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Background: The Veridex CellSearch is an FDA-approved technology for enumerating circulating tumor cells in blood samples of metastatic colorectal cancer mCRC) patients and has prognostic value. It is important to understand how counts of circulating tumor cells (CTCs), which are advocated to be tools for “liquid biopsy” of tumors, correlate with clinical and pathologic variables of significance in these patients. In this study, we have attempted to make such correlations along with evaluating how CTC counts change during the course of chemotherapy. Patients and methods: Following an IRB-approved protocol, blood samples were collected from 24 patients with mCRC along with relevant clinico-pathological data. Blood was collected at defined time-points both prior to as well as during the course of treatment with combination chemotherapy, and CTC counts were enumerated from7.5 ml of blood. Results: Seventeen out of 24 patients with mCRC showed a CTC count of 2 or less cells in 7.5 ml of blood at base-line assessment before chemotherapy while 7 patients showed 3 or more cells in 7.5 ml of blood at that point. A correlation was found between high carcino-embryonic antigen (CEA) levels and high CTC counts (P = 0.018) although it was also found that some patients had elevated CTCs without an elevated CEA. No correlation with the time interval between detection of primary tumor and appearance of secondary (metastatic) tumor(s) was found. CTC counts did not correlate with the presence of lung or liver metastases, i.e. a number of mCRC patients with lung or liver metastases had a count of zero CTCs at baseline. We also noted no correlation between CTC number and the status of KRAS or BRAF mutation. CTC counts dropped immediately after the start of chemotherapy in 11 out of 21 patients, and also reduced from the baseline at the end of chemotherapy in 5 out of 10 patients. Six of 7 patients who started with 3 or more CTCs in 7.5 ml at baseline also showed a final CTC reduction at the end of the therapy assessment. Conclusions: Analysis of circulating tumor cells may be of use in monitoring response to therapy in mCRC, either in combination with CEA monitoring or alone when CTCs are elevated but CEA level is not.

// Joshua E. Allen 1 , Bikramajit Singh Saroya 1 , Miriam Kunkel 1 , David T. Dicker 1 , Avisnata Das 1 , Kristi L. Peters 1 , Jamal Joudeh 1 , Junjia Zhu 1 , and Wafik S. El-Deiry 1 1 Penn State Hershey Cancer Institute, Penn State College of Medicine, Hershey, PA Correspondence: Wafik S. El-Deiry, email: // Keywords : CTC, colon cancer, colorectal cancer, metastasis, prognostic marker, apoptosis Received : October 8, 2013 Accepted : October 15, 2013 Published : October 17, 2013 Abstract Enumeration of circulating tumor cells (CTCs) by the CellSearch system provides prognostic information in metastatic colorectal cancer, regardless of metastatic site. We found that CTCs generally represent

The improved molecular understanding of cancer initiation, progression, and therapeutic resistance has yielded several novel molecular events that are being targeted by emerging therapies. While the treatment of ALL is a success story in the pediatric population, achieving a sustained remission in the adult population remains an area of investigation. Nevertheless, certain therapies have significantly improved the overall survival for adult ALL patients that should continue to improve with the discovery of better molecular targets and targeted agents. Here, we discuss novel therapeutic options under clinical investigation for the treatment of Philadephia chromosome negative ALL including immunotherapy, monoclonal antibodies, and small molecules that may be used as single agent or adjuvant therapy in the management of adult ALL.

Cytotoxic chemotherapy remains the mainstay of the medical -management of colorectal cancer (CRC). Research over the last two decades has led to a molecular understanding of the oncogenic mechanisms involved in CRC and has contributed to the rational development of antineoplastics that target these mechanisms. During carcinogenesis, genetic changes often occur in molecules that play key functional roles in cancer such as cell proliferation, angiogenesis, apoptosis, cell death and immune-mediated destruction of cancer cells. Here, we review novel antineoplastics that are approved or in development for CRC that target molecules associated with genetic aberrations in CRC. Some of these targeted antineoplastics have proven effective against other solid tumors and hold promise in treating CRC whereas others are now routinely used in combination with cytotoxic agents. This article reviews antineoplastics that target genetic changes in CRC, their antitumor mechanisms, and their stage of development.

Cytotoxic chemotherapy remains the mainstay of the medical ­management of colorectal cancer (CRC). Research over the last two decades has led to a molecular understanding of the oncogenic mechanisms involved in CRC and has contributed to the rational development of antineoplastics that target these mechanisms. During carcinogenesis, genetic changes often occur in molecules that play key functional roles in cancer such as cell proliferation, angiogenesis, apoptosis, cell death and immune-mediated destruction of cancer cells. Here, we review novel antineoplastics that are approved or in development for CRC that target molecules associated with genetic aberrations in CRC. Some of these targeted antineoplastics have proven effective against other solid tumors and hold promise in treating CRC whereas others are now routinely used in combination with cytotoxic agents. This article reviews antineoplastics that target genetic changes in CRC, their antitumor mechanisms, and their stage of development.

