Proliferative potential and resistance to immune checkpoint blockade in lung cancer patients.

BACKGROUND: Resistance to immune checkpoint inhibitors (ICIs) has been linked to local immunosuppression independent of major ICI targets (e.g., PD-1). Clinical experience with response prediction based on PD-L1 expression suggests that other factors influence sensitivity to ICIs in non-small cell lung cancer (NSCLC) patients. METHODS: Tumor specimens from 120 NSCLC patients from 10 institutions were evaluated for PD-L1 expression by immunohistochemistry, and global proliferative profile by targeted RNA-seq. RESULTS: Cell proliferation, derived from the mean expression of 10 proliferation-associated genes (namely BUB1, CCNB2, CDK1, CDKN3, FOXM1, KIAA0101, MAD2L1, MELK, MKI67, and TOP2A), was identified as a marker of response to ICIs in NSCLC. Poorly, moderately, and highly proliferative tumors were somewhat equally represented in NSCLC, with tumors with the highest PD-L1 expression being more frequently moderately proliferative as compared to lesser levels of PD-L1 expression. Proliferation status had an impact on survival in patients with both PD-L1 positive and negative tumors. There was a significant survival advantage for moderately proliferative tumors compared to their combined highly/poorly counterparts (p = 0.021). Moderately proliferative PD-L1 positive tumors had a median survival of 14.6 months that was almost twice that of PD-L1 negative highly/poorly proliferative at 7.6 months (p = 0.028). Median survival in moderately proliferative PD-L1 negative tumors at 12.6 months was comparable to that of highly/poorly proliferative PD-L1 positive tumors at 11.5 months, but in both instances less than that of moderately proliferative PD-L1 positive tumors. Similar to survival, proliferation status has impact on disease control (DC) in patients with both PD-L1 positive and negative tumors. Patients with moderately versus those with poorly or highly proliferative tumors have a superior DC rate when combined with any classification schema used to score PD-L1 as a positive result (i.e., TPS ≥ 50% or ≥ 1%), and best displayed by a DC rate for moderately proliferative tumors of no less than 40% for any classification of PD-L1 as a negative result. While there is an over representation of moderately proliferative tumors as PD-L1 expression increases this does not account for the improved survival or higher disease control rates seen in PD-L1 negative tumors. CONCLUSIONS: Cell proliferation is potentially a new biomarker of response to ICIs in NSCLC and is applicable to PD-L1 negative tumors.

Next generation sequencing of PD-L1 for predicting response to immune checkpoint inhibitors.

BACKGROUND: PD-L1 immunohistochemistry (IHC) has been traditionally used for predicting clinical responses to immune checkpoint inhibitors (ICIs). However, there are at least 4 different assays and antibodies used for PD-L1 IHC, each developed with a different ICI. We set to test if next generation RNA sequencing (RNA-seq) is a robust method to determine PD-L1 mRNA expression levels and furthermore, efficacy of predicting response to ICIs as compared to routinely used, standardized IHC procedures. METHODS: A total of 209 cancer patients treated on-label by FDA-approved ICIs, with evaluable responses were assessed for PD-L1 expression by RNA-seq and IHC, based on tumor proportion score (TPS) and immune cell staining (ICS). A subset of serially diluted cases was evaluated for RNA-seq assay performance across a broad range of PD-L1 expression levels. RESULTS: Assessment of PD-L1 mRNA levels by RNA-seq demonstrated robust linearity across high and low expression ranges. PD-L1 mRNA levels assessed by RNA-seq and IHC (TPS and ICS) were highly correlated (p < 2e-16). Sub-analyses showed sustained correlation when IHC results were classified as high or low by clinically accepted cut-offs (p < 0.01), and results did not differ by tumor type or anti-PD-L1 antibody used. Overall, a combined positive PD-L1 result (≥1% IHC TPS and high PD-L1 expression by RNA-Seq) was associated with a 2-to-5-fold higher overall response rate (ORR) compared to a double negative result. Standard assessments of sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV) showed that a PD-L1 positive assessment for melanoma samples by RNA-seq had the lowest sensitivity (25%) but the highest PPV (72.7%). Among the three tumor types analyzed in this study, the only non-overlapping confidence interval for predicting response was for "RNA-seq low vs high" in melanoma. CONCLUSIONS: Measurement of PD-L1 mRNA expression by RNA-seq is comparable to PD-L1 expression by IHC both analytically and clinically in predicting ICI response. RNA-seq has the added advantages of being amenable to standardization and avoidance of interpretation bias. PD-L1 by RNA-seq needs to be validated in future prospective ICI clinical studies across multiple histologies.

Predicting response to checkpoint inhibitors in melanoma beyond PD-L1 and mutational burden.

