Real-world outcomes of patients with resected stage III melanoma treated with adjuvant therapies.

BACKGROUND: Both immunotherapy (IO) and targeted therapy (TT) are used as adjuvant (adj) treatment for stage III melanoma, however, data describing real-world outcomes are limited. In addition, a significant proportion of patients relapse, for whom best management is unclear. The aim of our study was to assess the efficacy, and safety of adj anti-PD1 IO and TT in a real-world cohort of patients with resected stage III melanoma, and further delineate patterns of recurrence and treatment strategies. METHODS: We retrospectively analyzed 130 patients who received adj therapy (100 anti-PD1 IO and 30 TT). RESULTS: At a median follow-up of 30 months, median relapse-free survival (RFS) was 24.6 (95% CI, 17-not reached [NR]) versus 64 (95% CI, 29.5-NR) months for the TT and IO groups, respectively (p = 0.26). Median overall survival (OS) was NR for either subgroup. At data cutoff, 77% and 82% of patients in TT and IO arms were alive. A higher number of grade ≥3 treatment-related adverse events (AEs) were noted in the IO group (11% vs. 3%), however, a higher proportion of patients permanently discontinued adj therapy in the TT group (43% vs. 11%) due to toxicity. Strategies at relapse and outcomes were variable based on location and timing of recurrence. A significant number of patients who relapsed after adj IO received a second round of IO. Among them, patients who were off adj IO at relapse had superior second median RFS (mRFS2), compared to those who relapsed while on adj IO; mRFS2 was NR versus 5.1 months (95% CI, 2.5-NR), respectively, p = 0.02. CONCLUSION: In summary, both TT and IO yielded prolonged RFS in a real-world setting, however, longer follow-up is needed to determine any potential OS benefit. Adj therapy, particularly TT, may not be as well tolerated as suggested in clinical trials, with lower completion rates (59% vs. 74%) in a real-life setting. Overall, patients who relapse during adj therapy have poor outcomes, while patients who relapse after discontinuation of adj IO therapy appear to benefit from IO re-treatment.

Application of Near-Infrared Spectroscopy for Screening of Chlorothiazide Sodium Vials.

Chlorothiazide sodium for injection, USP, is a diuretic and antihypertensive medication in the form of a white or practically white, sterile, lyophilized powder. Each vial contains 500 mg of chlorothiazide sodium, equivalent to 500 mg of chlorothiazide, and 250 mg of mannitol as an inactive ingredient. The pH is adjusted with sodium hydroxide. Chlorothiazide sodium has a molecular weight of 317.71 amu. Since 2020 there have been multiple national shortages of chlorothiazide. Recent studies target chlorothiazide's low bioavailability, aiming to enhance it through nanoparticle production via a supercritical method. The drug's solubility in supercritical carbon dioxide (scCO2) is vital, with measurements ranging from 0.417×10-5 to 1.012×10-5 mole fraction under specific conditions. Adding co-solvents, like ethanol, DMSO, and acetone, to scCO2 boosts solubility, with ethanol proving most effective, enhancing solubility by 2.02-11.75 times. Intra-lot variability was discovered in a sample of a lot of chlorothiazide sodium by the University of Kentucky Drug Quality Task Force. Two vials of six screened in one lot were displaced from the center of the lot by 4.0 and 4.2 SDs, respectively. Inter-lot variability was confirmed in the near-IR spectra of 204 vials obtained from 28 different lots of chlorothiazide sodium. Using full spectrum BEST analysis 13 vials (6.4%) were outliers.

Screening for quality with process analytical technology in a health-system pharmacy: A primer.

