- Email: [email protected]
FUTURE DIRECTIONS IN LUNG CANCER RESEARCH AND THERAPEUTICS
MULTIDISCIPLINARY CARE OF LUNG CANCER PATIENTS, PART I1
0889-8588/97 $0.00
+ .20
FUTURE DIRECTIONS IN LUNG CANCER RESEARCH AND THERAPEUTICS Anthony D. Elias, MD
Lung cancer is preventable, common, lethal once it comes to clinical attention, and relatively resistant to current therapeutics. Much invasive and toxic therapy is offered in the name of hope. Should we discourage our present therapeutic incrementalism and instead concentrate our resources entirely on measures to prevent the development of overt lung cancer? Should we place every patient with lung cancer on clinical translational trials because standard therapy does not deserve its name? What can we afford? These are hard questions. Ultimately the field of oncology can only proceed with a sense of purpose and be optimistic that these obstacles to cure and prevention are surmountable.
EARLY DETECTION AND PREVENTION In Part I of this series, Drs. Miller and Franklin, Mulshine et al, and Emmons et a1 provided a thorough discussion of different aspects to the overall strategy of targeting early events (see also the article by Khuri et a1 in this issue). Detection and modulation of early events in lung carcinogenesis represents one of the most likely arenas to have an impact on the development of clinical disease. Despite the presence of a whole field of altered cells, the markedly lower degree of heterogeneity, adapt-
From the Department of Medical Oncology, Dana-Farber Cancer Institute, Boston, Massachusetts ~
~
~
~~~~~
HEMATOLOGY/ONCOLOGY CLINICS OF NORTH AMERICA VOLUME 11 * NUMBER 3 * JUNE1997
519
520
ELIAS
ability, and growth potential of these few cells compared with the billions of established malignant cells in the clinical disease state dramatically increases the potential impact of early intervention. An important lesson from the treatment of overt cancers, or as a parallel, infectious disease is that intervention should be targeted simultaneously against several identifiable molecular events that would otherwise produce a committed and surviving malignant cell. A single interruptiodintervention in this cascade might be easily side-stepped. What still must be identified are (1) which of the many molecular events are critically necessary for the survival of the malignant phenotype, and (2) how a premalignant cell can surmount intervention at those critical points. Not all genetically defined mutations lead to subsequent carcinoma, and many are irrelevant for their survival. An additional difficulty with intervention at this stage of preclinical disease is that millions of persons are at risk, but only a small percentage would actually go on to develop a clinically apparent tumor. Thus, any therapeutic intervention must have low host toxicity.
ADVANCES IN THE CURRENT MODALITIES: SURGERY, RADIATION THERAPY, AND CHEMOTHERAPY Minimally invasive surgical techniques appear to reduce the morbidity of resection, and they allow surgical approaches in those patients with poor respiratory reserve. If radiotherapy and systemic therapy improve, lung-sparing resections will become more common, as with surgery done for breast cancer. Improved staging, both invasive and noninvasive (of particular interest, positron emission tomography [PET] and/or radioligand scanning), and prognostic markers may help to define those few patients with early-stage disease who do not require systemic therapy. Radiation therapy to the chest is toxic and produces extremely poor local-regional control. Reduction of toxicity requires normal tissue sparing, and improved control requires increased dose. These goals might be achieved by three-dimensional conformal planning, respiratory cycle gating, use of tumor-selective tissue sensitizers, new fractionation schemes, or judicious surgical resection to limit radiation ports, as outlined by Arriagada. Present-day chemotherapy has some benefit. The ongoing randomized studies in each stage of disease to define the degree of benefit and the level of toxicity are critical to establish a benchmark from which to assess new therapeutics. Support for clinical trials is paramount. New systemic agents with greater efficacy are urgently required. The fact that
FUTURE DIRECTIONS IN LUNG CANCER RESEARCH AND THERAPEUTICS
521
a number of new agents are now in clinical evaluation is encouraging, and it suggests that the new screening methodology against solid tumor cell line panels at the National Cancer Institute and from the pharmaceutical industry is proving productive. New tools to disrupt resistance mechanisms to chemotherapy, to disrupt the metastatic process (such as inhibitors of proteolytic enzymes which penetrate intercellular matrices and basement membrane), cell adhesion molecules, growth factor receptors, or the tumor-induced host support of the microenvironment (eg, angiogenesis, matrix metalloproteinase inhibitors, hypoxic cell sensitizers, and so on) are now available for clinical trials. It is unlikely that any one of these approaches will work alone. The pharmaceutical industry must be encouraged to risk combining these with chemotherapy, other modalities, or each other.
