Challenges and Opportunities in Developing an Oncology Clinical Trial Network in the United States Veterans Affairs Health Care System: The VA STARPORT Experience.

The United States Veterans Affairs (VA) Health Care System has a strong history of conducting impactful oncology randomized clinical trials (RCTs). We developed a phase II/III RCT to test the use of metastasis-directed therapy in Veterans with oligometastatic prostate cancer (OMPC)-the first VA RCT in OMPC that leverages novel imaging and advanced radiotherapy techniques. To accomplish this, we developed a clinical trial network to conduct the study. In this manuscript, we describe several challenges we encountered in study development/conduct and our strategies to address them, with the goal of helping investigators establish robust study networks to conduct clinical trials. In the study start-up, we encountered challenges in timely site activation, and leveraged project management to maximize efficiency. Additionally, there were several changes in the clinical paradigms in imaging and treatment that led to protocol amendments to ensure maximum equipoise, recruitment, and impact of the study. Specifically, we amended the trial to add de novo OMPC patients (from initially only recurrent OMPC) and expanded the study to allow up to 10 metastases (from initially five). Finally, in order to maintain local study team engagement, we developed initiatives to maximize collaboration and add value to the overall clinical program through study participation.

Three-dimensional (3D) anatomic location, extension, and timing of severe osteoradionecrosis of the mandible.

Background: The purpose of this study was to describe the topography, extension (volume), and timing of severe osteoradionecrosis (ORN) that required mandible resection in patients previously treated for head and neck cancer at a high-volume Veterans Affairs Medical Center. Materials and methods: The records from a reference hyperbaric oxygen clinic were retrospectively analyzed (n = 50, 2018-2021). Inclusion criteria were: I) severe ORN defined as progressive ORN that required resection; II) pathologic confirmation of ORN; and III) availability of pre-operative CT-imaging. Using a radiotherapy (RT) imaging software, we performed a detailed volumetric (3D) analysis of the bone involvement by ORN. Time intervals from RT to surgery for ORN and from surgery to the last follow-up were calculated. Results: All patients that met inclusion criteria (n = 10) were male with significant smoking history (median 47.5 pack-years) and a median age of 57 years old at the time of RT. The primary tumors were: oropharynx (n = 6), oral cavity (n = 3) and nasopharynx (n = 1). The median time from RT to ORN surgery was 8 years. The most common ORN location was the posterior lateral body (molar) and six patients had associated fractures. The mean ORN volume was 3.6 cc (range: 0.6-8.3), corresponding to a mean 6.3% (range: 0.7-14) of the total mandibular volume. After a median follow-up of 13.5 months, no recurrence of ORN occurred. Three patients died of non-cancer and non-ORN-recurrence related causes (1 y OS 77.1%). Conclusion: Severe ORN occurred after a median of 8 years from the previous RT and usually affected the posterior lateral body. Surgical resection achieved excellent ORN control.

Persistent ethnicity-associated disparity in anti-tumor effectiveness of immune checkpoint inhibitors despite equal access.

We reviewed response to immune checkpoint inhibitors (ICI) of 207 patients with diagnoses of lung or head and neck cancer treated with chemotherapy/ICI combination therapy and ICI monotherapy between 2015 and 2020 at one of three clinical pavilions associated with the Dan L. Duncan Comprehensive Cancer Center at Baylor College of Medicine. Two of these pavilions (Harris Health System and the Michael E. DeBakey Veterans Affairs Medical Center) serve large minority populations and provide equal access to care regardless of means. 174 patients had a diagnosis of lung cancer (non-small cell or small cell) and 33 had a diagnosis of head and neck squamous cell carcinoma (HNSCC). 38% self-identified as Black, 45% as non-Hispanic White, and 18% as Hispanic. The objective response rate (ORR) was similar for lung cancer (35.057%) and HNSCC patients (30.3%) (p=0.894). The ORR for Hispanic and Black patients was lower compared to non-Hispanic White patients (H 27.0%, B 32.5%, W 38.7%; H vs. W p=0.209; B vs. W p=0.398). When considering only patients treated with ICI monotherapy, the ORR for Hispanic patients dropped further to 20.7% while the ORR of Black and non-Hispanic White patients remained about the same (B 29.3% and W 35.9%, H vs. W p=0.133; B vs. W p=0.419). Immune related adverse events were the lowest in the Hispanic population occurring in only 30% of patients compared to 40% of patients in the Black cohort and 50% of the non-Hispanic White cohorts.

