bone metastases. For patients with bone metastases only, besides radiation dose, T status was also correlated with OS, whereas the number of chemotherapy cycles was not. Therefore, aggressive thoracic radiation may play an important role in improving OS.

Edge Detection and Level Set Active Contour Model for the Segmentation of Cavity Present in Dental X-Ray Images

ABSTRACT In the field of medical science, one of the oldest and frequently used techniques, for detecting fractures/defects in human bones is X-Ray. This is worth to do research on segmentation of X-Ray images and inventing the new techniques for the same. In this paper we used X-Ray Dental Images and Segmentation is done using Level Set Active Contour. Keywords Edge detection, image segmentation, digital dental (dd) x-ray image, level set active contour (lsac). 1. INTRODUCTION Medical imaging system has been widely used in various medical applications like orthopedic, ultra sound and many more. Medical imaging is a process used to capture the images of human body, for clinical purposes like diagnosis, anatomy, physiology etc. In its broadest sense; medical imaging can also be said as biological imaging. A number of imaging technologies such as X-ray radiography, magnetic resonance imaging, medical ultra-sonography(generally called as ultrasound), endoscopy, elastography, tactile imaging, thermography etc. that are used to detect the bone structures, organs, DNA structures, blood cells and tissues etc. Apart from imaging, other techniques for measuring and recording purpose such as electrocardiography, electroencephalography, magneto encephalography, records the data in the form of graphs or maps that contain information about measurement locations. These measuring techniques may not be designed to produce images but can be considered as forms of medical imaging. Clinical imaging is generally equated to radiology which uses invisible light spectrum such as gamma rays (used in positron emission tomography), X-rays (used in orthopedics to detect bone structures), ultraviolet rays (used in lasers, photo chemotherapy and phototherapy for the treatment of cancer). Various researches have been done to identify the cavities present in the image and following terms are widely used for that purpose:

Parathyroid hormone receptor signaling induces bone resorption in the adult skeleton by directly regulating the RANKL gene in osteocytes.

PTH upregulates the expression of the receptor activator of nuclear factor κB ligand (Rankl) in cells of the osteoblastic lineage, but the precise differentiation stage of the PTH target cell responsible for RANKL-mediated stimulation of bone resorption remains undefined. We report that constitutive activation of PTH receptor signaling only in osteocytes in transgenic mice (DMP1-caPTHR1) was sufficient to increase Rankl expression and bone resorption. Resorption in DMP1-caPTHR1 mice crossed with mice lacking the distal control region regulated by PTH in the Rankl gene (DCR(-/-)) was similar to DMP1-caPTHR1 mice at 1 month of age, but progressively declined to reach values undistinguishable from wild-type (WT) mice at 5 months of age. Moreover, DMP1-caPTHR1 mice exhibited low tissue material density and increased serum alkaline phosphatase activity at 5 month of age, and these indices of high remodeling were partially and totally corrected in compound DMP1-caPTHR1;DCR(-/-) male mice, and less affected in female mice. Rankl expression in bones from DMP1-caPTHR1 mice was elevated at both 1 and 5 months of age, whereas it was high, similar to DMP1-caPTHR1 mice at 1 month, but low, similar to WT levels at 5 months in compound mice. Moreover, PTH increased Rankl and decreased Sost and Opg expression in ex vivo bone organ cultures established from WT mice, but only regulated Sost and Opg expression in cultures from DCR(-/-) mice. PTH also increased RANKL expression in osteocyte-containing primary cultures of calvarial cells, in isolated murine osteocytes, and in WT but not in DCR(-/-) osteocyte-enriched bones. Thus, PTH upregulates Rankl expression in osteocytes in vitro, ex vivo and in vivo, and resorption induced by PTH receptor signaling in the adult skeleton requires direct regulation of the Rankl gene in osteocytes.

