Construction of mismatched inverted repeat (IR) silencing vectors for maximizing IR stability and effective gene silencing in plants.

Inverted repeat (IR) RNA silencing vectors containing homologous fragments of target endogenous plant genes, or pathogen genes, are the most widely used vectors to either study the function of genes involved in biotic stress or silence pathogens to induce plant resistance, respectively. RNA silencing has been exploited to produce transgenic plants with resistance to viral pathogens via posttranscriptional gene silencing (PTGS). In some cases, this technology is difficult to apply due to the instability of IR constructs during cloning and plant transformation. We have therefore developed a robust method for the production of long IR vector constructs by introducing base pair mismatches in the form of cytosine to thymine mutations on the sense arm by exposure to sodium bisulfite prior to assembly of the IR.

Construction of effective inverted repeat silencing constructs using sodium bisulfite treatment coupled with strand-specific PCR.

RNA silencing has been exploited to produce transgenic plants with resistance to viral pathogens via posttranscriptional gene silencing (PTGS). In some cases, this technology is difficult to apply due to the instability of inverted repeat (IR) constructs during cloning and plant transformation. Although such constructs have been shown to be stabilized with introns and efficiently induce RNA silencing, we found that the Pdk intron did not stabilize South African cassava mosaic virus (SACMV) silencing constructs. Therefore, we developed a method for producing long SACMV IR constructs through bisulfite-induced base pair mismatches on the sense arm prior to IR assembly. Expression of SACMV BC1 mismatched IR constructs in the model test plant Nicotiana benthamiana resulted in a reduction in viral BC1 transcript levels, hence viral replication, upon SACMV infection. Mismatched SACMV AC1 IR constructs induced PTGS more efficiently in a N. benthamiana callus system than nonmismatched IR constructs. Our novel method for IR construct generation should be applicable to many sequences where the generation of these constructs has proven difficult in the past.

Potential impact of tumor registry rule changes for recording inflammatory breast cancer.

BACKGROUND: New tumor registry rules for abstracting multiple primaries and histologies include 1 specifically for inflammatory breast cancer (IBC), which states that the International Classification for Oncology (ICD-O) histology code 8530 (3) for IBC should be used only when it is on the pathology report. IBC is typically clinically diagnosed. The purpose of this project is to determine the potential impact of this new rule on identifying IBC cases by searching on the ICD-O histology code. METHODS: Two hundred forty patients were identified from The University of Texas M. D. Anderson Cancer Center Tumor Registry database, who had initially presented to the institution from 2005 through 2007 with a diagnosis of IBC. RESULTS: The basis of diagnosis of IBC was pathologic for 73 patients and clinical for 167. Of the 167 patients with a clinical diagnosis of IBC, 164 patients also had a pathologic diagnosis of a non-IBC histology. Following the new rule, the non-IBC histology would be recorded for these 164 patients. CONCLUSIONS: Based on the analysis of The University of Texas M. D. Anderson Cancer Center's patients with IBC, the new Surveillance, Epidemiology, and End Results multiple primary rule will result in the ICD-O IBC histology code being recorded for approximately 30% of patients with IBC. Clinically diagnosed cases are to be identified using the collaborative staging extension codes or the American Joint Committee on Cancer/TNM classification T value of 4 days. However, many researchers may continue to search only on the histology field, resulting in a perceived decrease in IBC.

Increased malignancy risk in the cutaneous T-cell lymphoma patient population.

BACKGROUND: Cutaneous T-cell lymphoma (CTCL) has been associated with increased risk for second malignancies. However, the degree of risk and types of second cancers detected have been inconsistent in previous studies. PATIENTS AND METHODS: To further characterize the risk for malignancy associated with CTCL, patients treated for CTCL at M. D. Anderson Cancer Center in Houston, Texas, between November 1979 and November 1999 were assessed for the occurrence of additional cancers by analysis of institutional tumor registry data. RESULTS: Of 672 patients with CTCL, 112 had > or = 1 additional cancer, 37 occurring after the diagnosis of CTCL. This represents a significant elevation in cancer prevalence and incidence, with a 1.79-fold risk (95% CI, 1.22-2.39) for developing cancer after CTCL. An excess of Hodgkin and non-Hodgkin lymphoma, acute myeloid leukemia, and vulvar cancers was seen. CONCLUSION: These data provide evidence for an increased overall incidence of second malignancy in CTCL, particularly with respect to other lymphoproliferative malignancies. Appropriate monitoring for the early detection of second cancers might be warranted in patients with CTCL.

Can digital rectal examination or transrectal ultrasonography biopsy findings predict the side of nodal metastasis in prostate cancer?