Endoscopic assessment of radiological stage IVA cervical cancer: A bivariate meta-analysis supporting an evidence-based staging algorithm proposal.

OBJECTIVE: To optimize the use of confirmatory endoscopic exams (cystoscopy/proctoscopy) in the staging of locally advanced cervical cancer (LACC), the present study evaluates the predictive value of radiological exams (CT and MRI) to detect bladder/rectum invasion. METHODS: A systematic search of databases (PubMed and EMBASE) was performed (CRD42021270329). The inclusion criteria were: a) cervix cancer diagnosis; b) staging CT and/or MRI (index test); c) staging cystoscopy and/or proctoscopy (standard test); and d) numbers of true positives (TP), true negatives (TN), false positives (FP), and false negatives (FN) provided. A random-effects bivariate meta-analysis of positive predictive value (PPV) and negative predictive value (NPV) was performed with moderator analyses by imaging modality (CT and MRI) and prevalence. RESULTS: Nineteen studies met the inclusion criteria, totaling 3480 and 1641 patients for bladder and rectum analyses, respectively. For bladder invasion (prevalence ranged from 0.9% to 34.5%), the overall PPV was 45% (95% confidence interval, 33%-57%, based on 19 studies). Per subgroup, the PPV was 31% for MRI/prevalence ≤6%, 33% for CT/prevalence ≤6%, and 69% for CT/prevalence >6%. For rectal invasion (prevalence ranged from 0.4% to 20.0%), the overall PPV was 30% (95% confidence interval, 17%-47%, based on 8 studies). Per subgroup, the PPV was 36% for MRI/prevalence ≤1%, 17% for MRI/prevalence >1%, and 38% for CT/prevalence >1%. The overall NPV for bladder invasion and rectal invasion were 98% (95% confidence interval, 97%-99%) and 100% (95% confidence interval, 99%-100%), respectively. Considering prevalence and radiological modality, the point estimate of NPV varied from 95% to 100% for bladder invasion and from 99% to 100% for rectum invasion. CONCLUSIONS: Due to low PPV (<50%) of radiological staging, endoscopic exams may be necessary to correctly assess radiological stage IVA LACC. However, they are not necessary after negative radiological exam (NPV ≥95%).

Consistent multimodality approach to oral cavity and high-risk oropharyngeal cancer in veterans.

PURPOSE: High-risk oropharyngeal squamous cell carcinoma (OPSCC) associated with tobacco exposure remains difficult to treat due to high rates of locoregional recurrence similar to oral cavity squamous cell carcinoma (OCSCC). Current NCCN guidelines allow for surgical management of this disease, but oncologic and functional data in the modern era remain scarce. We sought to compare and contrast oncologic and functional considerations for surgical management of OPSCC and OCSCC in a cohort of Veterans. MATERIALS AND METHODS: We conducted a retrospective review of patients treated at the Michael E. DeBakey Veterans Affairs Medical Center between 2017 and 2020, treated using a homogenous, multi-modality algorithm. RESULTS: OPSCC tumors presented with a higher rate of perineural invasion (p < 0.05) and extranodal extension (p = 0.02) compared to OCSCC tumors. Compliance with NCCN guidelines for adjuvant treatment were lower for OPSCC patients primarily due to a higher rate of previous irradiation; re-irradiation could be delivered in 75% of patients when recommended by NCCN guidelines. Total glossectomy was accompanied by concomitant total laryngectomy in 100% of OPSCC patients and 0% of OCSCC. CONCLUSION: Surgical resection and free flap reconstruction of high-risk OPSCC generates oncologic outcomes comparable to OCSCC with comparable complication rates but a lower overall functional status. Reconstruction focused on rapid healing allows for high-rates of re-irradiation and minimal treatment delays. LEVEL OF EVIDENCE: level 4.

Matrix Rigidity Controls Epithelial-Mesenchymal Plasticity and Tumor Metastasis via a Mechanoresponsive EPHA2/LYN Complex.