A noncanonical nuclear role for fibroblast growth factor signaling in skeletal development (921.6)

Fibroblast Growth Factor (FGF) signaling plays crucial roles in osteochondral progenitor cell proliferation and differentiation as seen by the multiple skeletal dysplasias resulting from abnormal FGF signaling. We identified that the skeletal dysplasia Bent Bone Dysplasia Syndrome (BBDS) results from unique FGF Receptor 2 (FGFR2) mutations that reduce canonical membrane signaling, while also enhancing a noncanonical nuclear role for the receptor in ribosome biogenesis. To determine how canonical and noncanonical FGFR2 signaling individually contribute to skeletal development and the pathogenesis of BBDS, we blocked membrane FGFR2 signaling with a chemical inhibitor and enhanced nuclear FGFR2 signaling by employing the BBDS mutations. In a bone organ culture assay, avian mandibles grown with inhibitors of membrane FGF signaling displayed reduced bone formation and exhibited uncharacteristic hypertrophy of Meckel’s cartilage. Expression of the BBDS FGFR2 mutations in developing chicken embryos caused increased proliferation and abnormal patterning in bone and cartilage. These results suggest that membrane FGF signaling is important in osteochondral progenitor differentiation and maturation while nuclear FGF signaling regulates proliferation. Combined our research shows that canonical and noncanonical FGF signaling have distinct roles during skeletal development.Grant Funding Source: Supported by NIH#5T32HD060549‐02 to JES, MOD #5‐FY12‐166 to AEM, NIH #5P30DE020750‐02

Effects of Housing Density in Five Inbred Strains of Mice

To evaluate the effect of increased mouse density in a cage, mice were housed at the density recommended by the 1996 Guide for the Care and Use of Laboratory Animals and at densities that were approximately 2, 2.6, and 3 times greater. Five strains of mice (129S1/SvImJ, A/J, BALB/cByJ, C57BL/6J, and DBA/2J) were evaluated throughout 3- and 8-month timeframes for health and well-being, including mortality, cardiac measures, plasma cholesterol, body weight, bone mineral density, organ weights, hematology, behavioral observations, and open field and light–dark tests. For 22 of the 27 traits measured, increased housing density had no significant effect. Kidney weight, adrenal weight, and heart rate decreased as mice were housed more densely, and some of the decreases were statistically significant. Reduced kidney weight, adrenal weight, and heart rate are not considered to be negative outcomes and may even indicate reduced stress. However, all measurements of these three traits were within normal physiological ranges. Percent fat increased slightly in strains 129S1/SvImJ, A/J, and DBA/2J, but did not increase in strains BALB/cByJ, and C57BL/6J. These results indicate that mice can be housed at higher densities than those currently recommended.

Bone and Other Organ Targeted Therapy

Bone and Other Organ Targeted Therapy

Species-specific homing mechanisms of human prostate cancer metastasis in tissue engineered bone

The development of effective therapeutic strategies against prostate cancer bone metastases has been impeded by the lack of adequate animal models that are able to recapitulate the biology of the disease in humans. Bioengineered approaches allow researchers to create sophisticated experimentally and physiologically relevant in vivo models to study interactions between cancer cells and their microenvironment under reproducible conditions. The aim of this study was to engineer a morphologically and functionally intact humanized organ bone which can serve as a homing site for human prostate cancer cells. Transplantation of biodegradable tubular composite scaffolds seeded with human mesenchymal progenitor cells and loaded with rhBMP-7 resulted in the development of a chimeric bone construct including a large number of human mesenchymal cells which were shown to be metabolically active and capable of producing extracellular matrix components. Micro-CT analysis demonstrated that the newly formed ossicle recapitulated the morphological features of a physiological organ bone with a trabecular network surrounded by a cortex-like outer structure. This microenvironment was supportive of the lodgement and maintenance of murine haematopoietic cell clusters, thus mimicking a functional organ bone. Bioluminescence imaging demonstrated that luciferase-transduced human PC3 cells reproducibly homed to the humanized tissue engineered bone constructs, proliferated, and developed macro-metastases. This model allows the analysis of interactions between human prostate cancer cells and a functional humanized bone organ within an immuno-incompetent murine host. The system can serve as a reproducible platform to study effects of therapeutics against prostate cancer bone metastases within a humanized microenvironment.

