Phase 1 study of twice weekly pulse dose and daily low-dose erlotinib as initial treatment for patients with EGFR-mutant lung cancers.

Background: Patients with EGFR-mutant lung cancers treated with EGFR tyrosine kinase inhibitors (TKIs) develop clinical resistance, most commonly with acquisition of EGFR T790M. Evolutionary modeling suggests that a schedule of twice weekly pulse and daily low-dose erlotinib may delay emergence of EGFR T790M. Pulse dose erlotinib has superior central nervous system (CNS) penetration and may result in superior CNS disease control. Methods: We evaluated toxicity, pharmacokinetics, and efficacy of twice weekly pulse and daily low-dose erlotinib. We assessed six escalating pulse doses of erlotinib. Results: We enrolled 34 patients; 11 patients (32%) had brain metastases at study entry. We observed 3 dose-limiting toxicities in dose escalation: transaminitis, mucositis, and rash. The MTD was erlotinib 1200 mg days 1-2 and 50 mg days 3-7 weekly. The most frequent toxicities (any grade) were rash, diarrhea, nausea, fatigue, and mucositis. 1 complete and 24 partial responses were observed (74%, 95% CI 60-84%). Median progression-free survival was 9.9 months (95% CI 5.8-15.4 months). No patient had progression of an untreated CNS metastasis or developed a new CNS lesion while on study (0%, 95% CI 0-13%). Of the 18 patients with biopsies at progression, EGFR T790M was identified in 78% (95% CI 54-91%). Conclusion: This is the first clinical implementation of an anti-cancer TKI regimen combining pulse and daily low-dose administration. This evolutionary modeling-based dosing schedule was well-tolerated but did not improve progression-free survival or prevent emergence of EGFR T790M, likely due to insufficient peak serum concentrations of erlotinib. This dosing schedule prevented progression of untreated or any new central nervous system metastases in all patients.

Nomogram to predict the presence of EGFR activating mutation in lung adenocarcinoma.

Epidermal growth factor receptor (EGFR) tumour genotyping is crucial to guide treatment decisions regarding the use of EGFR tyrosine kinase inhibitors in nonsmall cell lung cancer (NSCLC). However, some patients may not be able to obtain tumour testing, either because tissue is limited and/or tests are not routinely offered. Here, we aimed to build a model-based nomogram to allow for prediction of the presence of EGFR mutations in NSCLC. We retrospectively collected clinical and pathological data on 3,006 patients with NSCLC who had their tumours genotyped for EGFR mutations at five institutions worldwide. Variables of interest were integrated in a multivariate logistic regression model. In the 2,392 non-Asian patients with lung adenocarcinomas, the most important predictors of harbouring EGFR mutation were: lower tobacco smoking exposure (OR 0.41, 95% CI 0.37-0.46), longer time interval between smoking cessation and diagnosis (OR 2.19, 95% CI 1.71-2.80), advanced stage (OR 1.58, 95% CI 1.18-2.13), and papillary (OR 4.57, 95% CI 3.14-6.66) or bronchioloalveolar (OR 2.84, 95% CI 1.98-4.06) histologically predominant subtype. A nomogram was established and showed excellent discriminating accuracy: the concordance index on an independent validation dataset was 0.84. As clinical practices transition to incorporating genotyping as part of routine care, this nomogram could be highly useful to predict the presence of EGFR mutations in lung adenocarcinoma in non-Asian patients when mutational profiling is not available or possible.

Persistent infection with Theiler's virus leads to CNS autoimmunity via epitope spreading.

Multiple sclerosis (MS) is a T cell-mediated autoimmune demyelinating disease, which may be initiated by a virus infection. Theiler's murine encephalomyelitis virus (TMEV), a natural mouse pathogen, is a picornavirus that induces a chronic, CD4+ T cell-mediated demyelinating disease with a clinical course and histopathology similar to that of chronic progressive MS (ref. 3). Demyelination in TMEV-infected mice is initiated by a mononuclear inflammatory response mediated by virus-specific CD4+ T cells targeting virus, which chronically persists in the CNS (ref. 4-6). We show that beginning 3-4 weeks after disease onset, T-cell responses to multiple myelin autoepitopes arise in an ordered progression and may play a pathologic role in chronic disease. Kinetic and functional studies show that T-cell responses to the immunodominant myelin proteolipid protein epitope (PLP139-151) did not arise because of cross-reactivity between TMEV and self epitopes (that is, molecular mimicry), but because of de novo priming of self-reactive T cells to sequestered autoantigens released secondary to virus-specific T cell-mediated demyelination (that is, epitope spreading). Epitope spreading is an important alternate mechanism to explain the etiology of virus-induced organ-specific autoimmune diseases.

