Hereditary chronic neutrophilic leukemia in a four-generation family without transformation to acute leukemia.

Chronic neutrophilic leukemia (CNL) is a rare myeloproliferative neoplasm (MPN) characterized by peripheral blood neutrophilia, marrow granulocyte hyperplasia, hepatosplenomegaly, and driver mutations in the colony-stimulating factor 3 receptor (CSF3R). Designation of activating CSF3R mutations as a defining genomic abnormality for CNL has led to increased recognition of the disease. However, the natural history of CNL remains poorly understood with most patients reported being of older age, lacking germline data, and having poor survival, in part due to transformation to acute leukemia. CSF3R driver mutations in most patients with CNL have been reported to be acquired, although rare cases of germline mutations have been described. Here, we report the largest pedigree to date with familial CNL, spanning four generations with affected family members ranging in age from 4 to 53 years, none of whom have transformed to acute leukemia. A heterozygous T618I CSF3R mutation was identified in peripheral blood and mesenchymal stromal cells from the proband and in all affected living family members, while the unaffected family members tested were homozygous wild type. We show that the T618I mutation also confers a survival advantage to neutrophils in an MCL1-dependent manner. Collectively, these data provide additional insights into the natural history of familial CNL arising from T618I CSF3R mutations and suggest that enhanced neutrophil survival also contributes to the high neutrophil count observed in patients with CNL.

Real-World Impact of an In-House Dihydropyrimidine Dehydrogenase (DPYD) Genotype Test on Fluoropyrimidine Dosing, Toxicities, and Hospitalizations at a Multisite Cancer Center.

PURPOSE: Fluoropyrimidine-related toxicity and mortality risk increases significantly in patients carrying certain DPYD genetic variants with standard dosing. We implemented DPYD genotyping at a multisite cancer center and evaluated its impact on dosing, toxicity, and hospitalization. METHODS: In this prospective observational study, patients receiving (reactive) or planning to receive (pretreatment) fluoropyrimidine-based chemotherapy were genotyped for five DPYD variants as standard practice per provider discretion. The primary end point was the proportion of variant carriers receiving fluoropyrimidine modifications. Secondary end points included mean relative dose intensity, fluoropyrimidine-related grade 3+ toxicities, and hospitalizations. Fisher's exact test compared toxicity and hospitalization rates between pretreatment carriers, reactive carriers, and wild-type patients. Univariable and multivariable logistic regression identified factors associated with toxicity and hospitalization risk. Kaplan-Meier methods estimated time to event of first grade 3+ toxicity and hospitalization. RESULTS: Of the 757 patients who received DPYD genotyping (median age 63, 54% male, 74% White, 19% Black, 88% GI malignancy), 45 (5.9%) were heterozygous carriers. Fluoropyrimidine was modified in 93% of carriers who started treatment. In 442 patients with 3-month follow-up, 64%, 31%, and 30% of reactive carriers, pretreatment carriers, and wild-type patients had grade 3+ toxicity, respectively (P = .085); 64%, 25%, and 13% were hospitalized (P < .001). Reactive carriers had 10-fold higher odds of hospitalization compared with wild-type patients (P = .001), whereas no significant difference was noted between pretreatment carriers and wild-type patients. Time-to-event of toxicity and hospitalization were significantly different between genotype groups (P < .001), with reactive carriers having the earliest onset and highest incidence. CONCLUSION: DPYD genotyping prompted fluoropyrimidine modifications in most carriers. Pretreatment testing reduced toxicities and hospitalizations compared with reactive testing, thus normalizing the risk to that of wild-type patients, and should be considered standard practice.

Can a Liquid Biopsy Detect Circulating Tumor DNA With Low-passage Whole-genome Sequencing in Patients With a Sarcoma? A Pilot Evaluation.