The future of research in craniofacial biology and what this will mean for oral health professional education and clinical practice

Today, and looking to the future, scientific discoveries from cellular, developmental and molecular biology inform our understanding of cell, tissue and organ morphogenesis as exemplified in skin, bone, cartilage, dentine, enamel, muscle, nerve and many organs such as salivary glands and teeth. Present day biomedical science yields principles for the biomimetic design and fabrication of cells, tissues and organs. Bioengineering has become a strategy that can 'mimic' biological processes, and inform clinical procedures for tissue and organ replacements. The future of regenerative craniofacial biology holds enormous promise for the diagnosis and treatment of congenital birth defects, traumatic injuries, degenerative chronic diseases as well as for Mendelian single gene and complex multigene diseases and disorders. The past 50 years have heralded the completion of the human genome and the introduction of 'personalized medicine and dentistry', the utilization of stem cell therapy for an array of diseases and disorders, the 'proof of principle' to reverse select inherited diseases such as anhidrotic ectodermal dysplasia (ED), and the fruits from interdisciplinary research drawn from the diverse biomedical sciences. Looking to the future, we can readily anticipate as major goals to emphasize the clinician's role in identifying clinical phenotypes that can lead to differential diagnosis, and rejuvenate missing or damaged tissues by establishing processes for the utilization of gene, cell and/or protein therapies. The future is replete with remarkable opportunities to enhance clinical outcomes for congenital as well as acquired craniofacial malformations. Clinicians play a pivotal role because critical thinking and sound clinical acumen substantially improve diagnostic precision and thereby clinical health outcomes.

Targeted Cytotoxic Therapy Kills Persisting HIV Infected Cells During ART

Antiretroviral therapy (ART) can reduce HIV levels in plasma to undetectable levels, but rather little is known about the effects of ART outside of the peripheral blood regarding persistent virus production in tissue reservoirs. Understanding the dynamics of ART-induced reductions in viral RNA (vRNA) levels throughout the body is important for the development of strategies to eradicate infectious HIV from patients. Essential to a successful eradication therapy is a component capable of killing persisting HIV infected cells during ART. Therefore, we determined the in vivo efficacy of a targeted cytotoxic therapy to kill infected cells that persist despite long-term ART. For this purpose, we first characterized the impact of ART on HIV RNA levels in multiple organs of bone marrow-liver-thymus (BLT) humanized mice and found that antiretroviral drug penetration and activity was sufficient to reduce, but not eliminate, HIV production in each tissue tested. For targeted cytotoxic killing of these persistent vRNA+ cells, we treated BLT mice undergoing ART with an HIV-specific immunotoxin. We found that compared to ART alone, this agent profoundly depleted productively infected cells systemically. These results offer proof-of-concept that targeted cytotoxic therapies can be effective components of HIV eradication strategies.

Anaplastic lymphoma kinase-negative anaplastic large cell lymphoma with extranodal involvement of the thigh muscle: a case report

IntroductionAnaplastic large cell lymphoma is a rare type of non-Hodgkin lymphoma. The most common extranodal sites of anaplastic large cell lymphoma are skin, subcutaneous tissue, bone, lung, and gastrointestinal organs. The involvement of the skeletal muscle has been described rarely in extranodal anaplastic large cell lymphoma.Case presentationAn 89-year-old Japanese man visited our hospital with a three-month history of swelling of his left thigh and slight fever. The swelling had rapidly enlarged and become painful within the previous three days. Magnetic resonance imaging scans revealed two soft-tissue tumors in the intramuscular layer between the vastus medialis muscle and the adductor muscle. Extensive peritumoral inflammatory edema was obvious. As the results of physical and radiological examinations were highly suggestive of abscess formation, we prescribed antibiotics for two weeks. However, our patient’s symptoms did not improve. Therefore, we suspected a soft-tissue sarcoma, and our patient underwent an incision biopsy. Histological analysis revealed that the atypical cells were positive for CD3 and CD30 but negative for anaplastic lymphoma kinase. A computed tomography scan of the thorax revealed mediastinal lymphadenopathy and bilateral pleural effusions, suggestive of extranodal involvement of skeletal muscle in anaplastic lymphoma kinase-negative anaplastic large cell lymphoma. We planned to give our patient systemic chemotherapy. However, rapid systemic dissemination occurred and our patient died of multiple organ failure five weeks after his first visit to our hospital.ConclusionsHere, we present a case of anaplastic lymphoma kinase-negative anaplastic large cell lymphoma with extranodal involvement in the thigh muscle. The involvement of such a rare organ may lead to initial misdiagnosis and a delay in the onset of treatment.