HIGH-DOSE THERAPY The dose-response relationship in NSCLC has been insufficiently explored? No role for high-dose chemotherapy has been established or even suggested by the reported trials in metastatic disease. Indeed, given the poor overall activity of currently available chemotherapeutic agents as measured by response rates, an alchemist’s skills are required to generate a meaningful dose-survival benefit in widespread metastatic disease. Response rates increase when earlier-stage disease is treated, however, and are more likely to correlate with survival benefit. Thus, Shea’s phase I1 Cancer and Leukemia Group B (CALGB-9531) trial will evaluate the clinical and pathologic response to stem cell-supported high-dose carboplatin with Taxol as preoperative therapy for stage I11 NSCLC to indicate whether dose-intensive therapy is worth developing for stages I1 and I11 NSCLC. A number of new agents have demonstrated major activity against NSCLC. These include the taxanes, topoisomerase I inhibitors, vinorelbine, and gemcytabine, but their potential for use in high-dose therapy may be limited. For example, the topoisomerase I inhibitors are more dependent on schedule than dose, and they produce dose-limiting gastrointestinal toxicity at conventional doses. The dose range for new agents should be established as part of their phase I evaluation, with cytokine and progenitor cell support if necessary, to determine the nonhematologic dose-limiting toxicities, as well as their potential for high-dose development. An agent with both preclinical and clinical evidence for a dose-response relationship, a novel mechanism of action, and nonoverlapping nonhematopoietic dose-limiting toxicity encountered only after many-fold dose escalation should be considered a promising candidate to combine with other drugs into a high-dose regimen.
522
ELIAS
MINIMAL RESIDUAL TUMOR (MRT)
One of the most powerful predictors of outcome in most malignancies including lung cancer is the demonstration of metastatic involvement, either regionally to nodes or widespread to various organs. Thus, the development of rare event analysis technology to detect small numbers of malignant cells circulating in blood or residing in marrow or lymph nodes has become an exciting new area fostered by the field of high-dose therapy with stem cell support. Few clinical experiences are reported in MRT detection in patients with NSCLC. Pante15 screened 82 early stage I and I1 resected patients, evaluating bilateral iliac crest and sternal marrow aspirates. He used cytokeratin-18 (MoAb CK2) immunoperoxidase as a single detection reagent and reported a 1.7% false-positive rate from squamous cell contamination and other artifacts. An astonishing 22% of these earlystage patients, however, had abnormal keratin positive cells in their marrow. In a follow-up, 139 patients were screened, of whom 83 (60%) had keratin-positive cells in the marrow.6 Nodes were histologically uninvolved by tumor in 66 patients. Nine (75%) of 12 node-negative marrow keratin-positive patients relapsed, and 19 (35%) of 54 nodenegative marrow keratin-negative patients relapsed, with a median follow-up of 39 months ( P = .023). Keratin positivity in marrow did not change clinical prognosis in the 62 patients with node-positive disease. False positives were detected in 6 of 215 (2.8%)control patients. Brugger et all examined peripheral blood only following a first cycle of chemotherapy (VP-16, ifosfamide, cisplatin) and granulocyte colony-stimulating factor (G-CSF).One of 12 patients with stage IV NSCLC had keratinpositive cells circulating in the blood. Occult nodal metastases have been detected using similar immunohistochemical techniques in patients with histologically uninvolved regional nodes. Use of a polyclonal antikeratin antibody immunoperoxidase technique detected tumor cells in 38 of 60 (63%) patients with early-stage NSCLC.3No statistically significant survival impact was documented; median survival of 6.7 years and 5.4 years were observed for the keratin-negative and keratin-positive node patients, respectively. In another report, the Ber-Ep4 MoAb was used in 72 node-negative patients, of whom eleven (15./0) had isolated ber-ep4 positive cells detected.s Five of 10 (50%) Ber-Ep4-positive patients relapsed as compared with eight of 56 (14%) Ber-Epknegative patients, with a median followup of 26 months ( P = .005). In summary, limited information is available about the incidence of tumor contamination of hematopoietic tissues in lung cancer patients. Still less is known about the clinical impact of screening. With the burgeoning interest and an expanding technology in the detection of
FUTURE DIRECTIONS IN LUNG CANCER RESEARCH AND THERAPEUTICS
523
rare events, additional prospective trials to evaluate the clinical significance of marrow and/or nodal tumor contamination are needed. Heterogeneity of antigen expression in tumor cell populations requires the use of multiple reagents to detect all tumor cells. An elegant study by Pantel et a17 was performed in gastric, colon, and breast cancers, demonstrating the presence of tumor in marrow by keratin immunoperoxidase. Secondary chromophores tagged to antibodies to HER-2/neu, Ki-67, and p120 demonstrated the presence of several populations of tumor cells with presumably differing biologic behavior. Major questions remain. The methodology must improve to have sensitivity greater than one tumor cell in a million, to avoid falsepositive events, and to evaluate heterogeneity of antigen expression independently. Microscopic residual tumor represents therapy-resistant tumor and is likely to be heterogeneous. Detection and ultimately characterization of these cells by evaluating patterns of coexpression of biologic markers may guide pre- or post-transplant therapy. Although the demonstration of tumor cells in marrow or nodes indicates the ability of malignant cells to escape their local environment, this does not automatically imply that all the cells detected have mastered the biologic functions needed to grow into overt metastatic disease. In addition, critical thresholds of numbers of functional metastatic cells may still be needed to escape immune surveillance, systemic therapies such as chemotherapy, or cellular competition between normal tissues and these abnormal clones.