BACKGROUND: A liquid biopsy is a test that evaluates the status of a disease by analyzing a sample of bodily fluid, most commonly blood. In recent years, there has been progress in the development and clinical application of liquid biopsy methods to identify blood-based, tumor-specific biomarkers for many cancer types. However, the implementation of these technologies to aid in the treatment of patients who have a sarcoma remains behind other fields of cancer medicine. For this study, we chose to evaluate a sarcoma liquid biopsy based on circulating tumor DNA (ctDNA). All human beings have normal cell-free DNA (cfDNA) circulating in the blood. In contrast with cfDNA, ctDNA is genetic material present in the blood stream that is derived from a tumor. ctDNA carries the unique genomic fingerprint of the tumor with changes that are not present in normal circulating cfDNA. A successful ctDNA liquid biopsy must be able to target these tumor-specific genetic alterations. For instance, epidermal growth factor receptor (EGFR) mutations are common in lung cancers, and ctDNA liquid biopsies are currently in clinical use to evaluate the status of disease in patients who have a lung cancer by detecting EGFR mutations in the blood. As opposed to many carcinomas, sarcomas do not have common recurrent mutations that could serve as the foundation to a ctDNA liquid biopsy. However, many sarcomas have structural changes to their chromosomes, including gains and losses of portions or entire chromosomes, known as copy number alterations (CNAs), that could serve as a target for a ctDNA liquid biopsy. Murine double minute 2 (MDM2) amplification in select lipomatous tumors or parosteal osteosarcoma is an example of a CNA due to the presence of extra copies of a segment of the long arm of chromosome 12. Since a majority of sarcomas demonstrate a complex karyotype with numerous CNAs, a blood-based liquid biopsy strategy that searches for these CNAs may be able to detect the presence of sarcoma ctDNA. Whole-genome sequencing (WGS) is a next-generation sequencing technique that evaluates the entire genome. The depth of coverage of WGS refers to how detailed the sequencing is, like higher versus lower power on a microscope. WGS can be performed with high-depth sequencing (that is, > 60×), which can detect individual point mutations, or low-depth sequencing (that is, 0.1× to 5×), referred to as low-passage whole-genome sequencing (LP-WGS), which may not detect individual mutations but can detect structural chromosomal changes including gains and losses (that is, CNAs). While similar strategies have shown favorable early results for specific sarcoma subtypes, LP-WGS has not been evaluated for applicability to the broader population of patients who have a sarcoma. QUESTIONS/PURPOSES: Does an LP-WGS liquid biopsy evaluating for CNAs detect ctDNA in plasma samples from patients who have sarcomas representing a variety of histologic subtypes? METHODS: This was a retrospective study conducted at a community-based, tertiary referral center. Nine paired (plasma and formalin-fixed paraffin-embedded [FFPE] tissue) and four unpaired (plasma) specimens from patients who had a sarcoma were obtained from a commercial biospecimen bank. Three control specimens from individuals who did not have cancer were also obtained. The paired and unpaired specimens from patients who had a sarcoma represented a variety of sarcoma histologic subtypes. cfDNA was extracted, amplified, and quantified. Libraries were prepared, and LP-WGS was performed using a NextSeq 500 next-generation sequencing machine at a low depth of sequencing coverage (∼1×). The ichorCNA bioinformatics algorithm, which was designed to detect CNAs from low-depth genomic sequencing data, was used to analyze the data. In contrast with the gold standard for diagnosis in the form of histopathologic analysis of a tissue sample, this test does not discriminate between sarcoma subtypes but detects the presence of tumor-derived CNAs within the ctDNA in the blood that should not be present in a patient who does not have cancer. The liquid biopsy was positive for the detection of cancer if the ichorCNA algorithm detected the presence of ctDNA. The algorithm was also used to quantitatively estimate the percent ctDNA within the cfDNA. The concentration of ctDNA was then calculated from the percent ctDNA relative to the total concentration of cfDNA. The CNAs of the paired FFPE tissue and plasma samples were graphically visualized using aCNViewer software. RESULTS: This LP-WGS liquid biopsy detected ctDNA in 9 of 13 of the plasma specimens from patients with a sarcoma. The other four samples from patients with a sarcoma and all serum specimens from patients without cancer had no detectable ctDNA. Of those 9 patients with positive liquid biopsy results, the percent ctDNA ranged from 6% to 11%, and calculated ctDNA quantities were 0.04 to 5.6 ng/mL, which are levels to be expected when ctDNA is detectable. CONCLUSION: In this small pilot study, we were able to detect sarcoma ctDNA with an LP-WGS liquid biopsy searching for CNAs in the plasma of most patients who had a sarcoma representing a variety of histologic subtypes. CLINICAL RELEVANCE: These results suggest that an LP-WGS liquid biopsy evaluating for CNAs to identify ctDNA may be more broadly applicable to the population of patients who have a sarcoma than previously reported in studies focusing on specific subtypes. Large prospective clinical trials that gather samples at multiple time points during the process of diagnosis, treatment, and surveillance will be needed to further assess whether this technique can be clinically useful. At our institution, we are in the process of developing a large prospective clinical trial for this purpose.