The improved molecular understanding of cancer initiation, progression, and therapeutic resistance has yielded several novel molecular events that are being targeted by emerging therapies. While the treatment of ALL is a success story in the pediatric population, achieving a sustained remission in the adult population remains an area of investigation. Nevertheless, certain therapies have significantly improved the overall survival for adult ALL patients that should continue to improve with the discovery of better molecular targets and targeted agents. Here, we discuss novel therapeutic options under clinical investigation for the treatment of Philadephia chromosome negative ALL including immunotherapy, monoclonal antibodies, and small molecules that may be used as single agent or adjuvant therapy in the management of adult ALL.

Augmentation and acceleration of apoptosis for cancer therapy are logical therapeutic strategies given the increasing body of data suggesting the dysregulation of control of cell death in many neoplasms. Apoptosis is particularly well studied in hematological neoplasms, thus these varied diseases present opportunities for pro-apoptotic drug development both as single agents and in combination with established therapies. Accordingly, several agents targeting function of anti-apoptotic Bcl-2 family members have entered clinical trials in the last decade and are discussed.The pan Bcl-2 family member BH3 domain mimetic obatoclax (GX15-070) is currently under clinical evaluation in solid tumors and hematological neoplasms. This agent offers the attractive property of uniformly inhibiting all of the anti-apoptotic members of the Bcl-2 protein family. Its chemistry and preclinical development and activity are reviewed. Pharmacology, pharmacodynamics, drug resistance and clinical use of this agent in leukemias and lymphomas are discussed. The prospects for obatoclax in changing clinical practice are addressed.Obatoclax may not prove to have dramatic single agent activity for hematological neoplasms. It seems more likely that its activity will be manifest in combination therapy with other agents, particularly cytotoxic chemotherapies. Results of ongoing studies are awaited.

BACKGROUND Little is known about the degree of pain experienced by patients undergoing a bone marrow aspiration and biopsy (BMAB). OBJECTIVE To evaluate the effectiveness of several strategies aimed at reducing the pain score. METHODS We conducted a retrospective analysis of 258 consecutive adult patients who underwent BMAB via 6 different approaches, the first 5 of which were performed by one physician. Group A received local anesthesia with 1% lidocaine hydrochloride (5 mL) and a 5-minute wait time before the procedure; group B received local anesthesia with a double dose (10 mL) of lidocaine; group C received 5 mL of local anesthesia with a 10-minute wait; group D received 5 mL of local anesthesia plus a topical spray with ethyl chloride; group E received oral analgesia and anxiolysis 30 minutes before the procedure in addition to the group A dosage of lidocaine; and group F received the same anesthesia as did group A, but the BMAD was performed by a less experienced practitioner. RESULTS On a 0 to 10 scale, the mean pain level among the 258 patients was 3.2 (standard deviation = 2.6). Rate of complications was low (

TO THEEDITOR: We read with interest the article by Iinuma et al 1 that describes their study examining the clinical significance of cancer stem cells in the peripheral blood of patients with colorectal cancer (CRC). The defining characteristics of a circulating cancer stem cell (CTSC) are its capacity for self-renewal and for initiation of distant metastases; some of these cells are also resistant to traditional chemo- therapy. 2,3 The concept that circulating tumor stem cells can be iden- tified and targeted is attractive and has major diagnostic, prognostic, and therapeutic implications for patients with metastatic cancer. However, the identification of authentic CTSCs continues to be a challenge because of the lack of a clear understanding of the biologic heterogeneity of tumor stem-cell populations and also because of the lack of characterization of surface markers that predict clinically im- portant subsets of CTSCs, such as those with aggressive malignant potential or drug resistance.

Circulating tumor cells (CTCs) have prognostic significance for metastatic cancers including colorectal, breast and prostate cancer. We have been evaluating CTC numbers as well as molecular markers as prognostic, treatment and disease-monitoring tools. The FDA recommends a cutoff value of > 3 CTCs/7.5 ml of blood, using Johnson & Johnson Veridex Cellsearch technology as indicator of poor prognosis for metastatic colorectal cancer (mCRC). We report identifying a population of patients who have advanced disease and liver metastases (shown by CT/MRI/PET scan) but the CTC count using our CLIA-certified Veridex Cellsearch laboratory was Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 3413. doi:1538-7445.AM2012-3413

