Udo Rudloff

Nomogram for Predicting the Risk of Local Recurrence After Breast-Conserving Surgery for Ductal Carcinoma In Situ

PURPOSE While the mortality associated with ductal carcinoma in situ (DCIS) is minimal, the risk of ipsilateral breast tumor recurrence (IBTR) after breast-conserving surgery (BCS) is relatively high. Radiation therapy (RT) and antiestrogen agents reduce the risk of IBTR and are considered standard treatment options after BCS. However, they have never been proven to improve survival, and in themselves carry rare but serious risks. Individualized estimation of IBTR risk would assist in decision making regarding the various treatment options for women with DCIS. PATIENTS AND METHODS From 1991 to 2006, 1,868 consecutive patients treated with BCS for DCIS were identified. A multivariate Cox proportional hazards model was constructed using the 1,681 in whom data were complete. Ten clinical, pathologic, and treatment variables were built into a nomogram estimating probability of IBTR at 5 and 10 years after BCS. The model was validated for discrimination and calibration using bootstrap resampling. RESULTS The DCIS nomogram for prediction of 5- and 10-year IBTR probabilities demonstrated good calibration and discrimination, with a concordance index of 0.704 (bootstrap corrected, 0.688) and a concordance probability estimate of 0.686. Factors with the greatest influence on risk of IBTR in the model included adjuvant RT or endocrine therapy, age, margin status, number of excisions, and treatment time period. CONCLUSION The DCIS nomogram integrates 10 clinicopathologic variables to provide an individualized risk estimate of IBTR in a woman with DCIS treated with BCS. This tool may assist in individual decision making regarding various treatment options and help avoid over- and undertreatment of noninvasive breast cancer.

Central pancreatectomy revisited.

Central pancreatectomy is a surgical procedure that removes the middle segment of the pancreas and preserves the distal pancreas and spleen. This limited resection has the advantage of conserving normal, uninvolved pancreatic parenchyma, thus reducing the possibility of postoperative exocrine and endocrine dysfunction. While the incidence of postoperative endocrine insufficiency may be as low as 4%, procedural morbidity, specifically pancreatic fistula, appears to exceed the published rates for standard resections (i.e., distal/subtotal pancreatectomy or pancreaticoduodenectomy). We have reviewed our prospective pancreatic cancer database to determine the utilization of central pancreatectomy in a major cancer center with expertise in pancreatic surgery. We identified only 10 cases of central pancreatectomy over the past 13 years. Six (60%) had postoperative complications including three cases (30%) of pancreatic fistula. No patients died as a result of the procedure. At a median follow-up of 13.6 months (mean, 25.2 months), only one patient had mild endocrine insufficiency and no patients had clinically significant exocrine dysfunction. The associated morbidity of central pancreatectomy may outweigh any potential benefit in long-term pancreatic secretory function. We suggest that such a procedure be used selectively, where preservation of the pancreas appears essential.

DIFFERENT DOMAINS OF THE MURINE RNA POLYMERASE I-SPECIFIC TERMINATION FACTOR MTTF-I SERVE DISTINCT FUNCTIONS IN TRANSCRIPTION TERMINATION

Termination of mouse ribosomal gene transcription by RNA polymerase I (Pol I) requires the specific interaction of a DNA binding protein, mTTF‐I, with an 18 bp sequence element located downstream of the rRNA coding region. Here we describe the molecular cloning and functional characterization of the cDNA encoding this transcription termination factor. Recombinant mTTF‐I binds specifically to the murine terminator elements and terminates Pol I transcription in a reconstituted in vitro system. Deletion analysis has defined a modular structure of mTTF‐I comprising a dispensable N‐terminal half, a large C‐terminal DNA binding region and an internal domain which is required for transcription termination. Significantly, the C‐terminal region of mTTF‐I reveals striking homology to the DNA binding domains of the proto‐oncogene c‐Myb and the yeast transcription factor Reb1p. Site‐directed mutagenesis of one of the tryptophan residues that is conserved in the homology region of c‐Myb, Reb1p and mTTF‐I abolishes specific DNA binding, a finding which underscores the functional relevance of these residues in DNA‐protein interactions.