Mechanical cues from the extracellular matrix (ECM) regulate various cellular processes via distinct mechanotransduction pathways. In breast cancer, increased ECM stiffness promotes epithelial-to-mesenchymal transition (EMT), cell invasion, and metastasis. Here, we identify a mechanosensitive EPHA2/LYN protein complex regulating EMT and metastasis in response to increasing ECM stiffness during tumor progression. High ECM stiffness leads to ligand-independent phosphorylation of ephrin receptor EPHA2, which recruits and activates the LYN kinase. LYN phosphorylates the EMT transcription factor TWIST1 to release TWIST1 from its cytoplasmic anchor G3BP2 to enter the nucleus, thus triggering EMT and invasion. Genetic and pharmacological inhibition of this pathway prevents breast tumor invasion and metastasis in vivo. In human breast cancer samples, activation of this pathway correlates with collagen fiber alignment, a marker of increasing ECM stiffness. Our findings reveal an EPHA2/LYN/TWIST1 mechanotransduction pathway that responds to mechanical signals from the tumor microenvironment to drive EMT, invasion, and metastasis.

The impact of molecular status on survival outcomes for invasive micropapillary carcinoma of the breast.

Invasive micropapillary carcinoma (IMPC) is an uncommon variant of breast cancer. Previous studies demonstrated this subtype is often hormone receptor (HR)-positive, resulting in survival outcomes similar to invasive ductal carcinoma. However, many of these studies were conducted prior to HER2 testing availability. We aim to determine the impact of molecular marker status (including HER2 status) on IMPC survival outcomes. The National Cancer Data Base (NCDB) was used to retrieve patients with biopsy-proven IMPC from 2007 to 2012. Only patients with known HR and HER2 status were included. Cox multivariate regression was used to determine prognostic factors. In total, 865 patients were included; median follow-up was 2.5 years. Overall, 651 patients (75.3%) had HR + HER2- disease, 128 (14.8%) had HR + HER2+ disease, 41 (4.7%) had HR-HER2 + disease, and 45 (5.2%) had triple negative disease. Patients with triple negative disease were more likely to have poorly differentiated histology (66.7%), lymphovascular invasion (73.3%), stage 3 disease (37.8%), undergone mastectomy (68.9%), and positive surgical margins (15.6%). On Cox multivariate regression, those with triple negative disease had worse overall survival (hazard ratio [HR] 7.28, P < 0.001). Other adverse prognostic factors included African-American descent (HR 2.24, P = 0.018), comorbidity score of 1 (HR 2.50, P = 0.011), comorbidity score ≥2 (HR 3.27, P = 0.06), and ≥3 positive lymph nodes (HR 3.23, P = 0.007). Similar to invasive ductal carcinoma, triple negative disease in IMPC results in worse survival outcomes. This is the largest and first study to characterize molecular status (including HER2 status) in patients with IMPC and its impact on survival outcomes.

Comparison of height and weight after 12 vs. 18 Gy cranial radiation therapy in pediatric acute lymphoblastic leukemia (ALL) patients.

PURPOSE: To compare the effect of 12 versus 18 Gy cranial radiation therapy (RT) on height and weight indices among pediatric patients with acute lymphoblastic leukemia (ALL). METHODS AND MATERIALS: Records of children with ALL who were 2 to 14 years old at the time of RT and were treated at a single institution between 2000 and 2011 were reviewed. Patients' height, weight, and body mass index were converted into z-scores using the Centers for Disease Control growth charts to normalize the values to number of standard deviations from the mean. These values were measured at the pre-RT clinic visit and subsequent yearly intervals. The z-scores of the growth indices were fitted into a generalizing estimating equations model and analyzed by various clinical factors. RESULTS: A total of 48 patients met the study criteria, including 32 boys and 16 girls. The median age at the time of RT was 7 years (range, 2-14 years). Patients were separated into 2 dose groups: 12 Gy (n = 30) and 18 Gy (n = 18). Median follow-up was 4.9 years (range, 3.0-11.8 years) and 6.0 years (range, 3.1-10.5 years) and the median pre-RT height z-scores were -0.55 (range, -2.2 to 1.4) and -0.85 (range, -3.1 to 0.8) for the 2 groups, respectively (P = .65). Patients who received 18 Gy had a significant difference in change in height compared with those who received 12 Gy, who were able to maintain normal growth during the first 3 years of follow-up. This did not appear to be sex-specific, and there was no difference in change in weight or body mass index. CONCLUSIONS: Compared with 18 Gy, patients with ALL who received 12 Gy of cranial RT had less height impairment in the first 3 years post-RT, but further prospective studies are needed.