PURPOSE: To assess the use of several preoperative parameters in predicting the side of pelvic lymph node metastasis in patients with prostate cancer. MATERIALS AND METHODS: A retrospective chart review (January 1982 to February 2004) identified 106 men with pathology proven lymph node positive prostate cancer for whom complete medical records were available. RESULTS: The median serum prostate-specific antigen at diagnosis was 11 ng/ml with the clinical stage T1C in 9 patients, T2 in 68, and T3 in 29. The Gleason score on transrectal ultrasonography (TRUS) biopsy was < or =6 in 13, 7 in 41, and > or =8 in 52. A total of 93 patients had documented pretreatment digital rectal examination (DRE) findings: 54 had a unilaterally suspicious DRE, and 31 had a bilaterally suspicious DRE. Of patients with a unilaterally positive DRE, 30 had ipsilateral lymph node metastasis, 16 contralateral, and 8 bilateral. DRE showed a 71% sensitivity and 29% false-negative rate in predicting the side of nodal metastasis. A total of 98 patients had documented TRUS biopsy findings: 37 had unilaterally positive TRUS biopsies and 61 bilaterally positive biopsies. Of patients with unilaterally positive TRUS biopsies, 20 had ipsilateral lymph node metastasis, 11 contralateral, and 6 bilateral. TRUS biopsies showed an 86% sensitivity and 14% false-negative rate in predicting the side of nodal metastasis. CONCLUSIONS: DRE and TRUS biopsies do not accurately predict the side of pelvic lymph node metastasis and should not determine the extent of the pelvic lymphadenectomy.

Metastatic patterns in adenocarcinoma.

BACKGROUND: Unique metastatic patterns cited in the literature often arise from anecdotal clinical observations and autopsy reports. The authors analyzed clinical data from a large number of patients with histologically confirmed, distant-stage adenocarcinoma to evaluate metastatic patterns. METHODS: Tumor registry data were collected between 1994-1996 on 11 primary tumor sites and 15 metastatic sites from 4399 patients. The primary and metastatic sites were cross-tabulated in various ways to identify patterns, and the authors developed algorithms by using multinomial logistic regression analysis to predict the locations of primary tumors based on metastatic patterns. RESULTS: Three primary tumors had single, dominant metastatic sites: ovary to abdominal cavity (91%), prostate to bone (90%), and pancreas to liver (85%). The liver was the dominant metastatic site for gastrointestinal (GI) primary tumors (71% of patients), whereas bone and lung metastases were noted most frequently in non-GI primary tumors (43% and 29%, respectively). In a study of combinations of liver, abdominal cavity, and bone metastases, 86% of prostate primary tumors had only bone metastases, 80% of ovarian primary tumors had only abdominal cavity metastases, and 74% of pancreas primary tumors had only liver metastases. A single organ was the dominant source of metastases in 7 sites: axillary lymph node from the breast (97%), intestinal lymph node from the colon (84%), thoracic lymph node from the lung (66%), brain from the lung (64%), mediastinal lymph node from the lung (62%), supraclavicular lymph node from the breast (51%), and adrenal gland from the lung (51%). CONCLUSIONS: The algorithms that the authors developed achieved a cross-validated accuracy of 64% and an accuracy of 64% on an 1851-patient independent test set, compared with 9% accuracy when a random classifier was used.

Inadequacies of the current American Joint Committee on cancer staging system for prostate cancer.

BACKGROUND: Two major objectives of the American Joint Committee on Cancer (AJCC) staging system are to ensure appropriate treatments for patients and to determine prognosis. AJCC stage for distant prostate cancer includes patients with regional lymph node involvement. In the current study, the authors assessed whether patients with lymph node involvement and patients with distant metastasis, as determined using the Surveillance, Epidemiology, and End Results (SEER) staging system, had similar treatment and survival duration and, thus, were grouped together appropriately in the AJCC system. METHODS: In total, 4141 patients were selected from The University of Texas M. D. Anderson Cancer Center's Tumor Registry who initially had registered at the center between January 1, 1982, and December 31, 2001, with a diagnosis of prostate cancer; had received no treatment before presentation; and had received treatment at the center. Patients with unknown stage and patients with any other primary malignancies were excluded. Descriptive analyses of demographic and disease variables were performed. Using SEER stage groups, survival analyses and Cox proportional hazards regression analyses were performed. RESULTS: Treatments differed between patients with lymph node involvement and patients with distant metastasis. The median survival was 134 months for patients with lymph node involvement and 42 months for patients with distant metastasis. When these 2 groups were combined, as in the AJCC scheme, the median survival was 86 months. CONCLUSIONS: The treatment and median survival of patients with lymph node involvement differed substantially from those of patients with distant metastasis. The current AJCC scheme for prostate cancer appeared to be inappropriate when considering its purpose, and the authors concluded that it should be revised.