PDGFRA, KIT, and KDR Gene Amplification in Glioblastoma: Heterogeneity and Clinical Significance

Neurosurgery Oncology Medicine
glioblastoma PDGFRA KIT KDR genetic heterogeneity FISH

Brain Glioblastoma PDGFRA, KIT and KDR Gene Amplification: Heterogeneity and Its Clinical Significance

Academic Background

Glioblastoma (GBM) is the most common malignant tumor of the central nervous system, with its heterogeneity posing significant challenges in treatment. GBM exhibits cellular genetic and phenotypic diversity not only between different patients (heterogeneity) but also within the same tumor (homogeneity and interstitiality). This heterogeneity is a major obstacle in the development of solid tumor treatments, making GBM highly resistant to treatment and invasive. Moreover, intratumoral heterogeneity manifests as the coexistence of multiple subclones, which can exist in intermingled or spatially separated forms. Tumor heterogeneity is reflected in different genetic, molecular, and morphological expression patterns at the genetic, molecular, and cellular levels, all of which play key roles in tumor progression and treatment resistance.

Research Source

This article was written by Bianca Soares Carlotto et al., with authors from the Universidade Federal de Ciências da Saúde de Porto Alegre (UFCSPA) and its affiliated institutions in Brazil, and the Medical Oncology Division of the University of Colorado Anschutz Medical Campus in the USA. The study was published in “Neuromolecular Medicine” in 2023 (Volume 25:441-450).

Research Process

This study mainly investigated the heterogeneity and clinical significance of PDGFRA, KIT, and KDR gene amplifications in glioblastoma. Using fluorescence in situ hybridization (FISH) technology, the study detected homogeneous or heterogeneous co-amplification patterns of PDGFRA, KIT, and KDR genes within the 4q12 amplicon in 22 GBM samples, as well as the diffuse or localized distribution of cells carrying the amplifications.

  1. Sample Selection and Preprocessing

The study retrospectively selected 22 cases with 4q12 amplicon from 113 previously classified GBM NOS cases and detected gene amplification using FISH technology. Gene amplification was considered present when the gene-to-control copy number ratio was ≥2, or when ≥10% of tumor cells had ≥15 gene copies. All tumor samples were from formalin-fixed paraffin-embedded (FFPE) sections of surgically resected tissue.

  1. Fluorescence In Situ Hybridization

Multiple gene probe combinations were used for FISH detection, including 4-color probes (PDGFRA/KIT/KDR/CEP4) and 2-color probes (RB1/LSI 13q34). The labeled probes were red, cyan, gold, and green, respectively. The entire tumor area was then scanned with 40× and 100× objectives to determine the uniform or localized distribution of amplification patterns, and the clustering of gene-binding probes was recorded on all slides.

  1. Data and Statistical Analysis

Quantitative variables were described by means or medians, and categorical variables by absolute and relative frequencies. Chi-square tests and Fisher’s exact tests were used. Survival was assessed using the Kaplan-Meier method, and curves were compared using the Log-rank test. Multivariate analysis was performed using Cox proportional hazards model. Statistical significance was set at 5% (p < 0.05), and analyses were performed using SPSS 21.0.

Research Results

  1. Sample Description

Among the 22 patients, 59.1% were female, with ages ranging from 23 to 71 years, averaging 53.9 years. Focal functional deficits and increased intracranial pressure were the most common initial symptoms. Hospital stays ranged from 9 to 167 days, with a median of 18.5 days. The median overall survival was 7.2 months.

  1. Gene Amplification Analysis

All 22 GBM cases showed high-level amplification for at least one gene. 16 cases (72.7%) showed uniform distribution of gene amplification, while 6 (27.3%) showed localized specific amplification. 10 cases showed homogeneous co-amplification of all three genes PDGFRA, KIT, and KDR, 5 showed co-amplification of PDGFRA and KDR, and 1 showed amplification of only PDGFRA.

Among the 6 cases with heterogeneous amplification patterns, 3 showed coexistence of homogeneous co-amplification of PDGFRA, KIT, and KDR and co-amplification of two genes (PDGFRA and KDR), 1 showed coexistence of co-amplification of PDGFRA, KIT, and KDR and amplification of only PDGFRA, 1 showed coexistence of co-amplification of PDGFRA, KIT, and KDR and amplification of only KIT. Overall, 16 cases showed co-amplification of all three genes (72.8%), 5 showed co-amplification of two genes (22.7%), and 1 showed amplification of a single gene (4.6%).

  1. Association with Survival

After adjusting for confounding factors, homogeneous amplification of all three genes (PDGFRA, KIT, and KDR), age ≥60 years, and complete tumor resection were significantly associated with poorer survival. Patients with homogeneous amplification of these three genes had a 10.5-fold higher risk of death compared to patients with other amplification patterns. Additionally, patients aged 60 and above had a 13.9-fold higher risk of death compared to those under 60, and patients with complete resection had a 9.3-fold higher risk of death compared to those with partial resection.

Review and Conclusion

This study revealed a series of complex amplification patterns involving the RTK gene 4q12 amplicon in adult GBM, demonstrating a new level of tumor heterogeneity at the molecular level. Fluorescence in situ hybridization effectively resolved the genetic heterogeneity in these pathological samples and indicated that homogeneous co-amplification of PDGFRA, KIT, and KDR genes was significantly associated with poorer prognosis. Concurrently, the increased proportion of elderly patients in this specific GBM subgroup further supports the related survival data. Based on this research, more diversified and precise treatment strategies should be considered to address the complex heterogeneity of GBM.

Conclusion

This study, through a comprehensive assessment of PDGFRA, KIT, and KDR gene amplification patterns in GBM samples from southern Brazil, revealed high heterogeneity of the 4q12 amplicon and confirmed the effectiveness of FISH technology in identifying and interpreting heterogeneity in pathological samples in detail. Although the analysis was limited to a single surgical resection, the high levels of amplification found suggest the possibility of stable coexistence between these subclones. To improve treatment outcomes for GBM patients, such molecular heterogeneity must be systematically incorporated into the design of treatment plans.