James L. Peacock

Altered TGF-β Signaling in a Subpopulation of Human Stromal Cells Promotes Prostatic Carcinogenesis

Carcinoma-associated fibroblasts (CAF) play a critical role in malignant progression. Loss of TGF-β receptor II (TGFβR2) in the prostate stroma is correlated with prostatic tumorigenesis. To determine the mechanisms by which stromal heterogeneity because of loss of TGFβR2 might contribute to cancer progression, we attenuated transforming growth factor beta (TGF-β) signaling in a subpopulation of immortalized human prostate fibroblasts in a model of tumor progression. In a tissue recombination model, loss of TGFβR2 function in 50% of the stromal cell population resulted in malignant transformation of the nontumorigenic human prostate epithelial cell line BPH1. Mixing fibroblasts expressing the empty vector and dominant negative TGFβR2 increased the expression of markers of myofibroblast differentiation [coexpression of vimentin and alpha smooth muscle actin (αSMA)] through elevation of TGF-β1 and activation of the Akt pathway. In combination, these two populations of stromal cells recapitulated the tumor inductive activity of CAFs. TGFβR2 activity in mixed stromal cell populations cultured in vitro caused secretion of factors that are known to promote tumor progression, including TGF-β1, SDF1/CXCL12, and members of the fibroblast growth factor (FGF) and bone morphogenetic protein (BMP) families. In vivo, tissue recombination of fibroblasts overexpressing TGF-β1 and SDF1/CXCL12 not only induced transformation of BPH1 cells, but also promoted a robust growth of highly invasive cells, similar to effects produced by CAFs. While the precise nature and/or origin of the particular stromal cell populations in vivo remain unknown, these findings strongly link heterogeneity in TGF-β signaling to tumor promotion by tumor stromal cells.

Resistance to DNA-damaging agents is discordant from experimental metastatic capacity in MEF ras-transformants-expressing gain of function MTp53.

Tumor cells can acquire aggressive phenotypes secondary to the loss of expression of the wild-type p53 (WTp53) protein or by the gain of function for selected mutant p53 (MTp53) proteins. However, it is unclear as to whether the development of aggressive phenotypes is inter-related. Herein we report the radiosensitivity, chemosensitivity, and in vivo growth characteristics of isogenic p53−/− MEF ras-transformants that variably express an MTp53 protein. Initial experiments revealed significant clonal heterogeneity with respect to cellular sensitivity to DNA-damaging agents (i.e. ionizing radiation, ultraviolet radiation, cis-platinum, and methotrexate) within subclones of a pre-existing p53−/− MEF cell population. Moreover, this differential sensitivity was also observed within subclones of p53−/− MEF cells transformed with an activated ras allele, suggesting that secondary genetic events and clonal selection, but not cellular transformation per se, may drive the resistance patterns for certain null-p53 tumors. In contrast, uniform resistance was observed following the additional transfection of an MTp53 allele (MTp53pro193) into p53−/− MEF transformants and p53−/− DP-16 Friend erythroleukemia cells, consistent with a gain of MTp53 function for this allele. Relative tumor growth rate and experimental metastatic ability was not enhanced by MTp53pro193 expression. Our results support the concept that gain of MTp53pro193 function leads to the selection of dominant clones, which may exhibit cellular resistance following cancer therapy.

Cancer cachexia is transmissible in plasma

The cause of cancer cachexia is unclear. Tumors may be competing with the host for ingested nutrients or may be releasing some factor that actively inhibits energy utilization. To explore these questions, plasma was sterilely collected and pooled from 103 terminally cachectic Fischer 344 rats implanted with an experimental sarcoma. Control plasma was collected in similar fashion from 138 nontumor-bearing rats (NTBP). Plasma from tumor-bearing rats (TBP) or NTBP was continuously infused in a randomized, blinded fashion for 4 days into 20 normal rats. During infusion, food intake and nitrogen excretion were measured daily. At sacrifice, body weight and organ masses were determined. Rats receiving TBP demonstrated an immediate and profound anorexia compared with those receiving NTBP. Total food intake during treatment was 31.2 +/- 3.3 (g +/- SEM) in the TBP group versus 48.2 +/- 2.8 in the NTBP group (P less than 0.001 by t test). Likewise, the total decline in body weight was greater in the TBP group as compared with the NTBP group (-35.2 +/- 3.4 versus -14.6 +/- 4.0, P less than 0.001). Mean daily nitrogen balance during treatment was negative in the rats receiving TBP (-14.5 +/- 20.1 mg +/- SEM) while remaining highly positive in the rats receiving NTBP (110.7 +/- 19.3, P less than 0.002). Finally, cardiac and gastrocnemius muscle masses were decreased, while hepatic mass was unaffected. These data demonstrate that the syndrome of cancer-associated cachexia is transmissible in plasma and therefore may be mediated by a circulating molecule or molecules. Identification and purification of the molecule(s) responsible for this effect would have obvious clinical benefits.