Langerhans cell histiocytosis of the atlas in a female adult.

Langerhans cell histiocytosis (LCH) is an infrequent disorder complex. It is a disease of myeloid dendritic cells, lymphocytes, and macrophages mixed with eosinophils and neutrophils [1]. The accumulation of these cells causes the classic lytic bone lesions, skin rashes, lymphadenopathy, and organ dysfunction. Langerhaus cell histocytosis occurs at all ages from infancy up to senility with the peak incidence between 2 and 4 years of age. The male/female ratio is about 3.7: 1 [2]. We have recently treated a case of LCH of the atlas in a female adult, which may be the first case of atlas LCH in a female adult reported in the English literature. A 26-year-old woman complained of headache for 10 years, with aggravation for 3 months. No neurological deficits were observed, and no significant medical history or family history was reported. Physical examination demonstrated obvious restriction of the neck motion without neurological deficit or torticollis. Computed tomography (CT) examination confirmed a sharply osteolytic lesion of the right posterior lacerate foramen area. The atlas right lateral mass, anterior arch and squamous part of occipital bone had been eroded with attenuation. The gap between the atlas and dentate was enlarged. The tumor mass had a similar density as soft tissue such as muscles in a plain CT scan; however, after enhancement, the tumor mass density overall increased obviously, but not very uniformly. On magnetic resonsnce imaging MRI, the soft tissue mass was isointense on T1, but it enhanced homogeneously after contrast administration (Fig. 1A–D). The patient underwent surgical treatment. Under general anesthesia a grey-red tumor of approximately 3.5 × 5.5 × 6 cm was completely removed from the atlas. The tumor had a tough texture with clear border and abundant blood supply. Fig. 1 A) CT scan shows an osteolytic lesion of the right posterior lacerate foramen area, and the lesion's radiological density was similar to muscles nearby; B) after enhancement, the tumor density overall increased obviously, but not very uniformly; C) shows ... Histopathologically, it was proven to be LCH. At higher magnification, the proliferating cells had indented and folded or grooved nuclei resembling coffee beans. Multinuclear giant cells were frequently seen. Eosinophils and neutrophils were admixed with the proliferating cells. The main cells were positive for S-100 protein (Fig. 1E and ​andFF). Skull and mandible are common regions of LCH involvement, and there have been very few published cases describing Langerhans cell histiocytosis of the atlas. On radiological examination, sharply demarcated punch-out osteolytic lesions in bones are characteristic. Final LCH diagnosis is possible based only on histopathology. They are usually 10–15 µm in size with characteristically folded or multilobular nuclei with typical grooves. Langerhans cells are mixed with inflammatory cells, predominantly acidophilic granulocytes. A reliable immunological marker is S-100 protein, while CD1α is usually positive too. Treatment options include surgical excision and curettage of isolated or limited bone disease. External beam radiation with local doses ranging from 2.5 Gy to 120 Gy (usually 5–20 Gy) has also been used. Chemotherapy has been used with good success for disseminated or recalcitrant disease. Overall prognosis is good with survival rates greater than 90% in patients having limited organ involvement [3]. In our case, the delitescence of the course of disease was more than 10 years, and it showed a slow process of growth of LCH and a benign nature. After one year of follow-up, there is no evidence of recurrence.

