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Tumor Chemoresistance Assay, also known as the nonclonogenic cytotoxic drug resistance (NCDRA) and extreme drug resistance (EDR), is intended to identify cancer patients who are not likely to respond to a specific chemotherapeutic agent. The NCDRA is intended to provide information that allows patients to avoid the adverse effects of chemotherapy from which they would not benefit medically. In the absence of assay results, patients would generally receive the first-line chemotherapy of choice for their disease.

The Tumor Chemoresistance assay is performed by first culturing a tumor sample in vitro. Once viable tumor colonies have been established, each culture is exposed to a selected chemotherapeutic agent. After standard period of time the test cultures are analyzed for the number of remaining viable tumor cells. The relative decrease in number of viable cells in the test cultures compared to control cultures is taken as an indicator of tumor cell resistance. If the in vitro tumor-cell sample is resistant to a particular agent, then the agent may not be recommended for treating the patient.

 Tumor Chemosensitivity Assay, also known as the human tumor stem cell assay, is an in vitro technique designed to test the sensitivity of antineoplastic agents on cells from tumor specimens obtained from patients. The response of the tumor cell line in the laboratory is used to predict the effectiveness of the agent in the patient. More than one commercial method is available.

The assay is performed by first establishing in vitro colonies of stem cells isolated from a patient's tumor. Once viable tumor colonies have been established, they are exposed to selected single chemotherapeutic agents. After a predetermined period of time the number of colonies and number to colony forming cells in the culture are counted. The relative difference in cell number of colony forming cells in the culture are counted. The relative difference in cell proliferation in the test cultures compared to control cultures is taken as an indicator of tumor sensitivity. If an in vitro tumor-cell sample is sensitive to a particular chemotherapeutic agent, then the agent may be recommended for treating the patient.

In vitro chemoresistance and chemosensitivity assays have been developed to provide information about the characteristics of an individual patient’s malignancy to predict potential responsiveness of their cancer to specific drugs. Thus, these assays are sometimes used by oncologists to select treatment regimens for an individual patient. Several assays have been developed that differ with respect to processing of biological samples and detection methods. However, all involve the similar principles and they share protocol components including: 1) isolation of cells and establishment in an in vitro medium (sometimes in soft agar); 2) incubation of the cells with various drugs; 3) assessment of cell survival; and 4) interpretation of the result.

A variety of chemosensitivity and chemoresistance assays have been clinically evaluated in human trials. All assays use characteristics of cell physiology to distinguish between viable and non viable cells to quantify cell kill following exposure to a drug of interest. For the Oncotech Extreme Drug Resistance Assay (EDR®; Exiqon Diagnostics, Tustin, CA); and the ChemoFX® assay (Precision Therapeutics; Pittsburgh, PA) premarket approval from the U.S. Food and Drug Administration (FDA) is not required when the tests are performed in a laboratory that licensed by CLIA for high-complexity testing.

With few exceptions, drug doses used in the assays are highly variable depending on tumor type and drug class. But all assays require drug exposures ranging from several-fold below physiological relevance to several-fold above physiological relevance.

Although a variety of assays exist to examine chemosensitivity or chemoresistance, only a few are commercially available and currently used in the clinic periodically.

The DiSC assay uses dye exclusion by live cells.

• The Differential Staining Cytotoxicity (DiSC) Assay involves mechanical disaggregation of cells from surgical or biopsy specimens by centrifugation. Cells are then established in culture and treated with the drugs of interest at three dose levels; the middle dose is that which could be achieved in therapy; 10-fold lower than the physiologically relevant dose; and, 10-fold higher. Exposure time ranges from 4-6 days; then, cells are restained with fast green dye and counterstained with hematoxylin and eosin. The fast green dye is taken up by deadcells, and hematoxylin and eosin can differentiate tumor cells from normal cells. The intact cell membrane of a live cell precludes staining with the green dye. Drug sensitivity is measured by the ratio of live cells in the treated samples to the number of live cells in the untreated controls.

Several methods measure incorporation of radioactive precursors by macromolecues in viable cells.