NEW MODALITIES
Monoclonal Antibodies, Ligands, and Receptor Targets (f Linked to Toxic Agents) Monoclonal antibodies have not lived up to their early promise as the ”magic bullets.” Although on occasion this was due to cross-reaction with normal tissues causing toxicity, frequent causes were pharmacodynamic with poor drug delivery due to a bulky messenger, metabolic degradation, or rapid immunologic clearance due to human anti-mouse antibodies (HAMA). Other important causes included antigenic modulation and heterogeneity of antigenic density. Finally, cytotoxicity can be circumvented through a variety of mechanisms, including the fact that these therapies constitute passive transfer rather than represent the development of active immunity. Human chimeric antibodies linked to powerful toxins or radioactive ligands are currently in trials. Trials listed in the Physician Direct Query (PDQ) registry (Table 1)include yttrium90 linked to the MoAb B3, to the MoAb CC-49 (together with interferon-
524
ELIAS
Table 1. SELECTED TRIALS UTILIZING NOVEL MODALITIES LISTED IN PDQ OR RECUMBENT DNA ADVISORY COMITTEE REGISTRIES Target Population
Institute
Methods
Eady Detection:
Stage I resected NSCLC AtypialClS in sputum
SWOGlECOG Univ Colorado
sputumfbronchial washings white light versus autofluorescence bronchoscopy
Antibody:
Tumors (++ EGF-R)
MSKCC phase I
Tumor 30% + 83 Ag NSCLC 10% + TAG-72 Ag
NCI phase I Alabama phase I
MDX447 bispecific Ab ? GCSF link APC with EGF-R expressing tumor cells goY-labeledMoAb B3 g’Y-labeled MoAb CC-49 plus interferon g’Y-labeled-biotic and NR-LU10-streptavidin
Tumor 25% Gene Therapy:
Vaccine:
+
NR-LU-10 Ag
Virginia Mason phase I
Antisense mutated K-ras Wild-type p53
M.D. Anderson
IL-2
Harbin
CEA
NCI
AdenoCA
+ CEA)
NCI phase I
AdenoCA
+ ras mutation)
NCI phase I
Tumor ( + ras mutation)
NCI phase I
Tumor ( + ras or p53 mutation)
Vanderbilt phase I
Tumor ( + ras or p53 mutation)
NCI phase II
retroviral vector, direct injection into endobronchial tumor TIL cells reinfused into malignant pleural effusions vaccinia vector, subcutaneous injection CEA peptide-1 plus adjuvant Detox HLA-A2 restricted mutated ras protein plus adjuvant QS21 K-ras val-12, asp-12, cys-12, or asp-13 mutated ras protein plus adjuvant Detox K-ras Val-12, asp-12, cys-12 peripheral blood buffy coat cells pulsed with tumorspecific synthetic mutated peptide GM-CSF incubated peripheral blood buffy coat cells pulsed with tumor-specific synthetic mutated peptide plus interleukin-2
GM-CSF = granulocyte-macrophage colony-stimulating factor; CEA = carcinoernbryonic antigen; G-CSF lWyte colony-stimulatingfactor; CIS = carcinoma in situ; TIL = tumor intiltrating lymphocytes.
=
granu-
alpha as an adjuvant), or to biotin in patients preloaded with NR-LU10-streptavidin. Clinical results are not currently available. In addition, a bispecific antibody linking CD64 on macrophages to the EGF-receptor (MDX447) has the potential to generate active immunity by enhancing phagocytosis of tumor cells overexpressing EGF-R with subsequent tumor-associated peptide antigen presentation via HLA class I mechanisms. Because hepatocytes express high levels of EGF-R, liver toxicity is a potential. All these trials continue to use single agents to pursue single targets, which are unlikely to succeed at a phase I1 level.
FUTURE DIRECTIONS IN LUNG CANCER RESEARCH AND THERAPEUTICS
525
Of even greater potential are small receptor ligands bound to small toxins (eg, peptides bound to yttrium, maysantines, or the minor groove binder CC1065). These compounds are likely to have favorable pharmacologic delivery to tumor sites, less antigenicity, and greater biologic half-life.
Gene Therapy Gene therapy is still in its infancy from a clinical standpoint. Current vectors favor delivery and transduction of genetic material into proliferating cells preferentially. Strategies currently being pursued include the placement of cytokine genes (eg, IL-2) into tumor-stimulated lymphocytes to try to select for the T cells that proliferated in response to exposure to tumor-associated antigens. These cells are then reinjected into the patient. A second strategy is to overexpress MDR-1 into CD34 + hematopoietic stem cells to lessen hematologic toxicity from chemotherapy agents. A concern, however, is that hematologic stem cells already express high levels of MDR-1, tumor cells residing in the marrow might acquire the gene, and most of the chemotherapeutic agents affected by P-glycoprotein have mucosal, not hematologic, toxicity as a dose-limiting factor. An interesting approach is to place suicide genes, such as genes coding for P450s activating cyclophosphamide, viral thymidylate synthetase (sensitivity to ganciclovir), or those rendering sensitivity to 5-fluorouracil (5-FU) into tumor cells (cytosine deaminase) into vectors with tumor-specific targeting. This may be particularly useful for ex vivo applications such as marrow purging.2 Placement of cytokine genes into killed tumor cells, such as interferon-gamma to upregulate HLA class I, or granulocyte-macrophage colony-stimulating factor (GM-CSF) to attract antigen-presenting cells (APCs) to the tumor cells as a trigger to develop active immunity, are strategies in the clinic for renal cell carcinoma and for melanoma. Replacement of function genes such as wildtype p53 or RB-1, or administration of antisense oncogenes (such as antisense K-ras), are strategies designed to inhibit mutant expression and to replace function and thereby resume cellular controls as least in part. Currently, several gene therapy trials are specifically targeting NSCLC. At M.D. Anderson, either the antisense K-ras mutant or the p53 wild-type genes packaged in a retroviral vector with a p-actin promoter (supplied by Genetic Therapy) are injected directly into endobronchial tumors. Tumor regression has been reported in patients. Mutated K-ras is an attractive target because it is a commonly mutated gene in tumors, it is not wildly overexpressed, and it is centrally involved in signal transduction, and therefore is less likely to mutate further without loss
526
ELIAS
