Chemotherapy resistance and oncogene expression in non-small cell lung cancer
January 31st, 2007
Multi-drug chemotherapy resistance in non-small cell lung cancer tumor cultures is common, and associations between molecular markers and in vitro chemotherapy resistance are noted. Clinical validation through integration of such testing into clinical trials seems warranted.
At 1:29 pm on March 17th, 2007 Gregory D. Pawelski said:
It must be realized that these assays are complex procedures, fraught with the potential for error and misinterpretation. The results are only meaningful to the extent that the laboratory in question is experienced and diligent in its quality assurance practices. A patient interested in this testing should not hesitate to ask specific questions and to require specific answers of the laboratories under consideration.
The extreme drug resistance assays (EDR) cannot “accurately” predict for drug sensitivity. This kind of assay is specifically designed to identify “inactive” drugs and should not be used to identify “active” drugs. It has a very high specificity (>90%) for identification of “inactive” agents. However, many effective drugs do not test in the extreme drug resistance range. It has a relatively low sensitivity (>40%) for identification of “active” agents.
The “EDR” assay (a thymidine soft agarose assay) is much less useful and less well validated than are the various “cell-death” assays for the purpose of identifying “effective” treatment regimens. There are better ways of getting more useful information, namely: (1) higher evaluability rates, (2) more drugs tested per specimens, (3) multiple drug concentrations tested using multiple complementary endpoints, and (4) more extensive clinical validation of assay results. There is no data to indicate that the thymidine endpoint is in any way superior or advantageous over the cell-death endpoints.
The probability of a drug working depends on both the clinically expected historical experience and upon the drug resistance category (sensitive, intermediate, resistant, EDR) in which each drug falls, as determined by the assay. For example: If a given drug has a clinically expected response rate of 40% and the assay result is “sensitive,” then the drug has a 68% probability of providing clinical benefit (a clinically meaningful reduction in the amount of cancer). If the assay result is “resistant,” then the drug would have only an 8% probability of providing clinical benefit. If the assay result is “EDR,” then the drug would have only a 4% probability of providing clincal benefit.
The extreme drug resistance assay is a rather simple, straightforward test, requiring only about 5 minutes per assay of physician/lab work. DISC, MTT, and ATP assays are more geared to identifying active drugs (drugs which will work, as opposed to drugs which won’t work), and are a whole lot more labor intensive, requiring an average of 3 hours of physician/lab work. DISC, MTT, ATP and fluorescein diacetate assays, have a very high specificity (>90%) for identification of “inactive” agents, and a high sensitivity (>70%) for identification of “active” agents.
Different Types of Cell-Death Assays
The TCR Assay
ChemoFX Assay
The MiCK Assay
HDRA Assay
EVA Assay
DiSC Assay
MTT Assay
ATP Assay
Fluorescein Diacetate Assay
Some of the other assays focuse more on the discrimination between a relatively lower probability of response on one hand and a relatively higher response on the other hand. Basically, identify each and every drug as being either “good prognosis,” “poor prognosis,” or “average prognosis,” with the advantage of a good prognosis over a poor prognosis drug as being (on average) 7:1, meaning a good prognosis drug is 7 times as likely to work as a poor prognosis drug. Results used in a “positive” way to “select” the drugs most likely to work.
In order to have an assay done, tumor cells must be accessible through biopsy (needle biopsy is not enough). Tissue, blood, bone marrow, and ascites and pleural effusinos are possibilities, providing tumor cells are present, and only live cells can be used. At least one gram of fresh biopsy tissue is needed to perfom the tests, and a special kit must be gotten in advance from the lab. Arrangements have to be made with the surgeon and/or pathologist for preparation and sending of the specimen.
Good review papers exist and are increasingly appreciated, understood, and applied by private sector and European clinicians and scientists. This literature is not understood by many NCI investigators and by NCI-funded university investigators. NCI studies never determine if fresh tumor assays worked. All of the considerable literature which supports the use of these assays in patient management has been based on true fresh tumor (non-passaged) cell assays.
The NCI used “cell lines” because the major expertise of the investigators who carried out any study was in the creation of cancer cell lines, and they wanted to see if they could perform assays on these cell lines to use in patient therapy. The results were that they were able to test successfully only 22% of specimens received, including only 7% of primary lesions. This contrasts with a 75% overall success rate reported by earlier investigators who used the same assay system in fresh tumors and a routinely obtained > 95% success rate using improved methods available today.
Upgrading clinical therapy by using drug sensitivity assays can improve the conventional situation by allowing more drugs to be considered. Drug sensitivity tests support the idea that a marginal benefit in terms of overall survival is observed in cancer patients with normal prognoses, but there are marked survival benefits for cancer patients with poor prognoses.