Prognostic significance of myeloid-derived suppressor cells and systemic inflammation in newly diagnosed diffuse large B cell lymphoma treated with chemoimmunotherapy.

The revised International Prognostic Index (R-IPI) is an important prognostic tool in diffuse large B cell lymphoma (DLBCL); however, outcomes can vary markedly within R-IPI groups, and additional prognostic markers are needed. We conducted a prospective observational study to evaluate the circulating immature myeloid (IM) cell subsets and cytokine profiles of 31 patients with newly diagnosed DLBCL before and after chemoimmunotherapy. Among circulating IM cells, myeloid-derived suppressor cells (MDSCs) were the predominant cell type (73.8% ± 26%). At baseline, circulating monocytic MDSCs (M-MDSCs) and polymorphonuclear MDSCs (PMN-MDSCs) were predominantly mutually exclusive. Patients with DLBCL clustered into three distinct immunotypes according to MDSC levels and subtype predominance: M-MDSChigh, PMN-MDSChigh, and MDSClow. The M-MDSChigh immunotype was associated with the germinal center B cell-like (GCB) subtype and elevated serum IL-8 and MIP-1α levels. PMN-MDSChigh was associated with the non-GCB subtype and elevated IL-8, MCP-1, IP-10, TNFα, and IL-1Ra levels. Standard chemoimmunotherapy partially reduced M-MDSC distribution across the MDSClow and M-MDSChigh groups. By contrast, among the MDSClow and PMN-MDSChigh groups, PMN-MDSCs persisted after treatment. Two high-risk patients with non-GCB DLBCL and MDSClow immunotype experienced early disease recurrence within 12 months of treatment completion. This study demonstrates that distinct types of MDSCs are associated with subtypes of DLBCL. MDSC levels are dynamic and may be associated with disease status. Persistence of PMN-MDSCs among high-risk patients with DLBCL may be associated with early relapse.

Unveiling Discrepant and Rare Dihydropyrimidine Dehydrogenase (DPYD) Results Using an In-House Genotyping Test: A Case Series.

Fluoropyrimidine chemotherapy is a primary component of many solid tumor treatment regimens, particularly those for gastrointestinal malignancies. Approximately one-third of patients receiving fluoropyrimidine-based chemotherapies experience serious adverse effects. This risk is substantially higher in patients carrying DPYD genetic variants, which cause reduced fluoropyrimidine metabolism and inactivation (ie, dihydropyridine dehydrogenase [DPD] deficiency). Despite the known relationship between DPD deficiency and severe toxicity risk, including drug-related fatalities, pretreatment DPYD testing is not standard of care in the United States. We developed an in-house DPYD genotyping test that detects 5 clinically actionable variants associated with DPD deficiency, and genotyped 827 patients receiving fluoropyrimidines, of which 49 (6%) were identified as heterozygous carriers. We highlight 3 unique cases: (1) a patient with a false-negative result from a commercial laboratory that only tested for the c.1905 + 1G>A (*2A) variant, (2) a White patient in whom the c.557A>G variant (typically observed in people of African ancestry) was detected, and (3) a patient with the rare c.1679T>G (*13) variant. Lastly, we evaluated which DPYD variants are detected by commercial laboratories offering DPYD genotyping in the United States and found 6 of 13 (46%) did not test for all 5 variants included on our panel. We estimated that 20.4% to 81.6% of DPYD heterozygous carriers identified on our panel would have had a false-negative result if tested by 1 of these 6 laboratories. The sensitivity and negative predictive value of the diagnostic tests from these laboratories ranged from 18.4% to 79.6% and 95.1% to 98.7%, respectively. These cases underscore the importance of comprehensive DPYD genotyping to accurately identify patients with DPD deficiency who may require lower fluoropyrimidine doses to mitigate severe toxicities and hospitalizations. Clinicians should be aware of test limitations and variability in variant detection by commercial laboratories, and seek assistance by pharmacogenetic experts or available resources for test selection and result interpretation.