of function. Loss of p53 function in tumors is also a common pathway. Resolution of malignant pleural effusions has been reported by injection of genetically modified tumor infiltrating lymphocyte (TIL) cells (expressing IL-2; supplied by Anticancer) into the pleural cavity in a trial conducted in Harbin, China. At the NCI, part of the carcinoembryonic antigen (CEA) gene is being delivered in a vaccinia vector (supplied by Thereon). Other trials include the delivery of the cystic fibrosis gene in adenoviral vectors by inhalational therapy. Tumor Vaccines Vaccines against cancer cells are generally believed to be effective if they stimulate cellular immunity. Vaccines against peptides are HLA class I-restricted, because peptides are usually presented to T8 cytotoxic T cells in the context of the HLA class I molecule without internal processing by APCs. Thus, most clinical trials utilizing peptides are targeting HLA-A2, because this tissue type is present in about 40% of the US population. Vaccines against proteins are not host-HLA restricted, because the proteins are generally internalized by an APC, processed, and then presented on the cell surface as various peptides bound to the host HLA class I molecule. Alternative strategies to present proteins include the use of viral vectors (vaccinia and adenovirus are the two most common) or APCs which are genetically modified to produce specific gene products. These strategies generally present processed peptide antigens in the context of HLA class I1 molecules. To generate a greater array of potential antigens, whole cancer cells may be phagocytosed by or fused into APCs. Processed peptide antigens in the context of HLA class I1 molecules will trigger cellular immunity mediated by T4 cells, provided the appropriate co-stimulatory pathways (such as B7-1, B7-2, CD40 and CD40 ligand, and so on) are activated as well. Otherwise, anergy /tolerance might develop. Thus, various strategies exist to trigger these pathways, including direct vaccination of tumor-associated peptides and proteins with adjuvants, or killed tumor cells given subcutaneously. Genetic modification of the tumor cells to produce cytokines (such as GM-CSF, interleukin-2, tumor necrosis factor, and other interleukins), or to produce large amounts of the co-stimulatory molecules, are designed to trigger the immune system in vivo. Routes of administration include subcutaneous, inhalational, or cellular (eg, loaded onto dendritic cells or other APCs). Current endpoints of vaccine trials are to determine whether cellular or humoral immunity to the specific antigen can be documented prior to and after the vaccination, and whether toxicity and/or tumor response occurs.
FUTURE DIRECTIONS IN LUNG CANCER RESEARCH AND THERAPEUTICS
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It is likely that these new methods of gene therapy and vaccination will also work best in states of minimal tumor burden. Once safety is demonstrated in the advanced disease setting, these approaches need to be rapidly evaluated in the adjuvant settings and in the high-risk (prevention) settings.
SUMMARY
Although immune and genetic approaches do not yet have substantial clinical exposure in NSCLC, it is hoped that the future will bring less toxic, more specific, and more effective methods to prevent the development of end-stage NSCLC. The principles of multitargeted and multimodality therapy to overcome tumor heterogeneity and resistance, and drug delivery issues have been developed over years of effort using chemotherapy. The lessons learned should not be forgotten, because they will remain applicable to newer biologic and genetic modalities.
References 1. Brugger W, Bross KJ, Glatt M, et al: Mobilization of tumor cells and hematopoietic progenitor cells into peripheral blood of patients with solid tumors. Blood 8 3 : 6 3 M O , 1994
2. Chen L, Pulsipher M, Chen D, et a 1 Adenovirus mediated transgene expression for detection and elimination of contaminating carcinoma cells in hematopoietic stem cell sources. J Cell Invest 982539-2548, 1996 3. Chen Z, Perez S, Holmes EC, et a 1 Frequency and distribution of occult micrometastases in lymph nodes of patients with non-small cell lung carcinoma. J Natl Cancer Inst 85:493-498, 1993 4. Elias A, Cohen BF: Dose intensive therapy in lung cancer. In Armitage JO, Antman KH (eds): High-Dose Cancer Therapy: Pharmacology, Hematopoietins, Stem Cells, ed 2. Baltimore, Williams and Wilkins, 1995, pp 824-846 5. Pantel K, Izbicki JR, Angshvurm M, et al: Immunocytological detection of bone marrow micrometastasis in operable non-small cell lung cancer. Cancer Res 53:1027-1031, 1993 6. Pantel K, Izbicki J, Passlick B, et a 1 Frequency and prognostic significance of isolated tumor cell in bone marrow of patients with non-small cell lung cancer without overt methastases. Lancet 347649453,1996 7. Pantel K, Schlimok G, B r a n S, et a1 Differential expression of proliferation-associated molecules in individual micrometastatic carcinoma cells. J Natl Cancer Inst 85:14191424, 1993
8. Passlick B, Izbicki JR, Kubuschak B, et al: Immunocytochemical assessment of individual tumor cells in lymph nodes of patients with non-small cell lung cancer. J Clin Oncol 121827-1832, 1994
Address reprint requests to Anthony D. Elias, MD Dana-Farber Cancer Institute Department of Medical Oncology/STAMP 44 Binney Street Boston, MA 02115
0889-8588/97 $0.00
+ .20
FUTURE DIRECTIONS IN LUNG CANCER RESEARCH AND THERAPEUTICS Anthony D. Elias, MD
Lung cancer is preventable, common, lethal once it comes to clinical attention, and relatively resistant to current therapeutics. Much invasive and toxic therapy is offered in the name of hope. Should we discourage our present therapeutic incrementalism and instead concentrate our resources entirely on measures to prevent the development of overt lung cancer? Should we place every patient with lung cancer on clinical translational trials because standard therapy does not deserve its name? What can we afford? These are hard questions. Ultimately the field of oncology can only proceed with a sense of purpose and be optimistic that these obstacles to cure and prevention are surmountable.
