This phase 2 trial evaluated the tolerability and clinical efficacy of the combination of oxaliplatin and pemetrexed as an induction chemotherapy regimen in locally advanced head and neck squamous cell carcinoma (HNSCC).
This phase 2 trial evaluated the tolerability and clinical efficacy of the combination of oxaliplatin and pemetrexed as an induction chemotherapy regimen in locally advanced head and neck squamous cell carcinoma (HNSCC).
Forty-two patients were enrolled in the study. Patients received pemetrexed 300 mg/m2 intravenously (IV) and oxaliplatin 85 mg/m2 IV every 14 days for a total of 4 cycles. A subset of patients consented to correlative studies including tumor tissue for human papillomavirus (HPV) detection and expression of DNA repair genes that may be predictive of response or resistance to oxaliplatin or pemetrexed.
Response data were available for 40 patients. Eighteen had a partial response, and 1 had a complete response, for a response rate of 47.5%. Patients with HPV+ disease demonstrated superior response rates, progression-free survival, and overall survival. The regimen was well tolerated, with predominantly grade 1 or 2 alanine aminotransferase/aspartate aminotransferase elevation. One patient had grade 5 toxicity with neutropenia and sepsis. The authors did not identify genes predictive of response or toxicity, although HPV+ tumors demonstrated a unique gene expression signature.
Although the response rate of oxaliplatin and pemetrexed proved less than anticipated, the combination remains an active induction regimen in HNSCC. This regimen should be evaluated further in combination with targeted agents, such as cetuximab, especially in the HPV+ patient population. Cancer 2012;. © 2011 American Cancer Society.
Induction chemotherapy before definitive radiation therapy for locally advanced head and neck squamous cell carcinoma (HNSCC) continues to be an area of intense investigation. Theoretically, induction therapy may improve outcomes through enhancement of radiation efficacy, prediction of radiation response, eradication of micrometastatic disease, and early palliation of symptoms. Furthermore, there is considerable discussion as to whether induction therapy may be able to spare the use of aggressive concurrent chemotherapy, thus obviating the severe late effects that diminish long-term quality of life. Growing data support the use of induction therapy in a variety of settings.1-3 However, definitive data from randomized controlled trials are lacking. Furthermore, numerous important clinical questions remain to be answered including the identification of the optimal induction regimen for various cohorts of patients. Encouraging results using the 3-drug combination of docetaxel, cisplatin, and 5-fluorouracil have been reported.2 Thus, this regimen has become a platform for further investigation of the role and impact of induction therapy in the management of head and neck cancer. However, this regimen is associated with significant toxicity. Patients who are elderly or frail or who have multiple comorbidities are often poor candidates for 5-fluorouracil. Furthermore, for patients with tumors associated with better clinical outcome such as nonsmokers with human papillomavirus (HPV)-associated oropharyngeal carcinomas, aggressive induction therapy may not be necessary. Identification of efficacious, low-toxicity, cost-effective, and easy to administer regimens is therefore warranted.
We conducted a phase 2 trial of induction chemotherapy using oxaliplatin and pemetrexed in patients with previously untreated locally advanced HNSCC. The primary objective was clinical response rate by Response Evaluation Criteria in Solid Tumors (RECIST). Secondary endpoints included patient-reported outcomes (quality of life and symptom burden) and biological correlatives (HPV status, tumor global gene expression to detect differences in expression patterns between responders and nonresponders).
Between February 2007 and April of 2009, 42 patients were recruited through the Vanderbilt Ingram Cancer Center Affiliate Network. Patients were required to have previously untreated, histologically confirmed stage III/IV (M0) HNSCC. Two cohorts were included: 1) planned definitive radiation based treatment, or 2) planned surgical resection ± postoperative radiation. Additional eligibility criteria included: age ≥18 years; measurable disease defined by RECIST (version 1.0); Eastern Cooperative Oncology Group (ECOG) performance status of 0 to 2; adequate hematologic, hepatic, and renal function; and willingness to take folic acid, vitamin B12, and dexamethasone to minimize toxicity. Exclusion criteria included: grade 2 or higher peripheral neuropathy; pregnant or lactating; presence of clinically detectable third-space fluid collections; and history of prior malignancy within the past 5 years except for curatively treated cervical intraepithelial neoplasia, basal cell carcinoma of the skin, or localized prostate cancer. The study was conducted in accordance with the Helsinki Declaration and Good Clinical Practice guidelines and was approved by the Vanderbilt Institutional Review Board (IRB) and each of the IRBs of participating institutions of the Vanderbilt Ingram Cancer Center Associated Network. All patients signed informed consent before initiation of therapy.
