The roles of homologous recombination and the immune system in the genomic evolution of cancer.

A variety of factors, whether extracellular (mutagens/carcinogens and viruses in the environment, chronic inflammation and radiation associated with the environment and/or electronic devices/machines) and/or intracellular (oxidative metabolites of food, oxidative stress due to inflammation, acid production, replication stress, DNA replication/repair errors, and certain hormones, cytokines, growth factors), pose a constant threat to the genomic integrity of a living cell. However, in the normal cellular environment multiple biological pathways including DNA repair, cell cycle, apoptosis and the immune system work in a precise, regulated (tightly controlled), timely and concerted manner to ensure genomic integrity, stability and proper functioning of a cell. If damage to DNA takes place, it is efficiently and accurately repaired by the DNA repair systems. Homologous recombination (HR) which utilizes either a homologous chromosome (in G1 phase) or a sister chromatid (in G2) as a template to repair the damage, is known to be the most precise repair system. HR in G2 which utilizes a sister chromatid as a template is also called an error free repair system. If DNA damage in a cell is so extensive that it overwhelms the repair system/s, the cell is eliminated by apoptosis. Thus, multiple pathways ensure that genome of a cell is intact and stable. However, constant exposure to DNA damage and/or dysregulation of DNA repair mechanism/s poses a risk of mutation and cancer. Oncogenesis, which seems to be a multistep process, is associated with acquisition of a number of genomic changes that enable a normal cell to progress from benign to malignant transformation. Transformed/cancer cells are recognized and killed by the immune system. However, the ongoing acquisition of new genomic changes enables cancer cells to survive/escape immune attack, evolve into a more aggressive phenotype, and eventually develop resistance to therapy. Although DNA repair (especially the HR) and the immune system play unique roles in preserving genomic integrity of a cell, they can also contribute to DNA damage, genomic instability and oncogenesis. The purpose of this article is to highlight the roles of DNA repair (especially HR) and the immune system in genomic evolution, with special focus on gastrointestinal cancer.

Targeting IL-17A in multiple myeloma: a potential novel therapeutic approach in myeloma.

We have previously demonstrated that interleukin-17A (IL-17) producing T helper 17 cells are significantly elevated in blood and bone marrow (BM) in multiple myeloma (MM) and IL-17A promotes MM cell growth via the expression of IL-17 receptor. In this study, we evaluated anti-human IL-17A human monoclonal antibody (mAb), AIN457 in MM. We observe significant inhibition of MM cell growth by AIN457 both in the presence and the absence of BM stromal cells (BMSCs). Although IL-17A induces IL-6 production, AIN457 significantly downregulated IL-6 production and MM cell adhesion in MM-BMSC co-culture. AIN457 also significantly inhibited osteoclast cell differentiation. More importantly, in the SCIDhu model of human myeloma administration of AIN457 weekly for 4 weeks after the first detection of tumor in mice led to a significant inhibition of tumor growth and reduced bone damage compared with isotype control mice. To understand the mechanism of action of anti-IL-17A mAb, we report, here, that MM cells express IL-17A. We also observed that IL-17A knockdown inhibited MM cell growth and their ability to induce IL-6 production in co-cultures with BMSC. These pre-clinical observations suggest efficacy of AIN457 in myeloma and provide the rationale for its clinical evaluation for anti-myeloma effects and for improvement of bone disease.

Phase II trial of neoadjuvant temozolomide in resectable melanoma patients.

BACKGROUND: We treated melanoma patients with temozolomide (TMZ) in the neoadjuvant setting and collected cryopreserved tumor samples before and after treatment. The primary objective was to determine whether the response proportion was higher than previously reported in widely metastatic patients. A secondary objective was to test the feasibility of obtaining adequate tissue before and after treatment for genetic testing. MATERIALS AND METHODS: Chemotherapy-naive melanoma patients who were candidates for surgical resection were eligible. TMZ was administered orally at 75 mg/m(2)/day for 6 weeks of every 8-week cycle. Cycles were repeated until complete response (CR), progression, or stable disease (SD) for two cycles. RESULTS: Of 19 assessable patients, 2 had CRs and 1 had partial response. Four patients had SD; 12 progressed. Tumor O-6-methylguanine-DNA methyltransferase (MGMT) promoter was unmethylated in all nine patients analyzed including from the two CR patients. Pretreatment tumor microarray results were obtained in 16 of 19 patients. CONCLUSIONS: The response proportion to TMZ in the neoadjuvant setting was 16%, not different than in the metastatic setting. Responses were seen even in tumors with a methylated MGMT promoter. Pretreatment cryopreserved tumor adequate for microarray analysis could be obtained in most, but not all, patients. Post-treatment tumor was unavailable in complete responders.