Impact of Maximal Cytoreductive Surgery Plus Regional Heated Intraperitoneal Chemotherapy (HIPEC) on Outcome of Patients With Peritoneal Carcinomatosis of Gastric Origin: Results of the GYMSSA Trial

A prospective randomized trial was conducted to compare the impact of systemic chemotherapy versus multi‐modality therapy (complete cytoreductive surgery (CRS), hyperthermic intraperitoneal chemotherapy (HIPEC), and systemic chemotherapy) on overall survival (OS) in patients with gastric carcinomatosis.

A Novel Chimeric Antigen Receptor Against Prostate Stem Cell Antigen Mediates Tumor Destruction in a Humanized Mouse Model of Pancreatic Cancer

Despite advances in the understanding of its molecular pathophysiology, pancreatic cancer remains largely incurable, highlighting the need for novel therapies. We developed a chimeric antigen receptor (CAR) specific for prostate stem cell antigen (PSCA), a glycoprotein that is overexpressed in pancreatic cancer starting at early stages of malignant transformation. To optimize the CAR design, we used antigen-recognition domains derived from mouse or human antibodies, and intracellular signaling domains containing one or two T cell costimulatory elements, in addition to CD3zeta. Comparing multiple constructs established that the CAR based on human monoclonal antibody Ha1-4.117 had the greatest reactivity in vitro. To further analyze this CAR, we developed a human pancreatic cancer xenograft model and adoptively transferred CAR-engineered T cells into animals with established tumors. CAR-engineered human lymphocytes induced significant antitumor activity, and unlike what has been described for other CARs, a second-generation CAR (containing CD28 cosignaling domain) induced a more potent antitumor effect than a third-generation CAR (containing CD28 and 41BB cosignaling domains). While our results provide evidence to support PSCA as a target antigen for CAR-based immunotherapy of pancreatic cancer, the expression of PSCA on selected normal tissues could be a source of limiting toxicity.

TBP-associated factors interact with DNA and govern species specificity of RNA polymerase I transcription.

Unlike genes transcribed by RNA polymerases II and III, transcription by RNA polymerase I is highly species‐specific. Ribosomal promoter selectivity is brought about by a multisubunit transcription factor (SL1/TIF‐IB) which consists of the TATA‐binding protein (TBP) and three TBP‐associated factors (TAFs). To determine the basis for the inability of SL1/TIF‐IB to recognize heterologous rDNA, the transcriptional properties and the subunit composition of the murine and the human factor, as well as a chimeric complex containing epitope‐tagged human TBP and murine TAFs, have been compared. We show that TBP can be exchanged between the human and mouse factor indicating that the variable N‐terminal domain of TBP does not play a significant role in rDNA promoter selectivity. Instead, DNA binding is brought about by the TAFs. UV crosslinking experiments demonstrate that binding to the ribosomal gene promoter is mediated by two TAFs (TAFI48 and TAFI68) which have the same electrophoretic mobility in the human and mouse factor. The largest TAF is different in both species and is suggested to play a role in the species‐specific assembly of productive preinitiation complexes. Thus, evolutionary changes of rDNA promoter sequences have been accompanied by changes in specific TAFs.

Label-retaining liver cancer cells are relatively resistant to sorafenib.