Germinal center formation, immunoglobulin class switching, and autoantibody production driven by "non alpha/beta" T cells.

The production of class-switched antibodies, particularly immunoglobulin (Ig) G1 and IgE, occurs efficiently in T cell receptor (TCR) alpha-/- mice that are congenitally devoid of alpha/beta T cells. This finding runs counter to a wealth of data indicating that IgG1 and IgE synthesis are largely dependent on the collaboration between B and alpha/beta T cells. Furthermore, many of the antibodies synthesized in TCR alpha-/- mice are reactive to a similar spectrum of self-antigens as that targeted by autoantibodies characterizing human systemic lupus erythematosus (SLE). SLE, too, is most commonly regarded as an alpha/beta T cell-mediated condition. To distinguish whether the development of autoantibodies in TCR alpha-/- mice is due to an intrinsic de-regulation of B cells, or to a heretofore poorly characterized collaboration between B and "non-alpha/beta T" cells, the phenotype has been reconstituted by transfer of various populations of B and non-alpha/beta T cells including cloned gamma/delta T cells derived from TCR alpha-/- mice, to severe combined immunodeficient (SCID) mice. The results establish that the reproducible production of IgG1 (including autoantibodies) is a product of non-alpha/beta T cell help that can be provided by gamma/delta T cells. This type of B-T collaboration sustains the production of germinal centers, lymphoid follicles that ordinarily are anatomical signatures of alpha/beta T-B cell collaboration. Thus, non-alpha/beta T cell help may drive Ig synthesis and autoreactivity under various circumstances, especially in cases of alpha/beta T cell immunodeficiency.

Gamma delta T cell help of B cells is induced by repeated parasitic infection, in the absence of other T cells.

BACKGROUND: gamma delta T cells, like alpha beta T cells, are components of all well-studied vertebrate immune systems. Yet, the contribution of gamma delta T cells to immune responses is poorly characterized. In particular, it has not been resolved whether gamma delta cells, independent of any other T cells, can help B cells produce immunoglobulin and form germinal centers, anatomical foci of specialized T cell-B cell collaboration. RESULTS: TCR beta-/- mice, which lack all T cells except gamma delta T cells, routinely displayed higher levels of antibody than fully T cell-deficient mice. Repeated parasitic infection of TCR beta-/- mice, but not of T cell-deficient mice, increased antibody levels and induced germinal centers that contained B cells and monoclonal gamma delta cells in close juxtaposition. However, antibody specificities were more commonly against self than against the challenging pathogen. gamma delta T cell-B cell help was not induced by repeated inoculation of TCR beta-/- mice with mycobacterial antigens. CONCLUSIONS: In the absence of any other T cells, gamma delta T cell-B cell collaboration can be significantly enhanced by repeated infection. However, the lack of obvious enrichment for antibodies against the challenging pathogen distinguishes gamma delta T cell help from alpha beta T cell help induced under analogous circumstances. The increased production of generalized antibodies may be particularly relevant to the development of autoimmunity, which commonly occurs in patients suffering from alpha beta T cell deficiencies, such as AIDS.

Postoperative radiotherapy in the treatment of extremity soft tissue sarcomas.

From 1966 to 1983, 50 patients with extremity soft tissue sarcomas were treated with wide local excision and postoperative radiotherapy at the Mallinckrodt Institute of Radiology. The median follow-up was 70 months (range 28 to 168). Grade was the most significant factor affecting survival: all 11 patients with well differentiated tumor survived versus 6/8 patients with moderate and 17/31 patients with poorly differentiated tumors (p less than 0.01). In addition, lymph node involvement at diagnosis conferred a worse prognosis with only 2/5 patients alive after treatment (p less than 0.05). Eleven of 50 (22%) failed locally. Factors affecting local control included gross residual tumor after operation and limited treatment volume. Among the 35 patients who did not have gross residual tumor or limited treatment volume, two patients who received less than 5000 cGy failed locally versus 1/18 patients who received between 5000-6000 cGy and 2/15 patients who received more than 6000 cGy. Microscopically positive margins and a volume encompassing less than the total muscular compartment was not associated with an increase in the incidence of local failure. Eight patients developed local complication: five due to retreatment for local recurrence. Overall, 24/26 patients who are alive have had their limbs preserved with normal function.

