Quantitative Analysis of Drug Tablet Aging with Fast Hyper-Spectral Stimulated Raman Scattering Microscopy: An Interview with 2024 Craver Award Winner Conor L. Evans

Conor L. Evans of the Wellman Center for Photomedicine at Massachusetts General Hospital (Charlestown, Massachusetts) and associates, seeing a need for a new method of chemical imaging that delivers a quick acquisition time, low background, high resolution, and high chemical sensitivity recently published a study demonstrating sparse spectral sampling stimulated Raman scattering (S⁴RS). This new method is an extension of stimulated Raman scattering (SRS) microscopy that is particularly well suited for the imaging and quantification of active pharmaceutical ingredients (APIs). Evans will be receiving the Clara Carver Award, created by The Coblentz Society to recognize young individuals who have made significant contributions in applied analytical vibrational spectroscopy, at this year’s SciX conference, to be held from October 20 through October 25, in Raleigh, North Carolina. As part of an ongoing series of this year’s SciX conference honorees, Conor spoke to us about the motivation behind this study, his findings, and the next steps in its development.

In your paper (1), you discussed how chemical imaging (CI) has emerged as a crucial method for understanding the chemical and physical properties of pharmaceutical tablets. But you mention there are some gaps between what CI can deliver and what analysts need. Briefly discuss these.

Chemical imaging techniques are extraordinarily powerful tools in multiple areas of biomedicine, allowing for detailed characterization of molecular identity and spatial distribution. When applied to drugs and drug formulations, there are several scenarios where chemical imaging has significant value as a process analytical technology (PAT). One area is early in the formulation process where a drug is being combined with inactive ingredients, known as excipients, to create the formulation. For topical products such as those we put on the skin, these can be combinations of different creams, emollients, or solutions. For solid, oral tablets, the application is in understanding how and where the given drug is packaged with excipients to give the desired release kinetics. Tablet formulation can be a complicated process, involving solid or liquid processing, granulation steps where drugs and excipients are combined into granules, and then the actual mixing and compression of the tablets itself. Chemical imaging could provide chemical and spatial information on each step of this formulation process and connect what occurs at each of those steps to the performance of the final tablet.

A related, but important second application, is when tablets are made in different batches. Batch effects can also happen during drug development, such as when an initial batch of tablets are given an early phase clinical study, and then scaled up in later phase clinical studies. Chemical imaging can look at the microscale composition of the tablets created in these individual batches for comparison, which can be important to ensure that the manufactured tablets are consistent and have controlled drug release profiles.

Another area where chemical imaging has considerable potential, but I do not think it has reached yet, is in routine quality control. This would be a scenario where tablets are pulled off the manufacturing line and routinely analyzed to ensure that there’s consistency during the manufacturing process. This would be ideal to do, but current methodologies either do not meet quality control needs or are too time-consuming to be used in such a process. This has led other tools to be used during the manufacturing steps that can give indirect insight into composition, but do not offer the type of direct chemical mapping information that comes from chemical imaging.

To fill these gaps, you and your co-authors demonstrate fast chemical imaging by epi-detected sparse spectral sampling stimulated Raman scattering (S⁴RS), an extension of stimulated Raman scattering (SRS) microscopy, to quantify active pharmaceutical ingredients (API) and excipient degradation and distribution. What inspired you to choose this Raman technique?

We were approached by a company called UNION therapeutics who were facing several interesting challenges and their drug development pipeline. One of the things I really enjoy doing is speaking to scientists, clinicians, product managers—essentially end users—who are facing key challenges because it gives us the opportunity to think deeply about how we can innovate and how we can address issues outside of our immediate research domains. In speaking with folks from UNION, Michelle Wei, the work’s lead author, and I learned about difficulties in assessing the distribution of drugs with tablets, how challenging it is to capture quantitative information about how drugs are associated with different excipients, and how these factors can change during manufacturing and scale up. From these conversations, we learned that UNION needed the ability to do two things very well: 1) map the chemical distribution of compounds in their tablets with high resolution and 2) carry out mapping at a relatively rapid rate. Based on these two needs, we decided to develop and apply our S⁴RS technique, was an excellent fit for the problem at hand.