Onsite versus offsite radiation treatment of malignant spinal cord compression: lessons from a safety net health system.

OBJECTIVE: Metastatic spinal cord compression (MSCC) is an oncologic emergency that often warrants emergent treatment; but, it is unclear whether radiation treatment (RT) can be optimally managed from an offsite radiotherapy facility. METHODS: Patient charts from consecutive patients with MSCC who were treated with radiotherapy alone at either an onsite hospital radiation department (from 2008 to 2012) or an offsite radiotherapy centre (2012-2015) were reviewed. Patient clinical parameters were compared across groups with either the χ2 test or Fisher's exact test, while survival curves were compared with the log-rank test. The primary end points were ambulatory rate over time, overall survival and cancer-specific survival. RESULTS: A total of 45 patients were identified, with 19 patients treated onsite in the hospital department and 26 patients treated at the offsite radiotherapy centre with median follow-up of 42 days vs 48.5 days, respectively. The ambulatory rate over time, overall survival and cancer-specific survival were not significantly different between the two eras. Patients treated in-hospital were more likely to start treatment the same day as the consult ("sim and treat") (79% vs 27%, p = 0.006) and were more likely to not complete treatment (26% vs 4%, p = 0.029) as compared with those treated in the offsite centre. CONCLUSION: Patients with MSCC can be feasibly treated at an offsite radiotherapy centre with outcomes similar to those treated in-hospital. Advances in knowledge: This is the first study in literature to compare outcomes between onsite and offsite RT of MSCC.

Addition of Mesenchymal Stem Cells to Autologous Platelet-Enhanced Fibrin Scaffolds in Chondral Defects: Does It Enhance Repair?

BACKGROUND: The chondrogenic potential of culture-expanded bone-marrow-derived mesenchymal stem cells (BMDMSCs) is well described. Numerous studies have also shown enhanced repair when BMDMSCs, scaffolds, and growth factors are placed into chondral defects. Platelets provide a rich milieu of growth factors and, along with fibrin, are readily available for clinical use. The objective of this study was to determine if the addition of BMDMSCs to an autologous platelet-enriched fibrin (APEF) scaffold enhances chondral repair compared with APEF alone. METHODS: A 15-mm-diameter full-thickness chondral defect was created on the lateral trochlear ridge of both stifle joints of twelve adult horses. In each animal, one defect was randomly assigned to receive APEF+BMDMSCs and the contralateral defect received APEF alone. Repair tissues were evaluated one year later with arthroscopy, histological examination, magnetic resonance imaging (MRI), micro-computed tomography (micro-CT), and biomechanical testing. RESULTS: The arthroscopic findings, MRI T2 map, histological scores, structural stiffness, and material stiffness were similar (p > 0.05) between the APEF and APEF+BMDMSC-treated repairs at one year. Ectopic bone was observed within the repair tissue in four of twelve APEF+BMDMSC-treated defects. Defects repaired with APEF alone had less trabecular bone edema (as seen on MRI) compared with defects repaired with APEF+BMDMSCs. Micro-CT analysis showed thinner repair tissue in defects repaired with APEF+BMDMSCs than in those treated with APEF alone (p < 0.05). CONCLUSIONS: APEF alone resulted in thicker repair tissue than was seen with APEF+BMDMSCs. The addition of BMDMSCs to APEF did not enhance cartilage repair and stimulated bone formation in some cartilage defects. CLINICAL RELEVANCE: APEF supported repair of critical-size full-thickness chondral defects in horses, which was not improved by the addition of BMDMSCs. This work supports further investigation to determine whether APEF enhances cartilage repair in humans.

Hyaluronan concentration and size distribution in human knee synovial fluid: variations with age and cartilage degeneration.