Thrombosis Rates and Genetic Thrombophilia Risk Among Patients With Advanced Germ Cell Tumors Treated With Chemotherapy.

INTRODUCTION: Men with advanced germ cell tumors (GCT) treated with chemotherapy are at high risk of venous thromboembolism (VTE). Predictors of VTE may identify patients who would benefit from prophylactic anticoagulation. PATIENTS AND METHODS: Men with advanced GCT (Stage IS, II, III) treated with chemotherapy were identified at 2 centers. High genomic risk was defined from a 5 single nucleotide polymorphism (SNP) germline panel. Logistic regression was used to evaluate the impact of genomic risk on VTE within 6 months of chemotherapy initiation. Orthogonal Projection to Latent Structures Discriminant Analysis (OPLS-DA) was used to build models to predict VTE based on clinical variables and an 86 SNP panel. RESULTS: This 123-patient cohort experienced a VTE rate of 26% with an incidence of high genomic risk of 21%. Men with high genomic risk did not have a significantly higher VTE rate (31%, 8/26) than men with low genomic risk (25%, 24/97), unadjusted OR 1.4 (95% CI 0.5-3.5, P = .54). Incorporation of clinical variables (Khorana score, N3 status and elevated LDH) resulted in adjusted OR 2.1 (95% CI 0.7-6.5, P = .18). A combined model using clinical variables and 86 SNPs performed similarly (AUC 0.77) compared to clinical variables alone (AUC 0.72). CONCLUSIONS: A previously established 5-SNP panel was not associated with VTE among patients with GCT receiving chemotherapy. However, multivariable models based on clinical variables alone warrant further validation to inform prophylactic anticoagulation strategies.

Pharmacogenetic and clinical predictors of voriconazole concentration in hematopoietic stem cell transplant recipients receiving CYP2C19-guided dosing.

CYP2C19-guided voriconazole dosing reduces pharmacokinetic variability, but many patients remain subtherapeutic. The aim of this study was to evaluate the effect of candidate genes and a novel CYP2C haplotype on voriconazole trough concentrations in patients receiving CYP2C19-guided dosing. This is a retrospective candidate gene study in allogeneic hematopoietic cell transplant (HCT) patients receiving CYP2C19-guided voriconazole dosing. Patients were genotyped for ABCB1, ABCG2, CYP2C9, CYP3A4, CYP3A5, and the CYP2C haplotype. Of 185 patients, 36% were subtherapeutic (of which 79% were normal or intermediate metabolizers). In all patients, CYP2C19 (p < 0.001), age (p = 0.018), and letermovir use (p = 0.001) were associated with voriconazole concentrations. In the subset receiving 200 mg daily (non-RM/UMs), CYP2C19 (p = 0.004) and ABCG2 (p = 0.015) were associated with voriconazole concentrations; CYP2C19 (p = 0.028) and letermovir use (p = 0.001) were associated with subtherapeutic status. CYP2C19 phenotype and letermovir use were significantly associated with subtherapeutic voriconazole concentrations and may be used to improve voriconazole precision dosing, while further research is needed to clarify the role of ABCG2 in voriconazole dosing.

Evaluation of pentamidine tolerability and efficacy between CYP2C19 phenotypes.

Intravenous pentamidine is used for prophylaxis against Pneumocystis jirovecii pneumonia, an infection seen in hematopoietic stem cell transplant recipients. Pentamidine is partially metabolized by CYP2C19, which is vulnerable to pharmacogenetic variation. This retrospective study evaluated allogeneic hematopoietic stem cell transplant patients who received intravenous pentamidine as P. jirovecii pneumonia prophylaxis. The primary objective was the association between CYP2C19 phenotype and discontinuation of pentamidine due to drug-related side effects based on univariate logistic regression (N = 81). Ten patients (12.3%) discontinued pentamidine because of side effects. There was no difference in discontinuation between phenotype groups (p = 0.18) or discontinuation due to side effects (p = 0.76). Overall, no association was seen between phenotypes and pentamidine-related side effects (p = 0.475). Drug discontinuation rates and P. jirovecii pneumonia infection rates were low.

Genetic Predictors of Ibrutinib-related Cardiovascular Side Effects in Patients with Chronic Lymphocytic Leukemia.