PURPOSE: The University of Kentucky Drug Quality Study team briefly reviews the growing concerns over pharmaceutical manufacturing quality in the globalized environment, reviews the historical approach by the US Food and Drug Administration (FDA) that prioritizes process over product in enforcing quality with manufacturers, reviews the science of process analytical technology (PAT) such as near-infrared (NIR) spectroscopy, illustrates the use of PAT methods for assessing uniformity and quality in injectable pharmaceuticals, and demonstrates the application of NIR spectroscopy in a health-system pharmacy setting while maintaining current good practice quality guidelines and regulations (cGxP). SUMMARY: Given that the current approach to monitoring quality in pharmaceutical manufacturing was developed in the late 1960s at a time when manufacturing was mostly domestic, the current approach prioritizes process over product, and the global footprint of manufacturing is straining federal resources to fulfill their task of monitoring quality, an approach to augment the quality monitoring process has been developed. PAT methodologies are supported by FDA for monitoring quality and offer a fast, low-cost, nondestructive solution. Given that the Accreditation Council for Pharmacy Education has not required qualitative/quantitative analysis and drug assaying in the pharmacy curriculum for several decades, the authors spend time explaining the science behind one of these PAT methodologies, NIR spectroscopy. This primer reviews the application of this technology in the health-system pharmacy setting and the relevant clinical applications. CONCLUSION: Utilizing PAT methodologies such as NIR spectroscopy, health-system pharmacies can gain insights about whether process controls are in place or lacking in FDA-approved formulations.

Transcriptional and epigenetic regulators of human CD8+ T cell function identified through orthogonal CRISPR screens.

Clinical response to adoptive T cell therapies is associated with the transcriptional and epigenetic state of the cell product. Thus, discovery of regulators of T cell gene networks and their corresponding phenotypes has potential to improve T cell therapies. Here we developed pooled, epigenetic CRISPR screening approaches to systematically profile the effects of activating or repressing 120 transcriptional and epigenetic regulators on human CD8+ T cell state. We found that BATF3 overexpression promoted specific features of memory T cells and attenuated gene programs associated with cytotoxicity, regulatory T cell function, and exhaustion. Upon chronic antigen stimulation, BATF3 overexpression countered phenotypic and epigenetic signatures of T cell exhaustion. Moreover, BATF3 enhanced the potency of CAR T cells in both in vitro and in vivo tumor models and programmed a transcriptional profile that correlates with positive clinical response to adoptive T cell therapy. Finally, we performed CRISPR knockout screens that defined cofactors and downstream mediators of the BATF3 gene network.

Variability in Content of Hydrocortisone Sodium Succinate.

SOLU-CORTEFⓇ Sterile Powder is a type of anti-inflammatory glucocorticoid that contains hydrocortisone sodium succinate as its active ingredient. It can be administered intravenously or intramuscularly, and comes in several packages including 100 mg plain vials without diluent. The diluent, which is part of the ACT-O-VIAL system, contains only Water for Injection and no preservatives. The pH of each formula is adjusted with sodium hydroxide to ensure it falls within the specified range of 7 to 8 after reconstitution. Intralot variability was detected in lot GA6092. Measuring in the PC subspace using just PCs 4, 5 and 6, vial 12 plots 4.2 BEST SDs from the center of the cluster, and vial 7 is 3.7 SDs from the center. Vial 18 appears 3.1 SDS from the center of the cluster (3/18, 17%). Interlot variability was also found in the spectral library (lots GA6092, GK7048, GM6839, GR8925, FL8062, FN6860, FR1914, and FR5098) containing the spectra of 126 hydrocortisone sodium succinate vials.

Spectrometric Analysis of Dantrolene Sodium.

Dantrolene sodium is a direct-acting skeletal muscle relaxant. Dantrolene sodium for injection is indicated, along with suitable supportive measures, for the management of sudden, severe hypermetabolism of skeletal muscle typical of malignant hyperthermia crises in patients of any age. The formulation scanned in this work was designed to be injected intravenously. Intra-lot and inter-lot variability in the spectra of REVONTO™ (dantrolene sodium) was measured in the Drug Quality Study (DQS) using Fourier transform near-infrared spectrometry (FTNIR). Spectra of 69 vials from lot 20REV01A contained two groups (n1=56 vials, n2=13 vials) when scanned with an FTNIR. The two groups of spectra in lot 20REV01A were found to be 66.7 SDs apart using a subcluster detection test, suggesting that the two groups were manufactured differently. As a result, all available samples of dantrolene were examined. A library of spectra of 141 vials of dantrolene from 4 lots were found to contain 3 separate groups, also suggesting that different vials contain different materials.

Quality Variations in Thyrotropin Alfa.