BACKGROUND: One potential mechanism for early superficial cartilage wear in normal joints is alteration of the lubricant content and quality of synovial fluid. The purpose of this study was to determine if the concentration and quality of the lubricant, hyaluronan, in synovial fluid: (1) was similar in left and right knees; (2) exhibited similar age-associated trends, whether collected postmortem or antemortem; and (3) varied with age and grade of joint degeneration. METHODS: Human synovial fluid of donors (23-91 years) without osteoarthritis was analyzed for the concentrations of protein, hyaluronan, and hyaluronan in the molecular weight ranges of 2.5-7 MDa, 1-2.5 MDa, 0.5-1 MDa, and 0.03-0.5 MDa. Similarity of data between left and right knees was assessed by reduced major axis regression, paired t-test, and Bland-Altman analysis. The effect of antemortem versus postmortem collection on biochemical properties was assessed for age-matched samples by unpaired t-test. The relationships between age, joint grade, and each biochemical component were assessed by regression analysis. RESULTS: Joint grade and the concentrations of protein, hyaluronan, and hyaluronan in the molecular weight ranges of 2.5-7 MDa, 1-2.5 MDa, and 0.5-1 MDa in human synovial fluid showed good agreement between left and right knees and were similar between age-matched patient and cadaver knee joints. There was an age-associated decrease in overall joint grade (-15 %/decade) and concentrations of hyaluronan (-10.5 %/decade), and hyaluronan in the molecular weight ranges of 2.5-7 MDa (-9.4 %/decade), 1-2.5 MDa (-11.3 %/decade), 0.5-1 MDa (-12.5 %/decade), and 0.03-0.5 MDa (-13.0 %/decade). Hyaluronan concentration and quality was more strongly associated with age than with joint grade. CONCLUSIONS: The age-related increase in cartilage wear in non-osteoarthritic joints may be related to the altered hyaluronan content and quality of synovial fluid.

Combined Mapping of Multiple clUsteriNg ALgorithms (COMMUNAL): A Robust Method for Selection of Cluster Number, K.

In order to discover new subsets (clusters) of a data set, researchers often use algorithms that perform unsupervised clustering, namely, the algorithmic separation of a dataset into some number of distinct clusters. Deciding whether a particular separation (or number of clusters, K) is correct is a sort of 'dark art', with multiple techniques available for assessing the validity of unsupervised clustering algorithms. Here, we present a new technique for unsupervised clustering that uses multiple clustering algorithms, multiple validity metrics, and progressively bigger subsets of the data to produce an intuitive 3D map of cluster stability that can help determine the optimal number of clusters in a data set, a technique we call COmbined Mapping of Multiple clUsteriNg ALgorithms (COMMUNAL). COMMUNAL locally optimizes algorithms and validity measures for the data being used. We show its application to simulated data with a known K, and then apply this technique to several well-known cancer gene expression datasets, showing that COMMUNAL provides new insights into clustering behavior and stability in all tested cases. COMMUNAL is shown to be a useful tool for determining K in complex biological datasets, and is freely available as a package for R.

Matrix stiffness drives epithelial-mesenchymal transition and tumour metastasis through a TWIST1-G3BP2 mechanotransduction pathway.

Matrix stiffness potently regulates cellular behaviour in various biological contexts. In breast tumours, the presence of dense clusters of collagen fibrils indicates increased matrix stiffness and correlates with poor survival. It is unclear how mechanical inputs are transduced into transcriptional outputs to drive tumour progression. Here we report that TWIST1 is an essential mechanomediator that promotes epithelial-mesenchymal transition (EMT) in response to increasing matrix stiffness. High matrix stiffness promotes nuclear translocation of TWIST1 by releasing TWIST1 from its cytoplasmic binding partner G3BP2. Loss of G3BP2 leads to constitutive TWIST1 nuclear localization and synergizes with increasing matrix stiffness to induce EMT and promote tumour invasion and metastasis. In human breast tumours, collagen fibre alignment, a marker of increasing matrix stiffness, and reduced expression of G3BP2 together predict poor survival. Our findings reveal a TWIST1-G3BP2 mechanotransduction pathway that responds to biomechanical signals from the tumour microenvironment to drive EMT, invasion and metastasis.

Prognostic markers for invasive micropapillary carcinoma of the breast: a population-based analysis.