PURPOSE: Patients with chronic lymphocytic leukemia (CLL) treated with ibrutinib are at risk of developing cardiovascular side effects (CVSE). The molecular determinants of CVSEs have not been fully elucidated. We interrogated genetic polymorphisms in the Bruton tyrosine kinase (BTK) signaling pathway for their association with ibrutinib-related CVSEs. EXPERIMENTAL DESIGN: We conducted a retrospective/prospective observational pharmacogenetic study of 50 patients with newly diagnosed or relapsed CLL who received ibrutinib at a starting daily dose of 420 mg for at least 6 months. CVSEs, primarily atrial fibrillation and hypertension, occurred in 10 patients (20%), of whom 4 discontinued therapy. DNA was isolated from buccal swabs of all 50 patients and genotyped for 40 SNPs in GATA4, SGK1, KCNQ1, KCNA4, NPPA, and SCN5A using a customized next-generation sequencing panel. Univariate and multivariate logistic regression analysis were performed to determine genetic and clinical factors associated with the incidence of ibrutinib-related CVSEs. RESULTS: GATA4 rs804280 AA (P = 0.043), KCNQ1 rs163182 GG (P = 0.036), and KCNQ1 rs2237895 AA (P = 0.023) genotypes were univariately associated with ibrutinib-related CVSEs. On the basis of multivariate analysis, a high genetic risk score, defined as the presence of at least two of these genotypes, was associated with 11.5-fold increased odds of CVSEs (P = 0.019; 95% confidence interval, 1.79-119.73). CONCLUSIONS: Our findings suggest possible genetic determinants of ibrutinib-related CVSEs in CLL. If replicated in a larger study, pretreatment pharmacogenetic testing for GATA4 and KCNQ1 polymorphisms may be a useful clinical tool for personalizing treatment selection for CLL and/or instituting early risk mitigation strategies.

Human oral mucosa and oral microbiome interactions following supragingival plaque reconstitution in healthy volunteers: a diet-controlled balanced design proof-of-concept model to investigate oral pathologies.

Changes in the oral microbiome may contribute to oral pathologies, especially in patients undergoing cancer therapy. Interactions between oral microbiome and oral mucosa may exacerbate inflammation. We determined whether probiotic-controlled plaque formation could impact proximal oral mucosa gene expression profiles in healthy volunteers. A 3-weeks balanced sample collection design from healthy volunteers (HVs) was implemented. At Week-1 plaques samples and labial mucosa brush biopsies were obtained from HVs in the morning (N = 4) and/or in the afternoon (N = 4), and groups were flipped at Week-3. A fruit yogurt and tea diet were given 2-4hrs before sample collection. mRNA gene expression analysis was completed using RNA-Seq and DESeq2. Bacterial taxa relative abundance was determined by 16S HOMINGS. Bacterial diversity changes and metabolic pathway enrichment were determined using PRIMERv7 and LEfSe programs. Alpha- and beta-diversities did not differ morning (AM) vs. afternoon (PM). The most affected KEGG pathway was Toll-like receptor signaling in oral mucosa. Eighteen human genes and nine bacterial genes were differentially expressed in plaque samples. Increased activity for 'caries-free' health-associated calcifying Corynebacterium matruchotii and reduced activity for Aggregatibacter aphrophilus, an opportunistic pathogen, were observed. Microbial diversity was not altered after 8 hours plaque formation in healthy individuals as opposed to gene expression.

Addressing barriers to increased adoption of DPYD genotyping at a large multisite cancer center.