Impact of insulin on survival of cachectic tumor-bearing rats

The present study was performed to determine if a host nutritional treatment, insulin, in the absence of antitumor treatment could improve survival of cachectic tumor-bearing (TB) rats. Initially food intake and host weight were correlated with survival of untreated rats with similar size sarcomas (45-50 cm3). TB rat food intake (r = 0.69, p less than 0.0001) and host weight (r = 0.47, p less than 0.004) correlated positively with subsequent survival. Once daily neutral protamine hagedorn (NPH) insulin treatment (2 units/100 g) significantly improved food intake (p less than 0.01) and host weight (p less than 0.01) of cachectic TB rats without increasing tumor growth. Twice daily NPH insulin (2 units/100 g) maintained normal food intake of cachectic TB rats and turned a host weight loss into a host weight gain which was significantly greater than untreated controls (p less than 0.001) and all other methods of insulin administration including once daily (p less than 0.001). Twice daily NPH insulin maintained mild hypoglycemia (glucose = 84 +/- 12 mg/dl) compared to once daily NPH insulin which resulted in hyperglycemia (glucose = 140 +/- 8 mg/dl, p less than 0.001) prior to next dose. In addition, twice daily NPH insulin did not increase tumor growth. Once daily NPH insulin for 5 days during cachectic decline was well tolerated (no treatment deaths), and improved median survival of TB rats randomized to insulin (15 days) compared to controls (13 days, p = 0.06). However, twice daily NPH insulin during cachectic decline failed to improve survival because of treatment deaths.(ABSTRACT TRUNCATED AT 250 WORDS)

Surgeon beware: Many patients referred for parathyroidectomy are misdiagnosed with primary hyperparathyroidism

PURPOSE We hypothesized that patients referred for the evaluation and management of primary hyperparathyroidism (pHPT) often do not have pHPT and that they may be harmed by unwarranted parathyroidectomy (PTX). METHODS We reviewed all patients who were referred to our endocrine surgery practice between 2008 and 2011 with International Classification of Diseases, Ninth Revision codes for HPT (252.00), benign or malignant parathyroid tumors (227.1, 194.1, respectively), or hypercalcemia (275.42). Patients with renal failure were excluded. Clinical parameters for investigation included age, sex, presentation, laboratories, imaging studies, and referring physician. RESULTS Three hundred twenty-four patients were referred for pHPT. The diagnosis was confirmed in 265 (82%), of whom 211 (80%) underwent PTX. Misdiagnoses occurred in 60 of 324 patients (19%). Of these, 54 (90%) had secondary HPT and 6 (10%) had hypercalcemia but no pHPT. Before referral, 70% of misdiagnosed patients underwent localizing studies, 57% of which suggested a positive finding. CONCLUSION Considerable confusion exists regarding the differentiation of primary and secondary HPT. Surgeons should be cautioned that patients who are referred for parathyroidectomy, even those with complete laboratory and radiographic evaluations, might not have pHPT at all.

Cryosurgery for primary and metastatic liver tumors.

Cryosurgery is the application of extreme cold temperatures to achieve tumor cell death. Cryoablation, cryotherapy and cryosurgery are interchangeable terms which all apply to the process of tissue destruction using subzero temperatures. Topical application of cold substances to tumors was first reported in treatment of superfiicial tumors of skin, oropharynx and gynecologic systems. (1) The rationale for using extreme cold to treat tumors is the potential for in situ tumor destruction while sparing normal surrounding structures and without the complications of major surgical intervention. In 1963 Irving Cooper was the first to describe the feasibility of such an approach to the liver and other organs. (2) He devised a system of liquid nitrogen delivery through insulated cannulas that could be placed within the substance of the liver. Subsequently Andrew Gage and co-investigators performed in vivo studies of hepatic cryosurgery and described many of the physiologic responses to liver treatment. They showed that large areas of liver could be treated successfully and that large remnants of devitalized tissue did not produce toxic effects to the animal. (3)

High-grade angiosarcoma presenting with cytology-negative hemorrhagic ascites

Abstract Angiosarcomas are a rare subtype of soft-tissue sarcomas originating from the vascular endothelium. Both retroperitoneal and omental angiosarcomas tend to be aggressive and rapidly fatal if not amenable to early intervention. In this report, we describe an unusual case of high-grade angiosarcoma with cytology-negative hemorrhagic ascites and diffuse omental invasion. Multiple investigations into the origin of the hemorrhagic ascites, including cytological analysis, tumor marker measurements, serum-ascites albumin gradient calculation and frozen section pathological examination, failed to reveal a diagnosis. We conclude that malignancy should be considered in the differential diagnosis in the presence of suspicious cytology-negative hemorrhagic ascites and concomitant retroperitoneal and abdominal findings.