BACKGROUND: Invasive micropapillary carcinoma (IMPC) is a rare and distinct variant of breast carcinoma with a high propensity for regional lymph node involvement. Because of its lymphotropic nature, IMPC is considered to have an unfavorable prognosis when compared with invasive ductal carcinoma (IDC). PATIENTS AND METHODS: This study of 624 patients diagnosed with breast IMPC (2001-2008) listed in the National Cancer Institute's Surveillance, Epidemiology, and End Results (NCI SEER) database was performed to evaluate prognostic factors for disease-specific survival (DSS) and overall survival (OS). RESULTS: The 5-year DSS and OS for patients with IMPC were 91.9% and 83.8%, respectively. Of those with known estrogen receptor (ER) status, 84.8% were ER-positive (ER(+)), which was associated with better DSS (hazard ratio [HR], 0.27; P < .0002) and OS (HR 0.45; P < .006). At presentation, 52.9% of the patients with lymph node examinations had nodal involvement and 4.1% had distant metastases. Patients with 4 or more positive lymph nodes had worse DSS (HR 6.43; P = .0013) and OS (HR 3.47; P = .00067) than did patients with node-negative disease, but those with 1 to 3 positive lymph nodes had DSS and OS similar to those of patients with node-negative disease. CONCLUSION: Although IMPC has a high propensity for lymph node metastasis, it has a DSS and overall prognosis comparable to those of IDC. Patients with ER-negative (ER-) disease or those with 4 or more positive lymph nodes have the worst prognosis. This is the largest study of IMPC to date, and these findings will help address some of the inconsistencies regarding this rare histologic variant of breast cancer.

Treatment of articular cartilage defects in the goat with frozen versus fresh osteochondral allografts: effects on cartilage stiffness, zonal composition, and structure at six months.

BACKGROUND: Understanding the effectiveness of frozen as compared with fresh osteochondral allografts at six months after surgery and the resultant consequences of traditional freezing may facilitate in vivo maintenance of cartilage integrity. Our hypothesis was that the state of the allograft at implantation affects its performance after six months in vivo. METHODS: The effect of frozen as compared with fresh storage on in vivo allograft performance was determined for osteochondral allografts that were transplanted into seven recipient goats and analyzed at six months. Allograft performance was assessed by examining osteochondral structure (cartilage thickness, fill, surface location, surface degeneration, and bone-cartilage interface location), zonal cartilage composition (cellularity, matrix content), and cartilage biomechanical function (stiffness). Relationships between cartilage stiffness or cartilage composition and surface degeneration were assessed with use of linear regression. RESULTS: Fresh allografts maintained cartilage load-bearing function, while also maintaining zonal organization of cartilage cellularity and matrix content, compared with frozen allografts. Overall, allograft performance was similar between fresh allografts and nonoperative controls. However, cartilage stiffness was approximately 80% lower (95% confidence interval [CI], 73% to 87%) in the frozen allografts than in the nonoperative controls or fresh allografts. Concomitantly, in frozen allografts, matrix content and cellularity were approximately 55% (95% CI, 22% to 92%) and approximately 96% (95% CI, 94% to 99%) lower, respectively, than those in the nonoperative controls and fresh allografts. Cartilage stiffness correlated positively with cartilage cellularity and matrix content, and negatively with surface degeneration. CONCLUSIONS: Maintenance of cartilage load-bearing function in allografts is associated with zonal maintenance of cartilage cellularity and matrix content. In this animal model, frozen allografts displayed signs of failure at six months, with cartilage softening, loss of cells and matrix, and/or graft subsidence, supporting the importance of maintaining cell viability during allograft storage and suggesting that outcomes at six months may be indicative of long-term (dys)function. CLINICAL RELEVANCE: Fresh versus frozen allografts represent the "best versus worst" conditions with respect to chondrocyte viability, but "difficult versus simple" with respect to acquisition and distribution. The outcomes described from these two conditions expand the current understanding of in vivo cartilage remodeling and describe structural properties (initial graft subsidence), which may have implications for impending graft failure.

Cartilage-like mechanical properties of poly (ethylene glycol)-diacrylate hydrogels.