Patients were treated with pemetrexed 300 mg/m2 intravenously (IV) followed by oxaliplatin 85 mg/m2 IV every 14 days for a total of 4 cycles (8 weeks). After 2 cycles, patients underwent a clinical evaluation of measureable disease to determine response. Computed tomographic imaging was the primary modality used, and disease was measured by RECIST. However, direct laryngoscopy was performed for response assessment if no clinically measurable disease was seen on physical exam or on imaging. Partial response by direct laryngoscopy was defined as >50% reduction in tumor. Patients who had progression of tumor were taken off of the study. Patients with stable or responding disease continued until completion of the planned therapy. Patients were withdrawn from the study for unacceptable toxicity, treatment delay >2 weeks, or withdrawal of consent. After 4 cycles, patients underwent repeat staging with endoscopy and imaging. To limit treatment-related mucositis and neutropenia, patients were pretreated with an injection of vitamin B12 and daily folic acid starting 7 days before chemotherapy and continuing until 3 weeks after the last pemetrexed dose. In addition, patients were pretreated with dexamethasone to prevent pemetrexed-associated rash.
Toxicities were grading using National Cancer Institute Common Toxicity Criteria version 3.0.
If the absolute neutrophil count (ANC) was <1000/μL or platelet count was <100,000/μL on day 1 of a cycle, treatment was held until blood counts returned to above this level. Treatment was then resumed with a permanent 25% dose reduction. If treatment was held for >2 weeks because of count suppression, the patient was withdrawn from the study. If the ANC was between 1000 and 1500/μL or the platelet count was between 75,000 and 100,000/μL on day 1 of a cycle, the patient received a 25% dose reduction for that dose only.
Treatment was held for bilirubin greater than upper limits of normal (ULN) until the level was within the normal range. Treatment was then resumed at a 25% dose reduction. Treatment was held for an alkaline phosphatase >2.5× ULN. Aspartate aminotransferase (AST) or alanine aminotransferase (ALT) >1× to ≤2.5× ULN required a 25% dose reduction. Patients with an alkaline phosphatase >1× but <2.5× ULN plus AST or ALT >2.5× to 5× ULN received a 25% dose reduction for subsequent cycles.
For all other clinically significant grade 3 or 4 toxicities, doses were withheld until toxicity resolved to grade ≤1 and then resumed at a 25% dose reduction. If treatment was held for >2 weeks, the patient was withdrawn from the study. A second dose reduction was allowed for patients who experienced significant grade 3 or 4 toxicities on the reduced dose. Patients who did not tolerate the dose −2 level were taken off the study.
Blood for a subset of 24 patients enrolled in the correlative study was obtained at baseline and day 15, cycle 4. Tumor tissue was obtained from patients' original diagnostic biopsy and from the surgical specimen for those patients undergoing definitive resection. Tumor biopsy for research purposes was optional and performed at baseline and on day 15, cycle 4.