How much faster is the S⁴RS technique as compared to near-infrared microscopy, confocal Raman spectroscopy, and other imaging techniques?

There are three major chemical imaging technologies used today for the analysis of solid, oral tablets: near-infrared, confocal Raman, and laser direct infrared imaging (LDIR). Near-infrared imaging technologies are quite mature, and are rapid, offering tablet imaging in a period of minutes. The challenge is that the near-infrared wavelengths of light used result in poor optical imaging resolutions. This can be a problem when imaging the grains within some tablets, as they can be a micron or smaller and essentially are blurred or missed when using near-infrared imaging. LDIR offers a speed improvement over standard near-infrared microscopy, but similarly suffers from poor optical imaging resolution.

Raman imaging, confocal Raman in particular, offers excellent optical spatial resolutions but comes with two drawbacks. The first, largest drawback, is that spontaneous Raman is a weak process, meaning that the signal that one gets in a Raman microscope is small. To obtain high-quality spectral imaging, one needs to integrate at each pixel for a long period of time, which leads to slow imaging speeds. This means that imaging the face of a tablet can take hours or days, which is fine to do once and a while, but does not lend itself readily routine imaging or use as a quality control methodology. Raman spectroscopy can also suffer from background, such as auto fluorescence, which can be challenging, as several commonly used excipients are autofluorescent.
Stimulated Raman scattering (SRS) can overcome many of these challenges as it provides both rapid imaging speeds with high spatial resolutions. The key barrier to using standard SRS tools is that current SRS systems either can collect spectral information over narrow windows of the Raman spectrum or require long times to tune over the Raman spectral regions of interest. The sparse spectral sampling method S⁴RS was developed to address these key limitations, allowing for hyperspectral imaging across the entire Raman spectrum by only visiting the vibrational bands necessary. This allows for rapid imaging with chemical information at high spatial resolutions.

Other than the Raman technique being used, does your work differ from what has been previously done by yourself or others?

My team is certainly not the first to use SRS to image tablets. There has been excellent work by others in the field before us, including Ji-Xin Cheng’s and Dan Fu’s teams (2-4). These prior works really helped us identify some of the key challenges and opportunities and applying advanced Raman technologies for solid oral tablet studies. The innovation introduced by our recent work was in the application of S⁴RS as a means of rapidly capturing and mapping chemical information within tablets. I should also note that Michelle Wei also developed a method for cutting the tablets such that their cores could be imaged; this was essential for mapping the spatial changes in tablet composition due to different storage conditions.

Briefly state your findings.

The study was expertly led by Michelle Wei, an MD, PhD student in my team who I mentioned earlier. Using an advanced, rapidly tunable fiber laser, she was able to acquire hyperspectral SRS data from samples of the drugs and excipients used in the manufacturing of a solid oral tablet. Using these spectra as a library, we were able to determine a set of sparse Raman bands that could be sampled to obtain equivalent spectral SRS information, but at a much faster rate. Michelle was able to show that S⁴RS images acquired from tablets were essentially indistinguishable from continuous hyperspectral SRS images, proving the merit of this approach for tablet characterization.

Importantly, we studied whether our approach could detect changes in tablet structure and composition that were the result of sample handling conditions such as heat and humidity exposure. This was important for two main reasons. First, heating of tablets is generally a method used and what is called artificial aging, or accelerated aging, where tablets’ stability is examined at higher temperatures to simulate what occurs during storage. Second, it is important to understand how and if tablets are stable in the kind of conditions a patient might encounter at home. Michelle found that there were significant changes in tablet structure due to humidity that could be quantitatively gleaned from the imaging data. More surprisingly, she found that higher heat levels led to chemical composition changes of one of the excipients, lactose. Interestingly, lactose when stored on its own under the same condition did not experience this compositional change, indicating that it was something that specifically happened through chemical interactions with the compressed tablet. This is the type of information that is needed to understand the stability of tablet formulations for both safety and efficacy.