EARLY DETECTION AND PREVENTION In Part I of this series, Drs. Miller and Franklin, Mulshine et al, and Emmons et a1 provided a thorough discussion of different aspects to the overall strategy of targeting early events (see also the article by Khuri et a1 in this issue). Detection and modulation of early events in lung carcinogenesis represents one of the most likely arenas to have an impact on the development of clinical disease. Despite the presence of a whole field of altered cells, the markedly lower degree of heterogeneity, adapt-
From the Department of Medical Oncology, Dana-Farber Cancer Institute, Boston, Massachusetts ~
~
~
~~~~~
HEMATOLOGY/ONCOLOGY CLINICS OF NORTH AMERICA VOLUME 11 * NUMBER 3 * JUNE1997
519
520
ELIAS
ability, and growth potential of these few cells compared with the billions of established malignant cells in the clinical disease state dramatically increases the potential impact of early intervention. An important lesson from the treatment of overt cancers, or as a parallel, infectious disease is that intervention should be targeted simultaneously against several identifiable molecular events that would otherwise produce a committed and surviving malignant cell. A single interruptiodintervention in this cascade might be easily side-stepped. What still must be identified are (1) which of the many molecular events are critically necessary for the survival of the malignant phenotype, and (2) how a premalignant cell can surmount intervention at those critical points. Not all genetically defined mutations lead to subsequent carcinoma, and many are irrelevant for their survival. An additional difficulty with intervention at this stage of preclinical disease is that millions of persons are at risk, but only a small percentage would actually go on to develop a clinically apparent tumor. Thus, any therapeutic intervention must have low host toxicity.
ADVANCES IN THE CURRENT MODALITIES: SURGERY, RADIATION THERAPY, AND CHEMOTHERAPY Minimally invasive surgical techniques appear to reduce the morbidity of resection, and they allow surgical approaches in those patients with poor respiratory reserve. If radiotherapy and systemic therapy improve, lung-sparing resections will become more common, as with surgery done for breast cancer. Improved staging, both invasive and noninvasive (of particular interest, positron emission tomography [PET] and/or radioligand scanning), and prognostic markers may help to define those few patients with early-stage disease who do not require systemic therapy. Radiation therapy to the chest is toxic and produces extremely poor local-regional control. Reduction of toxicity requires normal tissue sparing, and improved control requires increased dose. These goals might be achieved by three-dimensional conformal planning, respiratory cycle gating, use of tumor-selective tissue sensitizers, new fractionation schemes, or judicious surgical resection to limit radiation ports, as outlined by Arriagada. Present-day chemotherapy has some benefit. The ongoing randomized studies in each stage of disease to define the degree of benefit and the level of toxicity are critical to establish a benchmark from which to assess new therapeutics. Support for clinical trials is paramount. New systemic agents with greater efficacy are urgently required. The fact that
FUTURE DIRECTIONS IN LUNG CANCER RESEARCH AND THERAPEUTICS
521
a number of new agents are now in clinical evaluation is encouraging, and it suggests that the new screening methodology against solid tumor cell line panels at the National Cancer Institute and from the pharmaceutical industry is proving productive. New tools to disrupt resistance mechanisms to chemotherapy, to disrupt the metastatic process (such as inhibitors of proteolytic enzymes which penetrate intercellular matrices and basement membrane), cell adhesion molecules, growth factor receptors, or the tumor-induced host support of the microenvironment (eg, angiogenesis, matrix metalloproteinase inhibitors, hypoxic cell sensitizers, and so on) are now available for clinical trials. It is unlikely that any one of these approaches will work alone. The pharmaceutical industry must be encouraged to risk combining these with chemotherapy, other modalities, or each other.