Thyrotropin alfa is a heterodimeric glycoprotein containing human thyroid stimulating hormone (TSH). It is used as an adjunctive diagnostic tool for serum thyroglobulin (Tg) testing with or without radioiodine imaging in the follow-up of patients with well-differentiated thyroid cancer who have previously undergone thyroidectomy. Inter-lot variability in the Fourier transform near-infrared spectra of 30 samples obtained from four separate lots of Thyrogen® was detected in the Drug Quality Study (DQS). The vials fell into two distinct groups (rtst = 0.90, rlim= 0.98, p=0.02). In addition, one vial of the 30 (3%) appeared 4.7 multidimensional SDs from all of the other vials, suggesting that it also represents a different material.

Spectrometric Assessment of Generic and Brand Drug Quality for a Sentinel Screening Network.

The U.S. Food and Drug Administration (FDA) is a worldwide leader among analogous regulatory organizations in other countries. The FDA uses current good manufacturing practices to regulate the processes that produce drugs. Nevertheless, investigative journalists have pointed out problems in the drug supply, and pharmacies are not required to test the drugs they receive. The University of Kentucky Drug Quality Study does perform screening on the sterile injectable drugs that it receives and regularly reports new findings to FDA, practitioners, and the public. A Sentinel Screening Network of academic health systems could provide independent data on drug quality to FDA not available through manufacturers.

Orthogonal CRISPR screens to identify transcriptional and epigenetic regulators of human CD8 T cell function.

The clinical response to adoptive T cell therapies is strongly associated with transcriptional and epigenetic state. Thus, technologies to discover regulators of T cell gene networks and their corresponding phenotypes have great potential to improve the efficacy of T cell therapies. We developed pooled CRISPR screening approaches with compact epigenome editors to systematically profile the effects of activation and repression of 120 transcription factors and epigenetic modifiers on human CD8+ T cell state. These screens nominated known and novel regulators of T cell phenotypes with BATF3 emerging as a high confidence gene in both screens. We found that BATF3 overexpression promoted specific features of memory T cells such as increased IL7R expression and glycolytic capacity, while attenuating gene programs associated with cytotoxicity, regulatory T cell function, and T cell exhaustion. In the context of chronic antigen stimulation, BATF3 overexpression countered phenotypic and epigenetic signatures of T cell exhaustion. CAR T cells overexpressing BATF3 significantly outperformed control CAR T cells in both in vitro and in vivo tumor models. Moreover, we found that BATF3 programmed a transcriptional profile that correlated with positive clinical response to adoptive T cell therapy. Finally, we performed CRISPR knockout screens with and without BATF3 overexpression to define co-factors and downstream factors of BATF3, as well as other therapeutic targets. These screens pointed to a model where BATF3 interacts with JUNB and IRF4 to regulate gene expression and illuminated several other novel targets for further investigation.

Spectral Intra-Lot and Inter-Lot Variability in Carfilzomib.

Carfilzomib is a prescription injectable drug approved for use by the FDA as an antineoplastic agent, part of a drug class of medications known as proteasome inhibitors, and used to stop and slow the growth and progression of cancer cells within the body. The drug is approved as an agent to treat multiple myeloma. It is provided as a single-use vial that contains 60 mg of carfilzomib as a sterile, white to off-white lyophilized cake or powder. Intra-lot and inter-lot variability in the spectra of carfilzomib vials was detected in the Drug Quality Study (DQS) using Fourier transform near-infrared spectrometry (FTNIR). One of 12 vials of lot 1143966 manufactured for Onyx Pharmaceuticals, Inc. appeared 4.7 multidimensional standard deviations (SDs) from the other 11 vials in a 3-D space formed by the first 3 principal components, which captured 81% of the total spectral variation. Spectra of 168 vials from 18 lots in the spectral library formed two groups in the 3-D space formed by the first 3 principal components. One group contained 155 vials and the other group contained 13 vials. The 2 groups had different locations and scales using a subcluster detection test at p=0.02.

Assessment of Vecuronium Quality Using Near-Infrared Spectrometry.

This study employed Fourier Transform near-infrared spectrometry to assess the quality of vecuronium bromide, a neuromuscular blocking agent. Spectral data from two lots of vecuronium were collected and analyzed using the BEST metric, principal component analysis (PCA) and other statistical techniques. The results showed that there was variability between the two lots and within each lot. Several outliers in the spectral data suggested potential differences in the chemical composition or sample condition of the vials. The outliers were identified and their spectral features were examined. A total of eight unique outliers were found in the PC space from PCs 1 to 9, so 22% of the total vials were outliers. The study findings suggest that the manufacturing process of vecuronium bromide may have been operating outside of a state of process control. Further investigation is needed to determine the source of these variations and their impact on the safety and efficacy of the drug product.