Efficacy of erlotinib in patients with relapsed gliobastoma multiforme who expressed EGFRVIII and PTEN determined by immunohistochemistry

Epidermal growth factor receptor gene (EGFR) alteration is a common feature in most of glioblastoma multiforme (GBM). Robust response of anti-EGFR treatments has been mostly associated with the EGFR deletion mutant variant III (EGFRvIII) and expression of PTEN. We have performed a prospective trial in order to confirm the efficacy of erlotinib treatment in patients with relapsed GBM who expressed EGFRvIII and PTEN. All patients included in the trial were required to be PTEN (+++), EGFR (+++) and EGFRvIII (+++) positives by immunohistochemistry. This new phase II trial enrolled 40 patients and was design to be stopped in case of fewer than two responses in the first 13 patients. Patient eligibility included histopathology criteria, radiological progression, more than 18 years old, Karnofsky performed status, KPS > 50, and adequate bone marrow and organ function. There was no limit to the number of prior treatments for relapses. No enzyme-inducing antiepileptic drugs were allowed. The primary endpoints were response and progression-free survival at 6 months (PFS6). Thirteen patients (6 men, 7 women) with recurrent GBM received erlotinib 150 mg/day. Median age was 53 years, median KPS was 80, and median prior treatments for relapses were 2. There was one partial response and three stable diseases (one at 18 months). PFS at 6 months was 20 %. Dose reduction for toxicity was not needed in any patient. Dermatitis was the main treatment-related toxicity, grade 1 in 8 patients and grade 2 in 5 patients. No grade 3 toxicity was observed. Median survival was 7 months (95 % IC 1.41–4.7). As conclusion, monotherapy with erlotinib in GBM relapses patients with high protein expression for PTEN (+++), EGFR (+++), and EGFRvlII (+++) showed low toxicity but minimal efficacy and the trial stopped.Electronic supplementary materialThe online version of this article (doi:10.1007/s11060-013-1316-y) contains supplementary material, which is available to authorized users.

A phase II evaluation of AMG 102 (rilotumumab) in the treatment of persistent or recurrent epithelial ovarian, fallopian tube or primary peritoneal carcinoma: A Gynecologic Oncology Group study

ObjectiveThis open-label, multi-institutional phase II trial evaluated activity and safety of rilotumumab (AMG 102), a monoclonal antibody that targets HGF (hepatocyte growth factor), the ligand for the MET receptor, in women with recurrent or persistent epithelial ovarian, fallopian tube or primary peritoneal cancer. Patients and methodsWomen were eligible for treatment with rilotumumab if they had measurable disease, a performance status of 0, 1 or 2, previously received platinum-based therapy with a progression-free interval of <12months or a second recurrence, and adequate bone marrow and organ function. Patients received rilotumumab 20mg/kg IV every 14days until evidence of unacceptable toxicity or disease progression. The study utilized co-dual primary endpoints of tumor response and six-month PFS to assess the efficacy of rilotumumab. Secondary endpoints included the frequency and severity of adverse events and the duration of progression-free and overall survival. ResultsThirty-one women enrolled and received rilotumumab. All were eligible for analysis. One patient achieved a complete response (3.2%; 90% CI 0.2–14%), and two women had 6-month PFS (6.5%; 90% CI 1.1–19%). Most adverse events were grade 1 or 2, with no grade 4 adverse events. Grade 3 adverse events were gastrointestinal (4), metabolic (3) anemia (3), a thromboembolic event (1), ventricular tachycardia (1), hypotension during infusion (1) and fatigue (1). The study was stopped after the first stage of accrual. ConclusionRilotumumab was well-tolerated, but had limited activity. The level of activity does not warrant further evaluation of rilotumumab as a single agent in patients with ovarian cancer.