HIGH-DOSE THERAPY The dose-response relationship in NSCLC has been insufficiently explored? No role for high-dose chemotherapy has been established or even suggested by the reported trials in metastatic disease. Indeed, given the poor overall activity of currently available chemotherapeutic agents as measured by response rates, an alchemist’s skills are required to generate a meaningful dose-survival benefit in widespread metastatic disease. Response rates increase when earlier-stage disease is treated, however, and are more likely to correlate with survival benefit. Thus, Shea’s phase I1 Cancer and Leukemia Group B (CALGB-9531) trial will evaluate the clinical and pathologic response to stem cell-supported high-dose carboplatin with Taxol as preoperative therapy for stage I11 NSCLC to indicate whether dose-intensive therapy is worth developing for stages I1 and I11 NSCLC. A number of new agents have demonstrated major activity against NSCLC. These include the taxanes, topoisomerase I inhibitors, vinorelbine, and gemcytabine, but their potential for use in high-dose therapy may be limited. For example, the topoisomerase I inhibitors are more dependent on schedule than dose, and they produce dose-limiting gastrointestinal toxicity at conventional doses. The dose range for new agents should be established as part of their phase I evaluation, with cytokine and progenitor cell support if necessary, to determine the nonhematologic dose-limiting toxicities, as well as their potential for high-dose development. An agent with both preclinical and clinical evidence for a dose-response relationship, a novel mechanism of action, and nonoverlapping nonhematopoietic dose-limiting toxicity encountered only after many-fold dose escalation should be considered a promising candidate to combine with other drugs into a high-dose regimen.
522
ELIAS
MINIMAL RESIDUAL TUMOR (MRT)
One of the most powerful predictors of outcome in most malignancies including lung cancer is the demonstration of metastatic involvement, either regionally to nodes or widespread to various organs. Thus, the development of rare event analysis technology to detect small numbers of malignant cells circulating in blood or residing in marrow or lymph nodes has become an exciting new area fostered by the field of high-dose therapy with stem cell support. Few clinical experiences are reported in MRT detection in patients with NSCLC. Pante15 screened 82 early stage I and I1 resected patients, evaluating bilateral iliac crest and sternal marrow aspirates. He used cytokeratin-18 (MoAb CK2) immunoperoxidase as a single detection reagent and reported a 1.7% false-positive rate from squamous cell contamination and other artifacts. An astonishing 22% of these earlystage patients, however, had abnormal keratin positive cells in their marrow. In a follow-up, 139 patients were screened, of whom 83 (60%) had keratin-positive cells in the marrow.6 Nodes were histologically uninvolved by tumor in 66 patients. Nine (75%) of 12 node-negative marrow keratin-positive patients relapsed, and 19 (35%) of 54 nodenegative marrow keratin-negative patients relapsed, with a median follow-up of 39 months ( P = .023). Keratin positivity in marrow did not change clinical prognosis in the 62 patients with node-positive disease. False positives were detected in 6 of 215 (2.8%)control patients. Brugger et all examined peripheral blood only following a first cycle of chemotherapy (VP-16, ifosfamide, cisplatin) and granulocyte colony-stimulating factor (G-CSF).One of 12 patients with stage IV NSCLC had keratinpositive cells circulating in the blood. Occult nodal metastases have been detected using similar immunohistochemical techniques in patients with histologically uninvolved regional nodes. Use of a polyclonal antikeratin antibody immunoperoxidase technique detected tumor cells in 38 of 60 (63%) patients with early-stage NSCLC.3No statistically significant survival impact was documented; median survival of 6.7 years and 5.4 years were observed for the keratin-negative and keratin-positive node patients, respectively. In another report, the Ber-Ep4 MoAb was used in 72 node-negative patients, of whom eleven (15./0) had isolated ber-ep4 positive cells detected.s Five of 10 (50%) Ber-Ep4-positive patients relapsed as compared with eight of 56 (14%) Ber-Epknegative patients, with a median followup of 26 months ( P = .005). In summary, limited information is available about the incidence of tumor contamination of hematopoietic tissues in lung cancer patients. Still less is known about the clinical impact of screening. With the burgeoning interest and an expanding technology in the detection of
FUTURE DIRECTIONS IN LUNG CANCER RESEARCH AND THERAPEUTICS
523
rare events, additional prospective trials to evaluate the clinical significance of marrow and/or nodal tumor contamination are needed. Heterogeneity of antigen expression in tumor cell populations requires the use of multiple reagents to detect all tumor cells. An elegant study by Pantel et a17 was performed in gastric, colon, and breast cancers, demonstrating the presence of tumor in marrow by keratin immunoperoxidase. Secondary chromophores tagged to antibodies to HER-2/neu, Ki-67, and p120 demonstrated the presence of several populations of tumor cells with presumably differing biologic behavior. Major questions remain. The methodology must improve to have sensitivity greater than one tumor cell in a million, to avoid falsepositive events, and to evaluate heterogeneity of antigen expression independently. Microscopic residual tumor represents therapy-resistant tumor and is likely to be heterogeneous. Detection and ultimately characterization of these cells by evaluating patterns of coexpression of biologic markers may guide pre- or post-transplant therapy. Although the demonstration of tumor cells in marrow or nodes indicates the ability of malignant cells to escape their local environment, this does not automatically imply that all the cells detected have mastered the biologic functions needed to grow into overt metastatic disease. In addition, critical thresholds of numbers of functional metastatic cells may still be needed to escape immune surveillance, systemic therapies such as chemotherapy, or cellular competition between normal tissues and these abnormal clones.