Pediatric nasopharyngeal carcinoma: long term follow-up of 29 patients.

Twenty-nine untreated children diagnosed with nasopharyngeal carcinoma were consecutively admitted to St. Jude Children's Research Hospital from 1962 to 1986. The age of the patients ranged from 6 to 19 years (median of 13) at diagnosis. Histologically, all had lymphoepithelioma. Patients were retrospectively staged in the American Joint Committee System. Disease extent was T1 (n = 5), T2 (n = 7), T3 (n = 9), T4 (n = 8); N0 (n = 1), N2 (n = 7), N3 (n = 21). Two patients had distant metastasis (M1) on admission, both ultimately succumbed to their disease. Twenty-seven patients were seen initially without metastatic disease: one received pre-irradiation vincristine, 17 were treated with concomitant radiotherapy and cyclophosphamide. From 1981 to the present, four patients received pre-irradiation and one received post-irradiation cisplatin-bleomycin, vinblastine (CDDP-BLEO-VLB) regimens. Four patients received radiotherapy alone. All patients completed chemotherapy and radiation therapy. Twenty-five patients had complete tumor clearance and four had a partial response. Overall, 14 patients are alive continuously without relapse with a median follow-up of 11 years (range 4 to 20). All patients who relapsed did so within 2 years postirradiation. Four patients failed locally--all had advanced (T3-T4) local disease at presentation and three of the failures were at the margin of treatment portals. Thirteen patients failed with distant metastasis. The major prognostic factor in these patients was the local extent of disease. Among the 27 M0 patients, all ten patients with T1-2 tumors are disease-free, whereas four of nine patients with T3 and two of eight patients with T4 tumors are alive and well. In the 16 patients who are long term survivors, eight have mild neck atrophy, two have shortening of the clavicles; except for one patient who required a neck brace for shoulder drop, all had normal function. Among the seven pre-pubertal patients who are long term survivors, three have decreased growth, including one with documented decreased growth hormone. Two patients developed irregular menstrual periods. One patient developed hypothyroidism, and another had a thyroid adenoma. One patient developed bleomycin pneumonitis and one patient who received pre- and post-irradiation chemotherapy died of laryngeal edema and fibrosis, in remission. Radiotherapy is the major modality in the therapy of childhood nasopharyngeal carcinoma. The long term toxicity of radiotherapy plus or minus chemotherapy is acceptable. In early stage tumors (T1-2, N1-2), radiotherapy alone (55-60 Gy) appears to be sufficient for disease control.(ABSTRACT TRUNCATED AT 400 WORDS)

Local-regional non-rhabdomyosarcomatous soft tissue sarcomas of the head and neck.

Between 1962 and 1987, 112 consecutive patients were treated at St. Jude Children's Research Hospital for soft tissue sarcomas of the head and neck region; 18 of these children (16%) had histologic subtypes other than rhabdomyosarcoma. We evaluated the impact of surgery, postoperative chemotherapy, and irradiation on local control and survival in these cases. Three patients who had complete resection of tumors and received no further treatment are alive without disease at 36, 42, and 162 months. Local control was achieved in 1 of 2 patients with microscopic residual tumor and 4 of 9 patients with gross residual tumor who were treated with irradiation (2500-5040 cGy). Chemotherapy was the only postoperative treatment in three patients; only one achieved lasting local control. One patient was treated with irradiation only; his primary site showed no tumor cells at autopsy following an automobile accident. Overall, local control was achieved in 50% of patients; the disease-free survival rate at 3 years was 44%. The prognosis for patients with nonresectable tumors remains unsatisfactory because of the difficulty in securing local control. A revised therapeutic approach to these patients is presented.

Radiation therapy and conservation surgery for primary and recurrent carcinoma of the vulva: report of 40 patients and a review of the literature.