PURPOSE: To describe the implementation of an in-house genotyping program to detect genetic variants linked to impaired dihydropyrimidine dehydrogenase (DPD) metabolism at a large multisite cancer center, including barriers to implementation and mechanisms to overcome barriers to facilitate test adoption. SUMMARY: Fluoropyrimidines, including fluorouracil and capecitabine, are commonly used chemotherapy agents in the treatment of solid tumors, such as gastrointestinal cancers. DPD is encoded by the DPYD gene, and individuals classified as DPYD intermediate and poor metabolizers due to certain genetic variations in DPYD can experience reduced fluoropyrimidine clearance and an increased risk of fluoropyrimidine-related adverse events. Although pharmacogenomic guidelines provide evidence-based recommendations for DPYD genotype-guided dosing, testing has not been widely adopted in the United States for numerous reasons, including limited education/awareness of clinical utility, lack of testing recommendations by oncology professional organizations, testing cost, lack of accessibility to a comprehensive in-house test and service, and prolonged test turnaround time. Based on stakeholder feedback regarding barriers to testing, we developed an in-house DPYD test and workflow to facilitate testing in multiple clinic locations at Levine Cancer Institute. Across 2 gastrointestinal oncology clinics from March 2020 through June 2022, 137 patients were genotyped, and 13 (9.5%) of those patients were heterozygous for a variant and identified as DPYD intermediate metabolizers. CONCLUSION: Implementation of DPYD genotyping at a multisite cancer center was feasible due to operationalization of workflows to overcome traditional barriers to testing and engagement from all stakeholders, including physicians, pharmacists, nurses, and laboratory personnel. Future directions to scale and sustain testing in all patients receiving a fluoropyrimidine across all Levine Cancer Institute locations include electronic medical record integration (eg, interruptive alerts), establishment of a billing infrastructure, and further refinement of workflows to improve the rate of pretreatment testing.

Genomic alterations and clinical outcomes in patients with dedifferentiated liposarcoma.

PURPOSE: Patients with unresectable dedifferentiated liposarcoma (DDLPS) have poor overall outcomes. Few genomic alterations have been identified with limited therapeutic options. EXPERIMENTAL DESIGN: Patients treated at Levine Cancer Institute with DDLPS were identified. Next generation sequencing (NGS), immunohistochemistry (IHC), and fluorescence in situ hybridization (FISH) testing were performed on tumor tissue collected at diagnosis or recurrence/progression. Confirmation of genomic alterations was performed by orthologous methods and correlated with clinical outcomes. Univariate Cox regression was used to identify genomic alterations associated with clinical outcomes. RESULTS: Thirty-eight DDLPS patients with adequate tissue for genomic profiling and clinical data were identified. Patient characteristics included: median age at diagnosis (66 years), race (84.2% Caucasian), and median follow-up time for the entire cohort was 12.1 years with a range from approximately 3.5 months to 14.1 years. Genes involved in cell cycle regulation, including MDM2 (74%) CDK4 (65%), and CDKN2A (23%), were amplified along with WNT/Notch pathway markers: HMGA2, LGR5, MCL1, and CALR (19%-29%). While common gene mutations were identified, PDE4DIP and FOXO3 were also mutated in 47% and 34% of patients, respectively, neither of which have been previously reported. FOXO3 was associated with improved overall survival (OS) (HR 0.37; p = 0.043) along with MAML2 (HR 0.30; p = 0.040). Mutations that portended worse prognosis included RECQL4 (disease-specific survival HR 4.67; p = 0.007), MN1 (OS HR = 3.38; p = 0.013), NOTCH1 (OS HR 2.28, p = 0.086), and CNTRL (OS HR 2.42; p = 0.090). CONCLUSIONS: This is one of the largest retrospective reports analyzing genomic aberrations in relation to clinical outcomes for patients with DDLPS. Our results suggest therapies targeting abnormalities should be explored and confirmation of prognostic markers is needed. Dedifferentiated liposarcoma is one of the most common subtypes of soft tissue sarcoma yet little is known of its molecular aberrations and possible impact on outcomes. The work presented here is an evaluation of genetic abnormalities among a population of patients with dedifferentiated liposarcoma and how they corresponded with survival and risk of metastases. There were notable gene mutations and amplifications commonly found, some of which had interesting prognostic implications.

Identification of potential biomarkers and novel therapeutic targets through genomic analysis of small cell bladder carcinoma and associated clinical outcomes.

