Genomic profile predicts the efficacy of neoadjuvant chemotherapy for cervical cancer patients

Despite the prevalence of screening and advancement of therapy, the mortality rate among women of reproductive age due to cervical cancer has increased over the last two decades in Japan [1]. As locally advanced cervical cancer (LACC) of FIGO stage Ib2 or IIb is frequently accompanied by lymph node metastasis, patients bearing LACC, treated only with excision or radiation, exhibit a high incidence of recurrence, resulting in a poor survival outcome [2]. Thus, radical hysterectomy (RH) coupled with platinum-based chemotherapy or radiation and concurrent chemo-radiation therapy are now employed as intensive treatments for LACC [3, 4]. Neoadjuvant chemotherapy (NAC) prior to RH is usually conducted to reduce the tumor volume and improve the safety and integrity of surgery, but the prognosis of NAC-refractory patients worsens with the delay of the main treatment [4]. Therefore, to optimize the efficacy of NAC-RH, a method is needed to exclude chemo-refractory cases before starting initial therapy.

Recently, genomic characterization by analyzing gene expression microarrays or genotyping onco-related/suppressive genes has been developed to evaluate characteristic profiles of chemo-refractory tumors and host patients [5, 6]. Single-sample Gene Set Enrichment Analysis (ssGSEA) is a bioinformatics method to characterize the biological features of individual samples as signature scores based on gene expression microarrays [7]. It was reported that the TP53 pathway ssGSEA score can be used to predict the response of lung cancer to radiation [8], but there has been no report suggesting that ssGSEA is useful to predict the chemo-susceptibility of LACC. On the other hand, it is well-known that irinotecan (CPT-11) causes severe side effects more frequently in patients with UGT1A1 polymorphisms than the wild-type [9], and, thus, UGT1A1 genotyping is a prerequisite before initiating chemotherapy using CPT-11 in a clinical setting. However, it remains unclear whether CPT-11 treatment is more effective in LACC patients with UGT1A1 polymorphisms. In this study, we assessed whether the chemo-susceptibility of LACC could be evaluated based on tumor expression microarray analysis and host UGT1A1 genotyping, in order to optimize the efficacy of NAC-RH.

The superiority of concurrent chemoradiation therapy (CCRT) over surgical excision for LACC due to its more favorable local control and reduced adverse event rates was described in a recent report [17], and CCRT is now designated as a standard therapy for LACC. However, CCRT has several difficulties in terms of preserving the ovarian function, reducing late radiation-based adverse effects, and managing pelvic lesions which remain or recur after CCRT [18]. Thus, a therapeutically intensive alternative has been investigated, and NAC-RH has been designated as a reasonable alternative for LACC in terms of not only avoiding primary radiation therapy, but also the therapeutic intensity [19]. In this study, the response rate to NAC using CPT-11 and NDP was 76.3 %, consistent with a previous report [10] and not inferior to the combination of cisplatin and CPT-11 [20] or paclitaxel [21], but nearly one quarter of the cases were non-responders, and those with tumor shrinkage rates following NAC below 50 % showed shorter PFS. This was in agreement with a previous report that non-responders had poor prognostic outcomes [22]. Thus, the shrinkage rate was an independent prognostic factor, and it should be predicted prior to NAC to reduce the risk of the main treatment being delayed without NAC benefits.

Pharmacogenomic analysis using gene expression microarrays with chemo-sensitivity profiles has successfully identified several “signatures” which are characteristic gene profiles in cancers resistant to specific anti-cancer drugs [5], and been employed to discover novel drugs effective against chemo-resistant cancers [23]. In this study, GSEA showed that a mismatch repair pathway, a well-known signal of platinum-resistance, was enriched in non-responders. ssGSEA, which quantitatively scores the activity of each “signature” pathway in every sample based on gene expression microarray [7], revealed that the GMP score significantly increased in non-responders, and that there was a positive correlation between GMP scores and IC50 values of CPT-11 among 57 SCC cell lines. These results suggest that GMP scores may represent the susceptibility of LACC to CPT-11.

CPT-11 shows a strong anti-tumor activity not only via the inhibition of DNA synthesis but also the inactivation of cystine transporters, leading to the accumulation of reactive oxygen species (ROS) in cancer cells [24]. As the presence of ROS is an apoptosis-inducing stress, this mechanism associating with glutathione synthesis may support the idea that GMP activity represents CPT-11 susceptibility. There is, however, a limitation of this study in that GMP was identified through the analysis of post-NAC tumors, and to apply this pathway as a predictive biomarker of NAC susceptibility, further validation based on pre-NAC tumors is necessary. Nonetheless, unsupervised hierarchical cluster analysis of a web-published SCC gene expression microarray, GSE 6213, which contains paired samples of identical patients before/after chemotherapy, revealed the following: The gene expression pattern was not markedly affected by chemotherapy, since cluster dendrograms did not divide samples of each individual patient before/after chemotherapy (Additional file 3: Figure S3). As the GMP score showed a strong positive correlation with the total glutathione concentration in SCC cells and SCC cells with low GMP scores showed high sensitivity to CPT-11, the GMP score might be expected to act as an NAC-susceptibility biomarker representing intracellular glutathione synthesis.

In contrast, patients with heterozygotic polymorphism of UGT1A1 *1/*6 or *1/*28 (UGT1A1-poly patients) showed a shrinkage rate > 50 % without a high incidence of G3/4 toxicities. The finding that SKGIIIa cells with a high GMP score showed UGT1A1 polymorphisms and a high susceptibility to CPT-11 suggests that host UGT1A1 polymorphism may constitute a complementary marker of CPT-11 susceptibility in an organ-specific manner. In colorectal cancers, FOLFIRI containing CPT-11 was more effective in patients with UGT1A1*28/*28 polymorphism than UGT1A1-wild patients accompanied with a decrease of SN38 glucuronidation in the liver [25]. Patients with homozygous polymorphism had a much higher AUC ratio of SN-38 than wild-type patients, whereas patients with heterozygous polymorphism had a slightly higher AUC ratio than wild-type patients [25]. This may be the reason why NAC containing CPT-11 showed a higher efficacy without marked toxicity in patients with heterozygous polymorphisms. The UGT1A1 gene was picked up as one of the chemo-refractory signature genes in a genome-wide analysis [26], and UGT activation was observed in a colon cancer cell line which acquired resistance to CPT-11 [27]. We considered that the UGT1A1 genotypes of cancerous tissues matched those of the host, although there has been no report to date on the correlation of UGT1A1 polymorphisms between the tumor and host. Although further prospective studies are needed, NAC using CPT-11 in combination with UGT1A1 genotyping might be performed more effectively and safely than other regimens without such markers.

Based on comprehensive analysis, the present study suggests that the GMP score and heterozygotic UGT1A1 polymorphisms may be complementary predictive markers of CPT-11 efficacy. Other than CPT-11, a comprehensive genomic analysis of both host and cancerous tissue might facilitate the establishment of a novel tailored therapy for LACC patients that exhibits high-level efficacy and feasibility.