Abstract OT1-4-04: A phase II randomized, open-label, neoadjuvant study of LCL161, an oral antagonist of inhibitor of apoptosis proteins, in combination with paclitaxel in patients with triple-negative breast cancer

Abstract Background: Inhibitor of apoptosis proteins (IAPs) negatively regulate cell death through a variety of mechanisms. LCL161 is an oral small-molecule antagonist of IAPs that has demonstrated single-agent activity and synergy with paclitaxel in breast cancer tumor models. In preclinical studies, a gene expression signature has been shown to enrich for response to LCL161. The recommended dose of LCL161 1800 mg once weekly has demonstrated preliminary antitumor activity with paclitaxel in an ongoing Phase Ib study in patients with breast cancer. Trial design: This is a Phase II, randomized, open-label study of neoadjuvant paclitaxel with or without LCL161 in women with operable, newly diagnosed triple-negative breast cancer (NCT01617668). Key inclusion criteria include women with histologically confirmed diagnosis of triple-negative breast cancer; clinical stages T2, N0–N2, M0; candidates for mastectomy or breast-conserving surgery; ECOG performance status ≤1; known status of the LCL161-predictive gene expression signature (positive and negative gene signature is a stratification factor); and adequate bone marrow and organ function. Key exclusion criteria are: bilateral or inflammatory breast cancer; locally recurrent breast cancer; patients currently receiving systemic therapy for any other malignancy, or having received systemic therapy for a malignancy in the preceding 3 months; impaired gastrointestinal function that may affect the absorption of LCL161; or uncontrolled cardiac disease. Patients are randomized 1:1 to receive paclitaxel IV (80 mg/m2 weekly) with or without oral LCL161 (1800 mg once weekly) for 12 weeks (corresponding to 4 treatment cycles). Each treatment arm is stratified 1:1 based on gene expression signature status (positive or negative). Endpoints: The primary endpoint is pathologic complete response (pCR), defined as the absence of invasive disease in the breast after 12 weeks of therapy, analyzed separately in the gene expression signature positive and negative groups. The key secondary endpoint is the pCR rate following treatment with LCL161 and paclitaxel in gene expression signature-positive or -negative tumors. Other secondary endpoints include: pCR rate in breast after 12 weeks of therapy in the full study population, and in patients with gene expression signature-positive and -negative tumors treated with paclitaxel alone; pCR rate in breast, regional nodes and axilla; biomarker evaluation including caspase 3 activation in tumor; safety; and pharmacokinetics of LCL161. Statistical methods: pCR analysis will be performed according to treatment group and gene expression signature status. An absolute increase of at least 7.5% in pCR rate of the experimental arm over the control arm will be considered as evidence of clinically relevant efficacy. Target accrual: Approximately 200 patients will be randomized into this study. Recruitment is ongoing across America, Europe, and Asia. Citation Information: Cancer Res 2013;73(24 Suppl): Abstract nr OT1-4-04.

Abstract OT2-6-07: Buparlisib (BKM120), an oral pan-PI3K inhibitor, in combination with paclitaxel in patients with HER2-negative, locally advanced or metastatic breast cancer, with or without PI3K pathway activation: A phase II, randomized, double-blind, placebo-controlled study – BELLE-4