Hydrogels prepared from poly-(ethylene glycol) (PEG) have been used in a variety of studies of cartilage tissue engineering. Such hydrogels may also be useful as a tunable mechanical material for cartilage repair. Previous studies have characterized the chemical and mechanical properties of PEG-based hydrogels, as modulated by precursor molecular weight and concentration. Cartilage mechanical properties vary substantially, with maturation, with depth from the articular surface, in health and disease, and in compression and tension. We hypothesized that PEG hydrogels could mimic a broad range of the compressive and tensile mechanical properties of articular cartilage. The objective of this study was to characterize the mechanical properties of PEG hydrogels over a broad range and with reference to articular cartilage. In particular, we assessed the effects of PEG precursor molecular weight (508 Da, 3.4 kDa, 6 kDa, and 10 kDa) and concentration (10-40%) on swelling property, equilibrium confined compressive modulus (H(A0)), compressive dynamic stiffness, and hydraulic permeability (k(p0)) of PEG hydrogels in static/dynamic confined compression tests, and equilibrium tensile modulus (E(ten)) in tension tests. As molecular weight of PEG decreased and concentration increased, hydrogels exhibited a decrease in swelling ratio (31.5-2.2), an increase in H(A0) (0.01-2.46 MPa) and E(ten) (0.02-3.5 MPa), an increase in dynamic compressive stiffness (0.055-42.9 MPa), and a decrease in k(p0) (1.2 × 10(-15) to 8.5 × 10(-15) m(2)/(Pa s)). The frequency-dependence of dynamic compressive stiffness amplitude and phase, as well as the strain-dependence of permeability, were typical of the time- and strain-dependent mechanical behavior of articular cartilage. H(A0) and E(ten) were positively correlated with the final PEG concentration, accounting for swelling. These results indicate that PEG hydrogels can be prepared to mimic many of the static and dynamic mechanical properties of articular cartilage.

SGK3 is associated with estrogen receptor expression in breast cancer.

While breast cancer mortality rate has seen a steady decline in the last few decades, advances in better treatment and diagnostic tools remain important as we come into the age of personalized therapy. In this report, we describe our studies of SGK3's role in breast cancer. SGK3 (also known as CISK) is a member of the AGC family of kinases. Our previous work indicates that SGK3 functions downstream of the PI 3-kinase cascade and shares molecular and biochemical similarities with Akt. Here, we show that SGK3 expression is linked to estrogen receptor (ER) both in breast caner cell lines and in primary tumor samples. Our analysis also indicated a positive correlation between SGK3 expression and tumor prognosis. Importantly, our immunochemistry analysis of human tumor samples established a clinical link between SGK3 expression and ER+ tumors. These findings implicate SGK3 as an additional component to a complex and heterogeneous disease, and point to the potential benefits of incorporating SGK3 into the process of breast cancer diagnosis and treatment.

Upregulation of mucin4 in ER-positive/HER2-overexpressing breast cancer xenografts with acquired resistance to endocrine and HER2-targeted therapies.

We studied resistance to endocrine and HER2-targeted therapies using a xenograft model of estrogen receptor positive (ER)/HER2-overexpressing breast cancer. Here, we report a novel phenotype of drug resistance in this model. MCF7/HER2-18 xenografts were treated with endocrine therapy alone or in combination with lapatinib and trastuzumab (LT) to inhibit HER2. Archival tumor tissues were stained with hematoxylin and eosin and with mucicarmine. RNA extracted from tumors at early time points and late after acquired resistance were analyzed for mucin4 (MUC4) expression by microarray and quantitative reverse transcriptase-PCR. Protein expression of the MUC4, ER, and HER2 signaling pathways was measured by immunohistochemistry and western blotting. The combination of the potent anti-HER2 regimen LT with either tamoxifen (Tam + LT) or estrogen deprivation (ED + LT) can cause complete eradication of ER-positive/HER2-overexpressing tumors in mice. Tumors developing resistance to this combination, as well as those acquiring resistance to endocrine therapy alone, exhibited a distinct histological and molecular phenotype-a striking increase in mucin-filled vacuoles and upregulation of several mucins including MUC4. At the onset of resistance, MUC4 mRNA and protein were increased. These tumors also showed upregulation and reactivation of HER2 signaling, while losing ER protein and the estrogen-regulated gene progesterone receptor. Mucins are upregulated in a preclinical model of ER-positive/HER2-overexpressing breast cancer as resistance develops to the combination of endocrine and anti-HER2 therapy. These mucin-rich tumors reactivate the HER2 pathway and shift their molecular phenotype to become more ER-negative/HER2-positive.