Do your findings correlate with what you had hypothesized?

Overall, yes, we determined that the sparse S⁴RS method was capable of rapid imaging and can be particularly capable for PAT applications.

Was there anything particularly unexpected that stands out from your perspective? Were there any chemicals or specific properties that were easier to detect or analyze as opposed to others?

The lactose decomposition finding was a surprise because Michelle had painstakingly tested the heating and humidity conditions on the individual components of the tablet but did not find anything altered under those conditions. We hypothesize that there must be a specific chemical interaction occurring with the compressed tablet, likely between lactose and magnesium stearate, that led to the specific structural and chemical changes found. Also, while the heat and humidity levels used for these parts of this work are not the kind we typically have in Boston, they are not uncommon in certain parts of the world and may indeed impact the shelf life and storage of tablets for many patients.

Were there any limitations or challenges you encountered in your work?

One potential limitation, at least for short term implementation of S⁴RS, is that results likely should be confirmed by another method such as Raman, NMR, or mass spectroscopy. This is because S⁴RS is still relatively new, and it would be important to ensure that results are properly benchmarked against a standardized method.

The S⁴RS approach uses a spectral decomposition method, which requires either a priori knowledge of the drug/excipient spectra or experimental acquisition of the spectra. This is usually not a challenge when working in a drug formulation context. Finally, like all vibrational imaging methodologies, it’s very helpful for a given drug or excipient to have a strong or distinct vibrational spectrum. The more unique, or orthogonal, the vibrational spectrum of the drugs and excipients, the fewer sparse spectral bands that are necessary, and the faster imaging can be.

What best practices can you recommend in this type of analysis for both instrument parameters and data analysis?

I believe we’re still very early in the phase of this research in terms of how it might be done on other instruments, but I think there’s a few things that could be helpful for other researchers. One aspect we found helpful for S⁴RS is having a rapidly tuning, narrowband picosecond light source. We have been extraordinarily fortunate to work closely with Refined Laser Systems (RLS), the developer of the rapid tuning fiber OPO that is at the heart of our S⁴RS system. This laser has unique capabilities including the ability to rapidly tune anywhere in the Raman spectrum, which was critical to make this S⁴RS study successful. The narrowband nature of the source was also extremely helpful to be able to have high vibrational spectrum resolution to resolve the individual chemical components of the tablets. The other part of this study that was very important, and was a key innovation from Michelle, was the ability to carefully section the tablets to expose the inner tablet across an extremely flat face.

Can you please summarize the feedback that you have received from others regarding this work?

The feedback from this work has been very interesting. On one side, we have received considerably positive feedback from contract manufacturing companies who create, and manufacture, oral tablets. There has been interest in our technology and companies are excited to work with us to further develop this technology for their specific needs. This really made us excited because it’s an opportunity to work directly with the individuals and the scientists who would most benefit from this technology in the short term.

We’ve also received helpful critical feedback from folks in the pharmaceutical industry who have tried to implement tools in PAT. This feedback was extremely useful, because a lot of the work that has been done in PAT has been done behind closed doors at companies and not been published or made public. As such, there is a huge body of work that is essentially proprietary to individual companies and not widely known. Being able to talk to people in the community who have worked in this area, have hit major barriers in implementing PAT, and have been frustrated by these limitations has been extremely eye-opening. These conversations have helped to focus our next steps on key areas where we hope to have the greatest impact.

Might these techniques work with other pharmaceutical formats, such as pastes, capsules, patches, or liquids, or for other pharma products which are not taken orally?