The status of human papillomavirus infection was determined by polymerase chain reaction and by p16 immunohistochemical (IHC) staining as previously described.4, 5 To determine the protein expression levels of TYMS (Invitrogen, Carlsbad, Calif; Cat#35-5800, dilution 1:30), DHFR (Abcam, Cambridge, Mass; ab49881, dilution 1:30), ERCC1 (Santa Cruz Biotechnology, Santa Cruz, Calif; FL-297, dilution 1:50), and XPF (Abcam; ab17798, dilution 1:50), IHC staining was performed for each protein. Briefly, the tumor sections were deparaffinized, hydrated, then antigen-retrieved using ethylenediaminetetraacetic acid followed by incubation with primary antibodies overnight. After thorough rinsing, the sections were incubated with secondary antibodies (Mouse Envision; DAKO, Carpentaria, Calif) followed by DAB chromogen and Mayer hematoxylin. The images were analyzed using the Ariol SL-50 automated slide scanner (Applied Imaging, San Jose, Calif) to quantify the amount of nuclear or non-nuclear staining for each slide by processing with the NuclearHighRes or CytoplasmHighRes scripts without knowledge of clinical data. These scripts provided the number of positive and negative stained cells per slide, which allowed calculation of the proportion of positive cells in each tissue sample. Cells were also classified as positive or negative based on predetermined thresholds that evaluate color and intensity of staining, as well as cell size, axis length, roundness, and compactness. Thus, blue hematoxylin staining of nuclei could be distinguished from brown DAB reaction product. Measured parameters were the mean intensity and percentage staining area. The staining index was determined by the mean intensity and percentage staining area divided by 100.
For gene expression analyses, total RNA was isolated from either frozen or formalin-fixed paraffin-embedded (FFPE) tissue, amplified, labeled, and hybridized onto Affymetrix 2.0 PLUS GeneChips as previously described.4 Differentially expressed genes with a false discovery rate <1% were selected from comparison of tumors from patients with partial response versus stable disease (SD) using significance analysis of microarrays and grouped using hierarchical clustering.6, 7 Differentially expressed pathways were determined by Ingenuity Pathway Analysis as previously described.8
The primary objective of the study was to evaluate the clinical response rate of the combination of pemetrexed and oxaliplatin in patients with locally advanced HNSCC. The 2 cohorts of pemetrexed and oxaliplatin patients represented similar populations; thus, the response data were combined. The intent to treat sample was defined as all subjects who received 1 cycle of oxaliplatin or pemetrexed. The per protocol sample was defined as all subjects who received at least 1 cycle of treatment, fulfilled all inclusion and exclusion criteria, and were evaluable for the primary efficacy criterion. The study sample evaluable for safety consisted of all the subjects who received any study medication for whom safety data were available. All safety analyses were carried out on that sample.
Response outcome was determined by RECIST. Clinical benefit was defined as that portion of patients experiencing complete response or partial response (PR). A Simon optimal 2-stage design was used to test the null hypothesis that the true response rate was 50% against the alternative that it was at least 70%.9 Descriptive statistics were used to summarize the patient characteristics in the primary and correlative studies. Mann-Whitney (continuous) and likelihood chi-square (nominal) tests were used to compare the demographic and clinical characteristics of the HPV status samples in the correlative study. Within the correlative study, logistic regressions were used to test the associations of biomarkers with response. Survival analysis was used to evaluate differences in progression-free and overall survival curves for the HPV-positive and HPV-negative groups. The Tarone-Ware statistic was used to evaluate differences between the curves, because the test is designed to have good power across a wide range of survival functions. Statistical significance was evaluated using a maximum alpha of .05.
The average age for the treated sample was 56 years (median, 58.5 years). Eighty-three percent of patients were men, and 17% were women. Other characteristics at baseline included an ECOG performance status of 0 in 29% of patients and 1 in 67% of patients. Other prevalent characteristics at baseline included: 71% had a oropharyngeal primary site, 81% had N2 or N3 disease, and 50% had T3 or T4 primary tumors (Table 1).
|Patient Characteristic||Overall, N=42||Correlative, n=24|
|Men||35 (83.3%)||21 (87.5%)|
|0||12 (28.6%)||5 (20.8%)|
|1||28 (66.7%)||18 (75.0%)|
|2||2 (4.7%)||1 (4.2%)|
|Oropharynx||27 (71.1%)||16 (69.6%)|
|Nonoropharynx||11 (28.9%)||7 (30.4%)|
|T1/T2||20 (50.0%)||13 (56.5%)|
|T3/T4||20 (50.0%)||10 (43.5%)|
|N0/N1||7 (16.7%)||6 (25.0%)|
|N2/N3||34 (81.0%)||18 (75.0%)|
|Nx||1 (2.3%)||0 (0.0%)|
|Never smoked||8 (19.0%)||4 (16.7%)|
|Former smoker||20 (47.7%)||12 (50.0%)|
|Current smoker||14 (33.3%)||8 (33.3%)|
The average number of cycles delivered was 3.7 (range, 1-4). Overall, 85% of patients received all 4 cycles of therapy. Ten patients required dose reductions, of which 9 were because of hepatotoxicity and 1 because of febrile neutropenia. Nine patients required a least 1 dose delay. In 8 patients, the dose delays were secondary to hepatotoxicity. Six patients went on to receive all 4 cycles of therapy. Dose delays occurred during cycles 3 and 4 for 8 of the 9 patients.