Association of 3-Dimensional Cartilage and Bone Structure with Articular Cartilage Properties in and Adjacent to Autologous Osteochondral Grafts after 6 and 12 months in a Goat Model.

OBJECTIVE: The articular cartilage of autologous osteochondral grafts is typically different in structure and function from local host cartilage and thereby presents a remodeling challenge. The hypothesis of this study was that properties of the articular cartilage of trochlear autografts and adjacent femoral condyle are associated with the 3-D geometrical match between grafted and contralateral joints at 6 and 12 months after surgery. DESIGN: Autografts were transferred unilaterally from the lateral trochlea (LT) to the medial femoral condyle (MFC) in adult Spanish goats. Operated and contralateral Non-Operated joints were harvested at 6 and 12 months, and analyzed by indentation testing, micro-computed tomography, and histology to compare (1) histological indices of repair, (2) 3-D structure (articular surface deviation, bone-cartilage interface deviation, cartilage thickness), (3) indentation stiffness, and (4) correlations between stiffness and 3-D structure. RESULTS: Cartilage deterioration was present in grafts at 6 months and more severe at 12 months. Cartilage thickness and normalized stiffness of Operated MFC were lower than Non-Operated MFC within the graft and proximal adjacent host regions. Operated MFC articular surfaces were recessed relative to Non-Operated MFC and exhibited lower cartilage stiffness with increasing recession. Sites with large bone-cartilage interface deviations, both proud and recessed, were associated with recessed articular surfaces and low cartilage stiffness. CONCLUSION: The effectiveness of cartilage repair by osteochondral grafting is associated with the match of 3-D cartilage and bone geometry to the native osteochondral structure.

Insulin-like growth factor-I and growth differentiation factor-5 promote the formation of tissue-engineered human nasal septal cartilage.

INTRODUCTION: Tissue engineering of human nasal septal chondrocytes offers the potential to create large quantities of autologous material for use in reconstructive surgery of the head and neck. Culture with recombinant human growth factors may improve the biochemical and biomechanical properties of engineered tissue. The objectives of this study were to (1) perform a high-throughput screen to assess multiple combinations of growth factors and (2) perform more detailed testing of candidates identified in part I. METHODS: In part I, human nasal septal chondrocytes from three donors were expanded in monolayer with pooled human serum (HS). Cells were then embedded in alginate beads for 2 weeks of culture in medium supplemented with 2% or 10% HS and 1 of 90 different growth factor combinations. Combinations of insulin-like growth factor-I (IGF-1), bone morphogenetic protein (BMP)-2, BMP-7, BMP-13, growth differentiation factor-5 (GDF-5), transforming growth factor ß (TGFß)-2, insulin, and dexamethasone were evaluated. Glycosaminoglycan (GAG) accumulation was measured. A combination of IGF-1 and GDF-5 was selected for further testing based on the results of part I. Chondrocytes from four donors underwent expansion followed by three-dimensional alginate culture for 2 weeks in medium supplemented with 2% or 10% HS with or without IGF-1 and GDF-5. Chondrocytes and their associated matrix were then recovered and cultured for 4 weeks in 12 mm transwells in medium supplemented with 2% or 10% HS with or without IGF-1 and GDF-5 (the same medium used for alginate culture). Biochemical and biomechanical properties of the neocartilage were measured. RESULTS: In part I, GAG accumulation was highest for growth factor combinations including both IGF-1 and GDF-5. In part II, the addition of IGF-1 and GDF-5 to 2% HS resulted in a 12-fold increase in construct thickness compared with 2% HS alone (p < 0.0001). GAG and type II collagen accumulation was significantly higher with IGF-1 and GDF-5. Confined compression modulus was greatest with 2% HS, IGF-1, and GDF-5. CONCLUSION: Supplementation of medium with IGF-1 and GDF-5 during creation of neocartilage constructs results in increased accumulation of GAG and type II collagen and improved biomechanical properties compared with constructs created without the growth factors.