Frequently Asked Questions

Is there data to support the use of laboratory assays for the selection of drug therapy?

There is a large and growing volume of clinical data that establishes the predictive validity of the EVA-PCD platform. Both solid and hematologic tumors have been evaluated with statistically significant results. An early report analyzed 1280 patients whose tumors were submitted for the measurement of drug induced cell death. The overall response rate, which included treated and untreated patients, was 56 percent. When the patients’ outcomes were examined including in vitro tumor sensitivity data, those patients who received “assay-sensitive” drugs had a clinical response of 81 percent versus 13 percent for those patients who received “assay-resistant” drugs (p<0.00000001).¹

More recently, a meta-analysis in human hematologic malignancies, correlated the outcome in 1929 patients who received chemotherapy with the results of Ex-Vivo Analysis of drug induced programmed cell death. The objective response rate for patients who received “assay-sensitive” drugs was 84.6 percent, compared with patients who received “assay-resistant” drugs of 28.3 percent (p<0.0001).²

1. Proc Japanese Chemotherapy Society, Weisenthal L, 1997.
2. Andrew G. Boasanquet, Gertjan J. Kapers, Rolf Larsson, Robert A. Nagourney, Peter Nygren, Rob Pieters, Peter Staib, Ulf Tidefelt, C Michael Zwaan and Larry Weisenthal.  Individualized Tumor Response (ITR) Profiling for Drug Selection in Tailored Therapy: Meta-Analysis of 1929 Cases of Leukemia and Lymphoma.  Blood (ASH Annual Meeting Abstracts) 110:3471, 2007.

Have prospective clinical trials been conducted to confirm results?

Numerous trials have been conducted to assess the predictive validity of these platforms. In addition to retrospective correlative analyses, prospective studies have also been conducted. Two trials reported by Robert Nagourney, MD, prospectively evaluated the results of EVA-PCD platform with response, time to progression and survival.

The results in breast cancer confirmed a correlation between drug sensitivity and progression-free survival in the evaluation of cisplatin plus gemcitabine.³ A second study conducted with ovarian cancer patients established the correlation between: sensitivity and response, time to progression and overall survival.⁴ More recently, a laboratory-directed protocol in NSCLC provided an objective response rate of 62 percent, statistically significantly superior to standard outcomes (p=0.003), with a median progression-free survival of 9.5 months and median overall survival of 22.3 months.

Despite these findings, no group has been willing to formally test laboratory-directed therapy against standard protocol treatment. Amongst the organizations approached to conduct such studies — NSABP, GOG, and CCG — none have undertaken confirmatory studies. The costs of prospective clinical trials (now in the range of millions of dollars) remain a hurdle for investigators in this field. Nonetheless, a fair test of these methodologies should be conducted.

We and other investigators in the field have made it abundantly clear that we are very willing to participate in these types of studies.

1. Nagourney RA, Link JS, Blitzer JB, Forsthoff C, Evans SS: Gemcitabine Plus Cisplatin Repeating Doublet Therapy In Relapsed, Pretreated Breast Cancer.  J Clin Oncol. Vol 18, Issue 11, 2000:2245-2249
2. Nagourney RA, Brewer CA, Radecki S, Kidder W, Sommers B, Evans SS, Minor D, DiSaia, P.: A Phase II Trial of Gemcitabine Plus Cisplatin Repeating Doublet Therapy In Previously Treated, Relapsed Ovarian Cancer Patients. Gynecol Oncol. 88, 35-39 (2003)

How does the EVA-PCD platform differ from older techniques?

The EVA-PCD analysis represents a significant advance over prior methodologies. Two fundamental discoveries underlie the unique success of this platform.

1. The finding that carcinogenesis represents dysregulation of cell death, not cell proliferation.
2. The recognition that cancer biology reflects cancer ecology.

By incorporating the modern tenets of programmed cell death (one form of which is apoptosis), into the EVA-PCD platform, we have developed a technique that accurately measures the processes associated with cancer development, progression and response to therapy. Applying cell death measurements in the human tumor microspheroid has, for the first time, provided a truly predictive methodology.

Older techniques that measured cell proliferation (e.g. clonogenic assays, thymidine incorporation assays and growth to confluence techniques) failed to incorporate the modern understanding of cancer biology. These techniques, by separating cancer cells from their stroma, inflammatory elements and vasculature, failed to capture the cell-cell interactions that are now known to be crucial for response prediction.

Can you test all forms of drugs?

No. It is important for scientists in this field to recognize the applications and limitations of these methodologies. There are several classes of therapeutics that are not well tested in the laboratory environment.

Cytostatic agents (tamoxifen) do not induce programmed cell death. As such, no signal can be obtained in short term cultures that measure cell death. Although many agents were originally believed to be cytostatic, it is now recognized that some of these drugs do accurately induce cell death, resulting in a slow diminution in tumor volume. Nonetheless, the remaining drugs, which truly function in growth inhibition, will not provide useful clinical information utilizing this platform.

Pro drugs (cyclophosphamide, ifosfamide) require metabolism for activity cannot be directly tested in short term, in vitro cultures. Drugs like cyclophosphamide require P-450 mixed function oxygenases to render them active as alkylators. To evaluate these classes of drugs, we use the final product — nitrogen mustard. Nitrogen mustard is one of the most potent types of chemotherapy. This parent compound is the active moiety for all members of the alkylating class.

Agents that require concentration or conjugation (methotrexate). The important anti-folate methotrexate, and the related pemetrexed, are primarily dependent upon membrane-based concentration proteins. Secondarily they depend upon conjugation with glutamic acid residues. These processes occur too slowly in vitro to provide a signal in the EVA-PCD platform. To overcome these hurdles, we utilize related drugs that are less dependent upon these activating events. Trimetrexate is an antifol that does not require concentration of polyglutamation. Thymitaq is a TS inhibitor that can provide a signal for pemetrexed. Using these alternative drugs provides the opportunity to analyze these important compounds.

Avastin (bevacizumab) is an antibody that binds to vascular endothelial growth factor. During this process it eliminates this important protein from the blood stream and influences the vasculature. In addition to the effect on endothelial growth, it also influences endothelial cell permeability. Pre-incubation with Avastin can influence human tumor microspheroids and enhance the effect of some chemotherapies. We have focused our attention on the related VEGF inhibitors that act by preventing the signaling through tyrosine kinases. These include sunitinib, sorafenib, cediranib and vandetanib. These compounds provide insights into the role of VEGF signaling in human tumor response.

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