NEW MODALITIES
Monoclonal Antibodies, Ligands, and Receptor Targets (f Linked to Toxic Agents) Monoclonal antibodies have not lived up to their early promise as the ”magic bullets.” Although on occasion this was due to cross-reaction with normal tissues causing toxicity, frequent causes were pharmacodynamic with poor drug delivery due to a bulky messenger, metabolic degradation, or rapid immunologic clearance due to human anti-mouse antibodies (HAMA). Other important causes included antigenic modulation and heterogeneity of antigenic density. Finally, cytotoxicity can be circumvented through a variety of mechanisms, including the fact that these therapies constitute passive transfer rather than represent the development of active immunity. Human chimeric antibodies linked to powerful toxins or radioactive ligands are currently in trials. Trials listed in the Physician Direct Query (PDQ) registry (Table 1)include yttrium90 linked to the MoAb B3, to the MoAb CC-49 (together with interferon-
524
ELIAS
Table 1. SELECTED TRIALS UTILIZING NOVEL MODALITIES LISTED IN PDQ OR RECUMBENT DNA ADVISORY COMITTEE REGISTRIES Target Population
Institute
Methods
Eady Detection:
Stage I resected NSCLC AtypialClS in sputum
SWOGlECOG Univ Colorado
sputumfbronchial washings white light versus autofluorescence bronchoscopy
Antibody:
Tumors (++ EGF-R)
MSKCC phase I
Tumor 30% + 83 Ag NSCLC 10% + TAG-72 Ag
NCI phase I Alabama phase I
MDX447 bispecific Ab ? GCSF link APC with EGF-R expressing tumor cells goY-labeledMoAb B3 g’Y-labeled MoAb CC-49 plus interferon g’Y-labeled-biotic and NR-LU10-streptavidin
Tumor 25% Gene Therapy:
Vaccine:
+
NR-LU-10 Ag
Virginia Mason phase I
Antisense mutated K-ras Wild-type p53
M.D. Anderson
IL-2
Harbin
CEA
NCI
AdenoCA
+ CEA)
NCI phase I
AdenoCA
+ ras mutation)
NCI phase I
Tumor ( + ras mutation)
NCI phase I
Tumor ( + ras or p53 mutation)
Vanderbilt phase I
Tumor ( + ras or p53 mutation)
NCI phase II
retroviral vector, direct injection into endobronchial tumor TIL cells reinfused into malignant pleural effusions vaccinia vector, subcutaneous injection CEA peptide-1 plus adjuvant Detox HLA-A2 restricted mutated ras protein plus adjuvant QS21 K-ras val-12, asp-12, cys-12, or asp-13 mutated ras protein plus adjuvant Detox K-ras Val-12, asp-12, cys-12 peripheral blood buffy coat cells pulsed with tumorspecific synthetic mutated peptide GM-CSF incubated peripheral blood buffy coat cells pulsed with tumor-specific synthetic mutated peptide plus interleukin-2
GM-CSF = granulocyte-macrophage colony-stimulating factor; CEA = carcinoernbryonic antigen; G-CSF lWyte colony-stimulatingfactor; CIS = carcinoma in situ; TIL = tumor intiltrating lymphocytes.
=
granu-
alpha as an adjuvant), or to biotin in patients preloaded with NR-LU10-streptavidin. Clinical results are not currently available. In addition, a bispecific antibody linking CD64 on macrophages to the EGF-receptor (MDX447) has the potential to generate active immunity by enhancing phagocytosis of tumor cells overexpressing EGF-R with subsequent tumor-associated peptide antigen presentation via HLA class I mechanisms. Because hepatocytes express high levels of EGF-R, liver toxicity is a potential. All these trials continue to use single agents to pursue single targets, which are unlikely to succeed at a phase I1 level.
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Of even greater potential are small receptor ligands bound to small toxins (eg, peptides bound to yttrium, maysantines, or the minor groove binder CC1065). These compounds are likely to have favorable pharmacologic delivery to tumor sites, less antigenicity, and greater biologic half-life.
Gene Therapy Gene therapy is still in its infancy from a clinical standpoint. Current vectors favor delivery and transduction of genetic material into proliferating cells preferentially. Strategies currently being pursued include the placement of cytokine genes (eg, IL-2) into tumor-stimulated lymphocytes to try to select for the T cells that proliferated in response to exposure to tumor-associated antigens. These cells are then reinjected into the patient. A second strategy is to overexpress MDR-1 into CD34 + hematopoietic stem cells to lessen hematologic toxicity from chemotherapy agents. A concern, however, is that hematologic stem cells already express high levels of MDR-1, tumor cells residing in the marrow might acquire the gene, and most of the chemotherapeutic agents affected by P-glycoprotein have mucosal, not hematologic, toxicity as a dose-limiting factor. An interesting approach is to place suicide genes, such as genes coding for P450s activating cyclophosphamide, viral thymidylate synthetase (sensitivity to ganciclovir), or those rendering sensitivity to 5-fluorouracil (5-FU) into tumor cells (cytosine deaminase) into vectors with tumor-specific targeting. This may be particularly useful for ex vivo applications such as marrow purging.2 Placement of cytokine genes into killed tumor cells, such as interferon-gamma to upregulate HLA class I, or granulocyte-macrophage colony-stimulating factor (GM-CSF) to attract antigen-presenting cells (APCs) to the tumor cells as a trigger to develop active immunity, are strategies in the clinic for renal cell carcinoma and for melanoma. Replacement of function genes such as wildtype p53 or RB-1, or administration of antisense oncogenes (such as antisense K-ras), are strategies designed to inhibit mutant expression and to replace function and thereby resume cellular controls as least in part. Currently, several gene therapy trials are specifically targeting NSCLC. At M.D. Anderson, either the antisense K-ras mutant or the p53 wild-type genes packaged in a retroviral vector with a p-actin promoter (supplied by Genetic Therapy) are injected directly into endobronchial tumors. Tumor regression has been reported in patients. Mutated K-ras is an attractive target because it is a commonly mutated gene in tumors, it is not wildly overexpressed, and it is centrally involved in signal transduction, and therefore is less likely to mutate further without loss