Resection of the sciatic, peroneal, or tibial nerves: assessment of functional status.

BACKGROUND: Lower-extremity tumors are often treated by amputation rather than limb-sparing excision that sacrifices the sciatic nerve or a branch. This study assessed the functional outcome of major nerve sacrifice during limb-sparing resections for lower-extremity soft tissue sarcoma. METHODS: Patients who underwent division of the sciatic, tibial, or peroneal nerve(s) during limb-sparing sarcoma surgery (January 1982 through June 2000) were identified. Eleven surviving patients evaluated their pre- and postoperative functional status by self-administered questionnaire (six sciatic, two tibial, and three peroneal nerve divisions). RESULTS: Eighteen patients (10 male, 8 female; 14-84 years old) had nine primary and nine locally recurrent tumors. Tumors were high (16) or low grade (two). Five patients died of disease and two died of other causes. Median overall survival was 50 months. One of 11 reported increased pain. Eight had new phantom sensations with a median intensity of 4.5 (1 = least; 10 = most). All patients used an ankle brace to walk after a sciatic (four) or peroneal (one) division. Walking ability and distance after surgery was unchanged (nine), improved (one), and worsened (one). Standing improved in 7 of 11 patients. Proprioception in the affected extremity was retained in six. The median postoperative leg functional score was 8 (1 = worst; 10 = best). No patient developed foot ulcers. One patient underwent amputation for recurrence. All patients preferred their status over having an amputation. CONCLUSIONS: Objectively and subjectively, division of the major lower-extremity nerves causes acceptable functional deficits in most patients. Resection of affected sciatic nerve (branches) during limb-sparing tumor surgery is an excellent alternative to amputation.

The present and future of appointment, tenure, and compensation policies for medical school clinical faculty.

The authors present data and information about appointment, tenure, and compensation policies to describe how medical schools are redefining the terms under which they relate to their full-time clinical faculties. First, the authors note the increasing differentiation of clinical faculty members into two groups, researchers and clinicians. The present-day competitive realities of both research and clinical enterprises have prompted this change and the principles of mission-based management are reinforcing it. Second, they document the long-term tendency of schools to appoint new clinical faculty members to contract-term (as opposed to tenure) appointments, as special non-tenure-eligible tracks for clinically oriented faculty proliferate. Third, they report on the policies of schools to limit the financial guarantees provided to clinical faculty members who are awarded tenure. For schools that have yet to address this issue, they discuss the various employment and pay arrangements that inform or confuse the question. Fourth, they describe historic problems with clinical faculty compensation arrangements and illustrate, with examples from ten schools, the characteristics of recently implemented performance- and risk-based compensation plans. While these trends in institutional policies and practices may initially concern faculty advocate groups, the authors argue that they may serve the long-term interests of those groups. The terms of relationships between medical schools and their clinical faculties are tied closely to the specifics of organizational structure, which are currently undergoing review and change. The challenge all schools face is to define these terms in ways that allow them to continue to attract high-quality clinical faculty while avoiding an insupportable financial liability.

Immunity against cancer: lessons learned from melanoma.

Most major advances in human cancer immunology and immunotherapy have come from studies in melanoma. We are beginning to understand the immune repertoire of T cells and antibodies that are active against melanoma, with recent glimpses of the CD4(+) T cell repertoire. The view of what the immune system can see is extending to mutations and parts of the genome that are normally invisible.

Clinicopathologic analysis of patients with adult rhabdomyosarcoma.