Forty patients with histologically confirmed primary or recurrent vulvar carcinoma were treated with radiation therapy for loco-regional disease. Nineteen of the patients with primary tumors received postoperative radiotherapy (5000 cGy in 6 weeks). Fifteen of the 19 exhibited local tumor control. Five patients with Stage III or IV disease were managed with radiotherapy alone. Four had a complete response with two currently NED. Two patients who received preoperative radiotherapy with local excision are also currently free of disease. The 4-year NED survival for the study population is 100%, 28%, 50%, 0% and 10% for Stage I, II, III, IV and recurrent tumors respectively. The poor results obtained in Stage II tumors is likely due to selection criteria since four of seven patients developed distant metastases. Two of the 14 patients treated for recurrent disease remain NED after local excision of their tumors prior to irradiation. Even though the number of patients is small no dose response for subclinical disease could be found between 4500 and 7000 cGy. Treatment morbidity was acceptable with two patients developing severe long-term complications requiring surgical intervention.

Wilms' tumor: reduced-dose radiotherapy in advanced-stage Wilms' tumor with favorable histology.

Fifty-two children with favorable histology Wilms' tumor who had residual abdominal disease (Surgical Stages III and IV) were treated from 1979 to 1988 on a protocol designed to assess the effectiveness of reduced radiation doses. All patients received three-agent chemotherapy, beginning within 1 week after surgery. To permit assessment of disease response to initial chemotherapy, radiation therapy was delayed for a median of 28 days after surgery (range, 14-71 days). Total doses of abdominal radiation were limited to 12 Gy, given as 150 cGy daily fractions; 18 patients with Stage IV disease received 12 Gy bilateral pulmonary irradiation. Two year disease-free survival was 85% and 71% for Stage III and IV, respectively (p = .24). Abdominal relapses occurred in 3 cases (5.7%). The interval between surgery and initiation of irradiation was not related to disease-free survival. Of several patient and disease-related factors analyzed, only patient age was related to outcome. Disease-free survival was 100% at 3 years for children under the age of 3 versus 78% for children greater than age 3 (p = .05). Reduced-dose abdominal radiotherapy in conjunction with multi-agent chemotherapy and surgery provided excellent disease control with minimal toxicity in advanced-stage, favorable histology Wilms' tumor.

Synthetic peptides which inhibit the interaction between C1q and immunoglobulin and prolong xenograft survival.

BACKGROUND: Acute vascular xenograft rejection (AVXR), also termed delayed xenograft rejection (DXR), occurs when hyperacute rejection (HAR) is prevented by strategies directed at xenoreactive natural antibodies and/or complement activation. We have hypothesized that AVXR/DXR is initiated in part by early components of the complement cascade, notably C1q. We have developed synthetic peptides (termed CBP2 and WY) that interfere with the interaction between C1q and antibody. METHODS: CBP2 and the WY-conjugates were used as inhibitors of immunoglobulin aggregate binding to solid phase C1q. Inhibition of complement activation by the peptides of the classical system was determined using lysis assays with sensitized sheep red blood cells or porcine aortic endothelial cells as targets and of the alternate complement pathway using guinea pig red blood cells as targets. Two transplant models were used to study the effects of administering peptides to recipients: rat heart transplant to presensitized mouse, and guinea heart transplant to PVG C6-deficient rats. RESULTS: CBP2 and WY-conjugates inhibited immunoglobulin aggregate binding to C1q. The peptides also inhibited human complement-mediated lysis of sensitized sheep red blood cells and porcine aortic endothelial cells in a dose-dependent manner and the WY-conjugates prevented activation of the alternate complement pathway as shown by inhibition of guinea pig red blood cells lysis with human serum. In addition, the use of the peptides and conjugates resulted in significant prolongation of xenograft survival. CONCLUSIONS: The CBP2 and WY peptides exhibit the functional activity of inhibition of complement activation. These peptides also prolong xenograft survival and thus provide reagents for the study of the importance of C1q and other complement components in transplant rejection mechanisms.

Hodgkin's disease in children.

This article discusses the presentation and evolution of therapy of pediatric Hodgkin's disease, contrasted with developments in adults. The issues of staging, treatment, and toxicity of radiotherapy, chemotherapy, and combined modality therapy are discussed. Recommendations for therapy are presented.

Simple purification methods for an alphagalactose-specific antibody from chicken eggs.