BACKGROUND: Immune checkpoint inhibitors (ICIs) have changed the clinical management of melanoma. However, not all patients respond, and current biomarkers including PD-L1 and mutational burden show incomplete predictive performance. The clinical validity and utility of complex biomarkers have not been studied in melanoma. METHODS: Cutaneous metastatic melanoma patients at eight institutions were evaluated for PD-L1 expression, CD8+ T-cell infiltration pattern, mutational burden, and 394 immune transcript expression. PD-L1 IHC and mutational burden were assessed for association with overall survival (OS) in 94 patients treated prior to ICI approval by the FDA (historical-controls), and in 137 patients treated with ICIs. Unsupervised analysis revealed distinct immune-clusters with separate response rates. This comprehensive immune profiling data were then integrated to generate a continuous Response Score (RS) based upon response criteria (RECIST v.1.1). RS was developed using a single institution training cohort (n = 48) and subsequently tested in a separate eight institution validation cohort (n = 29) to mimic a real-world clinical scenario. RESULTS: PD-L1 positivity ≥1% correlated with response and OS in ICI-treated patients, but demonstrated limited predictive performance. High mutational burden was associated with response in ICI-treated patients, but not with OS. Comprehensive immune profiling using RS demonstrated higher sensitivity (72.2%) compared to PD-L1 IHC (34.25%) and tumor mutational burden (32.5%), but with similar specificity. CONCLUSIONS: In this study, the response score derived from comprehensive immune profiling in a limited melanoma cohort showed improved predictive performance as compared to PD-L1 IHC and tumor mutational burden.

How to interpret thyroid biopsy results: a three-year retrospective interventional radiology experience.

Results of thyroid biopsy determine whether thyroid nodule resection is appropriate and the extent of thyroid surgery. At our institution we use 20/22-gauge core biopsy (CBx) in conjunction with fine-needle aspiration (FNA) to decrease the number of passes and improve adequacy. Occasionally, both ultrasound (US)-guided FNA and CBx yield unsatisfactory specimens. To justify clinical recommendations for these unsatisfactory thyroid biopsies, we compare rates of malignancy at surgical resection for unsatisfactory biopsy results against definitive biopsy results. We retrospectively reviewed a database of 1979 patients who had a total of 2677 FNA and 663 CBx performed by experienced interventional radiologists under US guidance from 2003 to 2006 at a tertiary-care academic center. In 451 patients who had surgery following biopsy, Fisher's exact test was used to compare surgical malignancy rates between unsatisfactory and malignant biopsy cohorts as well as between unsatisfactory and benign biopsy cohorts. We defined statistical significance at P = 0.05. We reported an overall unsatisfactory thyroid biopsy rate of 3.7% (100/2677). A statistically significant higher rate of surgically proven malignancies was found in malignant biopsy patients compared to unsatisfactory biopsy patients (P = 0.0001). The incidence of surgically proven malignancy in unsatisfactory biopsy patients was not significantly different from that in benign biopsy patients (P = 0.8625). In conclusion, an extremely low incidence of malignancy was associated with both benign and unsatisfactory thyroid biopsy results. The difference in incidence between these two groups was not statistically significant. Therefore, patients with unsatisfactory biopsy specimens can be reassured and counseled accordingly.

Adrenal hemangioma-adenoma: an exceedingly rare adrenal collision tumor.

Adrenal collision tumors are rare clinical entities referring to separate coexisting adjacent tumors involving an adrenal gland with sharp demarcation between the two and without a substantial histologic admixture at the interface. We report a case of a 60-year-old female patient with an exceedingly rare adrenal hemangioma-adenoma collision tumor. To our knowledge, this is the first report of a collision tumor comprising an adrenal hemangioma and an adenoma.

Cytologic features of primary chondroid tumors of bone in crush preparations.

Fine-needle aspiration biopsy can be utilized effectively in the diagnosis of many bone lesions. Often, these lesions are suspected to be foci of metastatic disease based on clinical and/or radiographic findings. On occasion, microscopic examination of the aspirate yields a diagnosis of a primary neoplasm of chondroid origin. We discuss the cytologic features observed in crush preparations of the most frequently seen primary chondroid tumors of bone and how these lesions can be differentiated from each other and from metastatic lesions.

Malignant biphasic pleural mesothelioma metastatic to the liver diagnosed by fine-needle aspiration.

As malignant pleural mesotheliomas are most often rapidly fatal, distant metastases are rarely detected. Here, we report a unique case in which the diagnosis of metastatic pleural mesothelioma was made via cytologic examination of a fine-needle aspiration (FNA) of the liver. Recognition of the cytomorphologic features inherent to mesothelioma cells on FNA material may become important for proper patient management. To the best of our knowledge, the diagnosis of malignant pleural mesothelioma metastatic to the liver made by FNA has not been previously documented.