BACKGROUND: Rhabdomyosarcoma (RMS) in adults (age > or = 16 years) is rare, accounting for less than 3% of adult soft tissue sarcomas. There is little information describing the disease biology or clinicopathologic factors that influence survival in adults with RMS. The objective of this study was to define the factors in patients with adult RMS that predict outcome, disease progression, and survival. METHODS: Eighty-four adult patients with a pathologic diagnosis of RMS that was confirmed by immunohistochemistry were identified by a prospective inpatient data base during the period 1982--1999 and were analyzed for disease specific survival and metastasis free survival using the Kaplan-Meier actuarial method. Statistical significance was evaluated using the log-rank test for univariate influence and a Cox regression model for multivariate influence. RESULTS: The median disease specific survival was 22 months. Patient age, extent of disease, tumor size at the time of diagnosis, and margin status after resection were significant predictors of disease specific survival. Patients who underwent a complete resection had a significantly longer median survival (105 months) compared with any other subgroup of patients. The histologic subtype did not predict patient survival but did vary with patient age. Most notably, the proportion of the pleomorphic subtype increased with advancing age, accounting for 42% of RMS in patients over the age of 40 years. CONCLUSIONS: The most important predictors of outcome in patients with adult RMS are patient age, tumor size, extent of disease, and margin status after resection. In contrast to patients with pediatric RMS, no association was noted between survival and histologic subtype in this group of patients with adult RMS. All histologic subtypes of RMS are aggressive malignancies with poor disease specific survival despite aggressive multimodality management.

Trends in US medical school faculty salaries, 1988-1989 to 1998-1999.

Expansion of managed care, intensified price competition, and the introduction of the Medicare Fee Schedule have all affected physician compensation during the past decade. We examine trends in the salaries of medical school faculty, particularly MD clinical faculty, based on a more extensive salary database than has been used previously. Data collected through the Association of American Medical Colleges' Faculty Salary Survey for the academic years 1988-1989, 1993-1994, and 1998-1999 were analyzed, and inflation-adjusted salary growth rates for clinical and basic science faculty during two 5-year periods, 1988-1993 and 1993-1998, compared across faculty ranks, departments, and various school characteristics. The comparison showed that, between 1988 and 1998, the actual median clinical faculty salary increased from $101,000 to $150,000, and the actual median basic science faculty salary increased from $52,000 to $78,000. Bivariate and multivariate analyses showed that the proportionate change in real mean salary (base year, 1988) in each 5-year period was related to department and faculty rank for clinical faculty (P<.001) and faculty rank for basic science faculty (P<.001). The inflation-adjusted annualized compound growth rate of clinical faculty salaries declined from 1.9% per year (1988-1993) to 0.2% per year (1993-1998), while the growth rate of basic science faculty salaries increased from 0.3% per year (1988-1993) to 1.3% per year (1993-1998). From 1993 to 1998, inflation-adjusted annualized salary growth rates in several clinical departments were negative (anesthesiology, -1.1%; obstetrics and gynecology, -0.5%; radiology, -0.4%; and neurology, -0.1%) but were positive for family practice (+2.7%). Significant differences in salary growth related to school characteristics (eg, geographic region, public vs private, community based vs non-community based, and research intensity) were specific to particular study periods. Overall, while actual average medical school faculty salaries are increasing, the real growth rate of average clinical faculty salaries is declining and that of basic science faculty increasing. JAMA. 2000;284:1130-1135

Immunization with DNA coding for gp100 results in CD4 T-cell independent antitumor immunity.

BACKGROUND: Xenogeneic DNA immunization can exploit small differences in expressed protein sequence resulting in immune recognition of self-molecules. We hypothesized that immunizing mice with xenogeneic DNA coding for the human melanosomal membrane glycoprotein gp100 would overcome immune ignorance or tolerance and result in tumor immunity. We also investigated the immunologic mechanisms of the antitumor immunity. METHODS: C57BL/6 mice were immunized with DNA coding for human gp100, mouse gp100, or control vector by gene gun. After immunization, mice were challenged with a syngeneic melanoma expressing gp100, and tumor growth was analyzed. Mice deficient in major histocompatibility complex class I or class II molecules were similarly studied to assess the immunologic mechanism of the tumor protection. RESULTS: There was significant tumor protection after vaccination with xenogeneic human gp100 DNA. Class I, but not class II, major histocompatibility complex molecules were required for tumor immunity. In addition, mice immunized with human gp100 demonstrated autoimmunity manifested as coat color depigmentation. CONCLUSIONS: Immunization with xenogeneic DNA coding for the melanosomal glycoprotein gp100 results in tumor protection and autoimmune depigmentation. These results show that xenogeneic DNA vaccines can lead to cancer immunity without CD4(+) T-cell help with potential implications for rational vaccine design.