Objective The standard therapy for advanced hepatocellular carcinoma (HCC) is sorafenib, with most patients experiencing disease progression within 6 months. Label-retaining cancer cells (LRCC) represent a novel subpopulation of cancer stem cells (CSC). The objective was to test whether LRCC are resistant to sorafenib. Methods We tested human HCC derived LRCC and non-LRCC before and after treatment with sorafenib. Results LRCC derived from human HCC are relatively resistant to sorafenib. The proportion of LRCC in HCC cell lines is increased after sorafenib while the general population of cancer cells undergoes growth suppression. We show that LRCC demonstrate improved viability and toxicity profiles, and reduced apoptosis, over non-LRCC. We show that after treatment with sorafenib, LRCC upregulate the CSC marker aldehyde dehydrogenase 1 family, wingless-type MMTV-integration-site family, cell survival and proliferation genes, and downregulate apoptosis, cell cycle arrest, cell adhesion and stem cells differentiation genes. This phenomenon was accompanied by non-uniform activation of specific isoforms of the sorafenib target proteins extracellular-signal-regulated kinases and v-akt-murine-thymoma-viral-oncogene homologue (AKT) in LRCC but not in non-LRCC. A molecular pathway map for sorafenib treated LRCC is proposed. Conclusions Our results suggest that HCC derived LRCC are relatively resistant to sorafenib. Since LRCC can generate tumours with as few as 10 cells, our data suggest a potential role for these cells in disease recurrence. Further investigation of this phenomenon might provide novel insights into cancer biology, cancer recurrence and drug resistance with important implications for the development of novel cancer therapies based on targeting LRCC.

The Influence of Margin Width and Volume of Disease Near Margin on Benefit of Radiation Therapy for Women With DCIS Treated With Breast-Conserving Therapy

Objective and Summary Background Data:There remains variation in the use of radiation therapy (RT) in women with ductal carcinoma in situ (DCIS), despite prospective randomized trials documenting its benefit in reducing the risk of ipsilateral breast tumor recurrence (IBTR). Methods:Patients with DCIS treated with excision alone or excision plus RT from 1991 to 1995 were identified. Margin width, number of involved ducts at closest margin, age, presence of palpable mass, presence of lobular neoplasia, nuclear grade, and necrosis were tested in uni- and multivariate analysis for association with risk of IBTR and added value of RT. Results:Two hundred ninety-four patients with a median follow-up of 11 years had actuarial 10- and 15-year overall IBTR rates of 22% and 29%, respectively. For lesions excised with margins of <1 mm, 1 to 9 mm, and ≥10 mm, the actuarial 10-year IBTR rates were 28%, 21%, and 19%, respectively. RT reduced adjusted IBTR rates by 62% (P = 0.002) for all patients; 83% for lesions with <1 mm margins (P = 0.002), 70% for 1 to 9 mm (P = 0.05), and 24% (P = 0.55) for ≥10 mm. After adjustment for other variables, higher volume of disease near the margin was associated with risk of IBTR in the no RT group (HR = 3.37, P = 0.002) and greater benefit of RT (HR 0.14; P = 0.004). Conclusion:Effect of RT on IBTR risk is influenced by both margin width and number of involved ducts at nearest margin. Patients with higher volume of disease near the margin derive a greater benefit from the addition of RT. Despite margins of ≥10 mm, the risk of IBTR remains substantial in patients with DCIS.

A Pilot Study Assessing the Potential Role of non-CD133 Colorectal Cancer Stem Cells as Biomarkers