Spectrometric Analysis of Process Variations in Remifentanil.

ULTIVA® (remifentanil hydrochloride) is a sterile, nonpyrogenic, preservative-free, white to off-white lyophilized powder for intravenous (IV) administration after reconstitution and dilution. Each vial contains 1, 2, or 5 mg of remifentanil base; 15 mg glycine; and hydrochloric acid to buffer the solutions to a nominal pH of 3 after reconstitution. ULTIVA® is a µ-opioid agonist with rapid onset and peak effect, and short duration of action. Intra-lot and inter-lot variability in the spectra of ULTIVA® was measured in the Drug Quality Study (DQS) using Fourier transform near-infrared spectrometry (FTNIR). In 6 vials sampled, 1 came from lot 220453F while 5 came from lot 30020BF. The 1 vial sampled from lot 220453F appeared 122 multidimensional SDs from the other vials from lot 30020BF, suggesting that it represents a different formulation or material. Consequently, additional spectra from other lots were analyzed. Spectra of 90 vials from 9 lots in the spectral library contained vials that were outside the main group (50.3 SDs using a subcluster detection test), suggesting that the 35 library vials (39% of the total) contain different materials from the other 55 vials.

Potential Process Control Issues with Pemetrexed.

Pemetrexed is a folate analog metabolic inhibitor used in treatment of locally advanced or metastatic nonsquamous non-small cell lung cancer and for mesothelioma. Intra-lot and inter-lot variability in the spectra of ALIMTA® was detected in the Drug Quality Study (DQS) using Fourier transform near-infrared spectrometry (FTNIR). One vial of 12 (8%) sampled from lot S20I013A appeared 3.0 multidimensional SDs from the other vials, suggesting that it represents a different material. Consequently, additional spectra from other lots were analyzed. Spectra of 147 vials from 23 lots in the spectral library contained 14 vials that were outside the main group (26.4 SDs using a subcluster detection test), suggesting that the 14 library vials (9.5% of the total) also contain differing materials.

FDA Approaches in Monitoring Drug Quality, Forces Impacting the Drug Quality, and Recent Alternative Strategies to Assess Quality in the US Drug Supply.

Since the US Food and Drug Administration (FDA) began monitoring the quality of pharmaceutical manufacturing by enforcing current good manufacturing practices roughly 60 years ago, forces related to the global economy have changed, rendering the task of monitoring quality more difficult. Alternative strategies by groups like Valisure, LLC, and the University of Kentucky Drug Quality Study to monitor the quality of the currently circulated US drug supply through end-product testing and screening have resulted in several concerning findings. Given the successful approaches of identifying quality defects in pharmaceuticals by non-regulatory bodies, and considering the changing landscape and pressures on manufacturing, the FDA, large buying groups, and the US Department of Defense should consider these alternative strategies as a means to augment current regulatory activities.

Patient-derived micro-organospheres enable clinical precision oncology.

Patient-derived xenografts (PDXs) and patient-derived organoids (PDOs) have been shown to model clinical response to cancer therapy. However, it remains challenging to use these models to guide timely clinical decisions for cancer patients. Here, we used droplet emulsion microfluidics with temperature control and dead-volume minimization to rapidly generate thousands of micro-organospheres (MOSs) from low-volume patient tissues, which serve as an ideal patient-derived model for clinical precision oncology. A clinical study of recently diagnosed metastatic colorectal cancer (CRC) patients using an MOS-based precision oncology pipeline reliably assessed tumor drug response within 14 days, a timeline suitable for guiding treatment decisions in the clinic. Furthermore, MOSs capture original stromal cells and allow T cell penetration, providing a clinical assay for testing immuno-oncology (IO) therapies such as PD-1 blockade, bispecific antibodies, and T cell therapies on patient tumors.

Neoadjuvant and Adjuvant Systemic Therapy for Early-Stage Non-small-Cell Lung Cancer.