Abstract Background: Phosphatidylinositol 3-kinase (PI3K)/Akt/mammalian target of rapamycin (mTOR) is the most frequently activated signaling pathway in breast cancer. Activation of this pathway has been implicated in tumor growth, progression, and resistance to anticancer therapies. Buparlisib (BKM120) is an oral pan-PI3K inhibitor that targets all four isoforms of class I PI3K (α, β, γ, Δ). Preliminary clinical activity was observed with buparlisib in combination with weekly paclitaxel in patients with advanced solid tumors. The maximum tolerated dose of this combination was declared as 100 mg/day and 80 mg/m2/week for buparlisib and paclitaxel, respectively (Dirix et al. ESMO 2012). Trial design: BELLE-4 (NCT01572727) is a randomized, double-blind, placebo-controlled, Phase II trial of buparlisib or placebo plus weekly paclitaxel in adult women with chemotherapy-naïve, HER2-negative, locally advanced or metastatic breast cancer. Key inclusion criteria are: known PI3K pathway activation status (activated vs non-activated; centrally determined); ECOG performance status ≤1; and adequate bone marrow and organ function. Key exclusion criteria are: no prior treatment with a PI3K or Akt inhibitor; no prior systemic chemotherapy for advanced breast cancer (prior endocrine and adjuvant chemotherapy allowed); and no symptomatic CNS metastases. Patients are randomized 1:1 to receive oral buparlisib (100 mg once daily) or matching placebo, and IV paclitaxel (80 mg/m2 once weekly) in a 28-day treatment cycle until disease progression, unacceptable toxicity, death or discontinuation from study treatment for any other reason. Stratification at randomization is based upon PI3K pathway activation and hormone receptor status. Tumor assessments are performed locally every 8 weeks from randomization until radiologic progression (RECIST 1.1). Endpoints: The primary endpoint is progression-free survival* (PFS; per local investigator) in the full and PI3K pathway-activated populations. Secondary endpoints include overall survival (OS), overall response rate (ORR)*, duration of response (DOR)*, clinical benefit rate (CBR)*, time to response*, buparlisib pharmacokinetics, and safety† (*RECIST 1.1, †CTCAE v4.03). Statistical methods: PFS analysis will be performed in PI3K pathway-activated sub-population, PI3K non-activated sub-population, and the full population, using Bayesian methodology. A stratified Cox proportional hazard model will estimate the hazard ratio of PFS. The Kaplan–Meier method will estimate PFS, OS and DOR (if relevant) for both full and PI3K pathway-activated populations. ORR and CBR will be summarized with exact 95% confidence intervals. Target accrual: Approximately 200 patients will be randomized onto BELLE-4 at around 80 centers across 18 countries. Recruitment is ongoing across America, Europe, and Asia. Contact: For further information about the study, contact your local Novartis representative. Citation Information: Cancer Res 2013;73(24 Suppl): Abstract nr OT2-6-07.

Abstract OT2-6-04: Dovitinib, a receptor tyrosine kinase inhibitor in combination with fulvestrant in postmenopausal endocrine resistant human epidermal growth factor receptor 2 negative (HER2-) / hormone receptor-positive (HR+) breast cancer: A phase II, randomized, double blind, placebo-controlled study

Abstract Background: Overcoming endocrine resistance is a critical goal in the treatment of HR+ breast cancer. Emerging in vitro evidence suggests that amplification and overexpression of fibroblast growth factor receptor 1 (FGFR1) is associated with endocrine resistance. Up to 8% of patients with HER2–/HR+ breast cancer have amplification of the FGFR1 gene. Dovitinib, a potent inhibitor of FGFR, as well as vascular endothelial growth factor receptor, and platelet-derived growth factor receptor, demonstrated antitumor activity in heavily pretreated breast cancer patients with FGF-pathway amplification. The objective of this study is to determine if dovitinib in combination with fulvestrant can improve outcomes in postmenopausal patients with endocrine resistant HER2-/HR+ breast cancer. Trial design: This multicenter, randomized, double-blind, placebo-controlled, phase II trial (NCT01528345) will enroll postmenopausal patients with HER2–/HR+ locally advanced or metastatic breast cancer. Patients are randomized 1:1 (stratified by FGF amplification [FGFR1, FGFR2, or FGF3] and presence of visceral disease) to receive intramuscular fulvestrant (500 mg q4w [with an additional dose 2 weeks after the initial dose]) in combination with oral dovitinib (500 mg, 5 days on/2 days off) or matching placebo until disease progression, unacceptable toxicity, death, or discontinuation (any reason). Crossover is not permitted. Eligibility criteria: Postmenopausal women with HER2-/HR+ breast cancer (N = 150) progressing within 12 months of completion of adjuvant endocrine therapy or after ≤ 1 prior endocrine therapy in the advanced setting are eligible. Other key inclusion criteria are: ECOG performance status ≤1; and adequate bone marrow and organ function; measurable disease as per RECIST 1.1 or ≥ 1 nonmeasurable lytic or mixed bone lesions in the absence of measurable disease. Patients with current or past evidence of CNS or leptomeningeal metastases, HER2 overexpression, and those who received previous treatment with fulvestrant or FGFR inhibitors are excluded. Specific aims: The primary endpoint is progression free survival (central review). Secondary endpoints include overall response rate, duration of response, overall survival, time to deterioration of ECOG performance status, patient-reported outcome scores, safety, and assessment of plasma pharmacokinetic concentrations of fulvestrant and dovitinib. Additionally, the pharmacodynamics of dovitinib on FGFR-associated angiogenic pathways and potential predictive biomarkers of response will be explored. Statistical methods: Analysis of primary endpoint will be carried out in FGF amplified subpopulation, FGF non-amplified subpopulation and the full population, using Bayesian methodology. Target accrual: Enrollment is ongoing. Target accrual is 150 patients (90 FGF amplified patients and 60 FGF non-amplified patients). Contact: For further information about the study please contact Gogov Sven (sven.gogov@novartis.com)/ Stephanie Deudon (stephanie.deudon@novartis.com). Citation Information: Cancer Res 2013;73(24 Suppl): Abstract nr OT2-6-04.