Abstract P1-08-37: Magee equations predict pathologic response to neoadjuvant chemotherapy

Introduction: Neoadjuvant chemotherapy is used in locally advanced breast cancer to downstage the tumor, facilitating surgical management. Oncotype DX (ODX) is used to estimate the risk of distant recurrence for ER-positive breast cancers, allowing selected patients to avoid the toxicity of chemotherapy. ODX is often not possible on the small core biopsy samples. Klein et al. have shown that standard histological variables, combined with semiquantitative ER, PR, HER-2, and Ki-67 results, can provide information similar to that with ODX, using equations derived by linear regression analysis (Magee equations). We applied a modification of these equations to pretreatment core biopsies in women who received neoadjuvant chemotherapy to determine if the risk scores were predictive of pathologic response. Methods: 25 patients who received chemotherapy for receptor positive locally advanced(21), inflammatory(3), or metastatic(1) breast cancer followed by surgical treatment of the primary site were identified from a prospective breast cancer database. Pretreatment core biopsies were reviewed by a breast pathologist and Nottingham grade, ER and PR status (% of cells staining and intensity of staining), and Her-2 status by IHC and/or FISH were recorded. Clinical tumor size was defined as the average of sizes derived from mammogram, ultrasound, MRI, PET-CT and clinical breast examination. Using these data in a modified Magee equation, the patient9s recurrence score was calculated. 0-18 was considered low risk (LR), >18- Results: Magee scores ranged from 13.8-41.6 (mean 27.4) and were significantly lower in the poor responders (mean = 23, range 13.8-41.6) compared to the good responders (mean = 33, range 22-41.3, p = 0.003). Table 1 shows the distribution of response by Risk Group (p = 0.018). 73% of patients with high risk Magee scores had a good response to chemotherapy, compared to 21% of patient with low or intermediate scores (p = 0.01). Conclusions: Modified Magee equations applied to pretreatment core biopsies seem to predict pathologic response to neoadjuvant chemotherapy. Use of these equations to assign risk scores may be a useful tool in deciding which ER positive breast cancer patients are likely to benefit from preoperative chemotherapy for cytoreduction, and who should go directly to surgery. These findings need to be validated in larger studies. Citation Information: Cancer Res 2013;73(24 Suppl): Abstract nr P1-08-37.

The surgeon’s impact on ER status.

153 Background: Long ischemic times prior to tissue fixation can lower the accuracy of immunohistochemical staining (IHC). The purpose of this study was to identify factors that impact the time to fixation (TTF), and to determine if TTF impacts prognostic factor (PF) patterns and if TTF can be changed with simple interventions. METHODS Surgeries performed between 2008 and June 2011 for which TTF was noted and PF testing was done were reviewed. TTF, patient age and race, surgeon, date and type of surgery, tumor size, nodal status, grade, and status of ER/PR and HER2 by IHC were noted. TTF was dichotomized into > or <60 minutes. In-Services (IS) for pathology and surgical staff were completed. TTF before and after the IS were compared. Associations between TTF and other factors were assessed using standard methods for contingency tables. Predictors of ER status were evaluated by multivariate analysis. RESULTS Mean TTF was 72 minutes (range 12-445) TTF was significantly associated with surgeon, type of specimen, and ER status (Table). TTF was an independent predictor of ER status by multivariate analysis. TTF was shorter and less variable after IS. CONCLUSIONS TTF varied significantly between surgeons. IS training of the surgeons and OR and pathology staff can decrease TTF. Longer TTF was associated with increased ER negativity, suggesting that longer ischemic time interferes with IHC for ER. If prolonged TTF leads to false negative ER staining, patients may not receive optimal therapies. To minimize this risk, the breast surgeon should take an active role in minimizing the TTF. [Table: see text].

Abstract 882: Serum Hsp27 levels may serve as a prognostic biomarker in patients with localized breast cancer: A preliminary report

Although there exist some prognostic biomarkers (eg, CA 15-3) for evaluation of responses to therapy in metastatic breast cancer patients, there is no reliable biomarker for early stage breast cancer patients. Heat shock protein 27 (Hsp27) has been reported as elevated in many human cancers of carcinoma origin. We have simultaneously tested serum levels of Hsp27 and CA 15-3 by specific ELISA in 49 untreated breast cancer patients (age→52.77±1.28) with early to locally advanced disease [Stage I (n=21), IIA and B (n=21), IIIA, B and C (n=7)] and 37 female controls (age→50.86±1.60). Hsp27 levels were significantly (p Work supported by ACS RSG-06-266-01-LIB awarded to A. De Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 882.