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ELIAS
of function. Loss of p53 function in tumors is also a common pathway. Resolution of malignant pleural effusions has been reported by injection of genetically modified tumor infiltrating lymphocyte (TIL) cells (expressing IL-2; supplied by Anticancer) into the pleural cavity in a trial conducted in Harbin, China. At the NCI, part of the carcinoembryonic antigen (CEA) gene is being delivered in a vaccinia vector (supplied by Thereon). Other trials include the delivery of the cystic fibrosis gene in adenoviral vectors by inhalational therapy. Tumor Vaccines Vaccines against cancer cells are generally believed to be effective if they stimulate cellular immunity. Vaccines against peptides are HLA class I-restricted, because peptides are usually presented to T8 cytotoxic T cells in the context of the HLA class I molecule without internal processing by APCs. Thus, most clinical trials utilizing peptides are targeting HLA-A2, because this tissue type is present in about 40% of the US population. Vaccines against proteins are not host-HLA restricted, because the proteins are generally internalized by an APC, processed, and then presented on the cell surface as various peptides bound to the host HLA class I molecule. Alternative strategies to present proteins include the use of viral vectors (vaccinia and adenovirus are the two most common) or APCs which are genetically modified to produce specific gene products. These strategies generally present processed peptide antigens in the context of HLA class I1 molecules. To generate a greater array of potential antigens, whole cancer cells may be phagocytosed by or fused into APCs. Processed peptide antigens in the context of HLA class I1 molecules will trigger cellular immunity mediated by T4 cells, provided the appropriate co-stimulatory pathways (such as B7-1, B7-2, CD40 and CD40 ligand, and so on) are activated as well. Otherwise, anergy /tolerance might develop. Thus, various strategies exist to trigger these pathways, including direct vaccination of tumor-associated peptides and proteins with adjuvants, or killed tumor cells given subcutaneously. Genetic modification of the tumor cells to produce cytokines (such as GM-CSF, interleukin-2, tumor necrosis factor, and other interleukins), or to produce large amounts of the co-stimulatory molecules, are designed to trigger the immune system in vivo. Routes of administration include subcutaneous, inhalational, or cellular (eg, loaded onto dendritic cells or other APCs). Current endpoints of vaccine trials are to determine whether cellular or humoral immunity to the specific antigen can be documented prior to and after the vaccination, and whether toxicity and/or tumor response occurs.
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It is likely that these new methods of gene therapy and vaccination will also work best in states of minimal tumor burden. Once safety is demonstrated in the advanced disease setting, these approaches need to be rapidly evaluated in the adjuvant settings and in the high-risk (prevention) settings.
SUMMARY
Although immune and genetic approaches do not yet have substantial clinical exposure in NSCLC, it is hoped that the future will bring less toxic, more specific, and more effective methods to prevent the development of end-stage NSCLC. The principles of multitargeted and multimodality therapy to overcome tumor heterogeneity and resistance, and drug delivery issues have been developed over years of effort using chemotherapy. The lessons learned should not be forgotten, because they will remain applicable to newer biologic and genetic modalities.
References 1. Brugger W, Bross KJ, Glatt M, et al: Mobilization of tumor cells and hematopoietic progenitor cells into peripheral blood of patients with solid tumors. Blood 8 3 : 6 3 M O , 1994
2. Chen L, Pulsipher M, Chen D, et a 1 Adenovirus mediated transgene expression for detection and elimination of contaminating carcinoma cells in hematopoietic stem cell sources. J Cell Invest 982539-2548, 1996 3. Chen Z, Perez S, Holmes EC, et a 1 Frequency and distribution of occult micrometastases in lymph nodes of patients with non-small cell lung carcinoma. J Natl Cancer Inst 85:493-498, 1993 4. Elias A, Cohen BF: Dose intensive therapy in lung cancer. In Armitage JO, Antman KH (eds): High-Dose Cancer Therapy: Pharmacology, Hematopoietins, Stem Cells, ed 2. Baltimore, Williams and Wilkins, 1995, pp 824-846 5. Pantel K, Izbicki JR, Angshvurm M, et al: Immunocytological detection of bone marrow micrometastasis in operable non-small cell lung cancer. Cancer Res 53:1027-1031, 1993 6. Pantel K, Izbicki J, Passlick B, et a 1 Frequency and prognostic significance of isolated tumor cell in bone marrow of patients with non-small cell lung cancer without overt methastases. Lancet 347649453,1996 7. Pantel K, Schlimok G, B r a n S, et a1 Differential expression of proliferation-associated molecules in individual micrometastatic carcinoma cells. J Natl Cancer Inst 85:14191424, 1993
8. Passlick B, Izbicki JR, Kubuschak B, et al: Immunocytochemical assessment of individual tumor cells in lymph nodes of patients with non-small cell lung cancer. J Clin Oncol 121827-1832, 1994
Address reprint requests to Anthony D. Elias, MD Dana-Farber Cancer Institute Department of Medical Oncology/STAMP 44 Binney Street Boston, MA 02115