• Tritiated thymine incorporation measures uptake of tritiated thymidine by DNA of viable cells. Using proteases and DNAse to disaggregate the tissue, samples are seeded into single cell suspension cultures on soft agar. They are then treated with the drug(s) of interest for four days. After three days, tritiated thymidine is added. After 24 hours of additional incubation cells are lysed and radioactivity is quantified and compared to a blank control consisting of cells that were treated with sodium azide. Only cells that are viable and proliferating will take up the radioactive thymidine. Therefore, there is an inverse relationship between update of radioactivity and sensitivity of the cells to the agent(s) of interest. 
• The Extreme Drug Resistance assay (EDR®) (commercially available at Exiqon Diagnostics, Tustin, CA) is methodologically similar to the thymidine incorporation assay, using metabolic incorporation of tritiated thymidine to measure cell viability; however, single cell suspensions are not required, so the assay simpler to perform. Small tissue samples are incubated with the drug(s) of interest for 5 days at doses ranging from 5-fold below to 80-fold above concentrations that would reflect physiological relevance. Subsequently, tritiated thymidine is added to the culture and update is quantified after various incubation times. Only live (resistant) cells will incorporate the compound. Therefore, the level of tritiated thymidine incorporation is directly related to chemoresistance. The interpretation of the results is unique in that resistance to the drugs is evaluated as opposed to evaluation of responsiveness. Tumors are considered to be highly resistant when thymidine incorporation is at least one standard deviation above reference samples. 
• The Histoculture Drug Resistance Assay (HDRA) is commercially available by AntiCancer, Inc. (San Diego, California) tests tissue fragments 1 to 2 millimeters in size. Samples are placed on a collagen matrix so they can grow 3 dimensionally and maintain signaling pathways mediated by cadherins and integrins, which control cell-cell and cell-matrix contact, respectively. After 24 hours, explants are incubated with drug for 3 days. Subsequently, they are fixed in formalin and embedded in paraffin. Radioactivity is quantified in slide sections using autoradiography.

Drug sensitivity is evaluated by quantification of cell growth in the 3-dimensional collagen matrix. There is an inverse relationship between the drug sensitivity of the tumor and cellgrowth. Concentrations of drug and incubation times are not standardized and vary depending on drug combination and tumor type.

• The Adenosine Triphosphate (ATP) Bioluminescence Assay relies on measurement of ATP to quantify the number of viable cells in a culture. Single cells or small aggregates are cultured, then exposed to drugs. Following incubation with drug the cells are lysed and the cytoplasmic components are solubilized under conditions that will not allow enzymatic metabolism of ATP. Luciferin and firefly luciferase are added to the cell lysis product. This catalyzes the conversion of ATP to adenosime di- and monophospate and light is emitted proportionally to metabolic activity. This is quantified with a luminometer. From the measurement of light, the number of cells can be calculated. A decrease in ATP indicates drug sensitivity, whereas no loss of ATP suggests that the tumor is resistant to the agent of interest. 
  • Precision Therapeutics (Pittsburgh, Pennsylvania) commercially markets ChemoFX®, which uses this technology http://www.chemofx.com/index.html. While the firefly luciferase and luciferin catalyze reduction of ATP and emitted light is quantified using a luminometer; cells must be grown in a monolayer rather than in a three-dimensional matrix.

The rationale for chemosensitivity assays is strongest where there are a variety of therapeutic options and there are no clear selection criteria for any particular regimen in an individual patient.

In vitro chemoresistance assays, including but not limited to extreme drug resistance assays, are considered investigational.

None

Investigative service is defined as the use of any treatment procedure, facility, equipment, drug, device, or supply not yet recognized by certifying boards and/or approving or licensing agencies or published peer review criteria as standard, effective medical practice for the treatment of the condition being treated and as such therefore is not considered medically necessary.

The coverage guidelines outlined in the Medical Policy Manual should not be used in lieu of the Member's specific benefit plan language.

12/18/2000: "and extreme drug resistance (EDR)" added to third paragraph under "description" section.

2/8/2002: Investigational definition added

5/1/2002: Type of Service and Place of Service deleted

9/2/2004: Code Reference section completed, non-covered table added

8/5/2005: Code Reference section reviewed, no changes

10/17/2006: Policy reviewed, no changes

1/4/2007: Policy reviewed, policy statement rewritten for clarity

2/19/2008: Policy reviewed, no changes

4/24/2009: Policy reviewed, no changes

04/27/2010: Policy title changed from “In Vitro Chemotherapeutic Drug Assays” to “In Vitro Chemoresistance and Chemosensitivity Assays” to reflect the scope of the policy. Policy description extensively re-written to add information on available tests and how they are performed. Policy statement unchanged. Deleted outdated references from the Sources section.

06/21/2011: Policy reviewed; no changes.

04/26/2012: Policy reviewed; no changes.

All codes billed for this procedure are considered investigational and not eligible for coverage.

Non-Covered Codes

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