Approximately a third of patients with non-small-cell lung cancer (NSCLC) present with surgically resectable disease. Patients who undergo surgical resection are at a high risk of relapse, and neoadjuvant and adjuvant chemotherapy improves disease-free survival (DFS) and overall survival (OS). The outcomes with neoadjuvant and adjuvant chemotherapy are similar, and both are used in clinical practice. Recent trials investigated the role of immunotherapy and targeted therapy in patients with early-stage disease. A phase III trial of adjuvant atezolizumab compared with standard of care (SOC) in patients with resected stage II or III disease and PD-L1 expression of 1% or greater, and a second trial of adjuvant pembrolizumab compared with placebo in patients with stage IB-III (regardless of tumor proportion score PD-L1 expression), both demonstrated an improvement in DFS. In the neoadjuvant setting, results of a phase III trial of chemotherapy and nivolumab compared with chemotherapy alone revealed an improvement in pathological complete response rate and event-free survival in patients with stage IB-IIIA disease. Finally, for epidermal growth factor receptor (EGFR) mutant NSCLC, a phase III trial of osimertinib compared with SOC revealed an improvement in DFS. The results of these and ongoing trials illustrate the integration of immunotherapy and targeted therapies into the treatment paradigm of patients with surgically resected NSCLC and have led to FDA and EMA approvals in selected populations. Neoadjuvant trials have investigated novel endpoints such as major and complete pathological response, which have the potential to serve as surrogate endpoints for future trials.

Variability in Content of Piperacillin and Tazobactam Injection.

Piperacillin and Tazobactam Injection is a combination product consisting of a penicillin-class antibacterial, piperacillin, and a beta-lactamase inhibitor, tazobactam, indicated for the treatment of patients with moderate to severe infections caused by susceptible isolates of bacteria. In the past decade some quality problems have been noted by the US Food and Drug Administration (FDA) with the Apotex Corp. manufacturing. Intra-lot and inter-lot variability in the spectra of Piperacillin and Tazobactam Injection 3.375 g was detected in the Drug Quality Study (DQS) using Fourier transform near-infrared spectrometry (FTNIR). One vial of 6 (17%) sampled from lot AD103008F3 appeared 14.4 multidimensional SDs from the other vials, suggesting that it represents a different material. Spectra of 132 vials from 19 lots in the spectral library contained 4 vials that were outside the group (21.0 SDs using a subcluster detection test), suggesting that the 4 library vials (3%) also contain differing materials.

Levothyroxine Variations by Process Analytical Technology.

Intra-lot and inter-lot variability in the spectra of levothyroxine was detected in the Drug Quality Study (DQS) using Fourier transform near-infrared spectrometry (FTNIR). Two vials of 12 vials sampled from Athenex lot AFN102 appeared 10.1 and 9.1 SDs from the center of the rest of the vials on the DQS FTNIR screening assay. Spectra of 108 vials from six lots in the library clustered in two groups (p=0.02), suggesting they represent different material.

FTNIR Spectrometry of Micafungin Sodium Quality.

Intra-lot and inter-lot variability in the spectra of micafungin was detected in the Drug Quality Study (DQS) using Fourier transform near-infrared spectrometry (FTNIR). Two vials of 6 vials sampled from Fresenius Kabi lot ACP106 appeared 7.9 and 14.0 standard deviations (SDs) from the center of the rest of the vials on the DQS FTNIR screening assay. Spectra of 48 vials from 7 lots in the library showed 2 outliers at 8.3 and 9.8 SDs from the center of the rest of the library, suggesting they represent different material.

Lack of Content Uniformity in MMR Vaccine.

The measles, mumps, and rubella (MMR) vaccine is a vaccine used to prevent measles, mumps, and rubella (German measles). The vaccine is mandated for children to attend public school in nearly all US states. However, measles cases have been increasing in the past decade, and quality problems have recently been noted by the US Food and Drug Administration (FDA) with the vaccine manufacturer. Intra-lot and inter-lot variability in the spectra of MMR (measles, mumps, and rubella) vaccine was detected in the Drug Quality Study (DQS) using Fourier transform near-infrared spectrometry (FTNIR). Six vials of 12 (50%) sampled from Merck lot U006488 appeared 14.5 SDs from the other vials on a subcluster detection test, suggesting that they represent different material. Spectra of 198 vials from 12 lots in the spectral library contained 140 vials in one tight ellipsoidal group, and 58 vials (30%) were outside that group (39.7 SDs using a subcluster detection test), suggesting that the library vials also contain differing materials.