For many applications avian antibody from egg yolk (IgY) offers advantages over the well-known mammalian antibodies. Different experimental techniques for the purification of IgY from chickens immunized with an alphagalactose-containing antigen (alphaGal-trisaccharide) were compared. These included ammonium sulfate precipitation, filtration with diatomaceous earth, treatment with deoxycholate, and thiophilic and affinity chromatography. Samples were tested for overall purity, protein and lipid content, and specific activity. Evaluated on the basis of these results and the simplicity of the process, the favored purification method is ammonium sulfate precipitation of diluted egg yolk directly followed by affinity chromatography. The high lipid content of IgY preparations is greatly reduced by either thiophilic or affinity chromatography. Affinity purification of ammonium sulfate precipitated material resulted in anti-alphaGal-trisaccharide IgY preparations with approximately 1% of the original protein content but approximately 100-fold higher specific activity for the alphaGal-trisaccharide epitope.

Inclusion of RBD improves the diagnostic classification of dementia with Lewy bodies.

OBJECTIVE: To determine whether adding REM sleep behavior disorder (RBD) to the dementia with Lewy bodies (DLB) diagnostic criteria improves classification accuracy of autopsy-confirmed DLB. METHODS: We followed 234 consecutive patients with dementia until autopsy with a mean of 4 annual visits. Clinical diagnoses included DLB, Alzheimer disease (AD), corticobasal syndrome, and frontotemporal dementia. Pathologic diagnoses used the 2005 DLB consensus criteria and included no/low likelihood DLB (non-DLB; n = 136) and intermediate/high likelihood DLB (DLB; n = 98). Regression modeling and sensitivity/specificity analyses were used to evaluate the diagnostic role of RBD. RESULTS: Each of the 3 core features increased the odds of autopsy-confirmed DLB up to 2-fold, and RBD increased the odds by 6-fold. When clinically probable DLB reflected dementia and 2 or more of the 3 core features, sensitivity was 85%, and specificity was 73%. When RBD was added and clinically probable DLB reflected 2 or more of 4 features, sensitivity improved to 88%. When dementia and RBD were also designated as probable DLB, sensitivity increased to 90% while specificity remained at 73%. The VH, parkinsonism, RBD model lowered sensitivity to 83%, but improved specificity to 85%. CONCLUSIONS: Inclusion of RBD as a core clinical feature improves the diagnostic accuracy of autopsy-confirmed DLB.

An integrated genomic analysis of lung cancer reveals loss of DUSP4 in EGFR-mutant tumors.

To address the biological heterogeneity of lung cancer, we studied 199 lung adenocarcinomas by integrating genome-wide data on copy number alterations and gene expression with full annotation for major known somatic mutations in this cancer. This showed non-random patterns of copy number alterations significantly linked to EGFR and KRAS mutation status and to distinct clinical outcomes, and led to the discovery of a striking association of EGFR mutations with underexpression of DUSP4, a gene within a broad region of frequent single-copy loss on 8p. DUSP4 is involved in negative feedback control of EGFR signaling, and we provide functional validation for its role as a growth suppressor in EGFR-mutant lung adenocarcinoma. DUSP4 loss also associates with p16/CDKN2A deletion and defines a distinct clinical subset of lung cancer patients. Another novel observation is that of a reciprocal relationship between EGFR and LKB1 mutations. These results highlight the power of integrated genomics to identify candidate driver genes within recurrent broad regions of copy number alteration and to delineate distinct oncogenetic pathways in genetically complex common epithelial cancers.

Oncogenes come of age.

Mutations of proto-oncogenes are common events in the pathogenesis of cancers, as shown in a wide range of studies during the 30 years since the discovery of these genes. The benefits of novel therapies that target the products of mutant alleles in human cancers, and the demonstrated dependence of cancers in mouse models on continued expression of initiating oncogenes, are especially promising signs that revolutionary improvements in cancer care are possible. Full realization of the promise of targeted therapies, however, will require better definitions of the genotypes of human cancers, new approaches to interrupt the biochemical consequences of oncogenic mutations, and a greater understanding of drug resistance and tumor progression. In this paper, we summarize recent efforts toward these goals in our laboratory and others.

Influence of site of expression on the fetal gamma delta T-cell receptor repertoire.

The sequences of productive T-cell gamma/delta receptor transcripts were compared in different murine fetal tissues. Differences from tissue to tissue suggest that the sequence repertoires are at least in part the products of selection, presumably through interaction of T cells bearing the gamma delta receptor with fetal self-ligands.