OBJECTIVES: Small cell bladder carcinoma (SCBC) represents a rare histologic variant with a poor prognosis and for which no routine biomarkers exist. Limited reports of genomic sequencing in SCBC have demonstrated a high prevalence of TP53 and RB1 gene mutations, though the prognostic value of these and other gene variants in SCBC remains undefined. In this study, we performed targeted genomic sequencing on a cohort of SCBC patients and correlated genomic findings with clinical outcomes to identify potential novel biomarkers. MATERIALS AND METHODS: Thirty-one patients with SCBC and available treatment-naïve tumor specimens were identified from an institutional database (23 limited stage [LS], 8 extensive stage [ES]). Small cell carcinoma specimens were microdissected and subjected to tumor next-generation whole-exon sequencing with a 592 gene panel. Kaplan-Meier techniques and Cox proportional hazards models were used to evaluate genomic aberration association with relapse-free survival (RFS) and overall survival (OS) in the limited stage cohort. RESULTS: The most common pathogenic gene variants included ARID1A (48%), TP53 (48%) and RB1 (48%). Mutations in genes with potential therapeutic targets not routinely evaluated in SCBC included BRCA1/2 (16%), POLE (13%), JAK2 (13%), PDGFB (13%) and FGFR3 (3%). Multiple novel biomarker candidates showed trends for improvements in OS in the LS subset including ERCC2 (HR 0.322, P = 0.122) and RB1 (HR 0.481, P = 0.182), while LS patients with TP53 mutations (HR 2.730, P = 0.056), and MCL1 gene amplification (HR 4.183, P = 0.018) suggested inferior OS. Additionally, gene or copy number variants with potential prognostic benefit included UBR5 and DAXX (P = 0.02, [hazard ratios nonestimable due to zero events in biomarker positive groups]). CONCLUSIONS: These results support the role for tumor genomic profiling in SCBC and identify multiple potential novel biomarkers and therapeutic targets in this rare disease. Efforts to validate these findings should lead to improved decision-making and treatment outcomes in SCBC.

Phase II Study of Pembrolizumab in Combination with Doxorubicin in Metastatic and Unresectable Soft-Tissue Sarcoma.

PURPOSE: Doxorubicin is standard therapy for advanced soft-tissue sarcoma (STS) with minimal improvement in efficacy and increased toxicity with addition of other cytotoxic agents. Pembrolizumab monotherapy has demonstrated modest activity and tolerability in previous advanced STS studies. This study combined pembrolizumab with doxorubicin to assess safety and efficacy in frontline and relapsed settings of advanced STS. PATIENTS AND METHODS: This single-center, single-arm, phase II trial enrolled patients with unresectable or metastatic STS with no prior anthracycline therapy. Patients received pembrolizumab 200 mg i.v. and doxorubicin (60 mg/m2 cycle 1 with subsequent escalation to 75 mg/m2 as tolerated). The primary endpoint was safety. Secondary endpoints included overall survival (OS), objective response rate (ORR), and progression-free survival (PFS) based on RECIST v1.1 guidelines. RESULTS: Thirty patients were enrolled (53.3% female; median age 61.5 years; 87% previously untreated) with 4 (13.3%) patients continuing treatment. The study met its primary safety endpoint by prespecified Bayesian stopping rules. The majority of grade 3+ treatment-emergent adverse events were hematologic (36.7% 3+ neutropenia). ORR was 36.7% [95% confidence interval (CI), 19.9-56.1%], with documented disease control in 80.0% (95% CI, 61.4-92.3%) of patients. Ten (33.3%) patients achieved partial response, 1 (3.3%) patient achieved complete response, and 13 (43.3%) patients had stable disease. Median PFS and OS were 5.7 months (6-month PFS rate: 44%) and 17 months (12-month OS rate: 62%), respectively. Programmed cell death ligand-1 (PD-L1) expression was associated with improved ORR, but not OS or PFS. CONCLUSIONS: Combination pembrolizumab and doxorubicin has manageable toxicity and preliminary promising activity in treatment of patients with anthracycline-naive advanced STS.

Genomic analysis of response to bacillus Calmette-Guérin (BCG) treatment in high-grade stage 1 bladder cancer patients.