The key to improving drug sensitivity tests is related to the number and types of drugs tested. The more anti-cancer drug types there are in the selective arsenal, the more likely the system is to prove beneficial. In order to acquire sufficient data, tumors should be tested with at least two assay endpoints, and most often three, for sensitivity tests in any one patient. On average, up to twenty drugs and combinations at two concentrations in three different assay systems, is an effective way to avoid false-positive or false-negative data. Careful choice of drug doses and administration intervals also improves outcomes.
Drug sensitivity assays do not harm patients in any way except in terms of cost. Every cancer patient should have his/her own unique chemotherapy trial based on consultation of pathogenic profiles and drug sensitivity testing data. Research and application of drug sensitivity assays are being encouraged by growing patient demands, scientific advances and medical ethics. Drug sensitivity tests are not a luxury but an absolute necessity, and a powerful strategy that cannot be overlooked. Having some foreknowledge of a given agent’s expected result before its administration would benefit the individual patient.
At 10:25 pm on July 6th, 2007 Gregory D. Pawelski said:
Medicare Contractor Establishes Reimbursement Coverage Policy for Cell Culture Assay Tests
National Heritage Insurance Company (NHIC), the contractor that administers Medicare programs in California, has established a positive coverage policy for Cell Culture Assay Tests known as Chemosensitivity (Resistance) Testing for a tumor specimen from a Medicare patient obtained anywhere within the United States, but submitted for testing by one of the approved laboratories located within California. Medicare bills for Chemosensitivity (Resistance) Testing are billed through NHIC because the test is conducted by the approved laboratories in California.
The Chemosensitivity (Resistance) Test can help see what treatments have the best opportunity of being successful for “high” risk cancer patients. The test measures the response of “live” tumor cells to drug exposure. Following this exposure, the assays measure both cell metabolism and cell morphology (Functional Profiling). The integrated effect of the drugs on the whole cell, resulting in a cellular response to the drug, measuring the interaction of the entire genome. Assays based on “cell-death” occur in the entire population of tumor cells.
This cell culture assay technology has been clinically validated for the selection of optimal chemotherapy regimens for individual patients. It is a laboratory analysis based on tumor tissue profiling that uses “fresh” human tumor biopsy or surgical specimen to determine which drugs or combinations of chemotherapeutic agents have the highest likelihood of response for individual cancer patients.
Following the collection of “fresh” tumor cells obtained from surgery or tru-cut needle biopsies, a cell culture assay is performed on the tumor sample to measure drug activity (sensitivity and resistance). This will pinpoint which drug(s) are most effective. Tissue, blood, bone marrow, and ascites and pleural effusions are possibilities, providing tumor cells are present. At least one gram of fresh tissue is needed to perform the tests, and a special kit is obtained in advance from the lab.The treatment program developed through this approach is known as assay-directed therapy.
Individualized assay-directed therapy is based on the premise that each patient’s cancer cells are unique and therefore will respond differently to a given treatment. This is in stark contrast to standard or empiric therapy, which chemotherapy for a specific patient is based on average population studies from prior clinical trials.
The decision had been made that the assay is a perfectly appropriate medical service, worthy of coverage on a non-investigational basis. What is of particular significance is that they abandoned the artificial distinction between “resistance” testing and “sensitivity” testing and are providing coverage for the whole FDA-approved kit. Drug “sensitivity” testing is merely a point a little farther along on the very same continuum which “resistance” testing resides.
Cell cuture assay tests based on “cell-death” have proven very effective in identifying novel treatment combinations for a variety of cancers. The value of cell-death assays is that they can and do accurately predict clinical outcomes and define novel chemotherapeutic synergies. It can help see what treatments will not have the best opportunity of being successful (resistant) and identify drugs that have the best opportunity of being successful (sensitive).
The current clinical applications of in vitro chemosensitivity testing is ever more important with the influx of new “targeted” therapies. Given the technical and conceptual advantages of “functional profiling” of cell culture assays together with their performance and the modest efficacy for therapy prediction on analysis of genome expression, there is reason for renewed interest in these assays for optimized use of medical treatment of malignant disease.
The payment provided will be sufficiently realistic that all Medicare patients for whom this testing is indicated will be able to get it with only the routine 20% co-payment, as Medi-gap insurance secondaries are mandated to provide payment for co-pays for Medicare-approved services.
The coverage became effective for claims for services performed on or after February 19, 2007. The decision is posted at:
http://www.medicarenhic.com/cal_prov/policies.shtml
http://www.medicarenhic.com/cal_prov/articles/chemoassaytest_0107.htm