Absolutely, yes—S⁴RS can serve as a PAT for a host of different pharmaceutical, and cosmeceutical formats. There has been some excellent work by my colleagues, such as Michael Roberts, who have used Raman imaging to look at the spatial distributions of drugs and excipients in topical product formulations. We believe that tools like S⁴RS, which offer improved speed over spontaneous Raman, can offer new advantages for pastes, capsules, patches, and topical product formulations.

What are the next steps in this research?

One of the exciting things about working at the Wellman Center for Photomedicine has been the laser focus on translational research. I have had the great pleasure of working with scientists such as Dr. Gabriela Apiou, the Director of Strategic Alliances at the MGH Research Institute, to assess our technologies, understand their limitations and opportunities, and then plan forward looking strategies to engage with partners.

For this technology, we have recently joined forces with Intek Scientific, a startup company here in Massachusetts, that will be working closely with us over the next several years to develop S⁴RS as a PAT. Intek Scientific is part of Intek Plus, an innovative South Korean company with significant capabilities in automation and image analysis. We are looking forward to working with our partners at Intek Scientific and Intek Plus to further develop S⁴RS as a PAT.

What are your thoughts regarding your being named the recipient of the Clara Craver Award?

I am extremely honored to have been selected to receive the Craver Award. Dr. Craver had an enormous impact to our field, one that is still being felt today. To be selected for this award named in her honor is humbling, as is being counted amongst the notable scientists who have been named the recipient for this award in years past.

References

(1) Wei, Y.; Pence, I. J.; Wiatrowski, A.; Slade, J. B.; Evans, C. L. Quantitative Analysis of Drug Tablet Aging by Fast Hyper-Spectral Stimulated Raman Scattering Microscopy. Analyst 2024, 149, 1436-1446. DOI: 10.1039/D3AN01527K

(2) Slipchenko, M. N.; Chen, H.; Ely, D.R.; Jung, Y.; Carvajal, M. T.; Chen, J. X. Vibrational Imaging of Tablets by Epi-Detected Stimulated Raman Scattering Microscopy. Analyst 2010, 135 (10), 2613-2619. DOI: 10.1039/C0AN00252F

(3) Francis, A. T.; Nguyen, T. T.; Lamm, M. S.; Teller, R.; Forster, S. P.; Xu, W.; Rhodes, T.; Smith, R. L.; Kuiper, J.; Su, Y.; Fu, D. In situ Stimulated Raman Scattering (SRS) Microscopy Study of the Dissolution of Sustained-Release Implant Formulation. Mol. Pharmaceutics 2018, 15 (12), 5793-5801. DOI: 10.1021/acs.molpharmaceut.8b00965

(4) Figueroa, B.; Nguyen, T.; Sotthivirat, S.; Xu, W.; Rhodes, T.; Lamm, M. S.; Smith, R. L.; John, C. T.; Su, Y.; Fu, D. Detecting and Quantifying Microscale Chemical Reactions in Pharmaceutical Tablets by Stimulated Raman Scattering Microscopy. Anal. Chem. 2019, 96 (10), 6894-6901. DOI: 10.1021/acs.analchem.9b01269

Conor L. Evans is an Associate Professor at the Harvard Medical School, an Affiliated Faculty member of the Harvard University Biophysics Program, a faculty member of the Laser Biomedical Research Center, and leads his lab at the Wellman Center for Photomedicine at Massachusetts General Hospital. Dr. Evans lab’s research is focused on the development and clinical translation of optical microscopy and spectroscopy tools. A recipient of the NIH Director’s New Innovator Award, his efforts have resulted in the creation of new technologies currently in multiple clinical trials. He is a Royce Fellow of Brown University, a Fellow of the SPIE, and has been honored with several awards, including the Goldwater Scholarship, NASA Space Grants, and the ASP New Investigator Award. Dr. Evans is an editor for Scientific Reports, academic editor for PLOS One, and has written more than 100 peer-reviewed publications. He holds 14 patents and patent applications and works to translate his technologies to the clinic. He additionally serves the Wellman Center as Faculty Liaison to the Department of Defense.