BACKGROUND: Intravesical bacillus Calmette-Guérin (BCG) therapy is standard treatment for high-risk non-muscle invasive bladder cancer (NMIBC) but overall efficacy is low, and no reliable predictive biomarkers currently exist to refine patient selection. We performed genomic analysis on high-grade (HG) T1 NMIBCs to determine if response to therapy is predicted by certain mutational and/or expressional changes. METHODS: Patients with HG T1 NMIBC treated with induction BCG were stratified by response into durable and non-durable responders. Baseline tumor samples were subjected to targeted DNA sequencing and whole-exome RNAseq. Genomic variants differing significantly between response groups were analyzed using Ingenuity Pathway Analysis (IPA) software. Variant selection was refined to target potential biomarker candidates for responsiveness to BCG. RESULTS: Among 42 patients, the median follow-up was 51.7 months and 40.5% (n=17) were durable BCG responders. Deleterious mutations in the RNA sequence of JCHAIN, S100A7, CLEC2B, and ANXA10 were more common in non-durable responders. Mutations in MCL1 and MSH6 detected on targeted sequencing were more commonly found in durable responders. Of all deleterious DNA and RNA mutations identified, only MCL1 was significantly associated with longer recurrence free survival (RFS) (P=0.031). CONCLUSIONS: Differences in the genomic profiles of HG T1 NMIBC tumors exist between those who show durable response to BCG and those who do not. Using pathway analysis, those differences imply upregulation of several interconnected inflammatory pathways among responders. Specific variants identified here, namely MCL1, are candidates for further study and, if clinically validated, may serve as useful biomarkers in the future.

Pharmacogenomics-guided supportive oncology: A tale of two trials.

Cancer-related symptoms, like depression, nausea, and pain, are common and negatively affect quality of life. Unfortunately, there is large inter-individual variability in response to supportive care medications for these symptoms. Pharmacogenomics may inform prescribing by identification of those genetically at risk for drug related adverse events or therapeutic failure. While such information can be applied to many drugs, there are specific oncology populations that could greatly benefit from pharmacogenomics-guided supportive care management due to high symptom burden, including those receiving palliative medicine and hematopoietic stem cell transplantation. The goal of this paper is to provide an overview of, and lessons learned from, the development of two prospective pharmacogenomics-guided interventional trials ("Supportive Care PGx Trial" and "Transplant PGx Trial") across two different clinical settings at the Levine Cancer Institute: the Department of Supportive Oncology and the Transplant and Cellular Therapy section. Key considerations included the appropriate study design and endpoints (balancing study goals and resources), dissemination and application of individual pharmacogenetics results, technical details about assay development, and overall care coordination to minimize clinic disruption.

North Carolina's multi-institutional pharmacogenomics efforts with the North Carolina Precision Health Collaborative.

The North Carolina Precision Health Collaborative is an interdisciplinary, public-private consortium of precision health experts who strategically align statewide resources and strengths to elevate precision health in the state and beyond. Pharmacogenomics (PGx) is a key area of focus for the North Carolina Precision Health Collaborative. Experts from Atrium Health's Levine Cancer Institute, Duke University/Duke Health System, Mission Health and the University of North Carolina (UNC) at Chapel Hill/UNC Health System have collaborated since 2017 to implement strategic PGx initiatives, including basic sciences research, translational research and clinical implementation of germline testing into practice and policy. This institutional profile highlights major PGx programs and initiatives across these organizations and how the collaborative is working together to advance PGx science and implementation.

Ex vivo efficacy of BCMA-bispecific antibody TNB-383B in relapsed/refractory multiple myeloma.

TNB-383B is a fully human BCMA-targeting T-cell engaging bispecific monoclonal antibody (T-BsAb). We assessed ex vivo efficacy of this drug to mediate killing of bone marrow mononuclear cells (BMMCs) freshly isolated from 10 patients with relapsed multiple myeloma (MM). BMMC were treated ex vivo with TNB-383B at doses ranging from 0.001-1 µg. Plasma cell (PC) lysis, viability, BCMA expression, CTL distribution, and degranulation were assessed by flow cytometry. Cytokine response to TNB-383B was quantified by multiplex protein assay. Dose-dependent PC lysis was triggered in all cases by TNB-383B at doses as low as 0.001 µg (P = .0102). Primary MM cells varied in BCMA expression. High BCMA+ PC count correlated with increased PC lysis (P = .005) and significant CTL degranulation specific to TNB-383B treatment (P = .0153 at 1 µg). High E:T ratio in bone marrow specimens led to lower viable and higher apoptotic PC compared with low E:T ratio (P < .001). Three cytokines were significantly modulated by TNB-383B: IL-2/TNFα increased by ∼4 ± 3.5-fold average (P < .005 at 1 µg) and IP10 increased by ∼50 ± 15-fold (P < .001 at 1 µg). We conclude that TNB-383B triggers primary PC lysis and CTL degranulation in a dose-dependent fashion ex vivo with no T cell expansion and mild increase of CRS-associated cytokines.