Faculty appointment and tenure policies in medical schools: a 1997 status report.

The authors present recent data on changes under way in and the current status of faculty appointment and tenure policies in U.S. medical schools. The data are drawn from a survey conducted by the Association of American Medical Colleges in 1997, to which deans at all 125 U.S. allopathic medical schools responded, supplemented by follow-up telephone and electronic mail inquiries. Faculty evaluation systems and faculty compensation systems top the list of areas in which medical schools are most frequently making policy changes, with approximately half of the schools involved in each area. Changes in evaluation systems reflect an increasing emphasis on post-tenure review. Changes in compensation systems are characterized by the division of pay into separate components, each with its own financial guarantees and with the level of compensation tied specifically to measures of individual and group productivity. Other policy changes include introducing new faculty tracks and career pathways, redefining or clarifying the portion of salary or compensation that is defined by tenure, lengthening the pre-tenure probationary period, and modifying the link between promotion and tenure. Of the 125 medical schools in the United States, only five do not award tenure, while another six effectively limit eligibility for tenure to basic science faculty. These numbers are unchanged from those reported in 1994. Only two schools indicated that eliminating tenure or ceasing to make tenure-eligible appointments was being considered, and neither reported that a policy change was imminent. Current data on the status of tenure guarantees, tenure probationary periods, other tenure eligibility criteria, and special clinical tracks are provided. Nearly three fourths of the medical schools in the United States now have a separate and distinct faculty track for full-time clinical faculty whose primary responsibilities are in patient care and teaching. The vast majority of these tracks do not permit faculty to be tenured, but 71% require evidence of scholarship for promotion. The authors conclude that faculty personnel policies in medical schools are likely to continue to evolve, consistent with a growing insinuation of the corporate culture into academia.

Loss of p120ctn in human colorectal cancer predicts metastasis and poor survival.

The p120ctn protein (formerly p120CAS) is an armadillo family member that associates directly with the cytoplasmic tail of E-cadherin and participates in the junctional complex responsible for cell-cell adhesion. Since reduced cell-cell adhesion is associated with metastasis in colorectal cancer and other neoplasms, we hypothesize that reduced expression of p120ctn may be related to metastasis in colorectal tumors. Here we describe a study of p120ctn expression in 44 primary human colorectal adenocarcinomas. As detected by immunohistochemical methods, we find altered p120ctn staining patterns in 86% of the cases. Regional complete loss of expression is seen in 18% of the cases, and it correlates with high stage disease and nodal metastasis as well as decreased survival. Although this is a preliminary study, it suggests that downregulation of p120ctn in colon cancer may be associated with metastasis and poor clinical outcome.

Localization and quantitation of expression of the cell motility-related protein thymosin beta15 in human breast tissue.

Thymosin beta15 is a newly discovered 5300-Da protein that binds actin monomers and inhibits actin polymerization and might thus increase cellular motility. Thymosin beta15 is upregulated at both the mRNA and protein levels in prostate cell lines in a manner directly related to their capacity to metastasize. We hypothesize that because this protein is upregulated in cells with a propensity to metastasize, it might be a useful prognostic marker in breast cancer. Because this is a newly described protein, neither the subcellular localization of thymosin beta15 or its expression in breast cancer has been examined. We describe the use of an affinity-purified polyclonal antibody to show that within breast epithelium, thymosin beta15 is localized diffusely throughout the cytoplasm and that thymosin beta15 is upregulated in malignant (compared with benign) breast tissue. In contrast to the prostate model, thymosin beta15 is upregulated in nonmetastatic breast cancer and even ductal carcinoma in situ (compared with benign breast tissue), and, consequently, it might represent a potential early marker for breast malignancy. Additional studies are needed to evaluate the precise role and prognostic value of thymosin beta15 in breast cancer.

Cathodic adsorptive stripping voltammetry of nickel complexed with hydroxynaphthol blue at a static mercury drop electrode.