The presence and number of circulating tumor cells (CTCs) in the blood of cancer patients is predictive of clinical outcome. To date only the CellSearch system by Veridex has been FDA approved for use in breast, prostate and colon cancer (cc). However, a limited number of biomarkers are examined in the CellSearch system. In the case of colorectal cancer, cytokeratin (CK) and epithelial cell adhesion molecule (EpCAM) expression is used to identify cells as CTCs. Additional analysis of other biomarkers by Cheshire technology is provided as fee for service and only uses the current fluorescent channels utilized by the CellSearch Analyzer. Our aim is to greatly enhance the number of biomarkers that can be tested in an economic and efficient manner and integrating the analysis into the existing CellSearch isolation and analytic platform. We combined Nuance multispectral imaging with quantum dot technology to expand the biomarkers analysis. Quantum dots (QD) have the advantage that they can be excited in the UV wavelengths while emitting across the visible spectrum. Only DAPI in the original CellSearch panel of reagents is excited in this range and its emission does not overlap with the QDs. This allows us to visualize additional biomarkers without interfering with CellSearch CTC analysis. Antibodies against a panel of known biomarkers were labeled with quantum dots. Each antibody was verified by standard immunofluorescence in appropriate tissue culture lines before quantum dot labeling. Antibody labeling and titration was confirmed on tissue culture cell lines before adding it to the panel of CTC staining reagents. To model multiplexing assays, blood is spiked with a known number of cells from appropriate cancer cell lines. Following standard CTC isolation and enumeration, cells are retained in the MagNest and allowed to dry. Cells are then stained with our biomarker panel. Currently 8 quantum dots are commercially available for simultaneous analysis. Our panel of biomarkers is under revision, but includes phospho-AKT, Ki-67, CD26, CD133 and EGFR. We are expanding the panel to include markers for death pathway signaling including Mcl-1, Bcl-XL, Flip, TRAIL, and death receptors. Separate panels are also being developed for breast cancer. Additional panels can include markers for sensitivity to chemotherapeutic reagents (TS, DPD, PTHFR, GSTP1, ERCC1, ERCC2, UGT1A1, BRAF), cell proliferation, known tumor susceptibility genes or tumor markers (APC, p53, HER2, PSA, IGF1-R) or probes for specific signal transduction pathways (NFκB, Wnt). Analysis of samples from patients undergoing chemotherapy -/+ targeted therapy is underway to evaluate the status of the stem cell, prognostic markers and cell death determinants in individual CTCs as a function of time. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 3818. doi:10.1158/1538-7445.AM2011-3818

Circulating tumor cells (CTCs) reflect disease burden in metastatic colorectal cancer (mCRC) and show reduced numbers in patients who are responding to chemotherapy. Current analysis is limited to counting CTCs and little additional marker analysis. We sought to develop methodology to allow multiplexing of marker detection in individual CTCs to assess tumor cell heterogeneity and to gain insight into relationships of prognostic and therapy response markers. This ongoing study is evaluating expression of up to 8 markers in a single CTC by a multispectral multiplex imaging technique using nano-particle type quantum dot (Q-Dot) dyes. We hypothesize that CTCs that are extremely rare (∼ 1 in 10 million RBCs) in the blood of cancer patients play an important role in metastasis. In view of the small number of such cells in the blood of mCRC patients (prognostication based on 3 CTCs/7.5 ml of blood) there is a need to derive more molecular correlations from single CTCs. We designed a clinical protocol to collect blood from mCRC patients receiving chemotherapy with the intent to evaluate multiplexed molecular marker expression in CTCs in responders versus non-responders using the FDA-approved Veridex CellSearch technology. We are evaluating CD26, CD133, EGFR, Ki 67, p27, c-Myc, ALDH1, Bcl-Xl, Mcl-1, phosho-Akt, phospho-ERK, TRAIL receptors, IAPs, and p53 using antibodies conjugated to different Q-Dots. Ongoing studies include prognostic marker, stem cell marker, and cell proliferation/death response marker analysis in human tumor xenografts of HCT116, p53-null HCT116, DLD1, SW620, and SW480. Initial profiling of tumor cell lines revealed that mutant p53 and KRAS-expressing SW620 derived from a patient with mCRC has CD26 and CD133 expression including a sub-population of tumor cells that co-express these two stem cell markers. HCT116 showed positive EGFR, CD26, but little or no CD133 expression. We are evaluating correlations between the primary tumors in mice as compared to the CTCs in mice and similar experiments are part of the clinical protocol. The studies should unravel the complexity of CTC heterogeneity, and define informative subsets of markers that can be routinely included in studies to assess prognosis and response to therapy. The ease with which CTCs can be isolated makes these studies of comparisons with primary tumors critical in order to establish and validate new methods for assessing tumor behavior and response to therapy in the clinic through this CTC “liquid biopsy” technique. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 5248. doi:10.1158/1538-7445.AM2011-5248