Introduction: Over 50% of patients with colorectal cancer (CRC) will progress and/or develop metastases. Biomarkers capable of predicting progression, risk stratification and therapeutic benefit are needed. Cancer stem cells are thought to be responsible for tumor initiation, dissemination and treatment failure. Therefore, we hypothesized that CRC cancer stem cell markers (CRCSC) will identify a group of patients at high risk for progression. Methods: Paraffin-embedded tissue cores of normal (n=8), and histopathologically well-defined primary (n= 30) and metastatic (n=10) CRC were arrayed in duplicate on tissue microarrays (TMAs). Expression profiles of non-CD133 CRCSC (CD29, CD44, ALDH1A1, ALDH1B1, EpCam, and CD166) were detected by immunohistochemistry and the association with clinicopathological data and patient outcomes was determined using standard statistical methodology. An independent pathologist, blinded to the clinical data scored the samples. Scoring included percent positive cells (0 to 4, 0 = <10%, 1 = 10 - 24%, 2 = 25 - 49%, 3 = 50 - 74%, 4 = 75 - 100%), and the intensity of positively stained cells (0 to 4; 0 = no staining, 1 = diminutive intensity, 2 = low intensity, 3 = intermediate intensity, 4 = high intensity). The pathologic score represents the sum of these two values, reported in this paper as a combined IHC staining score (CSS). Results: Of 30 patients 7 were AJCC stage IIA, 10 stage IIIB, 7 stage IIIC and 6 stage IV. Median follow-up was 113 months. DFI was 17 months. Median overall survival (OS) was not reached. Stage-specific OS was: II - not reached; III - not reached; IV - 11 months. In a univariate analysis, poor OS was associated with loss of CD29 expression; median OS, 32 months vs. not reached for CSS 3-7 vs. >7.5, respectively; p=0.052 comparing entire curves, after adjustment. In a Cox model analysis, loss of CD29 exhibited a trend toward association with survival (p=0.098) after adjusting for the effect of stage (p=0.0076). Greater expression of ALDH1A1 was associated with increasing stage (p=0.042 over stages 2, 3b, 3c, and 4) while loss of CD29 expression exhibited a trend toward being associated with stages 3 and 4 (p=0.08). Compared to normal colon tissue, primary tumors were associated with increased expression of ALDH1B1 (p=0.008). ALD1H1B1 expression level differed according to whether the tumor was moderately or poorly differentiated, well differentiated, or mucinous; the highest expression levels were associated with moderately or poorly differentiated tumors (p=0.011). Lymph node metastases were associated with a trend toward decreased expression of EpCAM (p = 0.06) when comparing 0 vs. 1 vs. 2+ positive lymph nodes, as was CD29 (p = 0.08) when comparing 0 vs. any positive lymph nodes. Compared to normal colon tissue metastatic colon cancers from different patients were associated with increased ALDH1B1 expression (p=0.001) whereas CD29 expression was higher in normal colonic tissue (p=0.014). Conclusion: CD29 may be associated with survival as well as clinical stage and number of lymph nodes. ALDH1B1 expression was associated with differentiation as well as type of tissue evaluated. ALDH1A1 was associated with clinical stage, and decreased EpCAM expression was found in patients with advanced lymph node stage. CRCSCs may be useful biomarkers to risk stratify, and estimate outcomes in CRC. Larger prospective studies are required to validate the current findings.

A growing family: Adding mutated Erbb4 as a novel cancer target

As the upward spiral of novel cancer gene discoveries and novel molecular compounds continues to accelerate, a repetitive theme in molecular drug development remains the lack of activity of initially promising agents when given to patients in clinical trials. It is however invigorating that a few targeted agents directed against a select group of a few ‘cancer gene superfamilies’ have escaped this all to common fate, and have evolved into novel, clinically meaningful molecular therapy strategies. Targeting dysregulated signaling of the epidermal growth factor family of transmembrane receptors (Erbb family) has encompassed over the last decade an ever increasing role in personalized treatment approaches in an increasing number of human malignancies. Erbbs are receptor tyrosine kinases that are important regulators of several signaling pathways. Two of its family members (Erbb1/EGFR and Erbb2/HER2) have previously been shown to be somatically mutated in large fraction of human cancers. To determine if this family is somatically mutated in melanoma, its sequences were recently analyzed and one of its members, Erbb4, was found to be somatically mutated in 19% of melanoma cases. Functional analysis of seven of its mutations was shown to increase its catalytic and transformation abilities as well as providing essential survival signals. Similar to other Erbb family members, mutant Erbb4 seems to confer ‘oncogene addiction’ on melanoma cells, making it an attractive therapeutic target. Gaining further understanding into the oncogenic mechanism of Erbb4 may not only help in the development of targeted therapy in melanoma patients but might accelerate the acceptance of a novel taxonomy of cancer which is based on the genomic perturbations in cancer genes and cancer gene families and their response to targeted agents.