A new method is described for the determination of Ni based on the cathodic adsorptive stripping of Ni(II) complexed with hydroxynaphthol blue (HNB) at a static mercury drop electrode. Optimal conditions were found to be: accumulation potential -0.50 V (vs. Ag/AgCl); final potential -1.10 V; accumulation time 50 sec; scan rate 200 mV/sec; linear scan mode; filter 0.1 sec; supporting electrolyte acetic acid/acetate (0.25M, pH = 6.0) and concentration of HNB 3.3 x 10(-5)M. The response of the system was found to be linear in a range of Ni concentrations from 25 ppb to the detection limit. The detection limit was found to be 1.7 nM (0.10 ppb) with 2 mins of accumulation time. The effect of various potential interferences (including a variety of cations, anions and organic surfactants) were also studied. With the exception of Co, at less than equimolar concentrations no significant interferences were observed. Al was found to interfere at high concentrations with respect to Ni, but Al concentrations up to 1000 ppb may be masked by sodium citrate or sodium fluoride. The utility of the method is demonstrated by the recovery of Ni in a doped sample of commercial mineral water.

Identification of cis-acting regulatory elements controlling interleukin-4 gene expression in T cells: roles for NF-Y and NF-ATc.

Activity of the murine interleukin-4 (IL-4) promoter was localized to several cis-acting elements present within the first 300 bp from the transcriptional initiation site. Five repeated elements, P0 to P4, that share the common consensus ATTTTCCNNT were located between -40 and -250, and each was shown to interact with the T-cell-specific factor NF(P). These distinct P sites appear functionally interchangeable and cooperatively confer cyclosporin A-sensitive and ionomycin-inducible promoter activity. NF(P) may be closely related to the cytoplasmic component of NF-AT (nuclear factor of activated T cells), a T-cell-specific factor essential for IL-2 gene transcription, as judged from indistinguishable molecular weights and protease fragmentation patterns of UV-photolabeled factors. Also, we identified an element in the IL-4 promoter with homology to the Y box common to all major histocompatibility complex class II gene promoters. Our data show that the IL-4 promoter Y box -114CTGATTGG-107 significantly enhances overall promoter activity, since point mutations within this element diminish promoter activity by 85%. The factor binding this region is indistinguishable from the cloned nuclear factor NF-Y, as judged from interactions with specific anti-NF-Y monoclonal and polyclonal antibodies. Last, we point out the presence of two sites that share sequence identity to the OAP region of the ARRE-1 site within the IL-2 promoter (K. S. Ullman, W. M. Flanagan, C. A. Edwards, and G. R. Crabtree, Science 254:558-562, 1991). These regions, -85GTGTAATA-78 and -245GTGTAATT-238, reside adjacent to the NF(P) binding sites P1 and P4 and bind a distinct nuclear factor.

Differential regulation of T helper phenotype development by interleukins 4 and 10 in an alpha beta T-cell-receptor transgenic system.

To address the mechanisms controlling T helper (Th) phenotype development, we used DO10, a transgenic mouse line that expresses the alpha beta T-cell receptor from an ovalbumin-reactive T hybridoma, as a source of naive T cells that can be stimulated in vitro with ovalbumin peptide presented by defined antigen-presenting cells (APCs). We have examined the role of cytokines and APCs in the regulation of Th phenotype development. Interleukin 4 (IL-4) directs development toward the Th2 phenotype, stimulating IL-4 and silencing IL-2 and interferon gamma production in developing T cells. Splenic APCs direct development toward the Th1 phenotype when endogenous IL-10 is neutralized with anti-IL-10 antibody. The splenic APCs mediating these effects are probably macrophages or dendritic cells and not B cells, since IL-10 is incapable of affecting Th phenotype development when the B-cell hybridoma TA3 is used as the APC. These results suggest that early regulation of IL-4 and IL-10 in a developing immune response and the identity of the initiating APCs are critical in determining the Th phenotype of the developing T cells.

Melting of diamond.

Radiation from a Q-switched YAG laser, focused on the (100) face of a single crystal diamond anvil in a high-pressure diamond cell, caused a portion of the diamond anvil face to melt. Potassium bromide mixed with graphite was under pressure between the anvils when melting occurred. The diamond surface melted at pressures greater than approximately 120 kilobars and graphitized at lower pressures. Evidence for the melting and graphitization of the diamond was obtained by optical and scanning electron microscopy.