All patients were evaluable for toxicity. Overall, the regimen was very well tolerated. Grade 3 and 4 adverse events are listed in Table 2. One patient experienced febrile neutropenia during cycle 3. The patient underwent subsequent treatment with a dose reduction without further incident. Nineteen patients had grade 1 or 2 elevations in ALT and/or AST. All but 2 of these increases occurred in cycles 3 or 4 or during end of treatment laboratory processes. All liver enzyme elevations resolved with dose delays or dose reductions per protocol or with completion of protocol treatment. The enzyme elevations were not associated with symptoms or signs of liver failure, and all patients resolved to baseline liver enzyme levels without sequelae.
|Neutropenia||7 (3%)||0 (0%)||0 (0%)|
|Diarrhea||2 (1%)||2 (1%)||0 (0%)|
|Anorexia||2 (1%)||0 (0%)||0 (0%)|
|Dehydration||2 (1%)||0 (0%)||0 (0%)|
|Hypokalemia||7 (3%)||0 (0%)||0 (0%)|
|Infection/acidosis/neutropenia||0 (0%)||0 (0%)||2 (1%)|
Response data were available for 40 of 42 patients. All responses were determined by imaging using RECIST. Of 40 evaluable patients, 18 had a PR, and 1 had a complete response, for an overall response rate of 47.5%. Nineteen patients had SD (47.5%), and 2 had progressive disease (5%). At a mean follow-up of 16 months (range, 2-36 months), 6 patients had relapsed or demonstrated progressive disease. Twenty-five patients (60%) were alive and free of disease at a minimum of 12 months follow-up, whereas 13 patients (25%) were alive with disease.
To identify potential biomarkers of response to oxaliplatin and pemetrexed, gene expression differences in tumors from responders and nonresponders were assessed. Ten tumors and 3 adjacent normal epithelia from 6 patients (4 PR and 2 SD) were available for gene expression analyses, and a supervised analysis was performed using PR versus SD as supervising parameters. Up-regulation of xenobiotic metabolism-related genes such as GSTA1, GSTA4, and ME1 was observed in tumors with SD (Fig. 1A). These are the key genes that metabolize xenobiotics and are induced during oxidative stress by Nrf2.10 Up-regulation of this class of genes has been associated with radiation, platinum, doxorubicin, and VP16 resistance.10
Additional top ranked differentially expressed genes were DHFR, TYMS, CDKN2C (p18), and CKDN2A (p16), which were key genes in the previously determined HPV signature.4 The tumors with PR contained the HPV signature when the gene expression profile was specifically queried for the HPV-associated genes (Fig. 1B). This prompted us to further examine the HPV status in additional tumors. Twenty-four FFPE tumor samples of the 42 enrolled patients were available for p16 IHC staining as a surrogate marker of HPV infection. Sixteen of 24 tumors (63%) were p16 positive, and of the cases with disease site information (n = 23), all of the oropharyngeal primary tumors (100%) but none of the 7 nonoropharyngeal tumors (P < .001) overexpressed p16. Comparisons of p16 status with other clinical information are detailed in Table 3. The association of p16 status with response to induction therapy was at the threshold of statistical significance set in this study (partial or complete response: p16 positive, 9 of 16, 56.3%; p16 negative, 1 or 7, 14.3%; likelihood chi-square(df = 1) = 3.82, P = .051). Median recurrence-free survival was 19.3 months (95% confidence interval [CI], 10.1-28.5) for p16-negative patients and 33.7 months (95% CI, 27.5-37.9) for the p16-positive group (Tarone-Ware = 4.59, P = .032). Median overall survival was 20.3 months (95% CI, 12.3-28.2) for p16-negative patients and 34.1 months (95% CI, 29.8-38.5) for the p16-positive group (Tarone-Ware = 4.24, P = .039) (Fig. 2).
