Blood, Sweat and Smears: "Modified Ham with Dr. Robert Brodsky"

Host: Dr. Brad Lewis, Machaon Diagnostics
Guest: Dr. Robert Brodsky
Date: December 6, 2024
Episode Theme: Introduction and in-depth discussion of the bioluminescent modified HAM test—a novel assay for diagnosing and monitoring complement-mediated thrombotic microangiopathies (TMAs) and related conditions.

Episode Overview

This episode centers on the newly developed bioluminescent modified HAM test, led by Dr. Robert Brodsky’s research group. Dr. Lewis and Dr. Brodsky explore the clinical potential, methodological advancements, and diagnostic/signaling utility of this assay, especially for challenging cases like atypical hemolytic uremic syndrome (aHUS), lupus, CAPS, and pregnancy-related TMAs.

Key Discussion Points & Insights

1. The Need for Improved Complement Diagnostics

• Current Diagnostic Gap: There has long been a lack of reliable, functional tests for diagnosing complement-mediated TMAs, particularly in complex cases such as pregnancy or lupus ([00:31]).
• Problems with Existing Methods: Genetic testing can be inconclusive (variants of unknown significance, or VUS) and slow, while functional assays for specific complement components are limited.

2. The Modified HAM Test: Evolution and Methodology

• Evolution of the Assay:

  • The original modified HAM test, developed about 10 years ago, was technically challenging and not widely adopted ([02:11]).
  • The new bioluminescent version overcomes these obstacles: using continuously glowing cells, no washes/reductions in cell number, and continuous readouts every five minutes for robust, reproducible data.

• Assay Design:

  • Cells with different combinations of complement regulators knocked out serve as “biosensors” for complement activity ([02:11]).

  "We often refer to this assay as kind of like a complement biosensor, depending on the complement regulators we knock out."
  —Dr. Robert Brodsky, [03:08]

3. Practical Applications and Clinical Value

• Diagnostic Inclusion Rather than Exclusion:

  • Could change aHUS diagnosis from being a diagnosis of exclusion to one of inclusion ([04:14]).
  • Enables simultaneous test ordering with other TMA workups (e.g., ADAMTS13).

• Functional Monitoring of Therapy:

  • Demonstrates in vitro susceptibility to complement inhibitors, predicting clinical response ([04:14]).
  • Distinguishes between pathway drivers (classical vs. alternative pathway)—a key insight from recent research ([04:14], [05:52]).

• Monitoring for Therapy Withdrawal/Failure:

  • May stratify risk for relapse after stopping complement inhibitors ([04:14]).

  "It may give insight into who you can discontinue the drug more safely in. It would make me feel a lot more comfortable stopping the drug in someone who doesn't still have a lot of activity..."
  —Dr. Robert Brodsky, [05:40]

• Utility Across Diseases:

  • Can be used in PNH, cold agglutinin disease, various TMAs, and potentially for complement deficiency disorders as well ([04:14], [26:50]).

4. Specific Insights on Disease Settings

a. Post-Transplant TMA ([09:40])

• Mixed Complement Involvement: Not all post-transplant TMAs are complement-driven. The assay is helping to stratify these cases.

b. Technical Distinction from Previous Assays ([10:27])

• Unique Functional Measurement: The assay directly measures complement-mediated cell death, rather than only fluid-phase complement products (like soluble C5b-9/MAC).

  "Soluble C5B9 is only fluid phase. Soluble C5B9 cannot land on a cell and can't insert in the cell and make a membrane attack complex."
  —Dr. Robert Brodsky, [11:18]

c. Specificity and False Positives ([13:55], [14:33])

• About 10% of healthy controls may test positive, likely reflecting underlying biology (lower disease threshold with strong triggers, multi-hit hypothesis), not true false positives.

  "They're as good as the clinical scenario that they're tested in. If the pretest probability is high ... and they're positive and it's blocked by a C5 inhibitor, that patient's going to respond."
  —Dr. Robert Brodsky, [15:18]

d. Complex Presentations: Lupus, CAPS, Pregnancy ([16:20])

• Lupus: Most lupus patients without TMA are negative; data on lupus with TMA still emerging.

• CAPS: Frequently positive, research ongoing on a simpler flow-cytometry adaptation for risk stratification ([19:59]).

• Pregnancy: Late 2nd/3rd trimester rises in autoreactive IgM may drive complement activity, explaining susceptibility to TMA/HELLP syndrome and PNH/CAPS exacerbation in pregnancy ([20:24]).

  "The autoreactive IgM increases in the third trimester of pregnancy and it goes back to normal shortly after delivery."
  —Dr. Robert Brodsky, [22:04]

e. Resolving Variants of Unknown Significance ([19:01])

• Isolating alternative pathway in the assay can clarify the functional impact of rare genetic variants.

f. Rare Renal Diseases and Complement Deficiency

• Potential to assess cases like C3 glomerulopathy (C3G) and functional impact of mutations linked to infections ([26:14], [26:50]).

Notable Quotes & Memorable Moments

• Dr. Brodsky on Paradigm Shift in Disease Mechanisms ([18:04]):

  "The whole point of the assay was really to show that what we thought was the pathophysiology of this disease for 50 years was only partly right. It was actually mostly wrong."

• Dr. Lewis on Utility in Clinical Decision-Making ([17:42]):

  "With the positive assay, it does give me the confidence to make the decision to start therapy."

• On New Research Directions ([25:37]):

  "40% of patients with macular degeneration have variants in factor VIII. Are they functional? We can answer that question now..." —Dr. Robert Brodsky

Important Timestamps

| Timestamp | Topic | |---------------|-----------| | 00:31 | Dr. Lewis introduces the topic and the clinical need for better complement testing | | 02:11 | Dr. Brodsky explains the evolution and methodology of the bioluminescent modified HAM assay | | 04:14 | Clinical applications: diagnosis, monitoring, and pathway distinction | | 09:40 | Post-transplant TMA relevance | | 10:27 | Differentiating this assay from previous complement activity tests | | 16:20 | Handling complex cases: lupus, HLH, CAPS, pregnancy | | 19:01 | Utility in resolving variants of unknown significance (VUS) | | 20:24 | Immunological changes in pregnancy explained via the assay | | 25:37 | Broader research implications, macular degeneration, nephrology | | 26:14 | C3G and complement deficiency states | | 27:17 | Final thoughts and gratitude |

Conclusion

This episode offered an engaging expert-to-expert deep dive into a transformative new assay for the diagnosis and management of complement-mediated diseases. Dr. Brodsky’s bioluminescent modified HAM test stands out for its functional, pathway-specific readouts and its potential to dramatically improve care in TMA, rare renal disease, complement deficiency, and scenarios with ambiguous genetics or clinical manifestations. Both Dr. Lewis and Dr. Brodsky anticipate a major clinical and research impact as this test becomes more widely available.

For a detailed understanding of complement-mediated disorders or information on the modified HAM test's availability, listeners are encouraged to contact Machaon Diagnostics.