Abstract P4-12-29: Assessment of safety and activity in an expanded phase 1 study of MM-302, a HER2-targeted liposomal doxorubicin, in patients with advanced HER2-positive (HER2+) breast cancer

Abstract Background: MM-302 is a novel antibody drug conjugate (ADC) combining PEGylated liposomal doxorubicin with anti-HER2 scFvs. MM-302 was designed to overcome the limitations associated with using anthracyclines in the treatment of HER2-positive breast cancer. Anthracyclines have been an effective backbone of breast cancer therapies for decades. However, cardiotoxicity issues associated with free anthracyclines have limited their effective use. While liposomal doxorubicin formulations have succeeded in reducing cardiotoxicity, they have failed to demonstrate clear-cut efficacy advantages. MM-302 specifically targets tumor cells overexpressing HER2 with minimal uptake into normal cells such as cardiomyocytes which express low levels of HER2. This Phase 1 study evaluates the safety of MM-302 in patients with HER2+ advanced breast cancer. Methods: Patients aged ≥ 18 years with histologically confirmed HER2+ advanced breast cancer that have progressed or recurred on standard therapy or for which no standard therapy exists having measurable disease, adequate performance status, bone marrow reserve and organ function, were eligible for the study. A post-treatment biopsy was required. A 3 + 3 dose escalation design (8, 16, 30, 40 and 50 mg/m2 administered i.v. q4w) is used to identify a Phase 2 dose followed by expansion arms at 40 and 50 mg/m2 for a preliminary indication of activity. Secondary endpoints included determination of dose-limiting toxicity, adverse event(s), and PK of MM-302, as well as overall response and clinical benefit rates of MM-302. Results: We report data from MM-302 dose levels (30, 40 and 50 mg/m2 as monotherapy) that are expected to be therapeutically relevant. Eight patients dosed at presumed sub-therapeutic dose levels of 8 and 16 mg/m2 were excluded from the efficacy analysis. Patients had received a median of 9.5 prior agents (range 3-19) in all settings. All patients had been treated with trastuzumab, 96% with prior taxanes, 67% lapatinib, 54% anthracyclines and 33% T-DM1. At the time of this analysis (n = 27), overall response rate was 17% (4 out of 24 evaluable patients) and the estimated median Progression Free Survival (PFS) was 5.7 months, with six patients still receiving treatment. Eight patients (30%) had a PFS of greater than 6.0 months. Neutropenia was the most common treatment-related grade 3/4 toxicity and occurred in 15% of patients. The most common adverse events include fatigue, nausea, vomiting, and decreased appetite. Constipation, stomatitis, headache and rash were also observed in greater than 20% of patients. No patients discontinued treatment for toxicity. There was no protocol defined cardiotoxicity observed across all dose cohorts and five patients have had cumulative anthracycline exposure exceeding 550 mg/m2 without a reduction in left ventricular ejection fraction. Conclusion: Overall MM-302 appears to be well tolerated with promising activity in this ongoing trial. Further development is continuing combining MM-302 with trastuzumab and/or cyclophosphamide. In addition, we are evaluating the use of PET imaging to correlate liposomal tumor deposition as an additional potential patient preselection strategy. Citation Information: Cancer Res 2013;73(24 Suppl): Abstract nr P4-12-29.