|Patient Characteristic||Negative, n=7||Positive, n=17||P|
|Men||5 (71.4%)||16 (94.1%)||.127|
|Oropharynx||0 (0.0%)||16 (100.0%)|
|Nonoropharynx||7 (100.0%)||0 (0.0%)|
|Never smoked||0 (0.0%)||4 (23.5%)|
|Former smoker||4 (57.1%)||8 (47.1%)|
|Current smoker||3 (42.9%)||5 (29.4%)|
|CR/PR||1 (14.3%)||9 (56.3%)|
|SD/PD||6 (85.7%)||7 (43.8%)|
Finally, we examined genes that were known to be important in DNA repair and found no statistically significant associations of those repair genes with the treatment response to pemetrexed and oxaliplatin (Fig. 1C). Consistent with the gene expression data, ERCC1 and XPF protein expression levels determined by IHC in 21 tumors were not associated with response or survival (data not shown).
The combination of oxaliplatin and pemetrexed is an active regimen in locally advanced HNSCC; however, the response rate using RECIST was lower than that reported by other combination regimens used in the induction setting. It should be noted that concerns were expressed by clinicians that RECIST was not adequately reflective of the clinical response. Because unidimensional shrinkage of a tumor may not be captured as a response measure, RECIST may underestimate the true response of a treatment regimen. This is especially true in an anatomically challenging area such as the head and neck, where measuring tumors across scan slices is a recognized challenge.
The combination of pemetrexed and oxaliplatin is easy to administer and generally well tolerated. Of note, the incidence of hepatotoxicity was high, with 45% of patients developing grade 1 or 2 toxicity and 21% requiring at least 1 dose reduction. Although the hepatotoxicity resolved in all patients, our experience brings into question the feasibility of administering this regimen over a more protracted treatment course in other clinical settings, such as patients with metastatic disease. That being said, the hepatotoxicity was asymptomatic. Patients can complete this induction regimen without the significant acute toxicities and mortality rates associated with cisplatin-based combinations.
We could not identify robust biomarkers that predicted response to oxaliplatin and pemetrexed. The failure to identify predictors of response may be because of the small sample size or measurement issues related to the use of RECIST. As expected, overall survival was strongly associated with p16 status reflecting HPV infection. In our study, 94% of oropharyngeal tumors were p16 positive, which is higher than other published series. In this study, the survival advantage noted in the patients with HPV-associated disease may be in part related to the mechanism of action of the therapeutic agents. HPV deregulates cell cycle and up-regulates dihydrofolate reductase as the result of loss of p53 and Rb mediated by viral oncogenes E6 and E7.11 Thus, HPV-associated tumors may be more responsive to chemotherapy drugs that target folate biosynthesis pathway or cell cycle. Targeting these specific deregulations with less toxic agents could be a novel approach to decrease treatment-related toxicities in this favorable group of patients.
Furthermore, our initial hypothesis was that the up-regulation of DNA repair genes would be associated with oxaliplatin resistance, but none of the genes reached statistical difference, and differential expression of DNA repair-related genes may not have a significant role in the predicted response to oxaliplatin. In addition, up-regulation of genes in xenobiotic metabolism may be a mechanism of oxaliplatin and pemetrexed resistance, as seen with cisplatin. Although these finding are promising, our sample size was very limited, and additional studies are warranted.
In summary, the combination of oxaliplatin and pemetrexed is active and well tolerated in the induction setting. The question remains whether we can improve on the clinical activity by the addition of a targeted agent, such as cetuximab. Moreover, we need to better understand whether induction chemotherapy adds clinical benefit to definitive concurrent chemotherapy. If so, it remains prudent to define the population in which the increased toxicities of more aggressive therapy are warranted.
C.H.C. received research funds from Lilly Oncology to conduct the laboratory correlative studies. The clinical trial was funded in part by Lilly Oncology and Sanofi-Aventis.
CONFLICT OF INTEREST DISCLOSURES
The authors made no disclosures.